Egf variants and methods of use thereof

Abstract

Described herein are compositions and methods for selective depletion of EGFR target molecules using a recyclable receptor-binding mediated complex to elicit uptake or endocytosis and cellular degradation of the target molecule. Exemplary compositions containing a peptide, such as a CDP peptide that binds a transferrin receptor, can be linked to a peptide that binds an EGFR target molecule. Such compositions can be used to selectively deplete EGFR from the cell surface or selectively recruit EGFR to endosomes via transferrin receptor-mediated endocytosis of the composition and the bound target molecule. Once inside the endosome, the acidic pH can lead to release of the EGFR from the composition due to pH-dependent binding of the composition for the target molecule, and the transferrin receptor portion is recycled back to the cell surface for "reloading". The EGFR can then be trafficked into lysosomes wherein it is degraded.

Description

EGF VARIANTS AND METHODS OF USE THEREOFCROSS-REFERENCE

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 470,404, entitled “EGF VARIANTS AND METHODS OF USE THEREOF,” filed June 1, 2023, which is incorporated herein by reference in its entirety.SEQUENCE LISTING

[0002] The instant application contains a Sequence Listing which has been submitted electronically in extensible Markup Language (XML) format and is hereby incorporated by reference in its entirety. Said XML copy, created on May 22, 2024, is named “438542- 725021_SL. xml” and is 757,899 bytes in size.BACKGROUND

[0003] Accumulation or over-expression of soluble and cell surface proteins is indicated in a variety of human diseases, ranging from neurodegenerative diseases to cancer. Furthermore, numerous diseases are associated with mutations in soluble or cell surface proteins resulting in constitutive activity, resistance to treatment, or dominant negative activity. However, many of these proteins have been deemed “undruggable,” “difficult to drug,” or “yet to be drugged” targets due to challenges in targeting them with small molecule therapeutics. For example, the oncoprotein EGFR drives cell growth through both scaffolding and kinase functions, and current therapeutic modalities typically address one but not the other, leaving cancer cells prone to continued signaling through mutation or other adaptations. There is a need for compositions and methods to target and selectively deplete soluble and cell surface proteins associated with disease. In addition, the oncoprotein EGFR drives cell growth through both scaffolding and kinase functions, and current therapeutic modalities typically address one but not the other, leaving cancer cells prone to continued signaling through mutation or other adaptations.Elimination of the entire EGFR protein would simultaneously disrupt both of its growth-driving functions. There is therefore a need for compositions and methods to target and selectively deplete EGFR associated with cancer, which can also be accomplished by engineering of EGF itself.SUMMARY

[0004] In various aspects, the present disclosure provides an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof,wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R.

[0005] In some aspects, the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof. In some aspects, the sequence further comprises one or more mutations relative to SEQ ID NO: 317 comprising DI 1R, I23S, V35E, E51P, L52E, R53E, or a combination thereof. In some aspects, the sequence further comprises one or more mutations relative to SEQ ID NO: 317 comprising E51H, L52H, R53H, or a combination thereof. In some aspects, the EGFR-binding peptide comprises a sequence having at least 80%, at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 458 - SEQ ID NO: 493 or a fragment thereof. In some aspects, the sequence comprises SEQ ID NO: 494. In some aspects, the sequence comprises any one of SEQ ID NO: 458 - SEQ ID NO: 493. In some aspects, the fragment comprises at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, or at least 50 amino acid residues.

[0006] In some aspects, the EGFR-binding peptide is capable of binding to EGFR without activating the EGFR. In some aspects, the EGFR-binding peptide blocks binding of EGF to EGFR when the EGFR-binding peptide is bound to the EGFR. In some aspects, the EGFR- binding peptide inhibits EGFR when the EGFR-binding peptide is bound to the EGFR. In some aspects, the EGFR-binding peptide prevents dimerization of EGFR when the EGFR-binding peptide is bound to the EGFR.

[0007] In various aspects, the present disclosure provides a peptide complex comprising: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-bindingpeptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR-binding peptide has affinity for a target molecule.

[0008] In some aspects, the EGFR-binding peptide is the EGFR-binding peptide as described herein. In some aspects, the affinity of the EGFR-binding peptide for the target molecule, the affinity of the cellular receptor binding peptide for the cellular receptor, or both is pH- independent. In some aspects, the affinity of the EGFR-binding peptide for the target molecule, the affinity of the cellular receptor binding peptide for the cellular receptor, or both is pH dependent. In some aspects, the affinity of the EGFR-binding peptide for the target molecule, the affinity of the cellular receptor-binding peptide for the cellular receptor, or both is ionic strength dependent.

[0009] In some aspects, the cellular receptor-binding peptide is a transferrin receptor-binding peptide or a PD-L1 -binding peptide. In some aspects, the cellular receptor is a transferrin receptor or PD-L1. In some aspects, the cellular receptor is a cation-independent mannose 6 phosphate receptor (CI-M6PR), an asialoglycoprotein receptor (ASGPR), CXCR7, folate receptor, or Fc receptor (including but not limited to neonatal Fc receptor (FcRn) or FcyRIIb).

[0010] In some aspects, the cellular receptor-binding peptide binds to the cellular receptor at a pH of from pH 4.5 to pH 7.4, from pH 5.5 to pH 7.4, from pH 5.8 to pH 7.4, or from pH 6.5 to pH 7.4. In some aspects, the cellular receptor-binding peptide is capable of binding the cellular receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no morethan 0.1 nM at pH 7.4. In some aspects, the cellular receptor-binding peptide is capable of binding the cellular receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 5.5. In some aspects, the cellular receptor-binding peptide is capable of binding the cellular receptor with a dissociation rate constant (koff or kd) of no more than 1 s'1, no more than 5x10'1s'1, no more than 2x10'1s'1, no more than 1x1 O'2s'1, no more than 5x1 O'2s'1, no more than 2x1 O'2s'1, no more than 1x1 O'2s'1, no more than 5x1 O'3s'1, no more than 2x1 O'3s'1, no more than 1x1 O'3s'1, no more than 5x1 O'4s'1, or no more than 2x1 O'4s'1at pH 5.5.

[0011] In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is pH-independent. In some aspects, the affinity of the EGFR-binding peptide for the target molecule is pH-dependent. In some aspects, the affinity of the EGFR-binding peptide for the target molecule is pH-independent. In some aspects, the affinity of the cellular receptorbinding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5 -fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some aspects, the dissociation rate constant (koff or kd) of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some aspects, the dissociation rate constant (koff or kd) of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

[0012] In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is pH dependent. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor decreases as pH decreases. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is higher at pH 7.4 than at pH 5.5. In someaspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is higher at pH 7.4 than at pH 5.8.

[0013] In some aspects, the affinity of the EGFR-binding peptide for the target molecule is pH dependent. In some aspects, the affinity of the EGFR-binding peptide for the target molecule decreases as pH decreases. In some aspects, the affinity of the EGFR-binding peptide for the target molecule is higher at a higher pH than at a lower pH. In some aspects, the higher pH is pH 7.4, pH 7.2, pH 7.0, or pH 6.8. In some aspects, the lower pH is pH 6.5, pH 6.0, pH 5.8, pH 5.5, pH 5.0, or pH 4.5. In some aspects, the affinity of the EGFR-binding peptide for the target molecule is higher at pH 7.4 than at pH 6.0. In some aspects, the affinity of the EGFR-binding peptide for the target molecule is higher at pH 7.4 than at pH 5.5. In some aspects, the affinity of the EGFR-binding peptide for the target molecule is higher at pH 7.4 than at pH 5.8.

[0014] In some aspects, the EGFR-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no more than 500 nM, no more than 200 nM, 100 nM, no more than 50 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, no more than 1 nM, or no more than 0.1 nM at pH 7.4.

[0015] In some aspects, the EGFR-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1x10'1s’1, 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, no more than 2x1 O’4s’1, no more than 1x1 O’4s’1, no more than 5xl0’5s’1, or no more than 2xl0’5s’1at pH 7.4. In some aspects, the EGFR-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 5.5. In some aspects, the EGFR-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x10’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 5.8.

[0016] In some aspects, the dissociation rate constant (koff or kd) for EGFR-binding peptide binding the target molecule is at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.5 than at pH 7.4. In some aspects, the dissociation rate constant (koff or kd) for EGFR-binding peptide binding the target molecule is at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.8 than at pH 7.4.

[0017] In some aspects, the EGFR-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.5. In some aspects, the EGFR-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.8.

[0018] In some aspects, the affinity of the EGFR-binding peptide for the target molecule at pH 7.4 is at least 1.5-fold, 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the EGFR-binding peptide for the target molecule at pH 5.5. In some aspects, the affinity of the EGFR-binding peptide for the target molecule at pH 7.4 is at least 1.5-fold, at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7- fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the EGFR-binding peptide for the target molecule at pH 5.8.

[0019] In some aspects, the affinity of the EGFR-binding peptide for the target molecule at pH 7.4 is less than 0.5-fold, less than 1-fold, less than, 1.5-fold, less than 2-fold, less than 3-fold, or less than 10-fold, greater than the affinity of the EGFR-binding peptide for the target molecule at pH 5.8.

[0020] In some aspects, the EGFR-binding peptide comprises one or more histidine amino acid residues. In some aspects, the affinity of the EGFR-binding peptide for the target molecule decreases as ionic strength increases. In some aspects, the EGFR-binding peptide comprises one or more polar or charged amino acid residues capable of forming polar or charge-charge interactions with the target molecule.

[0021] In some aspects, the cellular receptor-binding peptide is fused to, linked to, complexed with, or conjugated to the EGFR-binding peptide. In some aspects, the cellular receptor-binding peptide is fused to, linked to, complexed with, or conjugated to the EGFR-binding peptide via a polymer linker. In some aspects, the polymer linker is a polyethylene glycol (PEG), a hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a zwitterionic water soluble polymer, a water soluble poly(amino acid), a water soluble polymer comprising proline, alanine, serine, or a combination thereof, a water soluble polymer containing glycine, glutamic acid, and serine, an Fc region, a fatty acid, a palmitic acid, an albumin, or an albumin binding molecule.

[0022] In some aspects, the cellular receptor-binding peptide and the EGFR-binding peptide form a single polypeptide chain. In some aspects, the peptide complex comprises a dimer dimerized via a dimerization domain. In some aspects, a distance between the cellular receptorbinding peptide and the EGFR-binding peptide is at least 1 nm, at least 2 nm, at least 5 nm, at least 10 nm, at least 20 nm, at least 50 nm, or at least 100 nm. In some aspects, the dimerization domain comprises an Fc domain. In some aspects, the dimer is a homodimer dimerized via a homodimerization domain. In some aspects, the homodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 535, SEQ ID NO: 706, or SEQ ID NO: 246. In some aspects, the dimer is a heterodimer dimerized via a first heterodimerization domain and a second heterodimerization domain. In some aspects, the first heterodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 536, SEQ ID NO: 707, or SEQ ID NO: 709. In some aspects, the second hetero dimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 537, SEQ ID NO: 708, or SEQ ID NO: 710.

[0023] In some aspects, the EGFR-binding peptide is linked to the dimerization domain via a peptide linker. In some aspects, the cellular receptor-binding peptide is linked to the dimerization domain via a peptide linker. In some aspects, the cellular receptor-binding peptide is linked to the EGFR-binding peptide via a peptide linker. In some aspects, the peptide linker has a length of from 1 to 50 amino acid residues, from 2 to 40 amino acid residues, from 3 to 20 amino acid residues, or from 3 to 10 amino acid residues. In some aspects, the peptide linker comprises glycine and serine amino acids. In some aspects, the peptide linker has a persistencelength of no more than 6 A, no more than 8 A, no more than 10 A, no more than 12 A, no more than 15 A, no more than 20 A, no more than 25 A, no more than 30 A, no more than 40 A, no more than 50 A, no more than 75 A, no more than 100 A, no more than 150 A, no more than 200 A, no more than 250 A, or no more than 300 A. In some aspects, the peptide linker is derived from an immunoglobulin peptide. In some aspects, the peptide linker is derived from a doubleknot toxin peptide. In some aspects, the peptide linker comprises a sequence of any one of SEQ ID NO: 129 - SEQ ID NO: 141, SEQ ID NO: 195 - SEQ ID NO: 218, SEQ ID NO: 223 - SEQ ID NO: 223 - SEQ ID NO: 227, SEQ ID NO: 194, SEQ ID NO: 391, SEQ ID NO: 538, or SEQ ID NO: 540 - SEQ ID NO: 541.

[0024] In some aspects, the cellular receptor-binding peptide, the EGFR-binding peptide, peptide complex, or a combination thereof comprises a miniprotein, a nanobody, an antibody, an antibody fragment, an scFv, a DARPin, or an affibody. In some aspects, the antibody comprises an IgG, or wherein the antibody fragment comprises a Fab, a F(ab)2, an scFv, or an (scFv)2. In some aspects, the miniprotein comprises a cystine-dense peptide, an affitin, an adnectin, an avimer, a Kunitz domain, a nanofittin, a fynomer, a bicyclic peptide, a beta-hairpin, or a stapled peptide. In some aspects, the cellular receptor-binding peptide comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds. In some aspects, the EGFR-binding peptide comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds. In some aspects, the peptide complex comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds. In some aspects, the cellular receptorbinding peptide comprises at least six cysteine residues. In some aspects, the at least six cysteine residues are positioned at amino acid positions 4, 8, 18, 32, 42, and 46 of the cellular receptorbinding peptide. In some aspects, the at least six cysteine residues form at least three disulfide bonds.

[0025] In some aspects, the cellular receptor-binding peptide comprises a sequence of any one of SEQ ID NO: 148 - SEQ ID NO: 177. In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with afragment of any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64.

[0026] In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 96, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 96. In some aspects, the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 96.

[0027] In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 66, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 66. In some aspects, the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 66.

[0028] In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 65, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 65. In some aspects, the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 65.

[0029] In some aspects, the cellular receptor-binding peptide comprises a sequence of any one of SEQ ID NO: 392 - SEQ ID NO: 399. In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241.

[0030] In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 187, SEQ ID NO: 235, SEQ ID NO: 236, SEQ ID NO: 238, SEQ ID NO: 239, SEQ ID NO: 400, or SEQ ID NO: 401 or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ IDNO: 187. In some aspects, the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 187.

[0031] In some aspects, the fragment comprises at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36, at least 37, at least 38, at least 39, at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, or at least 50 amino acid residues.

[0032] In some aspects, the cellular receptor-binding peptide comprises one or more histidine residues at a cellular receptor-binding interface. In some aspects, the EGFR-binding peptide comprises one or more histidine residues at a EGFR-binding interface. In some aspects, the target molecule comprises an EGFR. In some aspects, the EGFR is wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximab-resistant EGFR, a panitumumab-resistant EGFR, or a combination thereof. In some aspects, the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

[0033] In some aspects, an off rate of the cellular receptor-binding peptide from the cellular receptor is slower than a recycling rate of the cellular receptor. In some aspects, a half-life of dissociation of the cellular receptor-binding peptide from the cellular receptor is no faster than 1 minute, no faster than 2 minutes, no faster than 3 minutes, no faster than 4 minutes, no faster than 5 minutes, no faster than 7 minutes, no faster than 10 minutes, no faster than 15 minutes, no faster than 20 minutes, no faster than 30 minutes, no faster than 45 minutes, no faster than 60 minutes, no faster than 90 minutes, or no faster than 120 minutes. In some aspects, a rate of dissociation of the EGFR-binding peptide from the target molecule is faster than a recycling rate of the cellular receptor. In some aspects, a half-life of dissociation of the target binding-binding peptide from the target molecule is less than 10 seconds, less than 20 seconds, less than 30 seconds, less than 1 minute, less than 2 minutes, less than 5 minutes, less than 10 minutes, lessthan 20 minutes, less than 30 minutes, less than 45 minutes, or less than 60 minutes in endosomal conditions.

[0034] In some aspects, the peptide complex is capable of being endocytosed via receptor- mediated endocytosis. In some aspects, the receptor-mediated endocytosis is transferrin receptor-mediated endocytosis. In some aspects, the receptor-mediated endocytosis is PD-L1- mediated endocytosis. In some aspects, the cellular receptor-binding peptide remains bound to the cellular receptor inside an endocytic vesicle. In some aspects, the peptide complex is recycled to the cell surface when the cellular receptor-binding peptide is bound to the cellular receptor and the cellular receptor is recycled. In some aspects, the target molecule is released or dissociated from the EGFR-binding peptide after the peptide complex is endocytosed via receptor-mediated endocytosis.

[0035] In some aspects, the target molecule is an extracellular protein, a circulating protein, or a soluble protein. In some aspects, the target molecule is a cell surface protein. In some aspects, the target molecule is a transmembrane protein.

[0036] In some aspects, the peptide complex further comprises a half-life modifying agent coupled to the cellular receptor-binding peptide, the EGFR-binding peptide, or both. In some aspects, the half-life modifying agent is a polymer, a polyethylene glycol (PEG), a hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a zwitterionic water soluble polymer, a water soluble poly(amino acid), a water soluble polymer of proline, alanine and serine, a water soluble polymer containing glycine, glutamic acid, and serine, an Fc region, a fatty acid, palmitic acid, an albumin, or a molecule that binds to albumin. In some aspects, the molecule that binds to albumin is a serum albumin-binding peptide. In some aspects, the serum albumin-binding peptide comprises a sequence of any one of SEQ ID NO: 178, SEQ ID NO: 179, or SEQ ID NO: 193.

[0037] In some aspects, the cellular receptor-binding peptide, the EGFR-binding peptide, or both is recombinantly expressed. In some aspects, the EGFR-binding peptide is configured to dissociate from the target molecule at pH 6.5, pH 6.0, pH 5.8, pH 5.5, pH 5.0, or pH 4.5. In some aspects, the cellular receptor-binding peptide is configured to dissociate from the cellular receptor at pH 6.5, pH 6.0, pH 5.5, pH 5.0, or pH 4.5.

[0038] In some aspects, the peptide complex comprises a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 499 - SEQ ID NO: 501, SEQ ID NO: 504, SEQ ID NO: 505, SEQ ID NO: 507, SEQ ID NO: 508, or SEQ ID NO: 511 -SEQ ID NO: 526. In some aspects, the peptide complex comprises a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 495 - SEQ ID NO: 497, SEQ ID NO: 503, or SEQ ID NO: 506.

[0039] In various aspects, the present disclosure provides a peptide-active agent complex comprising a peptide complexed to an active agent, wherein the peptide comprises the EGFR- binding peptide as described herein or the peptide complex as described herein.

[0040] In some aspects, the active agent comprises a peptide, a peptidomimetic, an oligonucleotide, a DNA, an RNA, an antibody, a single chain variable fragment (scFv), an antibody fragment, an aptamer, or a small molecule. In some aspects, the DNA comprises cDNA, ssDNA, or dsDNA. In some aspects, the RNA comprises RNAi, microRNA, snRNA, dsRNA, or an antisense oligonucleotide. In some aspects, the active agent is a therapeutic agent or a detectable agent. In some aspects, the detectable agent comprises a dye, a fluorophore, a fluorescent biotin compound, a luminescent compound, a chemiluminescent compound, a radioisotope, nanopartide, a paramagnetic metal ion, or a combination thereof. In some aspects, the therapeutic agent comprises a chemical agent, a small molecule, a therapeutic, a drug, a peptide, an antibody protein, any fragment thereof, or any combination thereof.

[0041] In some aspects, the therapeutic agent comprises an oncology agent, an autoimmune disease agent, an acute and chronic neurodegeneration agent, a pain management agent, or an anti-cancer agent. In some aspects, the anti-cancer agent comprises a radionuclide, radioisotope, a chemotherapeutic agent, a platinum therapeutic, a toxin, an enzyme, a sensitizing drug, an anti- angiogenic agent, cisplatin, an anti-metabolite, an anti-metabolic therapeutic, a mitotic inhibitor, a growth factor inhibitor, paclitaxel, temozolomide, topotecan, fluorouracil, vincristine, vinblastine, procarbazine, decarbazine, altretamine, methotrexate, mercaptopurine, thioguanine, fludarabine phosphate, cladribine, pentostatin, cytarabine, azacitidine, etoposide, teniposide, irinotecan, docetaxel, doxorubicin, daunorubicin, dactinomycin, idarubicin, plicamycin, mitomycin, bleomycin, tamoxifen, flutamide, leuprolide, goserelin, aminogluthimide, anastrozole, amsacrine, asparaginase, mitoxantrone, mitotane, or amifostine. In some aspects, the anti-cancer agent targets other oncogenic signaling pathways, targets immune response pathways, directly drives an immune response to cancer cells, or targets disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells.

[0042] In various aspects, the present disclosure provides a pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequenceidentity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent.

[0043] In some aspects, the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof. In some aspects, the EGFR-binding peptide is the EGFR-binding peptide as described herein.

[0044] In various aspects, the present disclosure provides a pharmaceutical composition comprising: a peptide complex comprising: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptideis complexed with the cellular receptor-binding peptide; and the EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent.

[0045] In some aspects, the peptide complex is the peptide complex as described herein or the peptide-active agent complex as described herein.

[0046] In various aspects, the present disclosure provides a method of administering a pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent, to a subject.

[0047] In some aspects, the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof. In some aspects, the EGFR-binding peptide is the EGFR-binding peptide as described herein.

[0048] In various aspects, the present disclosure provides a method of administering a pharmaceutical composition comprising: a peptide complex comprising: (i) a cellular receptorbinding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between thethird cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR- binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent, to a subject.

[0049] In some aspects, the peptide complex is the peptide complex as described herein or the peptide-active agent complex as described herein. In some aspects, the pharmaceutical composition is the pharmaceutical composition as described herein.

[0050] In various aspects, the present disclosure provides a method of inhibiting EGFR in a subject, the method comprising: administering to the subject a pharmaceutical composition, wherein the pharmaceutical composition comprises: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide binds to EGFR on the cell of the subject and inhibits activation of the EGFR.

[0051] In some aspects, the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof. In some aspects, the EGFR-binding peptide is the EGFR-binding peptide as described herein.

[0052] In various aspects, the present disclosure provides a method of inhibiting EGFR in a subject, the method comprising: administering to the subject a pharmaceutical composition, wherein the pharmaceutical composition comprises: a peptide complex comprising: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR- binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide binds to EGFR on the cell of the subject and inhibits activation of the EGFR.

[0053] In some aspects, the peptide complex is the peptide complex as described herein or the peptide-active agent complex as described herein. In some aspects, the pharmaceutical composition is the pharmaceutical composition as described herein. In some aspects, the EGFR- binding peptide inhibits activation of the EGFR by disrupting multimerization, dimerization, or heterodimerization of the EGFR on the cell of the subject that expresses EGFR.

[0054] In various aspects, the present disclosure provides a method of selectively depleting a target molecule, the method comprising: (a) contacting a peptide complex to a cell expressing a cellular receptor, wherein the peptide complex comprises: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine aminoacid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR- binding peptide has affinity for a target molecule; (b) binding the EGFR-binding peptide to the target molecule under extracellular conditions; (c) binding the cellular receptor-binding peptide to the cellular receptor under extracellular conditions; and (d) endocytosing the peptide complex, the target molecule, and the cellular receptor into an endocytic or lysosomal compartment, thereby depleting the target molecule.

[0055] In some aspects, the peptide complex as described herein or the peptide-active agent complex as described herein. In some aspects, the method further comprises (e) dissociating the EGFR-binding peptide from the target molecule, the cellular-receptor-binding peptide from the cellular receptor, or both, under endosomal or lysosomal conditions. In some aspects, the method further comprises (f) degrading the target molecule, thereby further depleting the target molecule. In some aspects, the method further comprises recycling the peptide complex and the cellular receptor to the cell surface.

[0056] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising administering a pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteineamino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent, to the subject, thereby treating the disease or condition.

[0057] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising: administering a pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide inhibits the EGFR on the cell of the subject, thereby treating the disease or condition.

[0058] In some aspects, the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof. In some aspects, the EGFR-binding peptide is the EGFR-binding peptide as described herein.

[0059] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising administering a pharmaceutical composition comprising: a peptide complex comprising: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, athird cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR- binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent, to the subject, thereby treating the disease or condition.

[0060] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising: administering a pharmaceutical composition comprising: a peptide complex comprising: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR- binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expressesEGFR, wherein the EGFR-b inding peptide inhibits the EGFR on the cell of the subject, thereby treating the disease or condition.

[0061] In some aspects, the peptide complex as described herein or the peptide-active agent complex as described herein. In some aspects, the pharmaceutical composition is the pharmaceutical composition as described herein.

[0062] In some aspects, the EGFR comprises wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximab-resistant EGFR, or a panitumumab- resistant EGFR. In some aspects, the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

[0063] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising: (a) administering a pharmaceutical composition to the subject, wherein the pharmaceutical composition comprises: a peptide complex comprising: (i) a cellular receptor-binding peptide; and (ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR-binding peptide has affinity for a targetmolecule; and a pharmaceutically acceptable excipient or diluent; (b) binding the EGFR-binding peptide under extracellular conditions to a target molecule associated with the disease or condition on a cell of the subject expressing the target molecule and a cellular receptor; (c) binding the cellular receptor-binding peptide under extracellular conditions to the cellular receptor on the cell of the subject; and (d) endocytosing the peptide complex, the target molecule, and the cellular receptor, thereby treating the disease or condition.

[0064] In some aspects, the peptide complex is the peptide complex as described herein or the peptide-active agent complex as described herein. In some aspects, the pharmaceutical composition is the pharmaceutical composition as described herein.

[0065] In some aspects, the method further comprises (e) dissociating the EGFR-binding peptide from the target molecule, the cellular-receptor-binding peptide from the cellular receptor, or both under endosomal conditions. In some aspects, the method further comprises (f) degrading the target molecule.

[0066] In some aspects, the target molecule comprises an EGFR. In some aspects, the EGFR comprises wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximab-resistant EGFR, or a panitumumab-resistant EGFR. In some aspects, the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

[0067] In some aspects, the disease or condition is a cancer. In some aspects, the cancer expresses EGFR, overexpresses EGFR, or contains mutant EGFR. In some aspects, the cancer is breast cancer, liver cancer, colon cancer, brain cancer, leukemia, lymphoma, non-Hodgkin lymphoma, myeloma, blood-cell-derived cancer, lung cancer, sarcoma, stomach cancer, a gastrointestinal cancer, glioblastoma, head and neck cancer, squamous head and neck cancer, non-small-cell lung cancer, squamous non-small cell lung cancer, pancreatic cancer, ovarian cancer, endometrial cancer, blood cancer, skin cancer, liver cancer, kidney cancer, or colorectal cancer. In some aspects, the cancer is TKI-resistant, cetuximab-resistant, necitumumab-resistant, or panitumumab-resistant. In some aspects, the cancer has one or more of the following:overexpresses EGFR, KRAS mutation, KRAS G12S mutation, KRAS G12C mutation, PTEN loss, EGFR exonl9 deletion, EGFR L858R mutation, EGFR T790M mutation, PIK3CA mutation, TP53 R273H mutation, PIK3CA amplification, PIK3CA G118D, TP53 R273H, EGFR C797X mutation, EGFR G724S mutation, EGFR L718Q mutation, EGFR S768I mutation, an EGFR mutation, a cetuximab-resistant EGFR, a panitumumab-resistant EGFR, or a combination thereof.

[0068] In some aspects, the cancer expresses or has upregulated c-MET, Her2, Her3 that heterodimerizes with EGFR. In some aspects, the cancer is a primary cancer, an advanced cancer, a metastatic cancer, a metastatic cancer in the central nervous system, a primary cancer in the central nervous system, metastatic colorectal cancer, metastatic head and neck cancer, metastatic non-small-cell lung cancer, metastatic breast cancer, metastatic skin cancer, a refractory cancer, a KRAS wild type cancer, a KRAS mutant cancer, or an exon20 mutant non- small-cell lung cancer.

[0069] In some aspects, the method further comprises administering an additional therapy to the subject. In some aspects, the additional therapy is adjuvant, first-line, or combination therapy. In some aspects, the additional therapy targets other oncogenic signaling pathways, targets immune response pathways, directly drives an immune response to cancer cells, or targets disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells. In some aspects, targeting of other oncogenic signaling pathways comprises administration of inhibitors of MEK / ERK pathway signaling, PI3K / AKT pathway signaling, JAK / STAT pathway signaling, or WNT / p-catenin pathway signaling. In some aspects, targeting of immune response pathways comprises PD-1 / PD-L1 checkpoint inhibition. In some aspects, directly driving an immune response to cancer cells comprises bispecific T cell engagers or chimeric antigen receptor expressing T cells. In some aspects, targeting disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells comprises administering seno lytic agents to a subject.

[0070] In some aspects, the additional therapy comprises administering fluorouracil, FOLFIRI, irinotecan, FOLFOX, gemcitabine, or cisplatin, irinotecan, oxiplatin, fluoropyrimidine to the subject.

[0071] In some aspects, the method further comprises forming a ternary complex between the peptide complex, the target molecule, and the cellular receptor. In some aspects, formation of the ternary complex increases, facilitates, or stabilizes recycling or turnover of the cellular receptor, the target molecule, or both. In some aspects, formation of the ternary complex increases, facilitates, or stabilizes binding of the target molecule to the cellular receptor. In someaspects, the peptide complex binds at higher levels to cells that overexpress the target molecule and the cellular receptor than to cells that have lower levels of the target molecule or the cellular receptor or both.

[0072] In some aspects, the peptide complex has a larger, longer, or wider therapeutic window as compared to an alternative therapy. In some aspects, the alternative therapy is not recycled to the cell surface. In some aspects, the alternative therapy is a lysosomal targeting therapy, a ubiquitin-proteosome system (UPS) targeting therapy, a non-selective therapeutic agent, an existing biologic, or a lysosomal delivery molecule. In some aspects, the peptide complex or the EGFR-binding peptide is administered at lower molar dosage than alternative therapies. In some aspects, the peptide complex or the EGFR-binding peptide binds at higher levels to cancer cells than to normal cells.

[0073] In some aspects, the peptide complex or the EGFR-binding peptide has a higher antiproliferative effect, a higher target molecule depletion effect, or a higher viability effect on cancer cells than on normal cells in vitro or in vivo. In some aspects, the peptide complex or the EGFR-binding peptide has a larger, longer, or wider therapeutic window than an anti-EGFR antibody or a TKI. In some aspects, the peptide complex or the EGFR-binding peptide has lower toxicity on skin or on keratinocytes than an anti-EGFR antibody or a TKI.

[0074] In some aspects, the method further comprises causing remission in, reducing, ameliorating, or ablating the disease or condition. In some aspects, the method further comprises causing remission in, reducing, ameliorating, or ablating the cancer.

[0075] In various aspects, the present disclosure provides an EGFR-binding peptide comprising a sequence having at least 70% sequence identity to SEQ ID NO: 317 and comprising at least one mutation relative to SEQ ID NO: 317.

[0076] In some aspects, the EGFR-binding peptide comprises a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue; wherein the EGFR-binding peptide comprises: seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue.

[0077] In some aspects, the first cysteine amino acid residue is at position 6 of the EGFR- binding peptide, the second cysteine amino acid residue is at position 14 of the EGFR-binding peptide, the third cysteine amino acid residue is at position 20 of the EGFR-binding peptide, the fourth cysteine amino acid residue is at position 31 of the EGFR-binding peptide, the fifth cysteine amino acid residue is at position 33 of the EGFR-binding peptide, and the sixth cysteine amino acid residue is at position 42 of the EGFR-binding peptide.

[0078] In some aspects, the at least one mutation comprises an amino acid substitution of DI 1R, I23S, V35E, S51P, L52E, R53E, M21R, A30W, I38D, W49R, V34S, Q43I, Q43V, Q43W, Q43Y, K48N, K48T, K48A, K48L, E51S, E51H, L52H, R53H, or a combination thereof. In some aspects, the at least one mutation comprises an amino acid substitution of M21R, A30W, 138D, W49R, or a combination thereof. In some aspects, the at least one mutation comprises an amino acid substitution of DI 1R, I23S, V35E, S51P, L52E, R53E, or a combination thereof. In some aspects, the at least one mutation comprises an amino acid substitution of E51H, L52H, R53H, or a combination thereof.

[0079] In various aspects, the present disclosure provides an EGFR-binding peptide comprising a sequence of SEQ ID NO: 314.

[0080] In various aspects, the present disclosure provides a n EGFR-binding peptide comprising a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 388 - SEQ ID NO: 390 or SEQ ID NO: 457 - SEQ ID NO: 494.

[0081] In some aspects, the EGFR-binding peptide comprises a sequence having at least 90% sequence identity with any one of SEQ ID NO: 388 - SEQ ID NO: 390 or SEQ ID NO: 457 - SEQ ID NO: 494. In some aspects, the EGFR-binding peptide is capable of binding to EGFR without activating the EGFR. In some aspects, the EGFR-binding peptide blocks binding of EGF to EGFR when the EGFR-binding peptide is bound to the EGFR. In some aspects, the EGFR-binding peptide inhibits EGFR when the EGFR-binding peptide is bound to the EGFR. In some aspects, the EGFR-binding peptide prevents dimerization of EGFR when the EGFR- binding peptide is bound to the EGFR.

[0082] In various aspects, the present disclosure provides a peptide complex comprising: a) a cellular receptor-binding peptide; and b) a target-binding peptide complexed with the cellular receptor-binding peptide, wherein the target-binding peptide has affinity for a target molecule, and wherein the target-binding peptide comprises the EGFR-binding peptide as described herein.

[0083] In various aspects, the present disclosure provides a peptide complex comprising: a) a cellular receptor-binding peptide; and b) a target-binding peptide complexed with the cellular receptor-binding peptide, wherein the target-binding peptide has affinity for a target molecule, and wherein the target-binding peptide comprises a sequence of SEQ ID NO: 314 or a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 388 - SEQ ID NO: 390 or SEQ ID NO: 457 - SEQ ID NO: 494.

[0084] In some aspects, the affinity of the target-binding peptide for the target molecule, the affinity of the cellular receptor binding peptide for the cellular receptor, or both is pH- independent. In some aspects, the affinity of the target-binding peptide for the target molecule, the affinity of the cellular receptor binding peptide for the cellular receptor, or both is pH dependent. In some aspects, the affinity of the target-binding peptide for the target molecule, the affinity of the cellular receptor-binding peptide for the cellular receptor, or both is ionic strength dependent.

[0085] In some aspects, the target binding peptide comprises a sequence having at least 90% sequence identity with any one of SEQ ID NO: 388 - SEQ ID NO: 390 or SEQ ID NO: 457 - SEQ ID NO: 494. In some aspects, the target binding peptide comprises a sequence of any one of SEQ ID NO: 388 - SEQ ID NO: 390 or SEQ ID NO: 457 - SEQ ID NO: 494.

[0086] In some aspects, the cellular receptor-binding peptide is a transferrin receptor-binding peptide or a PD-L1 -binding peptide. In some aspects, the cellular receptor is a transferrin receptor or PD-L1. In some aspects, the cellular receptor is a cation-independent mannose 6 phosphate receptor (CI-M6PR), an asialoglycoprotein receptor (ASGPR), CXCR7, folate receptor, or Fc receptor (including but not limited to neonatal Fc receptor (FcRn) or FcyRIIb).

[0087] In some aspects, the cellular receptor-binding peptide binds to the cellular receptor at a pH of from pH 4.5 to pH 7.4, from pH 5.5 to pH 7.4, from pH 5.8 to pH 7.4, or from pH 6.5 to pH 7.4. In some aspects, the cellular receptor-binding peptide is capable of binding the cellular receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 7.4. In some aspects, the cellular receptor-binding peptide is capable of binding the cellular receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no morethan 0.2 nM, or no more than 0.1 nM at pH 5.5. In some aspects, the cellular receptor-binding peptide is capable of binding the cellular receptor with a dissociation rate constant (koff or kd) of no more than 1 s'1, no more than 5x1 O'1s'1, no more than 2x1 O'1s'1, no more than 1x1 O'2s'1, no more than 5x1 O'2s'1, no more than 2x1 O'2s'1, no more than 1x1 O'2s'1, no more than 5x1 O'3s'1, no more than 2x1 O'3s'1, no more than 1x1 O'3s'1, no more than 5x1 O'4s'1, or no more than 2x1 O'4s'1at pH 5.5.

[0088] In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is pH-independent. In some aspects, the affinity of the target-binding peptide for the target molecule is pH-dependent. In some aspects, the affinity of the target-binding peptide for the target molecule is pH-independent.

[0089] In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some aspects, the dissociation rate constant (koff or kd) of the cellular receptorbinding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some aspects, the dissociation rate constant (koff or kd) of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

[0090] In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is pH dependent. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor decreases as pH decreases. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is higher at pH 7.4 than at pH 5.5. In some aspects, the affinity of the cellular receptor-binding peptide for the cellular receptor is higher at pH 7.4 than at pH 5.8. In some aspects, the affinity of the target-binding peptide for the target molecule is pH dependent. In some aspects, the affinity of the target-binding peptide for the target molecule decreases as pH decreases. In some aspects, the affinity of the target-bindingpeptide for the target molecule is higher at a higher pH than at a lower pH. In some aspects, the higher pH is pH 7.4, pH 7.2, pH 7.0, or pH 6.8. In some aspects, the lower pH is pH 6.5, pH 6.0, pH 5.8, pH 5.5, pH 5.0, or pH 4.5. In some aspects, the affinity of the target-binding peptide for the target molecule is higher at pH 7.4 than at pH 6.0. In some aspects, the affinity of the targetbinding peptide for the target molecule is higher at pH 7.4 than at pH 5.5. In some aspects, the affinity of the target-binding peptide for the target molecule is higher at pH 7.4 than at pH 5.8.

[0091] In some aspects, the target-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no more than 500 nM, no more than 200 nM, 100 nM, no more than 50 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, no more than 1 nM, or no more than 0.1 nM at pH 7.4. In some aspects, the target-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1x10'1s’1, 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, no more than 2x1 O’4s’1, no more than 1x1 O’4s’1, no more than 5x10’5s’1, or no more than 2x10’5s’1at pH 7.4.

[0092] In some aspects, the target-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 5.5. In some aspects, the target-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 5.8.

[0093] In some aspects, the dissociation rate constant (koff or kd) for target-binding peptide binding the target molecule is at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.5 than at pH 7.4. In some aspects, the dissociation rate constant (koff or kd) for target-binding peptide binding the target molecule is at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.8 than at pH 7.4.

[0094] In some aspects, the target-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.5. In some aspects, the target-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.8.

[0095] In some aspects, the affinity of the target-binding peptide for the target molecule at pH 7.4 is at least 1.5-fold, 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the target-binding peptide for the target molecule at pH 5.5. In some aspects, the affinity of the target-binding peptide for the target molecule at pH 7.4 is at least 1.5- fold, at least 2-fold, at least 3 -fold, at least 4-fold, at least 5 -fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the target-binding peptide for the target molecule at pH 5.8. In some aspects, the affinity of the target-binding peptide for the target molecule at pH 7.4 is less than 0.5-fold, less than 1-fold, less than, 1.5-fold, less than 2-fold, less than 3-fold, or less than 10-fold, greater than the affinity of the target-binding peptide for the target molecule at pH 5.8.

[0096] In some aspects, the target-binding peptide comprises one or more histidine amino acid residues. In some aspects, the affinity of the target-binding peptide for the target molecule decreases as ionic strength increases. In some aspects, the target-binding peptide comprises one or more polar or charged amino acid residues capable of forming polar or charge-charge interactions with the target molecule.

[0097] In some aspects, the cellular receptor-binding peptide is fused to, linked to, complexed with, or conjugated to the target-binding peptide. In some aspects, the cellular receptor-binding peptide is fused to, linked to, complexed with, or conjugated to the target-binding peptide via a polymer linker. In some aspects, the polymer linker is a polyethylene glycol (PEG), a hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a zwitterionic water soluble polymer, a water soluble poly(amino acid), a water soluble polymer comprising proline, alanine,serine, or a combination thereof, a water soluble polymer containing glycine, glutamic acid, and serine, an Fc region, a fatty acid, a palmitic acid, an albumin, or an albumin binding molecule.

[0098] In some aspects, the cellular receptor-binding peptide and the target-binding peptide form a single polypeptide chain. In some aspects, the peptide complex comprises a dimer dimerized via a dimerization domain. In some aspects, the distance between the cellular receptor-binding peptide and the target-binding peptide is at least 1 nm, at least 2 nm, at least 5 nm, at least 10 nm, at least 20 nm, at least 50 nm, or at least 100 nm. In some aspects, the dimerization domain comprises an Fc domain. In some aspects, the dimer is a homodimer dimerized via a homodimerization domain. In some aspects, the homodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 535, SEQ ID NO: 706, or SEQ ID NO: 246. In some aspects, the dimer is a heterodimer dimerized via a first hetero dimerization domain and a second heterodimerization domain. In some aspects, the first heterodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 536, SEQ ID NO: 707, or SEQ ID NO: 709. In some aspects, the second heterodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 537, SEQ ID NO: 708, or SEQ ID NO: 710.

[0099] In some aspects, the target-binding peptide is linked to the dimerization domain via a peptide linker. In some aspects, the cellular receptor-binding peptide is linked to the dimerization domain via a peptide linker. In some aspects, the cellular receptor-binding peptide is linked to the target-binding peptide via a peptide linker. In some aspects, the peptide linker has a length of from 1 to 50 amino acid residues, from 2 to 40 amino acid residues, from 3 to 20 amino acid residues, or from 3 to 10 amino acid residues. In some aspects, the peptide linker comprises glycine and serine amino acids. In some aspects, the peptide linker has a persistence length of no more than 6 A, no more than 8 A, no more than 10 A, no more than 12 A, no more than 15 A, no more than 20 A, no more than 25 A, no more than 30 A, no more than 40 A, no more than 50 A, no more than 75 A, no more than 100 A, no more than 150 A, no more than 200 A, no more than 250 A, or no more than 300 A. In some aspects, the peptide linker is derived from an immunoglobulin peptide. In some aspects, the peptide linker is derived from a double-knot toxin peptide. In some aspects, the peptide linker comprises a sequence of any one of SEQ ID NO: 129 - SEQ ID NO: 141, SEQ ID NO: 195 - SEQ ID NO: 218, SEQ ID NO: 223 - SEQ ID NO: 223 - SEQ ID NO: 227, SEQ ID NO: 194, SEQ ID NO: 391, SEQ ID NO: 538, or SEQ ID NO: 540 - SEQ ID NO: 541.

[0100] In some aspects, the cellular receptor-binding peptide, the target-binding peptide, peptide complex, or a combination thereof comprises a miniprotein, a nanobody, an antibody, an antibody fragment, an scFv, a DARPin, or an affibody. In some aspects, the antibody comprises an IgG, or wherein the antibody fragment comprises a Fab, a F(ab)2, an scFv, or an (scFv)2. In some aspects, the miniprotein comprises a cystine-dense peptide, an affitin, an adnectin, an avimer, a Kunitz domain, a nanofittin, a fynomer, a bicyclic peptide, a beta-hairpin, or a stapled peptide. In some aspects, the cellular receptor-binding peptide comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds. In some aspects, the target-binding peptide comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds. In some aspects, the peptide complex comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds. In some aspects, the cellular receptorbinding peptide comprises at least six cysteine residues. In some aspects, the at least six cysteine residues are positioned at amino acid positions 4, 8, 18, 32, 42, and 46 of the cellular receptorbinding peptide. In some aspects, the at least six cysteine residues form at least three disulfide bonds.

[0101] In some aspects, the cellular receptor-binding peptide comprises a sequence of any one of SEQ ID NO: 148 - SEQ ID NO: 177. In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identitywith SEQ ID NO: 96, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 96. In some aspects, the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 96.

[0102] In some aspects, the cellular receptor-binding peptide comprises a sequence of any one of SEQ ID NO: 392 - SEQ ID NO: 399. In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241. In some aspects, the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 187, SEQ ID NO: 235, SEQ ID NO: 236, SEQ ID NO: 238, SEQ ID NO: 239, SEQ ID NO: 400, or SEQ ID NO: 401 or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 187. In some aspects, the cellular receptorbinding peptide comprises a sequence of SEQ ID NO: 187.

[0103] In some aspects, the fragment comprises at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36, at least 37, at least 38, at least 39, at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, or at least 50 amino acid residues. In some aspects, the cellular receptor-binding peptide comprises one or more histidine residues at a cellular receptor-binding interface. In some aspects, the target-binding peptide comprises one or more histidine residues at a target-binding interface.

[0104] In some aspects, the target-binding peptide is an EGFR-binding peptide. In some aspects, the target molecule comprises an EGFR. In some aspects, the EGFR is wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, or a combination thereof. In some aspects, the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFRG796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

[0105] In some aspects, an off rate of the cellular receptor-binding peptide from the cellular receptor is slower than a recycling rate of the cellular receptor. In some aspects, a half-life of dissociation of the cellular receptor-binding peptide from the cellular receptor is no faster than 1 minute, no faster than 2 minutes, no faster than 3 minutes, no faster than 4 minutes, no faster than 5 minutes, no faster than 7 minutes, no faster than 10 minutes, no faster than 15 minutes, no faster than 20 minutes, no faster than 30 minutes, no faster than 45 minutes, no faster than 60 minutes, no faster than 90 minutes, or no faster than 120 minutes. In some aspects, a rate of dissociation of the target-binding peptide from the target molecule is faster than a recycling rate of the cellular receptor. In some aspects, a half-life of dissociation of the target binding-binding peptide from the target molecule is less than 10 seconds, less than 20 seconds, less than 30 seconds, less than 1 minute, less than 2 minutes, less than 5 minutes, less than 10 minutes, less than 20 minutes, less than 30 minutes, less than 45 minutes, or less than 60 minutes in endosomal conditions.

[0106] In some aspects, the peptide complex is capable of being endocytosed via receptor- mediated endocytosis. In some aspects, the receptor-mediated endocytosis is transferrin receptor-mediated endocytosis. In some aspects, the receptor-mediated endocytosis is PD-L1- mediated endocytosis. In some aspects, the cellular receptor-binding peptide remains bound to the cellular receptor inside an endocytic vesicle. In some aspects, the peptide complex is recycled to the cell surface when the cellular receptor-binding peptide is bound to the cellular receptor and the cellular receptor is recycled. In some aspects, the target molecule is released or dissociated from the target-binding peptide after the peptide complex is endocytosed via receptor-mediated endocytosis.

[0107] In some aspects, the target molecule is an extracellular protein, a circulating protein, or a soluble protein. In some aspects, the target molecule is a cell surface protein. In some aspects, the target molecule is a transmembrane protein.

[0108] In some aspects, the peptide complex further comprises a half-life modifying agent coupled to the cellular receptor-binding peptide, the target-binding peptide, or both. In some aspects, the half-life modifying agent is a polymer, a polyethylene glycol (PEG), a hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a zwitterionic water soluble polymer, a water soluble poly(amino acid), a water soluble polymer of proline, alanine and serine, a water solublepolymer containing glycine, glutamic acid, and serine, an Fc region, a fatty acid, palmitic acid, an albumin, or a molecule that binds to albumin. In some aspects, the molecule that binds to albumin is a serum albumin-binding peptide. In some aspects, the serum albumin-binding peptide comprises a sequence of any one of SEQ ID NO: 178, SEQ ID NO: 179, or SEQ ID NO: 193.

[0109] In some aspects, the cellular receptor-binding peptide, the target-binding peptide, or both is recombinantly expressed. In some aspects, the target-binding peptide is configured to dissociate from the target molecule at pH 6.5, pH 6.0, pH 5.8, pH 5.5, pH 5.0, or pH 4.5. In some aspects, the cellular receptor-binding peptide is configured to dissociate from the cellular receptor at pH 6.5, pH 6.0, pH 5.5, pH 5.0, or pH 4.5.

[0110] In some aspects, the peptide complex comprises a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 499 - SEQ ID NO: 501, SEQ ID NO: 504, SEQ ID NO: 505, SEQ ID NO: 507, SEQ ID NO: 508, or SEQ ID NO: 511 - SEQ ID NO: 526. In some aspects, the peptide complex comprises a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 495 - SEQ ID NO: 497, SEQ ID NO: 503, or SEQ ID NO: 506.

[0111] In various aspects, the present disclosure provides a peptide-active agent complex comprising a peptide complexed to an active agent, wherein the peptide comprises the EGFR- binding peptide as described herein or the peptide complex as described herein.

[0112] In some aspects, the active agent comprises a peptide, a peptidomimetic, an oligonucleotide, a DNA, an RNA, an antibody, a single chain variable fragment (scFv), an antibody fragment, an aptamer, or a small molecule. In some aspects, the DNA comprises cDNA, ssDNA, or dsDNA. In some aspects, the RNA comprises RNAi, microRNA, snRNA, dsRNA, or an antisense oligonucleotide. In some aspects, the active agent is a therapeutic agent or a detectable agent. In some aspects, the detectable agent comprises a dye, a fhiorophore, a fluorescent biotin compound, a luminescent compound, a chemiluminescent compound, a radioisotope, nanopartide, a paramagnetic metal ion, or a combination thereof. In some aspects, the therapeutic agent is an anti-cancer agent. In some aspects, the anti-cancer agent comprises a radionuclide, radioisotope, a chemotherapeutic agent, a platinum therapeutic, a toxin, an enzyme, a sensitizing drug, an anti-angiogenic agent, cisplatin, an anti-metabolite, an anti- metabolic therapeutic, a mitotic inhibitor, a growth factor inhibitor, paclitaxel, temozolomide,topotecan, fluorouracil, vincristine, vinblastine, procarbazine, decarbazine, altretamine, methotrexate, mercaptopurine, thioguanine, fludarabine phosphate, cladribine, pentostatin, cytarabine, azacitidine, etoposide, teniposide, irinotecan, docetaxel, doxorubicin, daunorubicin, dactinomycin, idarubicin, plicamycin, mitomycin, bleomycin, tamoxifen, flutamide, leuprolide, goserelin, aminogluthimide, anastrozole, amsacrine, asparaginase, mitoxantrone, mitotane, or amifostine. In some aspects, the anti-cancer agent targets other oncogenic signaling pathways, targets immune response pathways, directly drives an immune response to cancer cells, or targets disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells.

[0113] In various aspects, the present disclosure provides a pharmaceutical composition comprising the EGFR-binding peptide as described herein and a pharmaceutically acceptable excipient or diluent.

[0114] In various aspects, the present disclosure provides a pharmaceutical composition comprising the peptide complex as described herein or the peptide-active agent complex as described herein and a pharmaceutically acceptable excipient or diluent.

[0115] In various aspects, the present disclosure provides a method of inhibiting EGFR in a subject, the method comprising administering to the subject a composition comprising the EGFR-binding peptide as described herein and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide binds to EGFR on the cell of the subject and inhibits activation of the EGFR.

[0116] In some aspects, the EGFR-binding peptide inhibits activation of the EGFR by disrupting multimerization, dimerization, or heterodimerization of the EGFR on the cell of the subject that expresses EGFR.

[0117] In various aspects, the present disclosure provides a method of selectively depleting a target molecule, the method comprising: a) contacting the peptide complex as described herein to a cell expressing a cellular receptor; b) binding the target-binding peptide to the target molecule under extracellular conditions; c) binding the cellular receptor-binding peptide to the cellular receptor under extracellular conditions; and d) endocytosing the peptide complex, the target molecule, and the cellular receptor into an endocytic or lysosomal compartment, thereby depleting the target molecule.

[0118] In some aspects, the method further comprises: e) dissociating the target-binding peptide from the target molecule, the cellular-receptor-binding peptide from the cellular receptor, or both under endosomal or lysosomal conditions. In some aspects, the method further comprises: f) degrading the target molecule, thereby further depleting the target molecule. In some aspects,the method further comprises recycling the peptide complex and the cellular receptor to the cell surface.

[0119] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising administering to the subject the EGFR-binding peptide as described herein or the pharmaceutical composition as described herein and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide inhibits the EGFR on the cell of the subject, thereby treating the disease or condition.

[0120] In some aspects, the EGFR comprises wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximab-resistant EGFR, or a panitumumab- resistant EGFR. In some aspects, the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

[0121] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising: a) administering to the subject the EGFR-binding peptide as described herein, the peptide complex as described herein, the peptide-active agent complex as described herein, or the pharmaceutical composition as described herein; b) binding the target-binding peptide under extracellular conditions to a target molecule associated with the disease or condition on a cell of the subject expressing the target molecule and a cellular receptor; c) binding the cellular receptor-binding peptide under extracellular conditions to the cellular receptor on the cell of the subject; and d) endocytosing the peptide complex, the target molecule, and the cellular receptor.

[0122] In some aspects, the method further comprises: e) dissociating the target-binding peptide from the target molecule, the cellular-receptor-binding peptide from the cellular receptor, or both under endosomal conditions. In some aspects, the method further comprises: f) degrading the target molecule. In some aspects, the target molecule comprises an EGFR. In some aspects, the EGFR comprises wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, or EGFRmutant T790M. In some aspects, the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

[0123] In some aspects, the disease or condition is a cancer. In some aspects, the cancer expresses EGFR, overexpresses EGFR, or contains mutant EGFR. In some aspects, the cancer is breast cancer, liver cancer, colon cancer, brain cancer, leukemia, lymphoma, non-Hodgkin lymphoma, myeloma, blood-cell-derived cancer, lung cancer, sarcoma, stomach cancer, a gastrointestinal cancer, glioblastoma, head and neck cancer, squamous head and neck cancer, non-small-cell lung cancer, squamous non-small cell lung cancer, pancreatic cancer, ovarian cancer, endometrial cancer, blood cancer, skin cancer, liver cancer, kidney cancer, or colorectal cancer. In some aspects, the cancer is TKI-resistant, cetuximab-resistant, necitumumab-resistant, or panitumumab-resistant. In some aspects, the cancer has one or more of the following: overexpresses EGFR, KRAS mutation, KRAS G12S mutation, KRAS G12C mutation, PTEN loss, EGFR exonl9 deletion, EGFR L858R mutation, EGFR T790M mutation, a cetuximabresistant EGFR, a panitumumab-resistant EGFR, PIK3CA mutation, TP53 R273H mutation, PIK3CA amplification, PIK3CA G118D, TP53 R273H, EGFR C797X mutation, EGFR G724S mutation, EGFR L718Q mutation, EGFR S768I mutation, an EGFR mutation, or a combination thereof. In some aspects, the cancer expresses or has upregulated c-MET, Her2, Her3 that heterodimerizes with EGFR. In some aspects, the cancer is a primary cancer, an advanced cancer, a metastatic cancer, a metastatic cancer in the central nervous system, a primary cancer in the central nervous system, metastatic colorectal cancer, metastatic head and neck cancer, metastatic non-small-cell lung cancer, metastatic breast cancer, metastatic skin cancer, a refractory cancer, a KRAS wild type cancer, a KRAS mutant cancer, or an exon20 mutant non- small-cell lung cancer.

[0124] In some aspects, the method further comprises administering an additional therapy to the subject. In some aspects, the additional therapy is adjuvant, first-line, or combination therapy. In some aspects, the additional therapy targets other oncogenic signaling pathways, targets immune response pathways, directly drives an immune response to cancer cells, or targets disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells. In some aspects,targeting of other oncogenic signaling pathways comprises administration of inhibitors of MEK / ERK pathway signaling, PI3K / AKT pathway signaling, JAK / STAT pathway signaling, or WNT / p-catenin pathway signaling. In some aspects, targeting of immune response pathways comprises PD-1 / PD-L1 checkpoint inhibition. In some aspects, directly driving an immune response to cancer cells comprises bispecific T cell engagers or chimeric antigen receptor expressing T cells. In some aspects, the targeting disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells comprises administering senolytic agents to a subject. In some aspects, the additional therapy comprises administering fluorouracil, FOLFIRI, irinotecan, FOLFOX, gemcitabine, or cisplatin, irinotecan, oxiplatin, fluoropyrimidine to the subject.

[0125] In some aspects, the method further comprises forming a ternary complex between the selective depletion complex, the target molecule, and the cellular receptor. In some aspects, formation of the ternary complex increases, facilitates, or stabilizes recycling or turnover of the cellular receptor, the target molecule, or both. In some aspects, formation of the ternary complex increases, facilitates, or stabilizes binding of the target molecule to the cellular receptor. In some aspects, the peptide complex binds at higher levels to cells that overexpress the target molecule and the cellular receptor than to cells that have lower levels of the target molecule or the cellular receptor or both.

[0126] In some aspects, the peptide complex has a larger, longer, or wider therapeutic window as compared to an alternative therapy. In some aspects, the alternative therapy is not recycled to the cell surface. In some aspects, the alternative therapy is a lysosomal targeting therapy, a ubiquitin-proteosome system (UPS) targeting therapy, a non-selective therapeutic agent, an existing biologic, or a lysosomal delivery molecule. In some aspects, the peptide complex or the EGFR-binding peptide is administered at lower molar dosage than alternative therapies. In some aspects, the peptide complex or the EGFR-binding peptide binds at higher levels to cancer cells than to normal cells. In some aspects, the peptide complex or the EGFR-binding peptide has a higher antiproliferative effect, a higher target molecule depletion effect, or a higher viability effect on cancer cells than on normal cells in vitro or in vivo. In some aspects, the peptide complex or the EGFR-binding peptide has a larger, longer, or wider therapeutic window than an anti-EGFR antibody or a TKI. In some aspects, the peptide complex or the EGFR-binding peptide has lower toxicity on skin or on keratinocytes than an anti-EGFR antibody or a TKI.

[0127] In various aspects, the present disclosure provides a method of administering a peptide complex or an EGFR-binding peptide to a subject, the method comprising administering theEGFR-binding peptide as described herein, the peptide complex as described herein, the peptide-active agent complex as described herein, or the pharmaceutical composition as described herein.

[0128] In various aspects, the present disclosure provides a method of treating a disease or condition in a subject in need thereof, the method comprising administering to the subject the EGFR-binding peptide as described herein, the peptide complex as described herein, the peptide-active agent complex as described herein, or the pharmaceutical composition as described herein, thereby treating the disease or condition.INCORPORATION BY REFERENCE

[0129] All publications, patents, and patent applications cited in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.BRIEF DESCRIPTION OF THE DRAWINGS

[0130] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings of which:

[0131] FIG. 1 is an image illustrating domain binding in an EGF:EGFR complex. This crystal structure, publicly available and deposited into the RCSB as PDB ID: 1IVO, represents the ectodomains of an active EGFR homodimer (tetrameric due to the presence of two EGF molecules). EGF binding to both EGFR Domain I and Domain III brings Domain II away from Domain IV and into a conformation capable of association with another Domain II. In such a conformation, EGFR is dimerized and activated, leading to phosphorylation and activation of various pathways such as MEK / ERK and PI3K / AKT which leads to intracellular signaling and promote growth of cells, including promoting oncogenic potential and growth of cancer cells. Although the EGF variants herein bind EGFR, they also disrupt the dimerization of EGFR, and hence disrupt, or reduce, or ablate the promotion of growth and hence disrupt, or reduce, or ablate the oncogenic potential of the EGFR on cells. Such EGF variants herein can be used to kill target cancer cells through such disruption or via other means as described herein.

[0132] FIG. 2 illustrates a bar chart showing flow cytometry data for biotinylated soluble EGFR ectodomain and soluble full-length EGFR bound to the surface of a cell expressing wild typeEGF (SEQ ID NO: 317), using fluorescent (647 nm excitation) streptavidin as a co-stain. The chart shows that the wild type soluble EGFRvIII ectodomain (i.e., isolated EGFR Domain III, hereafter referred to simply as “EGFRvIII” when describing flow cytometry experiments) does not significantly interact with cell-surface-displayed wild type human EGF, while soluble full- length EGFR ectodomain (hereafter referred to simply as "full-length EGFR” when describing flow cytometry experiments) shows binding. This experiment shows the wild type EGFRvIII ectodomain does not bind to the EGF.

[0133] FIG. 3 illustrates a stepwise flow chart for engineering showing the high-level modification of EGF into a cell growth disrupting variant that binds EGFR but does not signal on the receptor that would also be ready for SDC incorporation or use as a direct EGFR inhibitor. This methodology is a general roadmap to select properties on variant or modified EGF that would be useful alone or in the context of an SDC without describing specific variants themselves. Specific EGF variants are described herein.

[0134] FIG. 4 illustrates the EGF:EGFR interface contacting residues subject to Rosetta design (Domain III) or intentional mutational disruption (Domain I). The publicly available crystal structure of EGFR, deposited into the RCSB as PDB ID: 1IVO, was used to analyze interactions between EGF and EGFR, which reveals EGF residues closely contacting either Domain I (grey font) or Domain III (black font) which are suitable for mutating in order to improve EGF:EGFR Domain III binding strength and / or disrupt EGF:EGFR Domain I binding strength. Specific residues used on the to improve EGF:EGFR Domain III binding strength and / or disrupt EGF:EGFR Domain I binding are depicted using single letter amino acid code. This methodology is a general roadmap to select properties on variant or modified EGF that would be useful alone or in the context of an SDC without describing specific variants themselves. Specific EGF variants are described herein.

[0135] FIG. 5 illustrates cells expressing pooled EGF variants screened for GFP fluorescence-x axis (EGF variant expression) and APC channel fluorescence-y-axis (biotinylated target protein + fluorescent streptavidin binding) using flow cytometry. The inset box in each graph shows that the EGF library has a high proportion of full-length EGFR binding variants (~23% of GFP+ cells), with a much smaller proportion capable of showing appreciable binding to Domain III alone (EGFRvIII, <2% of GFP+ cells), whereas control protein does not show appreciable APC channel fluorescence.

[0136] FIG. 6 illustrates successive enrichment or increase in cells expressing EGF variants capable of binding EGFRvIII using flow cytometry. Cells expressing EGF variants were stainedwith biotinylated EGFRvIII and streptavidin that fluoresces in the APC channel (approximately 647 nm excitation) at 100 nM each for 15 minutes on ice. GFP fluorescence (EGF variant expression) and APC channel fluorescence (target protein + streptavidin binding) were used to identify GFP+ cells with APC channel fluorescence above background levels, which were flow sorted and collected for further enrichment. The middle and bottom panels show same staining and flow sorting for a further two rounds of flow sorting enrichment.

[0137] FIG. 7 illustrates showing surface displayed EGF variant expression (x axis) and biotinylated EGFRvIII + fluorescent streptavidin binding (y-axis) in cells expressing a single EGF variant, including SEQ ID NO: 320, SEQ ID NO: 321, SEQ ID NO: 319, SEQ ID NO: 387, SEQ ID NO: 385, SEQ ID NO: 386, SEQ ID NO: 369, SEQ ID NO: 372, and SEQ ID NO: 383 measured using flow cytometry.

[0138] FIG. 8 illustrates amino acid sequences of several isolated EGF variants with demonstrable EGFRvIII binding and Domain 1 contact residues (depicted by “X”) identifying mutations (shaded) to Domain I-adjacent residues that do not eliminate binding to Domain III. Four such mutations are the EGF variants depicted as M21R (present in SEQ ID NO: 378), A30W (present in SEQ ID NO: 371), I38D (present in SEQ ID NO: 376), and W49R (present in SEQ ID NO: 383, SEQ ID NO: 384, and SEQ ID NO: 385). Three of these mutations (M21R, 138D, and W49R) replace hydrophobic residues with polar charged residues, potentially disrupting hydrophobic interactions strengthening the EGF:[EGFR Domain I] interaction. The other (A30W) replaces a small side chain with a large, bulky side chain, potentially disrupting EGF:[EGFR Domain I] interaction by steric hindrance.

[0139] FIG. 9 illustrates the EGF:[EGFR Domain I] interface from public crystallography data (deposited in the RCSB as PDB ID: 1IVO), with a focus on four proposed mutations M21R (present in SEQ ID NO: 378), A30W (present in SEQ ID NO: 371), I38D (present in SEQ ID NO: 376), and W49R (present in SEQ ID NO: 383, SEQ ID NO: 384, and SEQ ID NO: 385), and demonstrates their disruption of Domain I binding at specific residues in EGFR Domain I. In each panel, the original EGF residue is in black labeled with black font, while the modeled mutation is in grey labeled in grey font, depicting disrupting hydrophobic interactions strengthening the interaction with Domain 1 or disrupting Domain 1 though steric hindrance.

[0140] FIG. 10 illustrates flow cytometry data from surface-displayed EGF variants that exhibit EGFRvIII binding despite incorporation of one or more Domain I disrupting mutations (x-axis) and their EGFRvIII-binding capability (y-axis). FIG. 10 demonstrates maintenance of EGFR Domain III binding despite incorporation of one or more Domain I disrupting mutations.Variants of the lead EGFRvIII-binding EGF variant (SEQ ID NO: 319) were transfected individually into 293F cells and stained with 50 nM each biotinylated EGFRvIII and streptavidin that fluoresces in the APC channel (approximately 647 nm excitation). A subpopulation of cells with defined GFP expression was gated and fluorescence in these cells was measured. EGFRvIII binding was observed for all five variants, with the highest staining seen by EGF84v9 (SEQ ID NO: 458).

[0141] FIG. 11 illustrates a comparison of the wild type EGF and an EGF variant binding characteristics on full length EGFR and EGFRvIII (Domain III only). The binding characteristics of human EGF (SEQ ID NO: 317) control and EGF84v9 (SEQ ID NO: 458). Human EGF (SEQ ID NO: 317) and EGF84v9 (SEQ ID NO: 458) were transfected individually into wells of 293F cells and stained with either 50 nM biotinylated full-length EGFR + fluorescent streptavidin, or 50 nM EGFRvIII + fluorescent streptavidin. A subpopulation of cells with defined GFP expression was gated and fluorescence in these cells was measured. The variant EGF (SEQ ID NO: 458) could still bind to the full length EGFR in spite of its Domain 1 disruption but demonstrated much stronger binding to EGFRvIII (isolated EGFR Domain III), suggesting Domain I disruption was successful.

[0142] FIG. 12 illustrates, in tabular form, enrichment scores of mutations in a library of EGF84v9 (SEQ ID NO: 458) variants that have high EGFRvIII staining (high DAPI stain) but that lose this binding upon pH 5.5 incubation (low APC stain). Variant abundances were normalized to that of both EGF84v9 (SEQ ID NO: 458) itself and to the variant’s abundance in the initial pre-sort library, enabling evaluation of both overall enrichment / depletion and enrichment / depletion compared to the parent sequence. This double-normalized abundance is log2 -transformed and defined as the variant’s enrichment score; a negative enrichment score represents depletion from the population, while a positive enrichment score represents enrichment, the latter being a byproduct of improved EGFRvIII binding and a high degree of pH 5.5 release. The table shows these enrichment scores, rounded to the nearest integer. Figure discloses SEQ ID NO: 713.

[0143] FIG. 13 illustrates affinity maturation using EGFRvIII binding of EGF84v9 singleton variants analyzed by flow cytometry. While all of the matured variants demonstrated improved EGFRvIII staining compared to SEQ ID NO: 458, four EGF variants were selected for further analysis due to high EGFRvIII staining, including SEQ ID NO: 477, SEQ ID NO: 481, SEQ ID NO: 493, and SEQ ID NO: 494.

[0144] FIG. 14 illustrates EGFRvIII binding by EGF84v9 singleton variants SEQ ID NO: 477, SEQ ID NO: 481, SEQ ID NO: 493, and SEQ ID NO: 494 and the parent EGF84v9 (SEQ ID NO: 458) after a 10 minute buffer incubation (“rinse”) at pH 7.4 or pH 5.5. All variants demonstrated substantial reduction of EGFRvIII staining upon pH 5.5 rinse compared to pH 7.4 rinse, but SEQ ID NO: 494 had highest staining retention after pH 7.4 rinse while still very low staining after pH 5.5 rinse. Low or reduced staining at pH 5.5 rinse indicates that the binding strength of the tested singleton EGF variants to EGFRvIII is pH sensitive and the EGFRvIII is dissociated at the lower pH in the context of a bivalent interaction.

[0145] FIG. 15 illustrates EGF variant-based EGFR selective depletion complex (SDC) candidates. All seven molecules contain an Fc domain (homodimeric or heterodimeric knob- and-hole) and one or two domains constituting EGF variant SEQ ID NO: 494. Six also contain one or two domains constituting TfR-binding CDPs of SEQ ID NO: 96, making EGFR SDC candidate molecules (SEQ ID NO: 495, SEQ ID NO: 496, SEQ ID NO: 497, SEQ ID NO: 503 paired with SEQ ID NO: 504, SEQ ID NO: 503 paired with SEQ ID NO: 505, and SEQ ID NO: 504 paired with SEQ ID NO: 506). The differences between these SDC candidate molecules are the linker between the TfR-binding domain and the EGF variant domain, and also the variations in valence of EGFR or TfR binding capabilities. A control SDC without TfR-binding capabilities was also designed (SEQ ID NO: 498). Though the control SDC of SEQ ID NO: 498 does not have TfR binding capabilities and should therefore not drive TfR-cycling-dependent EGFR uptake, it does have EGFR binding capabilities and is predicted to bind EGFR without facilitating growth-signal-driving dimerization, which could render SEQ ID NO: 498 into an effective EGFR inhibitor. Ovals marked “N” and “C” represent homodimerization domains of SEQ ID NO: 535; ovals containing a convex or concave semicircular notch and marked “N” and “C” represent heterodimerization domains of SEQ ID NO: 536 and SEQ ID NO: 537, respectively; circles represent TfR-binding variants of SEQ ID NO: 96; 7-pointed stars represent EGF variants of SEQ ID NO: 494; curved and wavey lines represent linkers of SEQ ID NO: 223, SEQ ID NO: 540, SEQ ID NO: 541, or SEQ ID NO: 538.

[0146] FIG. 16 illustrates EGF variant-based EGFR selective depletion complex (SDC) candidates that use PD-L1 as the cellular receptor. All six molecules contain an Fc domain (homodimeric or heterodimeric knob-and-hole), one or two domains constituting EGF variant SEQ ID NO: 494, and one or two domains constituting PD-Ll-binding CDPs of SEQ ID NO: 187, making EGFR SDC candidate molecules (SEQ ID NO: 511, SEQ ID NO: 515, SEQ ID NO: 513, SEQ ID NO: 515 paired with SEQ ID NO: 516, SEQ ID NO: 515 paired with SEQ IDNO: 505, and SEQ ID NO: 506 paired with SEQ ID NO: 516). The differences between these SDC candidate molecules are the linker between the PD-L1 -binding domain and the EGF variant domain, and also the variations in valence of EGFR or PD-L1 binding capabilities. Ovals marked “N” and “C” represent homodimerization domains of SEQ ID NO: 535; ovals containing a convex or concave semicircular notch and marked “N” and “C” represent heterodimerization domains of SEQ ID NO: 536 and SEQ ID NO: 537, respectively; circles represent PD-Ll-binding variants of SEQ ID NO: 187; 7-ponited stars represent EGF variants of SEQ ID NO: 494; curved and wavey lines represent linkers of SEQ ID NO: 223, SEQ ID NO: 540, SEQ ID NO: 541, or SEQ ID NO: 538.

[0147] FIG. 17A-B illustrates the extent and duration of elimination of surface EGFR in the cells by selective depletion complexes (SDC) under different SDC concentrations and treatment regimens.

[0148] FIG. 17A illustrates the extent of elimination of surface EGFR in cells by a selective depletion complex (SDC) of SEQ ID NO: 495 evaluated by analyzing samples by flow cytometry to measure the surface EGFR under different SDC concentrations. A549 cancer cells were grown until approximately 30% confluence in RPMI media with 10% FBS and antibiotic / antimycotic supplementation. Cells were then left untreated (“Untreated”) or dosed with the SDC comprising SEQ ID NO: 495 at 2 nM, 10 nM, 50 nM, or 200 nM and incubated for a further 24 hours (untreated or in the presence of the SDC). After 24 hours, cells were rinsed, collected, stained with an antibody vs EGFR and a fluorescent co-stain, and subjected to flow cytometry to quantitate fluorescent signal corresponding to the amount of surface EGFR per cell in the population. The data for each population were averaged and plotted relative to the untreated population (Surface EGFR (% of untreated ± 95% confidence interval (CI)).

[0149] FIG. 17B illustrates the extent and duration of elimination of surface EGFR in the cells by a selective depletion complex (SDC) evaluated by analyzing samples by flow cytometry to measure the surface EGFR under different SDC treatment regimens. A549 cancer cells were grown until approximately 20% confluence in RPMI media with 10% FBS and antibiotic / antimycotic supplementation. Cells were then left untreated or dosed with 10 nM of the SDC comprising SEQ ID NO: 495 and incubated for a further 24 hours. After 24 hours, the media in the wells was replaced with fresh media. Three treatments were then performed and analyzed over the next 24 hours (summarized in the table in FIG. 17B): (1) the cells that had been subjected to treatment with the SDC comprising SEQ ID NO: 495 for 24 hours were then left untreated for an additional 24 hours (referred to as the “Withdrawal” sample, as it wassubjected to 24 hour treatment with the SDC followed by a 24 hour in untreated media (that is, treatment withdrawal of the SDC)), (2) previously-untreated cells were dosed with 10 nM of the SDC comprising SEQ ID NO: 495 for 24 hours (referred to as the “24 hr” sample, as its surface EGFR was analyzed immediately after a 24 hour SDC treatment, and (3) a third sample (“Untreated”) that was left untreated during the previous 24 hours was still left untreated for an additional 24 hours. After this second 24 hour incubation, cells in all samples were rinsed, collected, stained with an antibody vs EGFR and a fluorescent co-stain, and subjected to flow cytometry to quantitate fluorescent signal corresponding to the amount of surface EGFR per cell in the population. The data for each population were averaged and plotted relative to the untreated population (Surface EGFR (% of untreated ± 95% confidence interval (CI)).

[0150] FIG. 18 illustrates the uptake of soluble EGFR into the cells evaluated by analyzing samples by flow cytometry to measure the soluble EGFR uptake under different SDC treatment regimens. Hl 975 cancer cells were grown until approximately 40% confluence in RPMI media with 10% FBS and antibiotic / antimycotic supplementation. After this point, cells underwent one of three treatments to assess uptake of soluble biotinylated EGFRvIII (sEGFR; SEQ ID NO: 527). Each of the three treatments are summarized in the table in FIG. 18. Treatment (1), referred to as “No SDC”, incubated the cells for two hours in media with 20 nM sEGFR and 20 nM unlabeled monovalent streptavidin. After this 2 hour treatment, cells are rinsed twice with PBS and then incubated for 24 hours in media with 10 nM sEGFR and 10 nM fluorescent streptavidin. Treatment (2), referred to as “SEQ ID NO: 495 Pre-treat”, incubated the cells for 2 hours in media with 20 nM sEGFR, 20 nM unlabeled monovalent strepatavidin, and 5 nM of the SDC comprising SEQ ID NO: 495. This treatment first exposed cells to sEGFR-saturated SDC molecules. After this 2 hour treatment, cells were rinsed twice with PBS and then incubated for 24 hours with 10 nM sEGFR and 10 nM fluorescent streptavidin to assess entry of the fluorescent sEGFR into the cells. Treatment (3), referred to as “SEQ ID NO: 495 24 hr”, was the same treatment as “SEQ ID NO: 495 Pre-treat”, except the second incubation (24 hours with 10 nM sEGFR and 10 nM fluorescent streptavidin) also included 5 nM SDC comprising SEQ ID NO: 495.

[0151] FIG. 19 illustrates the assessment of disruption of EGFR signaling by an EGF -variant selective depletion complex (SDC) as well as the absence of EGFR activation by molecules that incorporate the EGF variant (designed to bind EGFR but not activate it) analyzed by Western blot. A549 cancer cells were grown until approximately 30% confluence in RPMI media with 10% FBS and antibiotic / antimycotic supplementation. Cells were left untreated or dosed witheither the SDC comprising SEQ ID NO: 495 or the control comprising SEQ ID NO: 498 at 10 nM and incubated for a further 24 hours. After 24 hours, and without exchanging media, cells were either left as-is or exposed to 50 ng / mL EGF (SEQ ID NO: 317). Cells were then lysed in buffer containing protease / phosphatase inhibitors. Lysate protein content was quantitated by bicinchoninic acid (BCA) assay and was ran on an SDS-PAGE gel prior to electrophoretic transfer to a polyvinylidine difluoride (PVDF) membrane and subsequent blocking and blotting with antibodies against actin, total EGFR, or phospho-Y1068 (pY1068) EGFR, the latter being a marker for ligand-activated EGFR. A fluorescent secondary antibody provides signal that can be read on a scanner.DETAILED DESCRIPTION

[0152] Described herein are compositions and methods for selective depletion of an EGFR target molecule using cellular endocytic pathways (e.g., transferrin receptor-mediated endocytosis). Epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase involved in cell signaling. EGFR signaling regulates cell growth and survival, and upregulation of EGFR is implicated in many types of cancer. EGFR proteins are regularly cycled through production, use, and degradation, and their degradation is typically within the endosomal-lysosomal pathway. In this pathway, endocytic vesicles containing material taken up from extracellular space as well as embedded membrane proteins become acidified and fuse with or enter lysosomes containing enzymes that degrade such proteins. Selective removal of certain cell proteins such as EGFR proteins, either from circulation or disease-associated tissues, such as by removing the proteins from the cell surface or from soluble forms, optionally with selective delivery to the lysosome, can be used to treat disease conditions, including diseases resulting from over-expression or mutations in EGFR. Alternatively or in addition, the peptide complexes described herein for selective depletion of a target molecule, also referred to as “selective depletion complexes”, can be used to deliver an administered therapeutic drug to a cell or an endosomal or lysosomal compartment in cells or tissues with increased EGFR expression, for example to treat lysosomal storage diseases like Gaucher’s Disease (deficiency of glucocerebrosidase) or Pompe Disease (deficiency of a-glucosidase) or a disease associated with EGFR (e.g., a cancer). A therapeutic molecule (e.g., a lysosomal enzyme for an enzyme replacement therapy or a chemotherapeutic agent) can be administered with a selective depletion complex comprising a target-binding peptide that binds the therapeutic molecule, thereby delivering the therapeutic molecule to the endosome or lysosome. In some embodiments, a selective depletion construct can function as aselective delivery complex and facilitate delivery of active enzymes to an endosome or lysosome. For example, a lysosomal enzyme can be delivered using a selective depletion complex and can retain enzymatic activity in the endosome or lysosome. Administration of a lysosomal enzyme in combination with a selective depletion complex comprising a targetbinding peptide that binds the lysosomal enzyme can increase the therapeutic response per dose of enzyme administered relative to administration of the lysosomal enzyme alone. For either selective depletion of target proteins from a cell, or delivery of lysosomal proteins, lysosomal delivery could be accomplished by taking advantage of existing protein uptake and recycling mechanisms, and engineering of pH-dependent binding domains into target-binding molecules.

[0153] A unique example of an endocytic pathway that can be used for selective depletion of target molecules is via transferrin receptor (TfR) internalization and trafficking, which is normally used for transferrin recycling via transferrin receptor (TfR) for iron delivery to cells and tissues. Transferrin is known as a serum chaperone for iron ions destined for redox sensitive intracellular enzymes. Iron-loaded transferrin (holo-transferrin) delivers iron to cells via specific binding to TfR, which is then trafficked to endosomes, where the pH is reduced by native proton pumps. Under acidic conditions, transferrin loses its iron binding affinity, releasing iron inside the cell, but maintains its TfR-binding affinity. The TfR:transferrin complex is natively recycled back to the cell surface, exposing transferrin to neutral pH conditions. Transferrin unbound by iron (apo-transferrin) no longer has TfR affinity under neutral pH conditions at the cell surface and is released back into circulation to pick up more iron, and repeat the process, in what is essentially a catalytic process for iron delivery to cells.

[0154] The compositions and methods of this disclosure exploit the transferrin receptor endocytic and recycling pathways to selectively deplete target molecules (e.g., EGFR) from the cell surface, disrupt EGFR multimerization, and / or to selectively deplete and deliver target molecules (e.g., EGFR) to endocytic vesicles for lysosomal degradation. The compositions and methods of this disclosure can be used to selectively deplete or degrade EGFR target receptor proteins or over-expressed in disease via this pathway. As a result of depleting target molecules from the cell surface, and / or through lysosomal degradation of the target receptors (e.g., EGFR), the compositions and methods described effectively reduce, diminish, eliminate, or deplete the target receptors from the cell surface which has many applications in medicine as described herein. Selective depletion complexes of the present disclosure comprising a TfR-binding peptide (e.g., a TfR-binding cystine-dense peptide) coupled to a target-binding peptide (e.g., a target-binding EGF variant) can recruit a target molecule to the TfR by binding to both the TfR(via the TfR-binding peptide) and to the target molecule (via the target-binding peptide). Upon endocytosis, the TfR can carry the selective depletion complex and the target molecule into the endocytic vesicle. In some embodiments, the TfR-binding peptide of the selective depletion complex can have high affinity for TfR at extracellular pH (about pH 7.4) to maturing endosomal pH (about pH 5.5), inclusive. The TfR-binding peptide can maintain its affinity for TfR upon internalization and as the endosomal compartment acidifies. The target-binding peptide of the selective depletion complex can have higher affinity for the target molecule at extracellular pH and lower affinity for the target molecule at a lower endosomal pH. Inside the endocytic vesicle, the selective depletion complex can remain bound to TfR and release the target molecule upon acidification of the endosome. Once the target molecule is released, the selective depletion complex can remain bound to TfR while TfR is recycled to the cell surface to be reloaded with another target molecule, and the target molecule can remain in the endosome and in some embodiments the target molecule is further delivered to a lysosome and degraded. In some embodiments, the TfR-binding peptide of the selective depletion complex can have higher affinity for TfR at extracellular pH and lower affinity for the target molecule at a lower endosomal pH. Inside the endocytic vesicle, the selective depletion complex can release from TfR upon acidification of the endosome. In some embodiments, the dissociation rate of the selective depletion complex from the target while in the endosome is faster than the rate of endosome recycling back to the cell surface. In this case, the target may be released from the selective depletion complex regardless of any or no variation in affinity to the target as a function of pH.

[0155] The methods of the present disclosure can comprise contacting a cell (e.g., a cell expressing TfR) with a selective depletion complex (e.g., a molecule comprising a TfR-binding peptide and a target-binding peptide). The selective depletion complex can recruit target molecules into endocytic vesicles via transferrin receptor-mediated (TfR-mediated) endocytosis. The target molecule can be released in the endocytic vesicle, and it may be further delivered to the lysosome and degraded. The selective depletion complex can remain bound to the TfR and can remain bound to TfR as TfR is recycled to the cell surface. Such methods can be used to deplete a target molecule (e.g., EGFR), such as a molecule associated with a disease or a condition (e.g., associated with cancer). For example, the methods of the present disclosure can be used to selectively deplete EGFR that is over-expressed, contains a disease-associated mutation (e.g., a mutation causing constitutive activity, resistance to treatment, or dominant negative activity), or accumulates in a disease or a condition. It is understood that selectivedepletion of a target molecule includes the depletion of the selected target from the cell surface or soluble target in circulation, each of which could result in a therapeutic effect of the selective depletion complex.

[0156] In some embodiments, the presently described selective depletion complex can comprise peptide conjugates, peptide complexes, peptide constructs, fusion peptides, or fusion molecules such as linked by chemical conjugation of any molecule type, such as small molecules, peptides, or proteins, or by recombinant fusions of peptides or proteins, respectively (e.g., a peptide construct or a peptide complex). The terms “fusion peptide” and “peptide fusion” are used interchangeably herein. In some embodiments, the peptide constructs or peptide complexes can be produced biologically or synthetically. Thus, in some cases, a selective depletion complex can comprise a TfR-binding peptide domain linked to another molecule or group of molecules such as small molecules, peptides, or proteins or other macromolecules such as nanoparticles.

[0157] In some embodiments, the presently described selective depletion complexes can be peptide complexes comprising one or more TfR-binding peptides as described herein conjugated to, linked to, or fused to. or complexed with one or more target-binding peptides (e.g., one or more EGFR-binding peptides), one or more active agents (e.g., therapeutic agents, detectable agents, or combinations thereof), or combinations thereof. Selective depletion complexes as described herein can include chemical conjugates and recombinant fusion molecules. In some cases, a chemical conjugate can comprise a TfR-binding peptide as described herein that is chemically conjugated to or linked to another peptide (e.g., an EGFR target-binding peptide), a molecule, an agent, or a combination thereof. Molecules can include small molecules, peptides, polypeptides, proteins, or other macromolecules (e.g., nanoparticles) and polymers (e.g., nucleic acids, polylysine, or polyethylene glycol). In some cases, a TfR-binding peptide of the present disclosure is conjugated to another peptide or a molecule via a linker. Linker moieties can include cleavable (e.g., pH sensitive or enzyme-labile linkers) or stable linkers. In some embodiments, a peptide complex is a fusion molecule (e.g., a fusion peptide or fusion protein) that can be recombinantly expressed, and wherein the fusion molecule can comprise one or more TfR-binding peptides fused to one or more other molecules peptides, polypeptides, proteins, or other macromolecules that can be recombinantly expressed.

[0158] The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can have a therapeutic effect at a lower dose or a longer lasting therapeutic effect as compared to lysosomal delivery molecules that are degraded and not recycled to the cell surface. Rather than being degraded in thelysosome, the selective depletion complexes of this disclosure can be recycled back to the cell surface to “reload” with the target molecule, meaning that the potential for one selective depletion complex of this disclosure can drive the degradation of multiple target molecules with a potentially catalytic effect. The selective depletion complex may also continue to have depletion activity even when the selective depletion complex is no longer present in serum but is present on or in a cell. A lysosomal delivery molecule that is not recycled to the cell surface can itself be degraded or can accumulate in the lysosome without being re-used or “reloaded”. The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can have a larger (e.g., longer or wider) therapeutic window (i.e., the dosage above which a therapeutic pharmacodynamic response is observed but below which toxicity is observed) or a higher potency or a longer duration of effect as compared to lysosomal delivery molecules that are not recycled to the cell surface. The therapeutic window of a drug (e.g., a selective depletion complex of the present disclosure) is the dose range at which the drug is effective without having unacceptable toxic effects. The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can be used with less risk of toxicity. The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can be used (e.g., administered) at lower molar dosage than alternative therapies (e.g., lysosomal delivery molecules) that are not recycled to the cell surface. Because of the selectivity and re-usable nature of the selective depletion complexes of this disclosure in the cell, as therapeutic agents they are advantageously not depleted as rapidly as non-recyclable delivery compositions targeted to lysosomes which are depleted as they are used. The selective depletion complexes can be more concentrated on tissues that express the receptor, such as TfR, or the target (such as EGFR) at higher levels than other tissues; as such, the selective depletion complexes and their effects can be more concentrated on diseased tissues that overexpress the applicable receptor or the target or both compared to normal or healthy tissues. Many solid tumors express TfR at high levels and thus selective depletion complexes that bind TfR may be more concentrated on solid tumor tissues, concentrating their depletion effect on the tumor tissues. Many human tumors express EGFR at high levels (e.g., the lung, head and neck, colon, pancreas, breast, ovary, bladder and kidney, and in glioma) and thus selective depletion complexes that bind EGFR may be more concentrated on such tumor tissues, concentrating their depletion effect on the tumor tissues. Moreover, because of the selectivity and recycling aspect of the selective depletion complexes of this disclosure (e.g., selectivedepletion complexes comprising a receptor-binding peptide and an EGFR target-binding peptide), as therapeutic agents they are advantageously less toxic than non-selective therapeutic agents. This is particularly advantageous for applications in cancer, where therapeutic agents can be non-selective and highly toxic and exhibit detrimental side effects on normal cells, organs, and tissues, or require lower than effective therapeutic doses less able to reduce, cure, ablate disease.

[0159] In other embodiments, a selective depletion complex can bind the PD-L1 receptor, rather than TfR, to enable uptake of the target and recycling of the selective depletion complex. PD-L1 is expressed by cells such as solid tumor cells, pancreatic beta cells, and certain cells of the immune system. PD-L1 can be taken up by endosomes, and then recycled back up to the cell surface. PD-L1 can co-localize with CMTM6 in recycling endosomes, where CMTM6 prevents PD-L1 from being targeted for lysosomal degradation. A selective depletion complex can bind to PD-L1 and to a target, such as EGFR, that is targeted for depletion. When PD-L1 is endocytosed into the cell, the target can be depleted from the cell surface. The selective depletion complex may recycle back up to the cell surface along with PD-L1 and the target may remain in the endosome and may proceed to the lysosome and be degraded. Selective depletion complexes that use PD-L1 for uptake have the potential to function in all the ways that selective depletion complexes that use TfR for uptake may function. Use of PD-L1 as a selective depletion complex receptor may permit selective targeting of selective depletion complexes to solid tumor cells, as PD-L1 is expressed in high levels on some solid tumors but is otherwise not commonly expressed in adult tissues except for some specific cell populations (e.g., certain cells of the immune system or pancreatic beta cells).

[0160] The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can have less immunogenicity than an alternative therapy (e.g., a lysosomal delivery molecule) that contains sugars, glycans, polymers containing sugar-like molecules, or other derivatives. A selective depletion complex of this disclosure can have less immunogenicity than an alternative therapy (e.g., a lysosomal delivery molecule) that targets the mannose-6-phosphate receptor, folate receptor, or the asialoglycoprotein receptor (ASGPR). A selective depletion complex of this disclosure can be manufactured by a single recombinant expression and can have improved manufacturing yield, purity, cost, or manufacturing time than a molecule that has multiple synthetic steps to generate a ligand for mannose-6-phosphate receptor, folate receptor, or the asialoglycoprotein receptor (ASGPR). A selective depletion complex of this disclosure can have a greater therapeutic effector a lower therapeutic dose due to the ability to design the linker for maximal ability to bind for the receptor (e.g., TfR) and the target molecule (e.g., EGFR) at the same time, including of the target molecule is bound in the cell surface. The TfR-binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides of this disclosure can have fewer epitopes to trigger an adaptive immune response, resulting in reduced immunogenicity as compared to TfR-binding antibody-based therapeutics. The TfR-binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides of this disclosure can exhibit more facile and less disruptive incorporation of active agents into protein fusion complexes as compared to TfR-binding antibody-based therapeutics. The TfR-binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides of this disclosure can have a smaller surface area, resulting in lower risk for off-target-binding, as compared to TfR-binding antibody-based therapeutics. The TfR- binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides of this disclosure can be formulated at a higher molar concentration than TfR-binding antibody-based therapeutics due to their lower molecule weight, lower hydrodynamic radius, or lower molar solution viscosity.

[0161] The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can, in some embodiments, cross the blood brain barrier. The selective depletion complexes of this disclosure may be able to transcytose across endothelial cells of the blood-brain barrier and thereby reach the central nervous system (CNS), brain, and associated cells. By this mechanism, the selective depletion complexes of this disclosure may be able to deplete targets that are in the CNS including tumors that are present in the brain. Because the blood-brain barrier excludes the great majority of molecules from the brain, alternative therapies (such as lysosomal delivery methods targeting receptors not known to facilitate blood brain barrier transcytosis or ubiquitin-proteosome system (UPS) targeting therapies) may be unable to reach targets in the CNS.

[0162] The selective depletion complexes of this disclosure (e.g., complexes comprising a receptor-binding peptide and an EGFR target-binding peptide) can function with a wide range of linker lengths. Selective depletion complexes of this disclosure can have a range of short to long linkers between the receptor-binding portion and the target binding portion of the SDC. and the structure of the SDC does not necessarily require close association of the target and the receptor in order cause depletion of the target so long as it forms a ternary complex with the receptor and target that is endocytosed when the receptor is endocytosed. Alternative approaches, such as inducing a ternary complex between a cell surface E3 ligases and a cell surface target to increaseor facilitate ubiquitination of the cell surface target and thereby leading to depletion of the target, typically require the target and the cell surface E3 ligase to come in close physical proximity in a suitable orientation in order for the target to be ubiquitinated. Of the dozens of cell surface E3 ligases, most are only expressed on some tissues and may not be expressed at high levels on solid tumors. Many targets (e.g., 55% of cell surface targets) may not have a suitable ubiquitination site accessible by the ligase domain. Because the ubiquitination domain of cell surface E3 ligases is on the intracellular portion of the ligase, it is unable to ubiquitinate extracellular soluble targets and thus the use of E3 ligases does not facilitate ubiquitin-mediated depletion of soluble targets for depletion. The selective depletion complexes of this disclosure do not require such proximity constraints and have been demonstrated to promote the cellular uptake of soluble targets (e.g., soluble EGFRvIII) and hence are an improvement on such prior systems.

[0163] In some embodiments, the TfR-binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides of this disclosure exhibit lower on- target toxicity than an anti-TfR antibody or other therapeutic agents when administered to a subject at the same molar dose or at a similarly effective dose. In some embodiments, the TfR-binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides exhibit lower off-target toxicity than an antibody or other therapeutic agent when administered to a subject at the same molar dose or a similarly effective dose. For example, the TfR-binding peptides, TfR-binding peptide conjugates, or TfR- binding fusion peptides of this disclosure can be administered to a subject at about 1-fold, 2- fold, 3-fold, 4-fold, 5-fold, 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold higher molar dose than an antibody while providing similar or lower observed toxicity. In some embodiments, the TfR-binding peptides, TfR-binding peptide conjugates, or TfR-binding fusion peptides of this disclosure exhibit higher efficacy than an anti-TfR antibody or other therapeutic agent when administered to a subject at the same dose by weight as the anti-TfR antibody or other therapeutic agent. The TfR-binding peptides of the present disclosure, when fused to a half-life extending moiety (e.g., Fc, SA21, PEG), can be delivered at even lower doses while preserving activity and efficacy and, thus, is far superior to administering an anti-TfR antibody or other therapeutic agent.

[0164] In some embodiments, the present disclosure provides peptides (e.g., CDPs, knotted peptides, or hitchins), chemical conjugates (e.g., comprising one or more TfR-binding peptides and one or more active agents), or recombinantly expressed fusion molecules (e.g., comprising one or more TfR-binding peptides and one or more active agents) that bind to TfR, PD-L1, otherreceptors, or the target. The TfR-binding peptides can be cystine-dense peptides (CDPs). The terms “peptides”, “miniproteins”, “proteins”, “CDPs”, “TfR-binding peptides,” “TfR-binding CDPs,” “TfR-binding peptides,” and “engineered TfR-binding peptides” are used interchangeably herein. The binding of peptides described in the present disclosure to TfR can facilitate transcytosis of the selective depletion complex, peptide, peptide complex or peptide construct (e.g., fusion protein, or peptide conjugated to, linked to, complexed with, or fused to an agent) across a cell barrier (e.g., the BBB). The binding of peptides described in the present disclosure to TfR can facilitate endocytosis of the selective depletion complex, peptide, or peptide complex in any cell that expresses TfR, or in cell that express TfR at higher levels, including some cancer cells, hepatic cells, spleen cells, and bone marrow cells. Also disclosed herein is the use of a mammalian surface display screening platform to screen a diverse library of CDPs and identify CDPs that specifically bind to human TfR. Such identified peptides can be modified to improve binding to TfR and used in selective depletion complexes as the peptide or peptide complex that binds TfR and is recycled to the cell surface (e.g., the pH-independent TFR-binding CDP). Also disclosed herein is the use of a mammalian surface display screening platform to screen a diverse library of CDPs and identify CDPs that specifically bind to a target molecule that is desired to be degraded. Such identified peptides can be optimized for binding to a selected target molecule and used in selective depletion complexes as the peptide or peptide complex that binds such selected target molecule and is released in the endosome for degradation within the cell (e.g., the pH-dependent target-binding EGF variant as shown in FIG. 14). Further affinity maturation can be subsequently implemented to produce an allelic series of TfR-binding CDPs or target-binding CDPs as appropriate with varying affinities. In some embodiments, TfR-binding CDPs or target-binding CDPs are identified, and binding can be determined by crystallography or other methods. Peptides of the present disclosure can have cross-reactivity across species. For example, the peptides disclosed herein, in some cases, bind to human and murine TfR. Peptides disclosed herein can accumulate in the CNS and can penetrated the BBB via engagement of the TfR, following intravenous administration. Disclosed herein are TfR-binding CDPs for use as therapeutic delivery agents in oncology, autoimmune disease, acute and chronic neurodegeneration, and pain management. Delivery of active or pharmaceutical agents via TfR-binding CDP can be advantageous over conventional anti-TfR antibodies due to simpler manufacturing (peptides can be made via biologic or synthetic means), improved stability, improved therapeutic window (e.g., a larger, longer, or wider therapeutic window), and smaller size (less potential for steric hindrance of cargo activity). Thus, themethods and compositions of the present disclosure can provide a solution to the problem of effectively transporting cargo molecules (e.g., therapeutic and / or diagnostic small molecules, peptides or proteins) into the CNS (e.g., the brain). For example, the peptides of the present disclosure aid in drug delivery to tumors located in the brain.

[0165] In some embodiments of the present disclosure, a diverse library of CDPs, knotted peptides, hitchins, or peptides derived from knotted peptides or hitchins can be used in combination with a mammalian surface display screening platform is used to identify peptides that specifically bind to human TfR or PD-L1 or other receptors desired for recycling or to a target molecule desired for degradation. (See e.g., Crook et al. (2017) Mammalian display screening of diverse cystine-dense peptides for difficult to drug targets. Nat Commun 8:2244). In some embodiments, a diverse library of CDPs, knotted peptides, hitchins, or peptides derived from knotted peptides or hitchins is mutagenized from endogenous peptide sequences to provide novel peptide sequences. Once TfR-binding or target-binding peptides have been identified, affinity maturation (e.g., site-saturation mutagenesis) can be performed to produce an allelic series of binders with varying (e.g., improved) affinities for TfR or a target molecule. These techniques can be used in combination with various other analytical methods (e.g., crystallography or spectroscopy) in order to determine the nature of peptide-receptor interaction (e.g., critical amino acid residues for receptor binding etc.). In some cases, the peptides of the present disclosure are developed to bind human TfR.

[0166] In some embodiments, the engineered peptides of the present disclosure (e.g., histidine- containing or histidine-enriched target-binding peptides) can have a high target-binding affinity at physiologic extracellular pH (e.g., a pH from about pH 7.2 to about pH 7.5, a pH of from about pH 6.5 to about 7.5, or a pH of from about pH 6.5 to about pH 6.9) but a significantly reduced binding affinity at lower pH levels such as endosomal pH of about 6.5, about 6.0, about 5.8, or about 5.5. Extracellular pH can be, for example pH 7.4. Extracellular pH can also be lower, including in the tumor microenvironment, such as pH 7.2, 7.0, or 6.8. In some embodiments, for example in a tumor environment, extracellular pH can be from about pH 6.5 to about pH 6.9. Upon endocytosis, the endosome undergoes a decrease in pH. Endosomal pH can decrease by the action of proton pumps or by merging with other vesicles with lower pH. The pH can decrease to 7.0, and then to 6.5, and then to 6.0, and then to 5.8, and then to 5.5 or lower. Some endosomes are called early endosomes and can have a pH around 6.5. Some of these endosomes become recycling endosomes. Some endosomes are called late endosomes and can have a pH around 5.5. Some endosomes become or merge with lysosomes, where the pHcan be 4.5. Enzymes and other factors in the lysosome can cause degradation of the contents of the lysosome. In some embodiments, the target-binding peptides release in the endosome at about pH 7.4, pH 7.3, pH 7.2, pH 7.1, pH 7.0, pH 6.9, pH 6.8, pH 6.7, pH 6.6, pH 6.5, pH 6.4, pH 6.3, pH 6.2, pH 6.1, pH 6.0, pH 5.9, pH 5.8, pH 5.7, pH 5.6, pH 5.5, pH 5.4, pH 5.3, pH 5.2, pH 5.1, pH 5.0, pH 4.9, pH 4.8, pH 4.7, pH 4.6, pH 4.5, or lower. In some embodiments, the target-binding peptide may release at any point during the endosomal maturation process upon a decrease in pH following endocytosis. In some cases, histidine scans and comparative binding experiments can be performed to develop and screen for such peptides. In some embodiments, an amino acid residue in a peptide of the present disclosure is substituted with a different amino acid residue to alter a pH-dependent binding affinity to the target molecule or to TfR or other receptors. The amino acid substitution can increase a binding affinity at low pH, increase a binding affinity at high pH, decrease a binding affinity at low pH, decrease a binding affinity at high pH, or a combination thereof. For example, a peptide that has high affinity to TfR and used in selective depletion complexes as the peptide or peptide complex that binds TfR for recycling to the cell surface can be a pH-independent TfR-binding peptide (e.g., a pH-independent TfR- binding CDP) such that it is not released in the endosome. In some embodiments, the TfR- binding peptide can remain bound to TfR as the ionic strength of the endosomal compartment increases upon acidification of the endosome. In some embodiments the TfR-binding peptides are stable at endosomal pH, and do not release in the endosome for example under acidic conditions, such as pH 6.9, pH 6.8, pH 6.7, pH 6.6, pH 6.5, pH 6.4, pH 6.3, pH6.2, pH 6.1, pH 6.0, pH 5.9, pH 5.8, pH 5.7, pH 5.6, pH 5.5, pH 5.4, pH 5.3, pH 5.2, pH 5.1, pH 5.0, pH 4.9, pH 4.8, pH 4.7, pH 4.6, pH 4.5, or lower. Conversely, a peptide that has high affinity for binding to a selected target molecule and used in selective depletion complexes as the peptide or peptide complex that binds such selected target molecule and is released in the endosome for degradation within the cell can be a pH-dependent target-binding CDP such that it is released in the endosome. In some embodiments, a target-binding peptide can release the target molecule as the ionic strength of the endosomal compartment increases upon acidification of the endosome. In some embodiments the target-binding peptides are less stable at endosomal pH, and release wholly or in part in the endosome for example under acidic conditions, such as pH 7.4, pH 7.3, pH 7.2, pH 7.1, pH 7.0, pH 6.9, pH 6.8, pH 6.7, pH 6.6, pH 6.5, pH 6.4, pH 6.3, pH 6.2, pH 6.1, pH 6.0, pH 5.9, pH 5.8, pH 5.7, pH 5.6, pH 5.5, pH 5.4, pH 5.3, pH 5.2, pH 5.1, pH 5.0, pH 4.9, pH 4.8, pH 4.7, pH 4.6, pH 4.5, or lower. In some cases, the TfR-binding peptides of the present disclosure can be optimized for improved intra- vesicular (e.g., intra-endosomal) function whileretaining high TfR binding capabilities. In some embodiments, the target-binding peptide may not release at any point during the endosomal maturation process, for example using designs that are not pH-sensitive (i.e., are pH-independent) to release the target molecule in the endosome or lysosome, but the selective depletion complex still results in selective depletion of the target molecule from the cell surface or soluble target molecule in circulation. Exemplary TfR-binding peptides of the present disclosure are shown in TABLE 1 with amino acid sequences comprising SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64.

[0167] Described herein are, in some embodiments, peptides and peptide complexes and methods of screening for peptides and peptide complexes that bind to a protein or molecule of interest, such as TfR, or bind to a target molecule for depletion, or both. Compared to wild type or endogenous molecules such as transferrin, the methods and compositions as described herein can provide peptides with improved TfR-binding capabilities, or peptides that exhibit improved transport capabilities across the BBB, or any combination thereof. In some cases, the presently described peptides efficiently transport cargo molecules (e.g., target-binding molecules) across endothelial cell layers (e.g., the BBB) or epithelial layers. In some embodiments, the TfR- binding peptides of the present disclosure bind to a TfR and promote vesicular transcytosis. In some cases, the TfR-binding peptides of the present disclosure bind to a cell that overexpress a TfR (e.g., a cancer cell) and promotes uptake of the peptide by the cell. In some aspects, a TfR binding peptide or peptide complexes as described herein promotes vesicular transcytosis and uptake by a TfR-overexpressing cell such as a cancer, or a combination thereof. In some cases, the TfR-binding peptides of the present disclosure facilitate TfR-mediated endocytosis of a selective depletion complex and a target molecule.

[0168] The TfR-binding peptides of the present disclosure can bind TfR of different species including human, monkey, mouse, and rat TfR. In some cases, variations or mutations in any of the amino acid residues of a TfR-binding peptide can influence cross-reactivity. In some cases, variations or mutations in any of the amino acid residues of a TfR-binding peptide that interact with the bindings site of TfR can influence cross-reactivity.

[0169] Described herein are peptides, including, but not limited to, designed or engineered peptides, recombinant peptides, and cystine-dense peptides (CDPs) / small disulfide-knotted peptides (e.g., knotted peptides, hitchins, and peptides derived therefrom), that can be large enough to carry a cargo molecule while retaining the ability to bind a target protein with highaffinity (e.g., TfR), but yet small enough to access cellular tissues, such as the center of cell agglomerates (e.g., solid tumors). In some cases, the peptides as described herein carry cargo molecules across the BBB into the CNS (e.g., the parenchyma) via vascular transcytosis. In some cases, the transcytosis is TfR-mediated.

[0170] Further described herein are methods and compositions for determining the nature of peptide-receptor interactions (e.g., using X-ray crystallography) as well as their pharmacodynamic and pharmacokinetic properties in vivo, including accumulation in the CNS (e.g., brain), or other affected organs and tissues. Some of the peptides described herein have the ability to target molecule and accumulate in tumor cells. In some cases, the tumor cells overexpress TfR, EGFR, or both. In some aspects, the peptides of the present disclosure have high in vivo stabilities, e.g., high protease stability, high tolerability of reducing agents such as glutathione (GSH), and tolerate elevated temperatures (e.g., up to 95 °C).

[0171] The present disclosure provides, in some embodiments, a peptide or protein design approach based on the 3D protein or receptor structure for identifying peptides or proteins capable of binding such receptor. In some cases, the receptor is a transferrin receptor.

[0172] As used herein, the abbreviations for the natural L-enantiomeric amino acids are conventional and are as follows: alanine (A, Ala); arginine (R, Arg); asparagine (N, Asn); aspartic acid (D, Asp); cysteine (C, Cys); glutamic acid (E, Glu); glutamine (Q, Gin); glycine (G, Gly); histidine (H, His); isoleucine (I, He); leucine (L, Leu); lysine (K, Lys); methionine (M, Met); phenylalanine (F, Phe); proline (P, Pro); serine (S, Ser); threonine (T, Thr); tryptophan (W, Trp); tyrosine (Y, Tyr); valine (V, Vai). Typically, Xaa can indicate any amino acid. In some embodiments, X can be asparagine (N), glutamine (Q), histidine (H), lysine (K), or arginine (R).

[0173] Some embodiments of the disclosure contemplate D-amino acid residues of any standard or non-standard amino acid or analogue thereof. When an amino acid sequence is represented as a series of three-letter or one-letter amino acid abbreviations, the left-hand direction is the amino terminal direction and the right-hand direction is the carboxy terminal direction, in accordance with standard usage and convention.

[0174] The terms “peptide”, “polypeptide”, “miniprotein”, “protein”, “hitchin”, “cystine-dense peptide”, “knotted peptides” or “CDP” can be used interchangeably herein to refer to a polymer of amino acid residues. In various embodiments, “peptides”, “polypeptides”, and “proteins” can be chains of amino acids whose alpha carbons are linked through peptide bonds. The terminal amino acid at one end of the chain (e.g., amino terminal, or N-terminal) therefore can have a freeamino group, while the terminal amino acid at the other end of the chain (e.g., carboxy terminal, or C-terminal) can have a free carboxyl group. As used herein, the term “amino terminus” (e.g., abbreviated N-terminus) can refer to the free a-amino group on an amino acid at the amino terminal of a peptide or to the a-amino group (e.g., imino group when participating in a peptide bond) of an amino acid at any other location within the peptide. Similarly, the term “carboxy terminus” can refer to the free carboxyl group on the carboxy terminus of a peptide or the carboxyl group of an amino acid at any other location within the peptide. Peptides also include essentially any polyamino acid including, but not limited to, peptide mimetics such as amino acids joined by an ether or thioether as opposed to an amide bond.

[0175] As used herein, the term “peptide construct” can refer to a molecule comprising one or more peptides of the present disclosure that can be conjugated to, linked to (including by complexation), or fused to one or more peptides or cargo molecules. In some cases, cargo molecules are active agents. The term “active agent” can refer to any molecule, e.g., any molecule that is capable of eliciting a biological effect and / or a physical effect (e.g., emission of radiation) which can allow the localization, detection, or visualization of the respective peptide construct. In various embodiments, the term “active agent” refers to a therapeutic and / or diagnostic agent. A peptide construct of the present disclosure can comprise a TfR-b inding peptide that is linked to one or more active agents via one or more linker moieties (e.g., cleavable or stable linker) as described herein.

[0176] As used herein, the term “peptide complex” can refer to one or more peptides of the present disclosure that are fused, linked, conjugated, or otherwise connected to form a complex. In some cases, the one or more peptides can comprise a TfR-b inding peptide, a target-binding peptide, a half-life modifying peptide, a peptide that modifies pharmacodynamics and / or pharmacokinetic properties, or combinations thereof. For example, a peptide complex comprising a TfR-b inding peptide and a target-binding peptide can be referred to herein as a selective depletion complex.

[0177] As used herein, the terms “comprising” and “having” can be used interchangeably. For example, the terms “a peptide comprising an amino acid sequence of SEQ ID NO: 32” and “a peptide having an amino acid sequence of SEQ ID NO: 32” can be used interchangeably.

[0178] As used herein, and unless otherwise stated, the term “TfR” or “transferrin receptor” is a class of protein used herein and can refer to a transferrin receptor from any species (e.g., human or murine TfR or any human or non-human animal TfR). In some cases, and as used herein, the term “TfR” or “transferrin receptor” refers to human TfR (hTfR) and can include TfR or any ofthe known TfR homologs or orthologs, including TfRl, TfR2, soluble TfR, or any combination or fragment (e.g., ectodomain) thereof.

[0179] As used herein, the terms “endosome,” “endosomal,” “endosomal compartment,” or “endocytic pathway” can be used interchangeably and may refer to any one or more components of the intracellular endosomal network or trans-Golgi network (TGN) that allows for the vesicular transcytosis or trafficking and transfer of peptides and cargoes between distinct membrane-bound compartments within a cell, including lysosomal degradation as well as recycling to the cell surface. It is understood that such pathway involves and includes the maturation and transition of vesicles commonly referred to as transport vesicles or early endosomes to late endosomes to lysosomes, and that endosomal compartment acidity increases upon acidification of the endosome throughout the maturation process. Lysosomes serving as the last vesicle in the matured endocytic pathway typically contain hydrolytic enzymes which digest the contents of the late endosomes. Other endosomes continue to a pathway of recycling endosomes, where the contents are recycled back to the cell surface.

[0180] As used herein “pH-independent,” when used in reference to a molecule or moiety, refer means that as the endosomal compartment is acidified, the binding affinity of the molecule or moiety to its target molecule does not change sufficiently to enable dissociation in the endosome with the target molecule. For example, the referenced molecule or moiety has the same or similar affinity to its target molecule at extracellular pH and at an endosomal pH. It is also understood that pH-independent molecules or moieties do not include pH-dependent molecules or moieties, since the binding affinity of pH-dependent molecules or moieties to its target molecule changes as it enters and proceeds through the endosomal pathway, for example, to enable dissociation in the endosome with the target molecule to some degree, or the referenced molecule or moiety has a different affinity at extracellular pH and at an endosomal pH.

[0181] The term “engineered,” when applied to a polynucleotide, denotes that the polynucleotide has been removed from its natural genetic milieu and is thus free of other extraneous or unwanted coding sequences and is in a form suitable for use within genetically engineered protein production systems. Such engineered molecules are those that are separated from their natural environment and include cDNA and genomic clones (i.e., a prokaryotic or eukaryotic cell with a vector containing a fragment of DNA from a different organism). Engineered DNA molecules of the present invention are free of other genes with which they are ordinarily associated but can include naturally occurring or non-naturally occurring 5 ’and 3’ untranslated regions such as enhancers, promoters, and terminators.

[0182] An “engineered” polypeptide or protein is a polypeptide or protein that is found in a condition other than its native environment, such as apart from blood and animal tissue. In a preferred form, the engineered polypeptide is substantially free of other polypeptides, particularly other polypeptides of animal origin. It is preferred to provide the polypeptides in a highly purified form, e.g., greater than 90% pure, greater than 95% pure, more preferably greater than 98% pure or greater than 99% pure. When used in this context, the term “engineered” does not exclude the presence of the same polypeptide in alternative physical forms, such as dimers, heterodimers and multimers, heteromultimers, or alternatively glycosylated, carboxylated, modified, or derivatized forms.

[0183] An “engineered” peptide or protein is a polypeptide that is distinct from a naturally occurring polypeptide structure, sequence, or composition. Engineered peptides include non- naturally occurring, artificial, isolated, synthetic, designed, modified, or recombinantly expressed peptides. Provided herein are engineered TfR-binding peptides, variants, or fragments thereof. These engineered TfR-binding peptides can be further linked to a target-binding moiety or a half-life extending moiety, or can be further linked to an active agent or detectable agent, or any combination of the foregoing.

[0184] Polypeptides of the disclosure include polypeptides that have been modified in any way, for example, to: (1) reduce susceptibility to proteolysis, (2) reduce susceptibility to oxidation, (3) alter binding affinity for forming protein complexes, (4) alter binding affinities, (5) alter binding affinity at certain pH values, and (6) confer or modify other physicochemical or functional properties. For example, single or multiple amino acid substitutions (e.g., conservative amino acid substitutions) are made in the naturally occurring sequence (e.g., in the portion of the polypeptide outside the domain(s) forming intermolecular contacts). A “conservative amino acid substitution” can refer to the substitution in a polypeptide of an amino acid with a functionally similar amino acid. The following six groups each contain amino acids that can be conservative substitutions for one another: i) Alanine (A), Serine (S), and Threonine (T); ii) Aspartic acid (D) and Glutamic acid (E); iii) Asparagine (N) and Glutamine (Q); iv) Arginine (R) and Lysine (K); v) Isoleucine (I), Leucine (L), Methionine (M), and Valine (V); vi) Phenylalanine (F), Tyrosine (Y), and Tryptophan (W). In some embodiments, a conserved amino acid substitution can comprise a non-natural amino acid. For example, substitution of an amino acid for a non-natural derivative of the same amino acid can be a conserved substitution.

[0185] The terms “polypeptide fragment” and “truncated polypeptide” as used herein can refer to a polypeptide that has an amino-terminal and / or carboxy-terminal deletion as compared to acorresponding full-length peptide or protein. In various embodiments, fragments are at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 100, at least 150, at least 200, at least 250, at least 300, at least 350, at least 400, at least 450, at least 500, at least 600, at least 700, at least 800, at least 900 or at least 1000 amino acids in length. In various embodiments, fragments can also be, e.g., at most 1000, at most 900, at most 800, at most 700, at most 600, at most 500, at most 450, at most 400, at most 350, at most 300, at most 250, at most 200, at most 150, at most 100, at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10, or at most 5 amino acids in length. A fragment can further comprise, at either or both of its ends, one or more additional amino acids, for example, a sequence of amino acids from a different naturally-occurring protein (e.g., an Fc or leucine zipper domain) or an artificial amino acid sequence (e.g., an artificial linker sequence).

[0186] As used herein, the terms “peptide” or “polypeptide” in conjunction with “variant”, “mutant”, or “enriched mutant”, or “permuted enriched mutant” can refer to a peptide or polypeptide that can comprise an amino acid sequence wherein one or more amino acid residues are inserted into, deleted from and / or substituted into the amino acid sequence relative to another polypeptide sequence. In various embodiments, the number of amino acid residues to be inserted, deleted, or substituted is at least 1, at least 2, at least 3, at least 4, at least 5, at least 10, at least 25, at least 50, at least 75, at least 100, at least 125, at least 150, at least 175, at least 200, at least 225, at least 250, at least 275, at least 300, at least 350, at least 400, at least 450 or at least 500 amino acids in length. Variants of the present disclosure include peptide conjugates or fusion molecules (e.g., peptide constructs or peptide complexes).

[0187] A “derivative” of a peptide or polypeptide can be a peptide or polypeptide that can have been chemically modified, e.g., conjugation to another chemical moiety such as, for example, polyethylene glycol, albumin (e.g., human serum albumin), phosphorylation, and glycosylation.

[0188] The term “% sequence identity” can be used interchangeably herein with the term “% identity” and can refer to the level of amino acid sequence identity between two or more peptide sequences or the level of nucleotide sequence identity between two or more nucleotide sequences, when aligned using a sequence alignment program. For example, as used herein, 80% identity means the same thing as 80% sequence identity determined by a defined algorithm, and means that a given sequence is at least 80% identical to another length of another sequence. In various embodiments, the % identity is selected from, e.g., at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 99% or more up to 100% sequence identity to a given sequence. In various embodiments, the % identity is in the range of, e.g., about 60% to about 70%, about 70% to about 80%, about 80% to about 85%, about 85% to about 90%, about 90% to about 95%, or about 95% to about 99%.

[0189] The terms “% sequence homology” or “percent sequence homology” or “percent sequence identity” can be used interchangeably herein with the terms “% homology,” “% sequence identity,” or “% identity” and can refer to the level of amino acid sequence homology between two or more peptide sequences or the level of nucleotide sequence homology between two or more nucleotide sequences, when aligned using a sequence alignment program. For example, as used herein, 80% homology means the same thing as 80% sequence homology determined by a defined algorithm, and accordingly a homologue of a given sequence has greater than 80% sequence homology over a length of the given sequence. In various embodiments, the % homology is selected from, e.g., at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% or more up to 100% sequence homology to a given sequence. In various embodiments, the % homology is in the range of, e.g., about 60% to about 70%, about 70% to about 80%, about 80% to about 85%, about 85% to about 90%, about 90% to about 95%, or about 95% to about 99%.

[0190] A protein or polypeptide can be “substantially pure,” “substantially homogeneous”, or “substantially purified” when at least about 60% to 75% of a sample exhibits a single species of polypeptide. The polypeptide or protein can be monomeric or multimeric. A substantially pure polypeptide or protein can typically comprise about 50%, 60%, 70%, 80% or 90% W / W of a protein sample, more usually about 95%, and e.g., will be over 98% or 99% pure. Protein purity or homogeneity can be indicated by a number of means well known in the art, such as polyacrylamide gel electrophoresis of a protein sample, followed by visualizing a single polypeptide band upon staining the gel with a stain well known in the art. For certain purposes, higher resolution is provided by using high-pressure liquid chromatography (e.g., HPLC) or other high-resolution analytical techniques (e.g., LC-mass spectrometry).

[0191] As used herein, the term “pharmaceutical composition” can generally refer to a composition suitable for pharmaceutical use in a subject such as an animal (e.g., human or mouse). A pharmaceutical composition can comprise a pharmacologically effective amount of an active agent and a pharmaceutically acceptable carrier. The term “pharmacologically effective amount” can refer to that amount of an agent effective to produce the intendedbiological or pharmacological result.

[0192] As used herein, the term “pharmaceutically acceptable carrier” can refer to any of the standard pharmaceutical carriers, vehicles, buffers, and excipients, such as a phosphate buffered saline solution, or a buffered saline solution, 5% aqueous solution of dextrose, and emulsions, such as an oil / water or water / oil emulsion, and various types of wetting agents and / or adjuvants. Suitable pharmaceutical carriers and formulations are described in Remington's Pharmaceutical Sciences, 21st Ed. 2005, Mack Publishing Co, Easton. A “pharmaceutically acceptable salt” can be a salt that can be formulated into a compound for pharmaceutical use including, e.g., metal salts (sodium, potassium, magnesium, calcium, etc.) and salts of ammonia or organic amines.

[0193] As used herein, the terms “treat”, “treating” and “treatment” can refer to a method of alleviating or abrogating a biological disorder and / or at least one of its attendant symptoms. As used herein, to “alleviate” a disease, disorder or condition, for example, means reducing the severity and / or occurrence frequency of the symptoms of the disease, disorder, or condition.Further, references herein to “treatment” can include references to curative, palliative, and prophylactic or diagnostic treatment.

[0194] Generally, a cell of the present disclosure can be a eukaryotic cell or a prokaryotic cell. A cell can be an epithelial cell. A cell can be a microorganism, bacterial, yeast, fungal or algae cell. A cell can be an animal cell or a plant cell. An animal cell can include a cell from a marine invertebrate, fish, insects, amphibian, reptile, or mammal. A mammalian cell can be obtained from a primate, ape, equine, bovine, porcine, canine, feline, or rodent. A mammal can be a primate, ape, dog, cat, rabbit, ferret, or the like. A rodent can be a mouse, rat, hamster, gerbil, hamster, chinchilla, or guinea pig. A bird cell can be from a canary, parakeet, or parrots. A reptile cell can be from a turtles, lizard, or snake. A fish cell can be from a tropical fish. For example, the fish cell can be from a zebrafish (e.g., Danino rerid). A worm cell can be from a nematode (e.g., C. elegans). An amphibian cell can be from a frog. An arthropod cell can be from a tarantula or hermit crab.

[0195] A mammalian cell can also include cells obtained from a primate (e.g., a human or a non-human primate). A mammalian cell can include a blood cell, a stem cell, an epithelial cell, connective tissue cell, hormone secreting cell, a nerve cell, a skeletal muscle cell, or an immune system cell.

[0196] As used herein, the term “vector,” generally refers to a DNA molecule capable of replication in a host cell and / or to which another DNA segment can be operatively linked so as to bring about replication of the attached segment. A plasmid is an exemplary vector.

[0197] As used herein, the term “subject,” generally refers to a human or to another animal. A subject can be of any age, for example, a subject can be prenatal, newborn, an infant, a toddler, a child, a pre-adolescent, an adolescent, an adult, or an elderly individual.

[0198] Ranges can be expressed herein as from “about” one particular value, and / or to “about” another particular value. When such a range is expressed, another embodiment includes from the one particular value and / or to the other particular value. Similarly, when values are expressed as approximations, by use of the antecedent “about,” it will be understood that the particular value forms another embodiment. It will be further understood that the endpoints of each of the ranges are in relation to the other endpoint, and independently of the other endpoint. The term “about” as used herein refers to a range that is 15% plus or minus from a stated numerical value within the context of the particular usage. For example, about 10 can include a range from 8.5 to 11.5.Peptides

[0199] The selective depletion complexes of the present disclosure can comprise one or more peptides. For example, a selective depletion complex of the present disclosure can comprise a receptor-binding peptide (e.g., a TfR-binding peptide or a PD-L1 -binding peptide) and a targetbinding peptide (e.g., a target-binding EGF variant). In some embodiments, two or more peptides can be connected via a linker. The peptides of the present disclosure (e.g., a TfR- binding peptide, a PD-L1 -binding peptide, an EGFR target-binding peptide, or a peptide comprising a TfR-binding peptide linked to an EGFR target-binding peptide) can be used in a method of selectively depleting a target molecule. The peptides of the present disclosure (e.g., TfR-binding peptide, an EGFR target-binding peptide, or a peptide comprising a TfR-binding peptide linked to an EGFR target-binding peptide) can be recycled to the cell surface following endocytosis.

[0200] In some instances, a peptide as disclosed herein can contain only one lysine residue, or no lysine residues. In some instances, one or more or all of the lysine residues in the peptide are replaced with arginine residues. In some instances, one or more or all of the methionine residues in the peptide are replaced by leucine or isoleucine. One or more or all of the tryptophan residues in the peptide can be replaced by phenylalanine or tyrosine. In some instances, one or more or all of the asparagine residues in the peptide are replaced by glutamine. In some embodiments, one or more or all of the aspartic acid residues can be replaced by glutamic acid residues. In some instances, one or more or all of the lysine residues in the peptide are replaced by alanine or arginine. In some embodiments, the N-terminus of the peptide is blocked orprotected, such as by an acetyl group or a te / 7-butyloxycarbonyl group. Alternatively or in combination, the C-terminus of the peptide can be blocked or protected, such as by an amide group or by the formation of an ester (e.g., a butyl or a benzyl ester). In some embodiments, the peptide is modified by methylation on free amines. For example, full methylation is accomplished through the use of reductive methylation with formaldehyde and sodium cyanoborohydride.

[0201] In some embodiments, the peptide comprises an amino acid sequence SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. In some embodiments, a dipeptide GS can be added as the first two N-terminal amino acids, for example as shown in SEQ ID NO: 1 - SEQ ID NO: 64, or such N-terminal dipeptide GS can be absent as shown in SEQ ID NO: 96 and SEQ ID NO: 65 - SEQ ID NO: 128 or can be substituted by any other one or two amino acids. In some embodiments, a dipeptide comprising GS is used as a linker or used to couple to a linker to form a peptide conjugate or fusion molecules such as a peptide construct or peptide complex. Similarly, in some embodiments, the peptide comprising the amino acid sequence of any one of SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241 can also comprise a dipeptide comprising GS used as a linker or used to couple to a linker to form a peptide conjugate or fusion molecules such as a peptide construct or peptide complex. In some embodiments, the linker comprises a GxSy(SEQ ID NO: 130) peptide, wherein x and y independently are any whole number, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 and the G and S residues are arranged in any order. In some embodiments, the peptide linker comprises (GS)x (SEQ ID NO: 131), wherein x can be any whole number, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10. In some embodiments, the peptide linker comprises GGSSG (SEQ ID NO: 132), GGGGG (SEQ ID NO: 133), GSGSGSGS (SEQ ID NO: 134), GSGG (SEQ ID NO: 135), GGGGS (SEQ ID NO: 136), GGGS (SEQ ID NO: 129), GGS (SEQ ID NO: 137), GGGSGGGSGGGS (SEQ ID NO: 138), or a variant or fragment thereof or any number of repeats and combinations thereof. Additionally, KKYKPYVPVTTN (SEQ ID NO: 139) from DkTx, and EPKSSDKTHT (SEQ ID NO: 140) from human IgG3 can be used as a peptide linker or any number of repeats and combinations thereof. In some embodiments, the peptide linker comprises GGGSGGSGGGS (SEQ ID NO: 141) or a variant or fragment thereof or any number of repeats and combinations thereof. It is understood that any of the foregoing linkers or a variant or fragment thereof can be used with any number of repeats or any combinations thereof. It is also understood that other peptidelinkers in the art or a variant or fragment thereof can be used with any number of repeats or any combinations thereof. The length of the linker can be tailored to maximize binding of the selective delivery complex to both TfR and the target molecule (e.g., EGFR) at the same time including accounting for steric access. In some embodiments, the linker between the TfR- binding and target-binding peptides (e.g., EGFR-b inding peptides) within the selective depletion complex is at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36 at least 37, at least 38, at least 39, at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, at least 50, at least 51, at least 52, at least 53, at least 54, at least 55, at least 56, at least 57, at least 58, at least 59, at least 60, at least 61, at least 62, at least 63, at least 64, at least 65 residues incrementally up to 100 residues long, particularly for example if the target molecule is not a soluble protein but rather a cell surface protein or cell receptor protein.

[0202] In some embodiments of the present disclosure, a peptide or peptide complex as described herein comprises a TfR-b inding peptide comprising an amino acid sequence set forth in any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. A TfR-binding peptide as disclosed herein can be a fragment comprising a contiguous fragment of any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64 that is at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36 at least 37, at least 38, at least 39, at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, at least 50, at least 51, at least 52, at least 53, at least 54, at least 55, at least 56, at least 57, at least 58, at least 59, at least 60, at least 61, at least 62, at least 63, at least 64, at least 65 residues long, wherein the peptide fragment is selected from any portion of the peptide. In some embodiments, the peptide sequence is flanked by additional amino acids. One or more additional amino acids, for example,confer a particular in vivo charge, isoelectric point, chemical conjugation site, stability, or physiologic property to a peptide.

[0203] In some instances, the peptides as described herein that are capable of targeting and binding to a TfR comprise no more than 80 amino acids in length, or no more than 70, no more than 60, no more than 40, no more than 35, no more than 30, no more than 25, no more than 20, no more than 15, or no more than 10 amino acids in length. In some instances, the peptides as described herein that are capable of targeting and binding to a target molecule comprise no more than 80 amino acids in length, or no more than 70, no more than 60, no more than 50, no more than 40, no more than 35, no more than 30, no more than 25, no more than 24, no more than 23, no more than 22, no more than 21, no more than 20, no more than 19, no more than 18, no more than 17, no more than 16, no more than 15, no more than 14, no more than 13, no more than 12, no more than 11, or no more than 10 amino acids in length.

[0204] In other embodiments, peptides can be conjugated to, linked to, or fused to a carrier or a molecule with targeting or homing function for a cell of interest or a target cell. In other embodiments, peptides can be conjugated to, linked to, or fused to a molecule that extends halflife or modifies the pharmacodynamic and / or pharmacokinetic properties of the peptides, or any combination thereof.

[0205] In some instances, a peptide comprises at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 positively charged residues, such as Arg or Lys, or any combination thereof. In some instances, one or more lysine residues in the peptide are replaced with arginine residues. In some embodiments, peptides comprise one or more Arg patches. In some embodiments, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, or 10 or more Arg or Lys residues are solvent exposed on a peptide. In some instances, a peptide comprises at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 histidine residues.

[0206] The peptides of the present disclosure can further comprise neutral amino acid residues. In some embodiments, the peptide has 35 or fewer neutral amino acid residues. In other embodiments, the peptide has 81 or fewer neutral amino acid residues, 70 or fewer neutral amino acid residues, 60 or fewer neutral amino acid residues, 50 or fewer neutral amino acid residues, 40 or fewer neutral amino acid residues, 36 or fewer neutral amino acid residues, 33 orfewer neutral amino acid residues, 30 or fewer neutral amino acid residues, 25 or fewer neutral amino acid residues, or 10 or fewer neutral amino acid residues.

[0207] The peptides of the present disclosure can further comprise negative amino acid residues. In some embodiments the peptide has 6 or fewer negative amino acid residues, 5 or fewer negative amino acid residues, 4 or fewer negative amino acid residues, 3 or fewer negative amino acid residues, 2 or fewer negative amino acid residues, or 1 or fewer negative amino acid residues. While negative amino acid residues can be selected from any negatively charged amino acid residues, in some embodiments, the negative amino acid residues are either E, or D, or a combination of both E and D.

[0208] In some embodiments of the present disclosure, a three-dimensional or tertiary structure of a peptide is primarily comprised of beta-sheets and / or alpha-helix structures. In some embodiments, designed or engineered TfR-b inding peptides or target-binding of the present disclosure are small, compact peptides or polypeptides stabilized by intra-chain disulfide bonds (e.g., mediated by cysteines) to form cystine and a hydrophobic core. In some embodiments, engineered TfR-b inding peptides have structures comprising helical bundles with at least one disulfide bridge between each of the alpha helices, thereby stabilizing the peptides. In other embodiments, the engineered TfR-b inding peptides or target-binding peptides comprise structures with three alpha helices and three intra-chain disulfide bonds, one between each of the three alpha helices in the bundle of alpha helices.Receptor-Binding Peptides

[0209] Disclosed herein are peptide sequences, such as those listed in TABLE 1 and TABLE 2, capable of binding to a receptor (e.g., a transferrin receptor or PD-L1 (also known as programmed death-ligand 1)). The peptide capable of binding a receptor may be referred to as a receptor-binding peptide. In some embodiments, a receptor-binding peptide may bind to a recycled receptor that undergoes recycling via a recycling pathway. The depletion of a selected target (e.g., EGFR) by the SDCs described herein is dependent on the normal trafficking and cycling behavior of the recycling receptor in cells to which the recycling receptor-binding peptide the in the SDC is bound. The recycled receptor may be endocytosed into an early endosome and packaged into a recycling endosome prior to maturation of the early endosome into a late endosome. The recycling endosome containing the recycled receptor may fuse with a cell membrane and return the recycled receptor to the cell surface. In some embodiments, a receptor-binding peptide of the present disclosure may remain bound to the receptor during therecycling process, thereby recycling the receptor-binding peptide as well. Examples of recycled receptors that may be targeted by a receptor-binding peptide include transferrin receptor, programmed death-ligand 1, cation-independent mannose 6 phosphate receptor (CI-M6PR), asialoglycoprotein receptor (ASGPR), CXCR7, folate receptor, or Fc receptors (including but not limited to neonatal Fc receptor (FcRn) or FcyRIIb). In some embodiments, a receptorbinding peptide of the present disclosure may comprise a miniprotein, a nanobody, an antibody, an IgG, an antibody fragment, a Fab, a F(ab)2, an scFv, an (scFv)2, a DARPin, or an affibody. In some embodiments, receptor binding can be achieved by engineering an Fc domain for improved binding to an existing Fc receptor, e.g., FcRn or FcyRIIb, or for novel binding to a non-native receptor, e.g., TfR. In some embodiments, the receptor-binding peptide may comprise a cystine-dense peptide, an affitin, an adnectin, an avimer, a Kunitz domain, a nanofittin, a fynomer, a bicyclic peptide, a beta-hairpin, or a stapled peptide. In some embodiments, receptor binding can be achieved by conjugation of a target-binding peptide or peptide complex with a sugar or other small molecule that is bound by the cellular receptor (e.g., mannose-6-phosphonate or N-acetylgalactosamine that bind with CI-M6PR and ASGPR, respectively).

[0210] In some embodiments, a receptor-binding peptide of the present disclosure can bind to the receptor (e.g., a recycled receptor) with an affinity that is pH-independent. For example, a receptor-binding peptide can bind the receptor at an extracellular pH (about pH 7.4) with an affinity that is substantially the same the binding affinity at an endocytic pH (such as about pH 5.5 or about pH 6.5). In some embodiments, a receptor-binding peptide can bind the receptor at an extracellular pH (about pH 7.4) with an affinity that is lower than the binding affinity at an endocytic pH (such as about pH 5.5 or about pH 6.5). In some embodiments, a receptor-binding peptide can bind the receptor at an extracellular pH (about pH 7.4) with an affinity that is higher than the binding affinity at an endocytic pH (such as about pH 5.5 or about pH 6.5). In some embodiments, the binding affinity of a receptor-binding peptide for the receptor at extracellular pH (about pH 7.4) and the binding affinity of a receptor-binding peptide for the receptor at endocytic pH (about pH 5.5) can differ by no more than about 1%, no more than about 2%, no more than about 3%, no more than about 4%, no more than about 5%, no more than about 6%, no more than about 7%, no more than about 8%, no more than about 9%, no more than about 10%, no more than about 12%, no more than about 15%, no more than about 17%, no more than about 20%, no more than about 25%, no more than about 30%, no more than about 35%, no more than about 40%, no more than about 45%, or no more than about 50%. In someembodiments, the affinity of the receptor-binding peptide for the receptor at pH 7.4 and at pH 5.5 can differ by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15 -fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40- fold, or no more than 50-fold. In some embodiments, a receptor-binding peptide (e.g., any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, SEQ ID NO: 1 - SEQ ID NO: 64, SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241) can be modified to remove one or more histidine amino acids in the receptor-binding interface, thereby reducing the pH-dependence of the binding affinity of the receptor-binding peptide for the receptor. In some embodiments, a receptor-binding peptide (e.g., any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, SEQ ID NO: 1 - SEQ ID NO: 64, SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241) can lack histidine amino acids in the receptor-binding interface.

[0211] In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with an equilibrium dissociation constant (KD) of less than 50 pM, less than 5 pM, less than 500 nM, less than 100 nM, less than 40 nM, less than 30 nM, less than 20 nM, less than 10 nM, less than 5 nM, less than 2 nM, less than 1 nM, less than 0.5 nM, less than 0.4 nM, less than 0.3 nM, less than 0.2 nM, or less than 0.1 nM at extracellular pH (about pH 7.4). In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with an equilibrium dissociation constant (KD) of less than 50 pM, less than 5 pM, less than 500 nM, less than 100 nM, less than 40 nM, less than 30 nM, less than 20 nM, less than 10 nM, less than 5 nM, less than 2 nM, less than 1 nM, less than 0.5 nM, less than 0.4 nM, less than 0.3 nM, less than 0.2 nM, or less than 0.1 nM at endosomal pH (about pH 5.5). In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with an equilibrium dissociation constant (KD) of less than 50 pM, less than 5 pM, less than 500 nM, less than 100 nM, less than 40 nM, less than 30 nM, less than 20 nM, less than 10 nM, less than 5 nM, less than 2 nM, less than 1 nM, less than 0.5 nM, less than 0.4 nM, less than 0.3 nM, less than 0.2 nM, or less than 0.1 nM at endosomal pH (about pH 5.8).

[0212] In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM,no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 7.4. In some embodiments, a receptor-binding peptide with pH- independent binding can bind to the receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 5.5. In some embodiments, a receptorbinding peptide with pH-independent binding can bind to the receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 5.8.

[0213] In some embodiments, the affinity of the receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some embodiments, the affinity of the receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

[0214] In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 7.4. In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’ no more than 5xl0’4s’1, or no more than 2xl0’4s’1at pH 5.5. In some embodiments, a receptor-binding peptide with pH-independent binding can bind to the receptor with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 5.8.

[0215] In some embodiments, the dissociation rate constant (koff or kd) of the receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25- fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some embodiments, the dissociation rate constant (koff or kd) of the receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

[0216] In some embodiments, the receptor-binding peptide can bind to the receptor with an affinity that is pH-dependent. For example, the receptor-binding molecule can bind to the receptor with higher affinity at extracellular pH (about pH 7.4) and with lower affinity at endosomal pH (about pH 5.5), thereby releasing the selective depletion complex from receptor upon internalization and acidification of the endosomal compartment.

[0217] In some embodiments, the recycling receptor may be TfR. A peptide capable of binding transferrin receptor (TfR) may bind TfR or any of the known TfR homologs, including TfRl, TfR2, soluble TfR, or any combination or fragment (e.g., ectodomain) thereof. A peptide capable of binding a transferrin receptor or a TfR homolog can be referred to herein as a transferrin receptor-binding peptide or a TfR-binding peptide. In some embodiments, peptides disclosed herein can penetrate, cross, or enter target cells in a TfR-mediated manner. These cell layers or cells can include TfR-expressing endothelial cells, epithelial cells, and TfR-expressing cells of various tissues or organs such as tumor cells, brain cells, cancerous or tumor cells, liver cells (e.g., hepatocytes (HCs), hepatic stellate cells (HSCs), Kupffer cells (KCs), or liver sinusoidal endothelial cells (LSECs)), pancreas cells, colon cells, ovarian cells, breast cells, spleen cells, bone marrow cells, and / or lung cells, or any combination thereof. In some embodiments, a TfR-binding peptide of the present disclosure may comprise a miniprotein, a nanobody, an antibody, an IgG, an antibody fragment, a Fab, a F(ab)2, an scFv, an (scFv)2, a DARPin, or an affibody. In some embodiments, the TfR-binding peptide may comprise a cystine-dense peptide, an affitin, an adnectin, an avimer, a Kunitz domain, a nanofittin, a fynomer, a bicyclic peptide, a beta-hairpin, or a stapled peptide.

[0218] In some embodiments, the peptides as discloses herein can cross cellular layers or barriers (e.g., BBB) or cell membranes via, for example, TfR-mediated vesicular transcytosis and TfR-mediated endocytosis, respectively. In addition to binding TfR and promote transcytosis and / or endocytosis, the peptides of the present disclosure can also bind to additionaltarget proteins on cells such as cancer cells. In some cases, a peptide is a peptide or peptide complex comprising a TfR-binding peptide conjugated to, linked to, or fused to a targeting moiety or an active agent (e.g., a therapeutic or diagnostic agent) such as a small molecule or a peptide that has an affinity for an additional target protein (e.g., receptor or enzyme). In some cases, the TfR-binding peptide is linked to a target-binding peptide and enables or promotes TfR-mediated transcytosis of the target-binding peptide across the BBB or TfR-mediated endocytosis into a cell. In some instances, and subsequent to transcytosis, a peptide complex comprising the TfR-binding peptide and a target-binding peptide can target a specific cell or tissue in the CNS and exert a biological effect (e.g., binding a target protein) upon reaching said cell or tissue. In some cases, a peptide complex of the present disclosure exerts a biological effect that is mediated by the TfR-binding peptide, the target-binding peptide, an active agent, or a combination thereof. In some cases, a TfR-binding peptide complex of the present disclosure comprising one target-binding peptides can transport and / or deliver target molecules into cells that express TfR (e.g., deliver target molecules into endosomes). In some cases, the TfR-binding peptide accumulates in tissues in the CNS. In some cases, off-target effects are reduced due to CNS-specific accumulation. In some cases, the TfR-binding peptide accumulates in tissue outside of the CNS (e.g., liver, kidney, spleen, or skin). In some cases, the cells expressing TfR are tumor cells and the TfR-binding peptide complex delivers anti-tumor agents to these tumor cells. In some cases, the anti-tumor agents alone show no or only very limited therapeutic efficacy against the tumor cells; however, when the anti -tumor agents are combined with the TfR-binding peptides of the present disclosure as, for example, a peptide complex, the therapeutic efficacy of these anti-tumor agents is significantly improved.

[0219] In some embodiments, the TfR-binding peptides of the present disclosure (e.g., SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, and SEQ ID NO: 1 - SEQ ID NO: 64) can induce a biologically relevant response. For example, a TfR-binding peptide conjugated to a target-binding peptide can selectively deplete a soluble target molecule or a cell surface target molecule. In some embodiments, the biologically relevant response can be induced after intravenous, subcutaneous, peritoneal, intracranial, or intramuscular dose, and in some embodiments, after a single intravenous, subcutaneous, peritoneal, intracranial, or intramuscular dose. In some embodiments, the TfR-binding peptides can be used in combination with various other classes of therapeutic compounds used to treat and / or prevent pain, neuropathic pain or other neurological disorders such as neurodegenerative disorders, infectious diseases,immunological disorders (e.g., autoimmune diseases) or lysosomal storage diseases. Binding of the herein described peptides and peptide complexes (e.g., peptide conjugates, fusion peptides, or recombinantly produced peptide complexes) to TfR and subsequent transport across a cell layer or barrier such as the BBB (e.g., via TfR-mediated vesicular transcytosis) or a cell membrane (e.g., via TfR-mediated endocytosis) can have implications in a number of diseases, conditions, or disorders associated with over-expression or accumulation of a target molecule (e.g., cancer, neurodegeneration, or lysosomal storage diseases) or diseases associated with mutations (e.g., mutations causing constitutive activity, resistance to treatment, or dominant negative activity) in soluble or surface proteins in a subject (e.g., a human).

[0220] Binding of the herein described peptides and peptide complexes (e.g., peptide conjugates, fusion peptides, or recombinantly produced peptide complexes) to TfR and subsequent transport across a cell layer or barrier such as the BBB (e.g., via vesicular transcytosis) or a cell membrane (e.g., via endocytosis) can have implications in a number of diseases, conditions, or disorders associated with EGFR-driven diseases in the CNS, such as EGFR-driven cancers.

[0221] In some embodiments, TfR-binding peptides of the present disclosure can bind to any of the known TfR homologs, including TfRl, TfR2, soluble TfR, or any combination or fragment (e.g., ectodomain) thereof. Thus, as used herein, “TfR” can refer to any known homolog, derivative, fragment, or member of the TfR family including TfRl, TfR2, and a soluble TfR. In other embodiments, peptides are capable of binding to one, one or more, or all TfR homologs. In some embodiments, peptides of the present disclosure can bind to a TfR and promote a particular biological effect such as vesicular transcytosis. In some embodiments, TfR-binding peptides of the present disclosure, including peptides and peptide complexes with amino acid sequences set forth in SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, and SEQ ID NO: 1 - SEQ ID NO: 64, and any derivatives or variant thereof, prevent or decrease the binding of endogenous TfR binders (e.g., transferrin or any derivatives such as apo-transferrin or holo-transferrin) to TfR. In some embodiments, peptides or peptide complexes of the present disclosure comprise derivatives and variants with at least 40% homology, at least 50% homology, at least 60% homology, at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 91% homology, at least 92% homology, at least 93% homology, at least 94% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least100% homology to amino acid sequences set forth in SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, and SEQ ID NO: 1 - SEQ ID NO: 64.

[0222] In various embodiments, the interface residues of the TfR-binding peptides of the present disclosure (e.g., those amino acid residues that interact with TfR for receptor binding) can be divided between two largely helical domains of the peptide. In some cases, the interface residues can comprise residues corresponding to residues 5-25 (e.g., and comprising corresponding residues G5, A7, S8, Ml 1, N14, L17, E18, and E21), with reference to SEQ ID NO: 32, or corresponding to residues 35-51 (e.g., and comprising corresponding residues L38, L41, L42, L45, D46, H47, H49, S50, and Q51), with reference to SEQ ID NO: 32, or both. For example, the interface residues can comprise residues corresponding to residues 5-25 (e.g., and comprising corresponding residues G5, A7, S8, Mi l, N14, L17, E18, and E21), with reference to SEQ ID NO: 32, or corresponding to residues 35-51 (e.g., and comprising corresponding residues L38, L41, L42, L45, D46, H47, H49, S50, and Q51), with reference to SEQ ID NO: 32. In some embodiments, a TfR-binding peptide can comprise a fragment of a peptide provided herein, wherein the fragment comprises the minimum interface residues for binding, for example residues corresponding to residues 5-25 (e.g., and comprising corresponding residues G5, A7, S8, Ml 1, N14, L17, E18, and E21), with reference to SEQ ID NO: 32, or corresponding to residues 35-51 (e.g., and comprising corresponding residues L38, L41, L42, L45, D46, H47, H49, S50, and Q51), with reference to SEQ ID NO: 32. In some cases, the TfR-binding peptide is a peptide having the sequence set forth in SEQ ID NO: 32 comprising the TfR-binding residues corresponding to residues G5, A7, S8, Mi l, N14, L17, E18, and E21 of the domain and corresponding to residues L38, L41, L42, L45, D46, H47, H49, S50, and Q51 of the second domain, with reference to SEQ ID NO: 32.

[0223] In some embodiments, TfR-binding peptides bind to TfR with equal, similar, or greater affinity (e.g., lower equilibrium dissociation constant KD) as compared to endogenous molecules (e.g., transferrin, holotransferrin (iron-bound transferrin), apotransferrin (transferrin not bound to iron), or any other endogenous TfR ligands) or other exogenous molecules. In some embodiments, the peptide can have an equilibrium dissociation constant (KD) of less than 50 pM, less than 5 pM, less than 500 nM, less than 100 nM, less than 40 nM, less than 30 nM, less than 20 nM, less than 10 nM, less than 5 nM, less than 2 nM, less than 1 nM, less than 0.5 nM, less than 0.4 nM, less than 0.3 nM, less than 0.2 nM, or less than 0.1 nM. In some embodiments, the peptide can have an equilibrium dissociation constant (KD) of no more than 100 nM, nomore than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM. In some embodiments, the peptide can have a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1. In some embodiments, peptide transport by TfR is improved by having a lower affinity (e.g., a higher equilibrium dissociation constant KD) as compared to endogenous molecules. In some embodiments, peptide transport by TfR is improved by having a faster off rate or higher koff than endogenous molecules. In some embodiments, the dissociation rate constant (kd or koff) is similar to that of transferrin. In some embodiments, peptide transport is improved by having a faster on rate or a higher kon, optionally such as higher than that of transferrin. In other embodiments, one or more conserved residues at the transferrin (Tf)-TfR- binding interface are also present in the amino acid sequences of the peptides described herein. In some embodiments, a TfR-binding peptide has an off rate that is slower than the recycling rate of TfR, such that the TfR-binding peptide is likely to remain bound to TfR during the recycling process. In some embodiments, the TfR-binding peptide may have a half-life of dissociation that is no faster than 1 minute, no faster than 2 minutes, no faster than 3 minutes, no faster than 4 minutes, no faster than 5 minutes, no faster than 7 minutes, no faster than 10 minutes, no faster than 15 minutes, or no faster than 20 minutes, no faster than 30 minutes, no faster than 45 minutes, no faster than 60 minutes, no faster than 90 minutes, or no faster than 120 minutes. In some embodiments, the TfR-binding peptide may have a half-life of dissociation that is from about 1 minute to about 20 minutes, from about 2 minutes to about 15 minutes, from about 2 minutes to about 10 minutes, or from about 5 minutes to about 10 minutes. In some embodiments, a rate of dissociation of the target-binding peptide from the target molecule is faster than a recycling rate of the cellular receptor. In some embodiments, a half-life of dissociation of the target molecule binding-binding peptide from the target molecule is less than 10 seconds, less than 20 seconds, less than 30 seconds, less than 1 minute, less than 2 minutes, less than 5 minutes, less than 10 minutes, less than 20 minutes, less than 30 minutes, less than 45 minutes, or less than 60 minutes in endosomal conditions.

[0224] In some embodiments, TfR-binding peptides that exhibit an improved TfR receptor binding show improved transcytosis function, improved endocytosis function, improved recycling, or combinations thereof. In some embodiments, TfR-binding peptides that exhibit animproved TfR receptor binding show no or small changes in transcytosis function, endocytosis function, recycling, or combinations thereof. In some embodiments, TfR-binding peptides that exhibit an improved TfR receptor binding show reduced transcytosis function, reduced endocytosis function, reduced recycling, or combinations thereof. In some embodiments, the TfR-b inding peptide binds at a site of high homology between human and murine TfR, including one or more, or all, of the amino acid domains corresponding to residues 506-510, 523-531, and 611-662 of the human TfR (SEQ ID NO: 190, MMDQARSAFSNLFGGEPLSYTRFSLARQVDGDNSHVEMKLAVDEEENADNNTKANVT KPKRCSGSICYGTIAVIVFFLIGFMIGYLGYCKGVEPKTECERLAGTESPVREEPGEDFPA ARRLYWDDLKRKLSEKLDSTDFTGTIKLLNENSYVPREAGSQKDENLALYVENQFREF KLSKVWRDQHFVKIQVKDSAQNSVIIVDKNGRLVYLVENPGGYVAYSKAATVTGKLV HANFGTKKDFEDLYTPVNGSIVIVRAGKITFAEKVANAESLNAIGVLIYMDQTKFPIVNA ELSFFGHAHLGTGDPYTPGFPSFNHTQFPPSRSSGLPNIPVQTISRAAAEKLFGNMEGDCP SDWKTDSTCRMVTSESKNVKLTVSNVLKEIKILNIFGVIKGFVEPDHYVWGAQRDAW GPGAAKSGVGTALLLKLAQMFSDMVLKDGFQPSRSIIFASWSAGDFGSVGATEWLEGY LSSLHLKAFTYINLDKAVLGTSNFKVSASPLLYTLIEKTMQNVKHPVTGQFLYQDSNWA SKVEKLTLDNAAFPFLAYSGIPAVSFCFCEDTDYPYLGTTMDTYKELIERIPELNKVARA AAEVAGQFVIKLTHDVELNLDYERYNSQLLSFVRDLNQYRADIKEMGLSLQWLYSARG DFFRATSRLTTDFGNAEKTDRFVMKKLNDRVMRVEYHFLSPYVSPKESPFRHVFWGSG SHTLPALLENLKLRKQNNGAFNETLFRNQLALATWTIQGAANALSGDVWDIDNEF). In some embodiments, the regions of TfR to which the peptides disclosed herein or variants thereof bind all or in part to such TfR domains. In some embodiments, the peptides disclosed herein bind to any one, any two, or all three of the TfR regions of high homology including the amino acid domains corresponding to residues 506-510, 523-531, and 611-662 of the human TfR (SEQ ID NO: 190). In some embodiments the peptides disclosed herein bind at least to the domain corresponding to residues 611-662 of the human TfR.

[0225] In some embodiments, the KA and KD values of a TfR-binding peptide can be modulated and optimized (e.g., via amino acid substitutions) to provide an optimal ratio of TfR-binding affinity and efficient transcytosis function.

[0226] In some embodiments, peptides disclosed herein or variants thereof bind to TfR at residues found in the binding interface (e.g., the binding domain or the binding pocket) of TfR with other exogenous or endogenous ligands (e.g., transferrin (Tf), Tf derivatives, or Tf-like peptides or proteins). In some embodiments, a peptide disclosed herein or a variant thereof,which binds to TfR, comprises at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to a sequence that binds residues of TfR, which makeup the binding pocket. In some embodiments, a peptide disclosed herein or a variant thereof, which binds to TfR, comprises at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to an endogenous or exogenous polypeptide known to bind TfR, for example, endogenous Transferrin or any one of the peptides listed in TABLE 1. In other embodiments, a peptide described herein binds to a protein of interest, which comprises at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to TfR, a fragment, homolog, or a variant thereof.

[0227] In some embodiments, peptides disclosed herein or variants thereof bind regions of TfR that comprise the amino acid residues corresponding to residues 506-510, 523-531, and 611-662 (the numbering of these amino acid residues is based on the following Uniprot reference protein sequence of endogenous human TFRC UniProtKB - P02786 (SEQ ID NO: 190, TFR1 HUMAN)). In some embodiments, the regions of TfR to which the peptides disclosed herein or variants thereof bind overlap with those of Tf, a fragment, homolog, or a variant thereof.

[0228] In other embodiments, a nucleic acid, vector, plasmid, or donor DNA comprises a sequence that encodes a peptide, peptide construct, a peptide complex, or variant or functional fragment thereof, as described in the present disclosure. In further embodiments, certain parts or fragments of TfR-binding motifs (e.g., conserved binding motifs) can be grafted onto a peptide or peptide complex with a sequence of any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. In some embodiments, peptides can cause TfR to be degraded, prevent TfR from localizing to a cell’s nucleus, or prevent TfR from interacting with transferrin or transferrin-like proteins.

[0229] In some embodiments, a peptide can be selected for further testing or use based upon its ability to bind to the certain amino acid residue or motif of amino acid residues. The certain amino acid residue or motif of amino acid residues in TfR can be identified an amino acidresidue or sequence of amino acid residues that are involved in the binding of TfR to Tf. A certain amino acid residue or motif of amino acid residues can be identified from a crystal structure of the TfR:Tf complex. In some embodiments, peptides (e.g., CDPs) demonstrate the resistance to heat, protease (pepsin), and reduction.

[0230] The peptides, peptide complexes (e.g., peptide conjugates or fusion peptides), and selective delivery complexes comprising one or more of the amino acid sequences set forth in SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64 can bind to a protein of interest. In some embodiments, the protein of interest is a TfR. In some embodiments, the peptides and peptide complexes (e.g., peptide conjugates or fusion peptides) that bind to a TfR comprise at least one of the amino acid sequences set forth in SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. In some embodiments, peptides, peptide complexes (e.g., peptide conjugates and fusion molecules) of the present disclosure that bind to a TfR comprise peptide derivatives or variants having at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to amino acid sequences set forth in SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. For example, peptides or peptide complexes (e.g., peptide conjugates and fusion molecules) of the present disclosure that bind to a TfR can comprise peptide derivatives or variants having at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to the amino acid sequence set forth in SEQ ID NO: 96. For example, peptides or peptide complexes (e.g., peptide conjugates and fusion molecules) of the present disclosure that bind to a TfR can comprise peptide derivatives or variants having at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to the amino acid sequence set forth in SEQ ID NO: 66. For example, peptides or peptide complexes (e.g., peptide conjugates and fusion molecules) of the present disclosure that bind to a TfR can comprisepeptide derivatives or variants having at least 70% homology, at least 75% homology, at least 80% homology, at least 85% homology, at least 90% homology, at least 95% homology, at least 96% homology, at least 97% homology, at least 98% homology, or at least 99% homology or at least 100% homology to the amino acid sequence set forth in SEQ ID NO: 65.

[0231] TABLE 1 lists exemplary peptide sequences according to the methods and compositions of the present disclosure.TABLE 1 - Exemplary TfR-Binding Peptide Sequences

[0232] In some embodiments, a TfR-binding peptide disclosed herein comprises REGCAX1RCX2KYX4DEX2X3KCX3ARMMSMSNTEEDCEQEX2EDX2X2YCX2X3X5CX5X1 X4(SEQ ID NO: 167) orGSREGCAX1RCX2KYX4DEX2X3KCX3ARMMSMSNTEEDCEQEX2EDX2X2YCX2X3X5CX5 X1X4 (SEQ ID NO: 148), wherein Xi can be independently selected from S, T, D, or N, X2 can be independently selected from A, M, I, L, or V, X3 can be independently selected from D, E, N, Q, S, or T, X4can be independently selected from D, E, H, K, R, N, Q, S, or T, and X5 can be independently selected from H, K, R, N, Q, S, or T.

[0233] In some embodiments, a TfR-binding peptide disclosed herein comprises REXICX2X3RCX4KYX5DEX6X7KCX8ARMMSMSNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 168) orGSREX1CX2X3RCX4KYX5DEX6X7KCX8ARMMSMSNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 149), wherein Xi, X2, X3, X4, X5, Xe, X7 and X8are TfR binding interface residues and can independently be any amino acid. In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCMKYNDELEKCEARMMSMSNTEEDCEQEXIEDX2X3YCX4X5X6CX7X8X9 (SEQ ID NO: 169) orGSREGCASRCMKYNDELEKCEARMMSMSNTEEDCEQEXIEDX2X3YCX4X5X6CX7X8X9 (SEQ ID NO: 150), wherein Xi, X2, X3, X4, X5, Xe, X7, X8, and X9 are TfR binding interfaceresidues and can independently be any amino acid. In some embodiments, a TfR-binding peptide disclosed herein comprises REX1CX2X3RCX4KYX5DEX6X7KCX8ARMMSMSNTEEDCEQEX9EDX10X11YCX12X13X13C Xi5Xi6Xi7 (SEQ ID NO: 170) or GSREX1CX2X3RCX4KYX5DEX6X7KCX8ARMMSMSNTEEDCEQEX9EDX10X11YCX12X13X13CX15X16X17 (SEQ ID NO: 151), wherein Xi, X2, X3, X4, X5, X6, X7, Xs, X9, X10, Xn, Xi2, Xi3, X14, X15, Xi6 and X17 are TfR binding interface residues and can independently be any amino acid. In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCMKYNDELEKCEARMMSMSNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 96) or GSREGCASRCMKYNDELEKCEARMMSMSNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 32). In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCTKYNAELEKCEARVMSMSNTEEDCEQELEDLLHCLDHCHSQ (SEQ ID NO: 66) or GSREGCASRCTKYNAELEKCEARVMSMSNTEEDCEQELEDLLHCLDHCHSQ (SEQ ID NO: 2). In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCTKYNAELEKCEARVSSMSNTEETCVQELFDLLHCVDHCVSQ (SEQ ID NO: 65) or GSREGCASRCTKYNAELEKCEARVSSMSNTEETCVQELFDLLHCVDHCVSQ (SEQ ID NO: 1).

[0234] In some embodiments, a TfR-binding peptide disclosed herein comprises X1X2X3X4GX5ASX6X7MX8X9NX10X11LEX12X13EX14X15X16X17X18X19X20X21X22X23X24X25X26 X27X28X29X30X31X32X33X34X35X36X37X38X39X40X41X42X43 (SEQ ID NO: 152), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, X10, Xn, X12, X13, X14,X15, X16, X17, X18, X19,X20, X21, X22, X23, X24, X25, X26, X27, X28, X29, X30, X31, X32, X33, X34, X35, X36, X37, X38, X39, X40, X41, X42, and X43 can independently be any amino acid.

[0235] In some embodiments, a TfR-binding peptide disclosed herein comprises XlX2X3X4X5X6X7X8X9XloXl 1X12X13X14X15X16X17X18X19X20X21X22X23X24X25X26X27X28X29X30 X31X32X33X34X35X36X37LX38X39LLX40X41LDHX42HSQ (SEQ ID NO: 153), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, X10, X11, X12,X13, X14, X15, X16, X17, X18,X19, X20, X21,X22, X23, X24, X25, X26, X27, X28, X29, X30, X31, X32, X33, X34, X35, X36, X37, X38, X39, X40, X41, and X42 can independently be any amino acid.

[0236] In some embodiments, a TfR-binding peptide disclosed herein comprisesX1X2X3X4GX5ASX6X7MX8X9NX10X11LEX12X13EX14X15X16X17X18X19X20X21X22X23X24X25X26X27X28X29LX30X31LLX32X33LDHX34HSQ (SEQ ID NO: 154), wherein Xi, X2, X3, X4, X5, X6,X7, X8, X9, X10, X11, X12, X13, X14, X15, X16, X17, X18, X19, X20,X21, X22, X23, X24, X25, X26, X27, X28, X29, X30, X31, X32, X33, and X34 can independently be any amino acid.

[0237] In some embodiments, a TfR-binding peptide or peptide complex disclosed herein comprises at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% sequence homology to any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO:64, or any variant, homolog, or functional fragment thereof. In some embodiments, a TfR- binding peptide or peptide complex disclosed herein comprises any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64, or any variant, homolog, or functional fragment thereof. In some embodiments, a peptide that binds to a TfR comprises the amino acid sequence set forth in SEQ ID NO: 96 or SEQ ID NO: 32. In some embodiments, a peptide that binds to a TfR comprises the amino acid sequence set forth in SEQ ID NO: 66 or SEQ ID NO: 2. In some embodiments, a peptide that binds to a TfR comprises the amino acid sequence set forth in SEQ ID NO: 65 or SEQ ID NO: 1.

[0238] In some embodiments, a TfR-binding peptide comprises canonical amino acid residues as surface interface residues at any one of the corresponding positions 5, 7, 8, 14, 17, 18, 21, 38, 42, 45, 46, 47, 50, 51, with reference to SEQ ID NO: 32 or a combination thereof. In some embodiments, a TfR-binding peptide comprises canonical amino acid residues as surface interface residues at any one of the corresponding positions G5, A7, S8, N14, L17, E18, E21, L38, L42, L45, D46, H47, S50, Q51, with reference to SEQ ID NO: 32 or a combination thereof. In some embodiments, the peptide or peptide complex of the present disclosure comprises at least one or more of these corresponding residues in SEQ ID NO: 96, SEQ ID NO:65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64. Such peptides can accordingly be engineered with enhanced binding to TfR. In some embodiments, a TfR-binding peptide disclosed herein comprises XlX2X3X4GX5ASX6X7X8X9X10NXllX12LEX13X14EXl5X16X17Xl8X19X20X21X22X23X24X25X26X27X28X29X3OLX3IX32X33LX34X35LDHX36X37SQ (SEQ ID NO: 155), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, X10, Xu, X12, X13, X14, X15, X16, X17, Xi8, X19, X20, X21, X22, X23, X24, X25, X26, X27, x28, X29, X30, X31, X32, X33, X34, X35, X36, and X37 can independently be any amino acid.

[0239] In some embodiments, surface-distal hydrophilic amino acid residues (e.g., D, E, H, K,R, N, Q, S, or T) present in the amino acid sequence of a peptide contribute to peptide solubility. In some embodiments, a peptide as disclosed herein comprises a hydrophilic amino acid residue at any one of the corresponding positions 3, 4, 9, 11, 15, 16, 19, 23, 26, 28, 29, 30, 31, 32, 33, 35, 36, 37, 39, 40, with reference to SEQ ID NO: 32, or any combination thereof. In some instances, a peptide of the present disclosure comprises hydrophilic amino acid residues at the following corresponding positions: R3, E4, R9, K12, D15, E16, K19, R23, S26, S28, N29, T30, E31, E32, D33, E35, Q36, E37, E39, D40, with reference to SEQ ID NO: 32, or any combination thereof. In some embodiments, any one of or any combination of corresponding positions R3, E4, R9, K12, D15, E16, K19, R23, S26, S28, N29, T30, E31, E32, D33, E35, Q36, E37, E39, D40 with reference to SEQ ID NO: 32, can be mutated to another hydrophilic residue without significantly impacting solubility or TfR-binding. In some embodiments, a TfR-binding peptide disclosed herein comprisesX1X2REX3X4X5X6RX7X8KX9X10DEX11X12KX13X14X15RX16X17SX18SNTEEDX19EQEX20EDX 21X22X23X24X25X26X27X28X29X30X31 (SEQ ID NO: 156), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, X10, Xu, X12, X13, X14, X15, X16, X17, X18, X19, X20, X21, X22, X23, X24, X25, X26, X27, X28, X29, X30, and X31 can independently be any amino acid. In some embodiments, a TfR-binding peptide disclosed herein comprises XiX2GCASX3CMX4YNX5X6LEX7CEAX8MMX9MXioXiiXi2Xi3Xi4Xi5CXi6Xi7Xi8LXi9X2oL LYCLDHCHSQ (SEQ ID NO: 171) or GSXiX2GCASX3CMX4YNX5X6LEX7CEAX8MMX9MXioXiiXi2Xi3Xi4Xi5CXi6Xi7Xi8LXi9X2 oLLYCLDHCHSQ (SEQ ID NO: 157), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, Xio, Xn, Xi2, X13, X14, X15, Xi6, X17, Xis, X19, and X20 can be independently selected from D, E, H, K, R, N, Q,S, or T.

[0240] In some embodiments, a peptide of the present disclosure comprises cysteine amino acid residues at corresponding positions 4, 8, 18, 32, 42, and 46 with reference to SEQ ID NO: 96. In some embodiments, a peptide of the present disclosure comprises cysteine amino acid residues at corresponding positions 6, 10, 20, 34, 44, and 48 with reference to SEQ ID NO: 32. In some embodiments, a peptide of the present disclosure comprises hydrophilic residues (e.g., D, E, H, K, R, N, Q, S, or T) at corresponding positions 15, 35, 39, 49, with reference to SEQ ID NO: 32, or any combination thereof. In some instances, a peptide of the present disclosure comprises hydrophilic amino acid residues at the following corresponding positions: D15, E35, E39, H49, with reference to SEQ ID NO: 32, or any combination thereof. In some embodiments, any oneof or any combination of corresponding positions D15, E35, E39, H49 with reference to SEQ ID NO: 32, can be mutated to another hydrophilic residue without significantly impacting solubility or TfR-binding. In some embodiments, a TfR-binding peptide disclosed herein comprises. In some embodiments, a TfR-binding peptide disclosed herein comprisesXlX2X3X4X5X6X7X8X9XloXl 1X12X13X14DX15X16X17X18X19X20X21X22X23X24X25X26X27X28X29X 30X31X32X33EX34X35X36EX37X38X39X40X41X42X43X44X45HX46X47 (SEQ ID NO: 158), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, X10, Xu, X12, X13, X14, X15, Xi6, X17, Xis, X19, X20, X21, X22, X23, X24, X25, X26, X27, X28, X29, X30, X31, X32, X33, X34, X35, X36, X37, X38, X39, X40, X41, X42, X43, X44, X45, X46, and X47 can independently be any amino acid. In some embodiments, a TfR- binding peptide disclosed herein comprises REGCASRCMKYNX1ELEKCEARMMSMSNTEEDCX2QELX3DLLYCLDHCX4SQ (SEQ ID NO: 172) orGSREGCASRCMKYNX1ELEKCEARMMSMSNTEEDCX2QELX3DLLYCLDHCX4SQ (SEQ ID NO: 159), wherein Xi, X2, X3, and X4 can be independently selected from D, E, H, K, R, N, Q, S, or T.

[0241] In some embodiments, a peptide of the present disclosure comprises hydrophobic residues (e.g., A, M, I, L, V, F, W, or Y) at corresponding positions 15, 35, 39, 49, with reference to SEQ ID NO: 32, or any combination thereof. In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCMKYNX1ELEKCEARMMSMSNTEEDCX2QELX3DLLYCLDHCX4SQ (SEQ ID NO: 173) or GSREGCASRCMKYNX1ELEKCEARMMSMSNTEEDCX2QELX3DLLYCLDHCX4SQ (SEQ ID NO: 160), wherein Xi, X2, X3, and X4 can be independently selected from A, M, I, L, V, F, W, or Y. In some embodiments, hydrophilic amino acid residues at any one of the corresponding positions 15, 35, 39, and 49, with reference to SEQ ID NO: 32, are associated with higher binding affinity for TfR (e.g., target engagement) and higher solubility. In some embodiments, mutation of an amino acid residue at any one of the corresponding positions 15, 35, 39, and 49, with reference to SEQ ID NO: 32, from a hydrophobic to a hydrophilic residue can lead to higher binding affinity for TfR (e.g., target engagement) and higher solubility.

[0242] In some embodiments, a peptide of the present disclosure comprises hydrophobic residues (e.g., A, M, I, L, V, F, W, or Y) at corresponding positions 11, 25, 27, with reference to SEQ ID NO: 32, or any combination thereof. In some embodiments, a peptide of the present disclosure comprises hydrophilic residues (e.g., D, E, H, K, R, N, Q, S, or T) at correspondingpositions 11, 25, 27, with reference to SEQ ID NO: 32, or any combination thereof. In some embodiments, hydrophobic amino acid residues at any one of the corresponding positions 11 , 25, and 27, with reference to SEQ ID NO: 32, are associated with higher binding affinity for TfR (e.g., target engagement) and higher solubility. In some embodiments, mutation of an amino acid residue at any one of the corresponding positions 11, 25, and 27, with reference to SEQ ID NO: 32, from a hydrophilic residue to a hydrophobic residue can lead to higher binding affinity for TfR (e.g., target engagement) and higher solubility. In some embodiments, a peptide of the present disclosure comprises hydrophobic amino acid residues at the corresponding positions Mi l, M25, M27, with reference to SEQ ID NO: 32, or any combination thereof. In some instances, a peptide comprises the hydrophobic amino acid residues at the corresponding positions Mi l, M25, and M27, with reference to SEQ ID NO: 32. In some embodiments, any combination of the corresponding positions Mi l, M25, and M27, with reference to SEQ ID NO: 32, can be mutated to another hydrophobic residue without significantly impacting solubility or TfR-binding. In some embodiments, a TfR-binding peptide disclosed herein comprises X1X2X3X4X5X6X7X8X9X1OMX11X12X13X14X15X16X17X18X19X2OX21X22X23MX24MX25X26X27X28 X29X30X31X32X33X34X35X36X37X38X39X40X41X42X43X44X45X46X47X48(SEQ ID NO: 161), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, Xio, Xu , X12, X13, Xi4, X15, Xi6,Xi7, Xi8, X19, X20, X21, X22, X23, X24, X25, X26, X27, X28, X29, X30, X3I , X32, X33, X34, X35, X36, X37, X38, X39, X40, X4I , X42, X43, X44, X45, X46, X47, and X48can independently be any amino acid. In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCX1KYNDELEKCEARMX2SX3SNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 174) or GSREGCASRCX1KYNDELEKCEARMX2SX3SNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 162), wherein Xi, X2, and X3 can be independently selected from A, M, I, L, V, F, W, or Y. In some embodiments, a TfR-binding peptide disclosed herein comprises REGCASRCX1KYNDELEKCEARMX2SX3SNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 175) or GSREGCASRCX1KYNDELEKCEARMX2SX3SNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 163), wherein Xi, X2, and X3 can be independently selected from D, E, H, K, R, N, Q, S, or T.

[0243] In some embodiments, a peptide of the present disclosure comprises an aliphatic amino acid residue (e.g., A, M, I, L, or V) at corresponding position 45, with reference to SEQ ID NO: 32. In some embodiments, a peptide of the present disclosure comprises an aromatic amino acidresidue (e.g., F, W, or Y) at corresponding position 45, with reference to SEQ ID NO: 32. In some embodiments, an aliphatic amino acid residue at corresponding position 45, with reference to SEQ ID NO: 32, is associated with higher binding affinity to TfR. In some instances, a peptide comprises the aliphatic amino acid residue corresponding to L45, with reference to SEQ ID NO: 32. In some embodiments, mutation of an amino acid residue at corresponding position 45 from an aromatic residue to an aliphatic reside can lead to higher binding affinity for TfR (e.g., target engagement) and higher solubility. In some embodiments, mutating corresponding position L45, with reference to SEQ ID NO: 32, to another aliphatic residue may not significantly impact solubility or TfR-binding. In some embodiments, a TfR-binding peptide disclosed herein comprisesXlX2X3X4X5X6X7X8X9XloXl 1X12X13X14X15X16X17X18X19X20X21X22X23X24X25X26X27X28X29X30 X31X32X33X34X35X36X37X38X39X40X41X42X43X44LX45X46X47X48X49X50 (SEQ ID NO: 164), wherein Xi, X2, X3, X4, X5, X6, X7, X8, X9, X10, Xu, X12, X13, Xi4, X15, Xi6,Xi7, Xis, X19, X20, X21, X22, X23, X24, X25, X26, X27, X28, X29, X30, X31, X32, X33, X34, X35, X36, X37, X38, X39, X40, X41, X42, X43, X44, X45, X46, X47, X48, X49, and X50 can independently be any amino acid. In some embodiments, a TfR-binding peptide disclosed herein comprises GSREGCASRCMKYNDELEKCEARMMSMSNTEEDCEQELEDLLYCXiDHCHSQ (SEQ ID NO: 165) or REGCASRCMKYNDELEKCEARMMSMSNTEEDCEQELEDLLYCXiDHCHSQ (SEQ ID NO: 176), wherein Xi can be independently selected from A, M, I, L, or V.

[0244] In some embodiments, a peptide of the present disclosure comprises GSREGCASRCMX1YNDELEX2CEARMMSMSNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 166) or REGCASRCMX1YNDELEX2CEARMMSMSNTEEDCEQELEDLLYCLDHCHSQ (SEQ ID NO: 177), wherein Xi and X2 can be independently selected from K or R. In some embodiments, these residues at corresponding position 12 and 19, with reference to SEQ ID NO: 32, can be used for chemical conjugation to another molecule (e.g., an active or a detectable agent). In some embodiments, Xi and X2 are both R and chemical conjugation occurs at the N-terminus of the peptide.

[0245] In some embodiments, a receptor-binding peptide may be derived from an antibody or antibody fragment. For example, a receptor-binding peptide may be derived from a single chain antibody fragment (scFv). Examples of TfR-binding peptides that may be incorporated into a selective depletion complex of the present disclosure include SEQ ID NO: 220(QVQLQESGGGWQPGRSLRLSCAASRFTFSSYAMHWVRQAPGKGLEWVAVISYDGSNKYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDLSGYGDYPDYWGQGT LVTVSSGGGGSGGGGSGGGGSSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQK PGQAPVLVMYGRNERPSGVPDRFSGSKSGTSASLAISGLQPEDEANYYCAGWDDSLTG PVFGGGTKLTVLG), SEQ ID NO: 221 (QVQLQESGGGWQPGRSLRLSCAASRFTFNNYAMHWVRQAPGKGLEWVAVISYDGS NKYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDLSGYGDYPDYWGQ GTLVTVSSGGGGSGGGGSGGGGSSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQ QKPGQAPVLVMYGRNERPSGVPDRFSGSKSGTSASLAISGLQPEDEANYYCAGWDDSL TGPVFGGGTKLTVLG), and SEQ ID NO: 222 (QVQLQESGGGWQPGRSLRLSCAASRYPFHHHDHHWVRQAPGKGLEWVAVISYDGS NKYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDLSGYGDYPDYWGQ GTLVTVSSGGGGSGGGGSGGGGSSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQ QKPGQAPVLVMYGRNERPSGVPDRFSGSKSGTSASLAISGLQPEDEANYYCAGWDDSL TGPVFGGGTKLTVLG). In some embodiments, a TfR-binding peptide may have a sequence of any one of SEQ ID NO: 220 - SEQ ID NO: 222, or a fragment thereof. In some embodiments, aTfR-binding peptide may have a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 220 - SEQ ID NO: 222, or a fragment thereof. In some embodiments, a peptide of SEQ ID NO: 220 or SEQ ID NO: 221 may function as a pH- independent TfR-binding peptide. In some embodiments, a peptide of SEQ ID NO: 222 may function as a pH-dependent TfR-binding peptide.

[0246] In some embodiments, mutations in any one or more of the amino acid residues of a peptide of the present disclosure can improve binding affinity of the peptide to TfR. In some embodiments, mutations in 5-80% of amino acid residues of a peptide of the present disclosure improve the binding affinity of the peptide to TfR. In some embodiments, mutations in 1-100%, 5-100%, or 5-50% of amino acid residues of a peptide of the present disclosure improve binding affinity of the peptide to TfR. In some embodiments, mutations in 15-50% of amino acid residues of a peptide of the present disclosure improve binding affinity of the peptide to TfR. In some embodiments, mutations in 15-30% of amino acid residues of a peptide of the present disclosure improve binding affinity of the peptide to TfR. In some embodiments, mutations in 25-30% of amino acid residues of a peptide of the present disclosure improve binding affinity of the peptide to TfR. For example, mutations in 14 of the 51 amino acid residues (27.5%) of apeptide having a sequence of SEQ ID NO: 32 can improve binding affinity of the peptide to TfR.

[0247] In some embodiments, mutations in any one or more of the amino acid residues of a peptide of the present disclosure can lie at the binding interface of TfR. In some embodiments, a mutation to a peptide can improve binding affinity, which can be beneficial to binding and transcytosis of a peptide or peptide complex disclosed herein. In some embodiments, the peptides provided herein can have many mutations or few mutations to obtain optimal activity, wherein optimal activity is sufficient binding for engagement of the TfR, but not necessarily binding that is so strong as to preclude release of the peptide and / or peptide complex after transcytosis. Thus, peptides of the present disclosure can comprise a number of mutations (also referred to as % mutated amino acid residues) that tune binding affinity and off rate to obtain optimal binding, function (e.g., transcytosis, BBB-penetration, cell membrane penetration, transport across a biological barrier, endocytosis, recycling, or combinations thereof), and release of the peptide or peptide complex. Thus, mutations that result in the highest possible affinity may not necessarily correlate to a superior peptide having optimal binding and transcytosis.

[0248] In some embodiments, 1-100% or 5-100% of amino acid residues of a peptide of the present disclosure lie at the binding interface of TfR. In some embodiments, 10-90% of amino acid residues of a peptide of the present disclosure lie at the binding interface of TfR. In some embodiments, 20-80% of amino acid residues of a peptide of the present disclosure lie at the binding interface of TfR. In some embodiments, 30-70% of amino acid residues of a peptide of the present disclosure lie at the binding interface of TfR. In some embodiments, 40-60% of amino acid residues of a peptide of the present disclosure lie at the binding interface of TfR. In some embodiments, 30-35% of amino acid residues of a peptide of the present disclosure lie at the binding interface of TfR. For example, 17 of the 51 amino acid residues (33%) of a peptide having a sequence of SEQ ID NO: 32 can lie at the binding interface of TfR.

[0249] In some embodiments, mutations in any one or more of the amino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR can improve binding affinity of the peptide to TfR. In some embodiments, mutations in 1-100% or 5-100% of amino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 5-80% of amino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 10-70% ofamino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 15-60% of amino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 20-50% of amino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 25-30% of amino acid residues of a peptide of the present disclosure that lie at the binding interface of TfR improve binding affinity of the peptide to TfR. For example, mutations in 5 of the 17 amino acid residues (29%) of a peptide having a sequence of SEQ ID NO: 32 that lie at the binding interface of TfR and can improve binding affinity of the peptide to TfR.

[0250] In some embodiments, mutations in any one or more of the amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. In some embodiments, 1-100% or 5-100% of amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. In some embodiments, 10-90% of amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. In some embodiments, 20-80% of amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. In some embodiments, 30-70% of amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. In some embodiments, 40- 60% of amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. In some embodiments, 65-70% of amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR. For example, 34 of the 51 amino acid residues (66%) of a peptide having a sequence of SEQ ID NO: 32 can lie at the binding interface of TfR.

[0251] In some embodiments, mutations in any one or more of the amino acid residues of a peptide of the present disclosure are distal to the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 1-100% or 5-100% of amino acid residues of a peptide of the present disclosure that are distal to the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 5-80% of amino acid residues of a peptide of the present disclosure that are distal to the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 10- 70% of amino acid residues of a peptide of the present disclosure that are distal to the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 15-60% of amino acid residues of a peptide of the present disclosure that are distalto the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 20-50% of amino acid residues of a peptide of the present disclosure that are distal to the binding interface of TfR improve binding affinity of the peptide to TfR. In some embodiments, mutations in 25-30% of amino acid residues of a peptide of the present disclosure that are distal to the binding interface of TfR improve binding affinity of the peptide to TfR. For example, mutations in 5 of the 17 amino acid residues that are distal to the binding interface of TfR can improve binding affinity of the peptide to TfR. For example, mutations in 9 of the 34 amino acid residues (26.5%) of a peptide having a sequence of SEQ ID NO: 32 that are distal to the binding interface of TfR can improve binding affinity of the peptide to TfR. In some embodiments, and without being bound to any theory, one or more mutations in the amino acid residues of the peptide that are distal to the binding interface of TfR can improve protein folding, enhance protein solubility, and / or alter the backbone geometry that can improve binding through an optimized interface shape complementarity.

[0252] It is understood that for any of the foregoing peptide or peptide complex of the present disclosure describing the mutations and amino acid substitutions and revisions with reference to SEQ ID NO: 32, that the mutations and amino acid substitutions comprise at least one or more of the corresponding residues in SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64.

[0253] In some embodiments, a receptor-binding peptide of the present disclosure may be a PD- L1 -binding peptide. The PD-L1 -binding peptide may be incorporated into a selective depletion complex of the present disclosure to facilitate selective depletion of a target molecule via PD- Ll-mediated endocytosis. In some embodiments, the PD-Ll-binding peptide that is a receptorbinding peptide may bind PD-L1 with an affinity that is pH-independent (for example, a similar affinity at extracellular pH and at an endosomal pH) or may bind PD-L1 with an affinity that is pH-dependent (for example, a higher affinity at extracellular pH and a lower affinity at an endosomal pH). Examples of PD-Ll-binding peptides are provided in TABLE 2.TABLE 2 - Exemplary PD-Ll-Binding Peptides

[0254] In some embodiments, a PD-L1 -binding peptide disclosed herein comprises a sequence of X1X2X3CX4X5X6CX7X8X9X10X11X12X13X14X15CX16X17X18X19X20X21X22X23X24X25X26X27X28C X29X30X31X32X33X34X35X36X37CX38X39X40CX41X42X43(SEQ ID NO: 392), wherein X1can independently be selected from E, M, V, or W; X2can independently be selected from G, E, L, or F; X3can independently be selected from D, E, or S; X4can independently be selected from K, R, or V; X5can independently be selected from E, Q, S, M, L, or V; X6can independently be selected from D, E, H, K, R, N, Q, S, or Y; X7can independently be selected from D, M, or V;X8can independently be selected from A, K, R, Q, S, or T; X9can independently be selected from A, D, E, H, Q, S, T, M, I, L, V, or W; X10can independently be selected from A, E, R, Q, S, T, W, or P; X11can independently be selected from A, E, K, R, N, Q, T, M, I, L, V, or W; X12can independently be selected from G, A, E, K, N, T, or Y; X13can independently be selected from G, A, D, E, H, K, R, N, Q, S, T, M, I, L, V, W, Y, or P; X14can independently be selected from D, K, R, N, L, or V; X15can independently be selected from G, A, D, T, L, W, or P; X16can independently be selected from G, A, E, H, K, N, S, F, or P; X17can independently be selected from G, A, D, E, N, or P; X18can independently be selected from G, D, H, K, R, N, Q, S, T, V, or Y; X19can independently be selected from G, D, E, H, K, N, Q, S, T, M, I, F, W, Y, or P; X20can independently be selected from G, A, D, E, H, K, R, N, Q, S, Y, or P; X21can independently be selected from G, A, D, H, N, Q, S, V, F, or P; X22can independently be selected from A, D, H, N, Q, S, T, M, I, V, Y, or P; X23can independently be selected from G, A, D, K, R, T, W, or Y; X24can independently be selected from G, A, E, N, Q, T, I, V, or P; X25can independently be selected from G, D, N, Q, T, L, V, F, or P; X26can independently be selected from G, A, E, K, R, N, Q, S, T, I, Y, or P; X27can independently be selected from A, D, N, or I; X28can independently be selected from G, D, E, H, N, F, or W; X29can independently be selected from G, A, E, N, S, Y, or P; X30can independently be selected from G, M, or L; X31can independently be selected from G, A, D, K, N, Q, or W; X32can independently be selected from D, E, H, K, N, Q, S, T, L, V, F, Y, or P; X33can independently be selected from G, E, Q, or F; X34can independently be selected from D or K; X35can independently be selected from G, V, or P; X36can independently be selected from G, H, R, V, F, W, or P; X37can independently be selected from A, D, or K; X38can independently be selected from E, H, Q, L, or F; X39can independently be selected from D, E, R, S, T, M, L, or F; X40can independently be selected from G, A, D, E, H, K, R, M, L, or P; X41can independently be selected from G, A, K, S, I, or L; X42can independently be selected from G, A, D, E, R, Q, T, or F; and X43can independently be selected from A, H, N, Q, S, F, or P.

[0255] In some embodiments, a binding peptide disclosed herein comprises a sequence of EEDCKVX1CVX1X1X1X1X2X3KX1CX1EX1X4X1X1X1X1X1X1X1AX1CX1GX1X5FX6VFX6CLX 'X'CX'X'X1(SEQ ID NO: 393), wherein X1can independently be selected from any noncysteine amino acid; X2can independently be selected from M, I, L, or V; X3can independently be selected from Y, A, H, K, R, N, Q, S, or T; X4can independently be selected from D, E, N, Q, or P; X5can independently be selected from K or P; and X6can independently be selected from D or K.

[0256] A PD-L1 -binding peptide may comprise a PD-L1 -binding motif that forms part or all of a binding interface with PD-L1. One or more residues of a PD-L1 -binding motif may interact with one or more residues of PD-L1 at the binding interface between the PD-L1 -binding peptide and PD-L1. In some embodiments, multiple PD-L1 -binding motifs may be present in a PD-L1- binding peptide. A PD-L1 -binding motif may comprise a sequence of CX1X2X3CX4X5X6X7X8X9X10X11X12C (SEQ ID NO: 394), wherein X1can independently be selected from K, R, or V; X2can independently be selected from E, Q, S, M, L, or V; X3can independently be selected from D, E, H, K, R, N, Q, S, or Y; X4can independently be selected from D, M, or V; X5can independently be selected from A, K, R, Q, S, or T; X6can independently be selected from A, D, E, H, Q, S, T, M, I, L, V, or W; X7can independently be selected from A, E, R, Q, S, T, W, or P; X8can independently be selected from A, E, K, R, N, Q, T, M, I, L, V, or W; X9can independently be selected from G, A, E, K, N, T, or Y; X10can independently be selected from G, A, D, E, H, K, R, N, Q, S, T, M, I, L, V, W, Y, or P; X11can independently be selected from D, K, R, N, L, or V; and X12can independently be selected from G, A, D, T, L, W, or P. In some embodiments, a PD-Ll-binding motif may comprise a sequence of CKVX1CVX1X1X1X1X2X3KX1C (SEQ ID NO: 396), wherein X1can independently be selected from any non-cysteine amino acid; X2can independently be selected from M, I, L, or V; and X3can independently be selected from Y, A, H, K, R, N, Q, S, or T. In some embodiments, a PD-Ll-binding motif may comprise a sequence of CKVHCVKEWMAGKAC (SEQ ID NO: 398). In some embodiments, a PD-Ll-binding motif may comprise at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to SEQ ID NO: 398.

[0257] A PD-Ll-binding motif may comprise a sequence of X1X2X3X4X5X6CX7X8X9C (SEQ ID NO: 395), wherein X1can independently be selected from D, E, H, K, N, Q, S, T, L, V, F, Y, or P; X2can independently be selected from G, E, Q, or F; X3can independently be selected from D or K; X4can independently be selected from G, V, or P; X5can independently be selected from G, H, R, V, F, W, or P; X6can independently be selected from A, D, or K; X7can independently be selected from E, H, Q, L, or F; X8can independently be selected from D, E, R, S, T, M, L, or F; and X9can independently be selected from G, A, D, E, H, K, R, M, L, or P. In some embodiments, a PD-Ll-binding motif may comprise a sequence of X1FX2VFX2CLX3X3C (SEQ ID NO: 397), wherein X1can independently be selected from K or P; X2can independently be selected from D or K; and X3can independently be selected from any noncysteine amino acid. In some embodiments, a PD-Ll-binding motif may comprise a sequence ofKFDVFKCLDHC (SEQ ID NO: 399). In some embodiments, a PD-L1 -binding motif may comprise at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% identity to SEQ ID NO: 399.

[0258] A PD-Ll-binding peptide (e.g., any one of SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241, or a pH-independent variant thereof) with high affinity PD-Ll-binding at endosomal pH may be complexed with a target-binding peptide as described herein to form a selective depletion complex for selective depletion of the target molecule. The selective depletion complex can be used to selectively deliver a target molecule across a cellular layer or membrane. For example, the selective depletion complex can be used to selectively deliver the target molecule to an endocytic compartment via PD-L1 -mediated endocytosis. The target molecule can be selectively depleted upon binding to the target-binding peptide of the selective depletion complex and endocytosis via PD-L1 -mediated endocytosis as described.

[0259] Selective depletion of a target molecule using PD-L1 -mediated endocytosis may be used to selectively deplete the target molecule specifically in tissues that express PD-L1. In some embodiments, a selective depletion complex comprising a receptor-binding peptide that binds PD-L1 may be used to selectively deplete a target molecule in a PD-L1 positive cancer, a lung tissue, a pancreatic islet tissue, a lymphoid tissue, an immune cell, a gastrointestinal tissue, a bone marrow tissue, a reproductive tissue, a muscle tissue, an adipose tissue, or any other PD-L1 positive tissue. For example, a selective depletion complex comprising a PD-Ll-binding peptide and an ACE2-binding peptide may be used to selectively deplete ACE2 in lung tissue to prevent a viral infection (e.g., a SARS-CoV-2 infection). In another example, a selective depletion complex comprising a PD-Ll-binding peptide and an HLA-b inding peptide may be used to selectively deplete HLA in pancreatic islet cells to prevent T-cell attack of insulin-expressing cells in type I diabetes.Target-Binding Peptides

[0260] Peptides, peptide complexes, or selective depletion complexes of the present disclosure can comprise a target-binding peptide (e.g., a target-binding EGF variant). The target-binding peptide can be capable of binding a target molecule (e.g., EGFR). In some embodiments, the target-binding peptide can bind to the target molecule with an affinity that is pH-dependent. For example, the target-binding peptide can bind the target molecule with a higher affinity at anextracellular pH (such as about pH 7.4) than at an endosomal pH (such as about pH 5.5). A target-binding peptide can be conjugated to a receptor-binding peptide of the present disclosure (e.g., a TfR-binding peptide any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64 or a PD-Ll-binding peptide of any one of SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241) to form a selective depletion complex. The selective depletion complex can be used to selectively deliver a target molecule across a cellular layer or membrane (e.g., BBB or cell membrane). For example, the selective depletion complex can be used to selectively deliver the target molecule (e.g., EGFR) to an endocytic compartment via receptor-mediated endocytosis (e.g., PD-L1 -mediated endocytosis or TfR-mediated endocytosis). The target molecule (e.g., EGFR) can be selectively depleted upon binding to the target-binding peptide of the selective depletion complex and endocytosis via receptor-mediated endocytosis. The target molecule can be an EGFR protein. Selective depletion of a cell surface molecule (e.g., a receptor such as EGFR) using a selective depletion complex comprising a target-binding peptide that binds to the cell surface molecule can result in a reduction of the cell surface molecule (e.g., a surface exposed protein). The surface exposed protein can be associated with a disease or a condition. In some embodiments, a selective depletion complex of the present disclosure can comprise two or more target-binding peptides to promote dimerization of a target molecule. Promoting dimerization can increase internalization of the target molecule, resulting in selective depletion of the target molecule. For example, a selective depletion complex comprising two copies of a target-binding peptide can promote homodimerization of the target molecule.

[0261] In some embodiments, the EGF variant peptide of the present disclosure may further comprise a miniprotein, a nanobody, an antibody, an IgG, an antibody fragment, a Fab, a F(ab)2, an scFv, an (scFv)2, a DARPin, or an affibody. In some embodiments, the target-binding peptide may comprise a cystine-dense peptide, an affitin, an adnectin, an avimer, a Kunitz domain, a nanofittin, a fynomer, a bicyclic peptide, a beta-hairpin, or a stapled peptide. In some embodiments, an EGF variant peptide can further comprise an antibody (e.g., IgG or other antibody), an antibody fragment, (e.g., scFv, scFv2, Fab, F(ab)2, or other antibody fragment), or a nanobody (e.g., a VHH-domain nanobody or VNAR-domain nanobody from camelids or sharks), which can be stable at a low pH.

[0262] In some embodiments, a target-binding peptide of the present disclosure (e.g., an EGFR- binding peptide) can bind to the target molecule (e.g., EGFR) with an affinity that is pH-dependent. For example, the target-binding peptide can bind the target molecule at an extracellular pH (such as about pH 7.4) with an affinity that is higher than the binding affinity at an endocytic pH (such as about pH 7.0, pH 6.5, pH 6.0, pH 5.8, or pH 5.5). In some embodiments, the binding affinity of the target-binding peptide for the target molecule at an extracellular pH (about pH 7.4) can be at least about 1.1 -fold, at least about 1.2-fold, at least about 1.3-fold, at least about 1.4-fold, at least about 1.5-fold, at least about 1.6-fold, at least about 1.7-fold, at least about 1.8-fold, at least about 1.9-fold, at least about 2-fold, at least about 2.5-fold, at least about 3-fold, at least about 3.5-fold, at least about 4-fold, at least about 4.5- fold, at least about 5-fold, at least about 6-fold, at least about 7-fold, at least about 8-fold, at least about 9-fold, at least about 10-fold, at least about 15-fold, at least about 20-fold, at least about 25-fold, at least about 30-fold, at least about 35-fold, at least about 40-fold, at least about 45- fold, at least about 50-fold, at least about 60-fold, at least about 70-fold, at least about 80-fold, at least about 90-fold, at least about 100-fold, at least about 200-fold, at least about 500-fold, at least about 1000-fold, at least about 10,000-fold the binding affinity of the target-binding peptide for the target molecule at an endosomal pH (such as about pH 7.0, pH 6.5, pH 6.0, pH 5.8, pH 5.5, or pH 5.0). In some embodiments, the affinity of the target-binding peptide for the target at pH 6.5 or pH 5.5 is no greater than about 0.1%, about 0.5%, about 1%, about 5%, about 10%, about 20%, about 30%, about 40%, or about 50% the affinity of the target-binding peptide for the target at pH 7.4. In some embodiments, the affinity of the target-binding peptide for the target at pH 7.4 is at least 2-fold, at least 3 -fold, at least 4-fold, at least 5 -fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the target-binding peptide for the target molecule at pH 6.5 or pH 5.5

[0263] In some embodiments, a target-binding peptide with pH-dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of less than 50 pM, less than 5 pM, less than 500 nM, less than 100 nM, less than 40 nM, less than 30 nM, less than 20 nM, less than 10 nM, less than 5 nM, less than 2 nM, less than 1 nM, less than 0.5 nM, less than 0.4 nM, less than 0.3 nM, less than 0.2 nM, or less than 0.1 nM at extracellular pH (such as about pH 7.4). In some embodiments, a target-binding peptide with pH-dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of at least 1 nM, at least 2 nM, at least 5 nM, at least 10 nM, at least 20 nM, at least 50 nM, at least 100 nM, at least 200 nM, at least 500 nM, at least 1 pM, at least 2 pM, at least 5 pM, at least 10 pM, at least 20 pM, at least 50 pM, at least 100 pM, at least 500 pM, at least 1 mM, at least 2 mM, at least 5 mM, at least 10 mM, at least 20 mM, at least 50 mM, at least 100 mM, at least 200 mM, at least 500 mM, or atleast 1 M at endosomal pH (about pH 5.5 or about pH 6.5). In some embodiments, a targetbinding peptide with pH-dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of at least 1 nM, at least 2 nM, at least 5 nM, at least 10 nM, at least 20 nM, at least 50 nM, at least 100 nM, at least 200 nM, at least 500 nM, at least 1 pM, at least 2 pM, at least 5 pM, at least 10 pM, at least 20 pM, at least 50 pM, at least 100 pM, at least 500 pM, at least 1 mM, at least 2 mM, at least 5 mM, at least 10 mM, at least 20 mM, at least 50 mM, at least 100 mM, at least 200 mM, at least 500 mM, or at least 1 M at endosomal pH (about pH 5.8).

[0264] In some embodiments, a target-binding peptide with pH-dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 7.4. In some embodiments, a target-binding peptide with pH- dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.5. In some embodiments, a targetbinding peptide with pH-dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.8.

[0265] In some embodiments, the affinity of the target-binding peptide with pH-dependent binding to the target molecule at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25- fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some embodiments, the affinity of the target-binding peptide with pH-dependent binding to the target molecule at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25-fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

[0266] In some embodiments, a target-binding peptide with pH-dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, orno more than 0.1 nM at pH 7.4. In some embodiments, a target-binding peptide with pH- dependent binding can bind a target molecule with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 6.5.

[0267] In some embodiments, a target-binding peptide with pH-dependent binding can bind a target molecule with a dissociation rate constant (koff or kd) of no more than 1x10'1s’1, 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’ no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, no more than 2x1 O’4s’1, no more than 1x1 O’4s’1, no more than 5xl0’5s’1, or no more than 2xl0’5s’1at pH of 7.4. In some embodiments, a target-binding peptide with pH-dependent binding can bind a target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 1x10’1s’1, 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, no more than 2x1 O’4s’1, no more than 1x1 O’4s’1, no more than 5xl0’5s’1, or no more than 2xl0’5s’1at pH 6.5. In some embodiments, a targetbinding peptide with pH-dependent binding can bind a target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’ or no more than 2xl0’4s’1at pH 5.5. In some embodiments, a target-binding peptide with pH- dependent binding can bind a target molecule with a dissociation rate constant (koff or kd) of no more than 1 s’1, no more than 5x10’1s’1, no more than 2x10’1s’1, no more than 1x1 O’2s’1, no more than 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, or no more than 2x1 O’4s’1at pH 5.8.

[0268] In some embodiments, the dissociation rate constant (koff or kd) of the target-binding peptide with pH-dependent binding to the target molecule at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold. In some embodiments, the dissociation rate constant (koff or kd) of the target-binding peptide with pH-dependent binding to the target molecule at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no morethan 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

[0269] In some embodiments, the dissociation rate constant (koff or kd) of the target-binding peptide with pH-dependent binding to the target molecule at pH 7.4 and at pH 5.5 at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.5 than at pH 7.4. In some embodiments, the dissociation rate constant (koff or kd) of the target-binding peptide with pH-dependent binding to the target molecule at pH 7.4 and at pH 5.5 at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.8 than at pH 7.4.

[0270] In some embodiments, the target-binding molecule can release the target molecule upon internalization into an endosomal compartment and acidification of the endosome. Such release the target molecule upon acidification of the endosome can occur at about pH 7.3, pH 7.2, pH 7.1, pH 7.0, pH 6.9, pH 6.8, pH 6.7, pH 6.6, pH 6.5, pH 6.4, pH 6.3, pH 6.2, pH 6.1, pH 6.0, pH 5.9, pH 5.8, pH 5.7, pH 5.6, pH 5.5, pH 5.4, pH 5.3, pH 5.2, pH 5.1, pH 5.0, pH 4.9, pH 4.8, pH 4.7, pH 4.6, pH 4.5, or lower. In some embodiments, release of the target molecule can occur at a pH of from about pH 7.0 to about pH 4.5, from about pH 6.5 to about pH 5.0, or from about pH 6.0 to about pH 5.5 or lower.

[0271] Target-binding peptides with pH-dependent binding affinity can be engineered by selective integration of histidine (His) amino acid residues in the target-binding interface. In some instances, a target-binding peptide with pH-dependent binding affinity comprises at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 histidine residues in the target-binding interface. Since the side chain of histidine is predominantly uncharged at pH between about 6.0 and about 9.2 and predominantly positively charged at pH below about 6.0, selectively inserting or removing His residues in a target-binding peptide can impart pH-dependent binding properties. A target-binding peptide can be stable at low pH (e.g., at endosomal pH).

[0272] In some embodiments, release of the target molecule by the target-binding peptide upon internalization into an endosomal compartment can be affected by differences in the ionic strength between the extracellular physiologic environment and endosomal cellularcompartments. In some embodiments, the ionic strength of the endosomal compartment is higher than the ionic strength of the extracellular physiologic environment. Ionic strength, which varies with salt concentration, may depend on the concentrations of various electrolytes in solution, for example hydrogen (H+), hydroxide (OH’), hydronium (H3O+), sodium (Na+), potassium (K+), calcium (Ca2+), magnesium (Mg2+), manganese (Mn2+), chloride (Cl’), carbonate (CO32’), cobalt (Co2+), phosphate (PCU3'), or nitrate (NO,-). In some embodiments, targetbinding peptides with salt-dependent or ionic strength-dependent binding affinity can be engineered by selective integration of salt labile moieties (e.g., polar or charged amino acid side chains) in the target-binding interface that would enable dissociation of the target-binding molecule in the endosome. For example, the target-binding interface of the target-binding peptide may form one or more polar or charge-charge interactions with the target-binding peptide that can be disrupted as the ionic strength of the environment increases.

[0273] In some instances, a target-binding peptide with a binding affinity dependent on ionic strength (e.g., dependent on hydrogen, hydroxide, hydronium, sodium, potassium, calcium, magnesium, manganese, chloride, carbonate, cobalt, phosphate, and / or nitrate concentration) could dissociate over a range of ionic strengths, for example ionic strengths from about 30 mM to about 1 M. In some embodiments, an ionic strength-dependent target-binding peptide with a binding affinity dependent on ionic strength could dissociate at an ionic strength of from about 50 mM to about from about 50 mM to about 1 M, from about 60 mM to about 950 mM, from about 70 mM to about 900 mM, from about 80 mM to about 850 mM, from about 90 mM to about 800 mM, from about 100 mM to about 750 mM, from about 110 mM to about 700 mM, from about 120 mM to about 650 mM, from about 130 mM to about 600 mM, from about 140 mM to about 550 mM, from about 150 mM to about 500 mM, from about 160 mM to about 450 mM, from about 170 mM to about 400 mM, from about 180 mM to about 350 mM, from about 190 mM to about 300 mM, or from about 200 mM to about 250 mM. In some embodiments, the ionic strength-dependent target-binding peptide comprises at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 polar or charge-charge interactions in the target-binding interface (e.g., the interface between EGFR and the EGFR-b inding peptide).

[0274] A target-binding peptide of the present disclosure may bind to a target (e.g., a target molecule), such as EGFR, EGFRvIII, or TKI-resistant EGFR. The target molecule (e.g., EGFR, EGFRvIII, or TKI-resistant EGFR) may be endocytosed and degraded upon binding to thetarget-binding peptide of a selective depletion complex. The amino acid sequence of soluble EGFRvIII ectodomain (EGFRvIII) is provided as follows:LEEKKGNYWTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKVCNGIGIGEFKDS LSINATNIKHFKNCTSISGDLHILPVAFRGDSFTHTPPLDPQELDILKTVKEITGFLLIQAW PENRTDLHAFENLEIIRGRTKQHGQFSLAWSLNITSLGLRSLKEISDGDVIISGNKNLCY ANTINWKKLFGTSGQKTKIISNRGENSCKATGQVCHALCSPEGCWGPEPRDCVSCRNVS RGRECVDKCNLLEGEPREFVENSECIQCHPECLPQAMNITCTGRGPDNCIQCAHYIDGPH CVKTCPAGVMGENNTLVWKYADAGHVCHLCHPNCTYGCTGPGLEGCPTNGPKIPS (SEQ ID NO: 527). In some embodiments, soluble EGFRvIII (e.g., SEQ ID NO: 527) may also comprise a labelling or affinity tag. In some embodiments, soluble EGFRvIII (e.g., SEQ ID NO: 527) may comprise one or more of a G4S spacer (e.g., SEQ ID NO: 136), a poly-His tag (e.g., SEQ ID NO: 142), a biotinylation tag (e.g., SEQ ID NO: 543), or any combination thereof. In some embodiments, soluble EGFRvIII (e.g., SEQ ID NO: 527) may be in a biotinylated form. In some embodiments, soluble EGFRvIII (e.g., SEQ ID NO: 527) may comprise a biotinylation tag (e.g., SEQ ID NO: 543) and undergo biotinylation by a BirA enzyme.

[0275] A target-binding peptide of the present disclosure may bind to a target molecule, such as a target molecule with clinical relevance. In some embodiments, a target molecule may be a protein that is over-expressed or over-activated in a disease or condition. For example, a target molecule may be an EGFR transmembrane protein involved in oncogenic signaling. The target molecule (e.g., EGFR) may be endocytosed and degraded upon binding to the target-binding peptide of a selective depletion complex.

[0276] In some embodiments, a target molecule may be a transmembrane protein, such as a receptor tyrosine kinase. Examples of receptor tyrosine kinases that may be targeted using a selective depletion complex include EGF receptor, or ErbB. In some embodiments, the receptor tyrosine kinase may be EGFR. Targeting the transmembrane protein using a selective depletion complex may lead to internalization and degradation of the transmembrane protein

[0277] Endocytosis and subsequent degradation of the target molecule may treat (e.g., eliminate, reduce, slow progression of, or treat symptoms of) a disease or condition associated with the target molecule. In some embodiments, the degradation of the target molecule may cause remission in, reduce, ameliorate, or ablate a disease or condition (e.g., a cancer). In some embodiments, the degradation of the target molecule may cause remission in, reduce, ameliorate, or ablate a cancer. In some embodiments, targeting and degradation of a receptor tyrosine kinase with a selective depletion complex may be beneficial in treating a variant ofcancers. For example, targeting and degrading EGFR with a selective depletion complex comprising an EGFR-binding peptide may be beneficial in treating cancers, such as non-small- cell lung cancer, primary non-small-cell lung cancer, metastatic non-small-cell lung cancer, head and neck cancer, head and neck squamous cell carcinoma, glioblastoma, brain cancer, metastatic brain cancer, colorectal cancer, colon cancer, tyrosine kinase inhibitor (TKI)-resistant cancer, cetuximab-resistant cancer, necitumumab-resistant cancer, panitumumab-resistant cancer, local cancer, regionally advanced cancer, recurrent cancer, metastatic cancer, refractory cancer, KRAS wildtype cancer, KRAS mutant cancers, or exon20 mutant non-small-cell lung cancer.

[0278] Additionally, the selective depletion complexes of the present disclosure may be well- suited for treatment of CNS-associated disorders such as EGFR-driven brain cancers due to the ability of the selective depletion complexes to penetrate the blood-brain barrier (BBB) and access the CNS via TfR-binding. A selective depletion complex (e.g., comprising a TfR-binding peptide) may facilitate higher BBB penetration.

[0279] In some embodiments, binding and subsequently depleting a target molecule using a selective depletion complex of the present disclosure comprising a target-binding peptide may be used to treat a disease or condition wherein the target molecule is a cell-based or soluble moiety associated with a disease or condition and is expressed or present in diseased tissues or cells. In some embodiments, depletion of the target molecule may be cell type or tissue dependent. For example, depletion of a target molecule may be specific to cells or tissues expressing both the target molecule targeted by the target-binding peptide of the selective depletion complex and the cell surface receptor targeted by the receptor-binding peptide of the selective depletion complex. The degradation and depletion and of the target molecule using a selective depletion complex may prevent, treat, or ameliorate the disease or condition.

[0280] In some embodiments, a target-binding peptide (e.g., a target-binding EGF variant) may comprise a sequence of any one of SEQ ID NO: 317 - SEQ ID NO: 390, or SEQ ID NO: 457 - SEQ ID NO: 494. In some embodiments, a target-binding peptide may comprise a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 318 - SEQ ID NO: 390, or SEQ ID NO: 457 - SEQ ID NO: 494, or a fragment thereof. For example, a target-binding peptide may comprise a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 318 - SEQ ID NO: 390, or SEQ ID NO: 457 -SEQ ID NO: 494. Examples of target-binding peptides and their corresponding target molecules are provided in TABLE 3.

[0281] In some embodiments, a target-binding peptide (e.g., an EGFR target-binding peptide) may comprise a sequence of any one of SEQ ID NO: 318 - SEQ ID NO: 390, or SEQ ID NO: 457 - SEQ ID NO: 494. In some embodiments, a target-binding peptide (e.g., an EGFR targetbinding peptide) may comprise a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 318 - SEQ ID NO: 390, or SEQ ID NO: 457 - SEQ ID NO: 494, or a fragment thereof.

[0282] In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 458 - SEQ ID NO: 494, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of any one of SEQ ID NO: 458 - SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has at least 80% sequence identity with any one of SEQ ID NO: 458 - SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has at least 90% sequence identity with any one of SEQ ID NO: 458 - SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has at least 95% sequence identity with SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence of any one of SEQ ID NO: 458 - SEQ ID NO: 494.

[0283] In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 494, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR targetbinding peptide) comprises a sequence that has at least 80% sequence identity with SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has at least 90% sequence identity with SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence that has atleast 95% sequence identity with SEQ ID NO: 494. In some aspects, a target-binding peptide (e.g., an EGFR target-binding peptide) comprises a sequence of SEQ ID NO: 494.

[0284] Examples of EGFR target-binding peptides are provided in TABLE 3.TABLE 3 - Exemplary Target-Binding Peptides that Bind EGER as a Target-Ill-

[0285] In some embodiments, a target-binding peptide disclosed herein (e.g., an EGFR targetbinding peptide) comprises a sequence of XlX2X3X4X5CX6X7X8X9X10XllX12CX13X14Xl5GX16CX17YSXl8X19LX20X21YX22CNCX23X24G YX25GX26RCQX27X28X29X3oX3iX32X33 (SEQ ID NO: 314) wherein Xi is any amino acid; X2is any amino acid; X3 is any amino acid; X4 is any of Lys, Ser, He, or Vai; X5 is any of Glu, Lys, or Asn; Xe is Asp or Pro; X7 is any amino acid; X8is any of Gly, Ser, or Met; X9 is any of His, Leu, or Trp; X10 is Arg or Vai; Xu is any of Gly, Asp, Lys, Arg, Asn, or Met; X12 is Phe or Tyr; X13 is any of Met, Leu, or Phe; X14 is His or Gin; X15 is any of Asp, Arg, Asn; Xi6 is any of Gin, Met, Vai; X17 is any of His, Arg, Asn, Vai; Xi8is any of Glu, Lys, Gin, Ser, Ala, Vai; X19 is any of Glu, Ser, or Ala; X20 is Asp or Asn; X21 is any of Lys, Thr, He, or Leu; X22 is He or Trp; X23 is Ser or Thr; X24 is any of Glu, He, or Phe; X25 is any amino acid; X26 is any of Asp, Glu, or Pro; X27 is Ser or Tyr; X28is any of His, Lys, Arg, or Gin; X29 is Asp or Thr; X30 is Leu or Phe; X31 is any of Gly, Asn, Ser, Thr; X32 is any amino acid; and X33 is any of Met, Leu, Phe, Trp.

[0286] An EGFR-binding peptide of the present disclosure may comprise a configuration (e.g., a cysteine configuration), one or more amino acid substitutions, or a combination thereof, relative to an EGF peptide (e.g., a human EGF peptide of SEQ ID NO: 317), wherein “relative to” the EGF peptide describes the amino acid positions of the EGFR-binding peptide relative to the amino acid positions of the EGF peptide. For example, an EGFR-binding peptide of the present disclosure may comprise 53 amino acids described by positions 1-53 relative to positions 1-53 of SEQ ID NO: 317. In another example, an EGFR-binding peptide of the present disclosure may comprise 50 amino acids described by positions 1-50 relative to positions 1-50 of SEQ ID NO: 317. In some embodiments, an EGFR-binding peptide may comprise additional amino acid positions on the N terminus of the peptide relative to an EGF peptide (e.g., a human EGF peptide of SEQ ID NO: 317) which can be described as amino acid positions 0 through -n relative to the EGF peptide (e.g., a human EGF peptide of SEQ ID NO: 317). For example, an EGFR-binding peptide may comprise an additional 5 amino acids on the N terminus as compared to an EGF peptide (e.g., a human EGF peptide of SEQ ID NO: 317) that can be described as amino acid positions 0, -1, -2, -3, and -4 relative to the EGF peptide (e.g., a humanEGF peptide of SEQ ID NO: 317). In some embodiments, an EGFR-binding peptide may comprise additional amino acid positions on the C terminus of the peptide which can be described as amino acid positions of the last position number +1 through the last position number +n (e.g., 54 through 53+n relative to SEQ ID NO: 317) relative to the EGF peptide (e.g., a human EGF peptide of SEQ ID NO: 317). For example, an EGFR-binding peptide may comprise an additional 5 amino acids on the C terminus as compared to an EGF peptide of SEQ ID NO: 317 that can be described as amino acid positions 54, 55, 56, 57, and 58 relative to the EGF peptide of SEQ ID NO: 317. The amino acid positions of an EGFR-binding peptide relative to an EGF variant may describe a position of a specific amino acid (e.g., a position of a cysteine amino acid). The amino acid positions of an EGFR-binding peptide relative to an EGF variant may describe a position of an amino acid substitution using amino acid substitution nomenclature of “(the original amino acid)(the position of the amino acid substitution)(the substituted amino acid).” For example, an EGFR-binding peptide with a DI 1R amino acid substitution relative to a human EGF peptide of SEQ ID NO: 317 describes the amino acid substitution of R in place of D at position 11 of the EGFR-binding peptide.

[0287] In some embodiments, an EGFR target-binding peptide may comprise a cysteine configuration of that of an EGF peptide (e.g., a human EGF peptide). For example, an EGFR target-binding peptide may comprise cysteine amino acids at positions 6, 14, 20, 31, 33, and 42 of the EGFR target-binding peptide, not counting tags, linkers, or other components complexed with the EGFR target-binding peptide. In some embodiments, an EGFR target-binding peptide may comprise cysteine residues spaced by 7 amino acid residues between a first cysteine and a second cysteine, 5 amino acid residues between the second cysteine and a third cysteine, 10 amino acid residues between the third cysteine and a fourth cysteine, 1 amino acid residue between the fourth cysteine and a fifth cysteine, and 8 amino acid residues between the fifth cysteine and a sixth cysteine (SEQ ID NO: 711). An EGFR target-binding peptide may comprise a truncation relative to an EGF peptide. In some embodiments, an EGFR target-binding peptide may comprise 1, 2, 3, 4, or 5 amino acids truncated from the C-terminus relative to EGF. In some embodiments, an EGFR target-binding peptide may comprise 1, 2, 3, 4, or 5 amino acids truncated from the N-terminus relative to EGF. An EGFR target-binding peptide may comprise at least 70%, at least 75%, at least 77%, at least 80%, at least 82%, at least 85%, at least 87%, at least 90%, at least 92%, at least 94%, or at least 96% sequence identity to an EGF peptide (e.g., a human EGF) and may further comprise at least 1 , at least 2, at least 3, at least 4, or at least 5 amino acid substitutions relative to the EGF peptide. The EGFR target-binding peptide maycomprise amino acid substitutions DI 1R, I23S, V35E, S51P, L52E, R53E, M21R, A30W, I38D, W49R, V34S, Q43I, Q43V, Q43W, Q43Y, K48N, K48T, K48A, K48L, E51S, E51H, L52H, R53H, or combinations thereof, relative to human EGF. For example, an EGFR target-binding peptide may comprise substitutions V34S, K48T, E51S, or any combination thereof, relative to human EGF. In another example, an EGFR target-binding peptide may comprise substitutions M21R, A30W, I38D, W49R, or combinations thereof, relative to human EGF. In another example, an EGFR target-binding peptide may comprise substitutions DI 1R, I23S, V35E, S51P, L52E, R53E, or combinations thereof, relative to human EGF. In another example, an EGFR target-binding peptide may comprise substitutions E51H, L52H, R53H, or combinations thereof, relative to human EGF.Cystine-Dense Peptides

[0288] In some embodiments, TfR-binding peptides or PD-Ll-binding peptides or targetbinding peptides (e.g., EGFR target-binding peptides) of...

Claims

CLAIMSWHAT IS CLAIMED IS:

1. An EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R.

2. The EGFR-binding peptide of claim 1, wherein the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof.

3. The EGFR-binding peptide of claim 1 or claim 2, wherein the sequence further comprises one or more mutations relative to SEQ ID NO: 317 comprising DI 1R, I23S, V35E, E51P, L52E, R53E, or a combination thereof.

4. The EGFR-binding peptide of any one of claims 1-3, wherein the sequence further comprises one or more mutations relative to SEQ ID NO: 317 comprising E51H, L52H, R53H, or a combination thereof.

5. The EGFR-binding peptide of any one of claims 1-4, wherein the EGFR-binding peptide comprises a sequence having at least 80%, at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 458 - SEQ ID NO: 493 or a fragment thereof.

6. The EGFR-binding peptide of any one of claims 1-5, wherein the sequence comprises SEQ ID NO: 494.

7. The EGFR-binding peptide of any one of claims 1-5, wherein the sequence comprises any one of SEQ ID NO: 458 - SEQ ID NO: 493.

8. The EGFR-binding peptide of any one of claims 1-7, wherein the fragment comprises at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, or at least 50 amino acid residues.

9. The EGFR-binding peptide of any one of claims 1-8, wherein the EGFR-binding peptide is capable of binding to EGFR without activating the EGFR.

10. The EGFR-binding peptide of any one of claims 1-9, wherein the EGFR-binding peptide blocks binding of EGF to EGFR when the EGFR-binding peptide is bound to the EGFR.

11. The EGFR-binding peptide of any one of claims 1-10, wherein the EGFR-binding peptide inhibits EGFR when the EGFR-binding peptide is bound to the EGFR.

12. The EGFR-binding peptide of any one of claims 1-11, wherein the EGFR-binding peptide prevents dimerization of EGFR when the EGFR-binding peptide is bound to the EGFR.

13. A peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein:there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptor-binding peptide; and the EGFR-binding peptide has affinity for a target molecule.

14. The peptide complex of claim 13, wherein the EGFR-binding peptide is the EGFR- binding peptide of any one of claims 1-11.

15. The peptide complex of claim 13 or claim 14, wherein the affinity of the EGFR-binding peptide for the target molecule, the affinity of the cellular receptor-binding peptide for a cellular receptor, or both is pH-independent.

16. The peptide complex of any one of claims 13-15, wherein the affinity of the EGFR- binding peptide for the target molecule, the affinity of the cellular receptor-binding peptide for the cellular receptor, or both is pH dependent.

17. The peptide complex of any one of claims 13-16, wherein the affinity of the EGFR- binding peptide for the target molecule, the affinity of the cellular receptor-binding peptide for the cellular receptor, or both is ionic strength dependent.

18. The peptide complex of any one of claims 13-17, wherein the cellular receptor-binding peptide is a transferrin receptor-binding peptide or a PD-L1 -binding peptide.

19. The peptide complex of any one of claims 13-18, wherein the cellular receptor is a transferrin receptor or PD-L1.

20. The peptide complex of any one of claims 13-19, wherein the cellular receptor is a cation-independent mannose 6 phosphate receptor (CI-M6PR), an asialoglycoprotein receptor (ASGPR), CXCR7, folate receptor, or Fc receptor (including but not limited to neonatal Fc receptor (FcRn) or FcyRIIb).

21. The peptide complex of any one of claims 13-20, wherein the cellular receptor-binding peptide binds to the cellular receptor at a pH of from pH 4.5 to pH 7.4, from pH 5.5 to pH 7.4, from pH 5.8 to pH 7.4, or from pH 6.5 to pH 7.4.

22. The peptide complex of any one of claims 13-21, wherein the cellular receptor-binding peptide is capable of binding the cellular receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 7.4.

23. The peptide complex of any one of claims 13-22, wherein the cellular receptor-binding peptide is capable of binding the cellular receptor with an equilibrium dissociation constant (KD) of no more than 100 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, or no more than 0.1 nM at pH 5.5.

24. The peptide complex of any one of claims 13-23, wherein the cellular receptor-binding peptide is capable of binding the cellular receptor with a dissociation rate constant (koff or kd) of no more than 1 s'1, no more than 5x10'1s'1, no more than 2x10'1s'1, no more than 1x1 O'2s'1, no more than 5x1 O'2s'1, no more than 2x1 O'2s'1, no more than 1x1 O'2s'1, no more than 5x1 O'3s'1, no more than 2x1 O'3s'1, no more than 1x1 O'3s'1, no more than 5x1 O'4s'1, or no more than 2x1 O'4s'1at pH 5.5.

25. The peptide complex of any one of claims 13-24, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor is pH-independent.

26. The peptide complex of any one of claims 13-25, wherein the affinity of the EGFR- binding peptide for the target molecule is pH-dependent.

27. The peptide complex of any one of claims 13-26, wherein the affinity of the EGFR- binding peptide for the target molecule is pH-independent.

28. The peptide complex of any one of claims 13-27, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

29. The peptide complex of any one of claims 13-28, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15-fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

30. The peptide complex of any one of claims 13-29, wherein the dissociation rate constant (koff or kd) of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.5 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15- fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

31. The peptide complex of any one of claims 13-30, wherein the dissociation rate constant (koff or kd) of the cellular receptor-binding peptide for the cellular receptor at pH 7.4 and at pH 5.8 differs by no more than 2-fold, no more than 5-fold, no more than 10-fold, no more than 15- fold, no more than 20-fold, no more than 25 -fold, no more than 30-fold, no more than 40-fold, or no more than 50-fold.

32. The peptide complex of any one of claims 13-31, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor is pH dependent.

33. The peptide complex of claim 32, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor decreases as pH decreases.

34. The peptide complex of claim 32 or claim 33, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor is higher at pH 7.4 than at pH 5.5.

35. The peptide complex of claim 32 or claim 33, wherein the affinity of the cellular receptor-binding peptide for the cellular receptor is higher at pH 7.4 than at pH 5.8.

36. The peptide complex of any one of claims 13-35, wherein the affinity of the EGFR- binding peptide for the target molecule is pH dependent.

37. The peptide complex of any one of claims 13-36, wherein the affinity of the EGFR- binding peptide for the target molecule decreases as pH decreases.

38. The peptide complex of any one of claims 13-37, wherein the affinity of the EGFR- binding peptide for the target molecule is higher at a higher pH than at a lower pH.

39. The peptide complex of claim 38, wherein the higher pH is pH 7.4, pH 7.2, pH 7.0, or pH 6.8.

40. The peptide complex of claim 38 or claim 39, wherein the lower pH is pH 6.5, pH 6.0, pH 5.8, pH 5.5, pH 5.0, or pH 4.5.

41. The peptide complex of any one of claims 13-40, wherein the affinity of the EGFR- b inding peptide for the target molecule is higher at pH 7.4 than at pH 6.0.

42. The peptide complex of any one of claims 13-41, wherein the affinity of the EGFR- binding peptide for the target molecule is higher at pH 7.4 than at pH 5.5.

43. The peptide complex of any one of claims 13-42, wherein the affinity of the EGFR- b inding peptide for the target molecule is higher at pH 7.4 than at pH 5.8.

44. The peptide complex of any one of claims 13-43, wherein the EGFR-b inding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no more than 500 nM, no more than 200 nM, 100 nM, no more than 50 nM, no more than 20 nM, no more than 10 nM, no more than 5 nM, no more than 2 nM, no more than 1 nM, no more than 0.5 nM, no more than 0.4 nM, no more than 0.3 nM, no more than 0.2 nM, no more than 1 nM, or no more than 0.1 nM at pH 7.4.

45. The peptide complex of any one of claims 13-44, wherein the EGFR-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1x10'1s’1, 5x1 O’2s’1, no more than 2x1 O’2s’1, no more than 1x1 O’2s’1, no more than 5x1 O’3s’1, no more than 2x1 O’3s’1, no more than 1x1 O’3s’1, no more than 5x1 O’4s’1, no more than 2x10’4s’1, no more than IxlO’4s’1, no more than 5xl0’5s’1, or no more than 2xl0’5s’1at pH 7.4.

46. The peptide complex of any one of claims 13-45, wherein the EGFR-b inding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1 s'1, no more than 5x1 O'1s'1, no more than 2x1 O'1s'1, no more than 1x1 O'2s'1, no more than 5x1 O'2s'1, no more than 2x1 O'2s'1, no more than 1x1 O'2s'1, no more than 5x1 O'3s'1, no more than 2x1 O'3s'1, no more than 1x1 O'3s'1, no more than 5x1 O'4s'1, or no more than 2x1 O'4s'1at pH 5.5.

47. The peptide complex of any one of claims 13-46, wherein the EGFR-binding peptide is capable of binding the target molecule with a dissociation rate constant (koff or kd) of no more than 1 s'1, no more than 5x1 O'1s'1, no more than 2x1 O'1s'1, no more than 1x1 O'2s'1, no more than 5x1 O'2s'1, no more than 2x1 O'2s'1, no more than 1x1 O'2s'1, no more than 5x1 O'3s'1, no more than 2x1 O'3s'1, no more than 1x1 O'3s'1, no more than 5x1 O'4s'1, or no more than 2x1 O'4s'1at pH 5.8.

48. The peptide complex of any one of claims 13-47, wherein the dissociation rate constant (koff or kd) for EGFR-binding peptide binding the target molecule is at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.5 than at pH 7.4.

49. The peptide complex of any one of claims 13-48, wherein the dissociation rate constant (koff or kd) for EGFR-binding peptide binding the target molecule is at least 2 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 50 fold, at least 100 fold, at least 200 fold, at least 500 fold, at least 1,000 fold, at least 2,000 fold, at least 5,000 fold, at least 10,000 fold, at least 20,000 fold, or at least 50,000 fold higher at pH 5.8 than at pH 7.4.

50. The peptide complex of any one of claims 13-49, wherein the EGFR-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10 nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.5.

51. The peptide complex of any one of claims 13-50, wherein the EGFR-binding peptide is capable of binding the target molecule with an equilibrium dissociation constant (KD) of no less than 0.1 nM, no less than 0.5 nM, 1 nM, no less than 2 nM, no less than 5 nM, no less than 10nM, no less than 20 nM, no less than 50 nM, no less than 100 nM, no less than 200 nM, or no less than 500 nM, or no less than 1000 nM at pH 5.8.

52. The peptide complex of any one of claims 13-51, wherein the affinity of the EGFR- binding peptide for the target molecule at pH 7.4 is at least 1.5-fold, 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the EGFR-binding peptide for the target molecule at pH 5.5.

53. The peptide complex of any one of claims 13-51, wherein the affinity of the EGFR- binding peptide for the target molecule at pH 7.4 is at least 1.5-fold, at least 2-fold, at least 3- fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 15-fold, or at least 20-fold greater than the affinity of the EGFR-binding peptide for the target molecule at pH 5.8.

54. The peptide complex of any one of claims 13-53, wherein the affinity of the EGFR- binding peptide for the target molecule at pH 7.4 is less than 0.5-fold, less than 1-fold, less than, 1.5-fold, less than 2-fold, less than 3-fold, or less than 10-fold, greater than the affinity of the EGFR-binding peptide for the target molecule at pH 5.8.

55. The peptide complex of any one of claims 13-54, wherein the EGFR-binding peptide comprises one or more histidine amino acid residues.

56. The peptide complex of any one of claims 13-55, wherein the affinity of the EGFR- binding peptide for the target molecule decreases as ionic strength increases.

57. The peptide complex of any one of claims 13-56, wherein the EGFR-binding peptide comprises one or more polar or charged amino acid residues capable of forming polar or chargecharge interactions with the target molecule.

58. The peptide complex of any one of claims 13-57, wherein the cellular receptor-binding peptide is fused to, linked to, complexed with, or conjugated to the EGFR-binding peptide.

59. The peptide complex of any one of claims 13-58, wherein the cellular receptor-binding peptide is fused to, linked to, complexed with, or conjugated to the EGFR-binding peptide via a polymer linker.

60. The peptide complex of claim 59, wherein the polymer linker is a polyethylene glycol (PEG), a hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a zwitterionic water soluble polymer, a water soluble poly(amino acid), a water soluble polymer comprising proline, alanine, serine, or a combination thereof, a water soluble polymer containing glycine, glutamic acid, and serine, an Fc region, a fatty acid, a palmitic acid, an albumin, or an albumin binding molecule.

61. The peptide complex of any one of claims 13-60 wherein the cellular receptor-binding peptide and the EGFR-binding peptide form a single polypeptide chain.

62. The peptide complex of any one of claims 13-61, wherein the peptide complex comprises a dimer dimerized via a dimerization domain.

63. The peptide complex of any one of claims 13-62, wherein a distance between the cellular receptor-binding peptide and the EGFR-binding peptide is at least 1 nm, at least 2 nm, at least 5 nm, at least 10 nm, at least 20 nm, at least 50 nm, or at least 100 nm.

64. The peptide complex of claim 62, wherein the dimerization domain comprises an Fc domain.

65. The peptide complex of claim 62, wherein the dimer is a homodimer dimerized via a homodimerization domain.

66. The peptide complex of claim 65, wherein the homodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 535, SEQ ID NO: 706, or SEQ ID NO: 246.

67. The peptide complex of claim 62, wherein the dimer is a heterodimer dimerized via a first heterodimerization domain and a second heterodimerization domain.

68. The peptide complex of claim 67, wherein the first heterodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 536, SEQ ID NO: 707, or SEQ ID NO: 709.

69. The peptide complex of claim 67 or claim 68, wherein the second heterodimerization domain comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 537, SEQ ID NO: 708, or SEQ ID NO: 710.

70. The peptide complex of any one of claims 62-69, wherein the EGFR-binding peptide is linked to the dimerization domain via a peptide linker.

71. The peptide complex of any one of claims 62-70, wherein the cellular receptor-binding peptide is linked to the dimerization domain via a peptide linker.

72. The peptide complex of any one of claims 13-71, wherein the cellular receptor-binding peptide is linked to the EGFR-binding peptide via a peptide linker.

73. The peptide complex of claim 70-72, wherein the peptide linker has a length of from 1 to 50 amino acid residues, from 2 to 40 amino acid residues, from 3 to 20 amino acid residues, or from 3 to 10 amino acid residues.

74. The peptide complex of any one of claims 71-73, wherein the peptide linker comprises glycine and serine amino acids.

75. The peptide complex of any one of claims 71-74, wherein the peptide linker has a persistence length of no more than 6 A, no more than 8 A, no more than 10 A, no more than 12 A, no more than 15 A, no more than 20 A, no more than 25 A, no more than 30 A, no more than 40 A, no more than 50 A, no more than 75 A, no more than 100 A, no more than 150 A, no more than 200 A, no more than 250 A, or no more than 300 A.

76. The peptide complex of any one of claims 71-75, wherein the peptide linker is derived from an immunoglobulin peptide.

77. The peptide complex of any one of claims 71-75, wherein the peptide linker is derived from a double-knot toxin peptide.

78. The peptide complex of any one of claims 71-77, wherein the peptide linker comprises a sequence of any one of SEQ ID NO: 129 - SEQ ID NO: 141, SEQ ID NO: 195 - SEQ ID NO: 218, SEQ ID NO: 223 - SEQ ID NO: 223 - SEQ ID NO: 227, SEQ ID NO: 194, SEQ ID NO: 391, SEQ ID NO: 538, or SEQ ID NO: 540 - SEQ ID NO: 541.

79. The peptide complex of any one of claims 13-78, wherein the cellular receptor-binding peptide, the EGFR-binding peptide, peptide complex, or a combination thereof comprises a miniprotein, a nanobody, an antibody, an antibody fragment, an scFv, a DARPin, or an affibody.

80. The peptide complex of claim 79, wherein the antibody comprises an IgG, or wherein the antibody fragment comprises a Fab, a F(ab)2, an scFv, or an (scFv)2.

81. The peptide complex of claim 79 or claim 80, wherein the miniprotein comprises a cystine-dense peptide, an affitin, an adnectin, an avimer, a Kunitz domain, a nanofittin, a fynomer, a bicyclic peptide, a beta-hairpin, or a stapled peptide.

82. The peptide complex of any one of claims 13-81, wherein the cellular receptor-binding peptide comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds.

83. The peptide complex of any one of claims 13-82, wherein the EGFR-binding peptide comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds.

84. The peptide complex of any one of claims 13-83, wherein the peptide complex comprises at least one disulfide bond, at least two disulfide bonds, at least three disulfide bonds, or at least four disulfide bonds.

85. The peptide complex of any one of claims 13-84, wherein the cellular receptor-binding peptide comprises at least six cysteine residues.

86. The peptide complex of claim 85, wherein the at least six cysteine residues are positioned at amino acid positions 4, 8, 18, 32, 42, and 46 of the cellular receptor-binding peptide.

87. The peptide complex of claim 85 or claim 86, wherein the at least six cysteine residues form at least three disulfide bonds.

88. The peptide complex of any one of claims 13-87, wherein the cellular receptor-binding peptide comprises a sequence of any one of SEQ ID NO: 148 - SEQ ID NO: 177.

89. The peptide complex of any one of claims 13-88, wherein the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of any one of SEQ ID NO: 96, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67 - SEQ ID NO: 95, SEQ ID NO: 97 - SEQ ID NO: 128, SEQ ID NO: 220 - SEQ ID NO: 222, or SEQ ID NO: 1 - SEQ ID NO: 64.

90. The peptide complex of any one of claims 13-89, wherein the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 96, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 96.

91. The peptide complex of any one of claims 13-90, wherein the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 96.

92. The peptide complex of any one of claims 13-90, wherein the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 66, or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 66.

93. The peptide complex of any one of claims 13-90 or claim 92, wherein the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 66.

94. The peptide complex of any one of claims 13-90, wherein the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 65, or at least 80%, at least 90%, at least 92%, at least 93%, at least94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 65.

95. The peptide complex of any one of claims 13-90 or claim 94, wherein the cellular receptor-binding peptide comprises a sequence of SEQ ID NO: 65.

96. The peptide complex of any one of claims 13-87, wherein the cellular receptor-binding peptide comprises a sequence of any one of SEQ ID NO: 392 - SEQ ID NO: 399.

97. The peptide complex of any one of claims 13-87 or claim 96, wherein the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 187, SEQ ID NO: 233 - SEQ ID NO: 239, SEQ ID NO: 400 - SEQ ID NO: 456, or SEQ ID NO: 241.

98. The peptide complex of any one of claims 13-87, 96, or 97, wherein the cellular receptor-binding peptide comprises a sequence that has at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 187, SEQ ID NO: 235, SEQ ID NO: 236, SEQ ID NO: 238, SEQ ID NO: 239, SEQ ID NO: 400, or SEQ ID NO: 401 or at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with a fragment of SEQ ID NO: 187.

99. The peptide complex of any one of claims 13-87 or 96-98, wherein the cellular receptorbinding peptide comprises a sequence of SEQ ID NO: 187.

100. The peptide complex of any one of claims 89-99, wherein the fragment comprises at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36, at least 37, at least 38, at least 39, at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, or at least 50 amino acid residues.

101. The peptide complex of any one of claims 13-100, wherein the cellular receptor-binding peptide comprises one or more histidine residues at a cellular receptor-binding interface.

102. The peptide complex of any one of claims 13-101, wherein the EGFR-binding peptide comprises one or more histidine residues at a EGFR-binding interface.

103. The peptide complex of any one of claims 13-102, wherein the target molecule comprises an EGFR.

104. The peptide complex of claim 103, wherein the EGFR is wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximab-resistant EGFR, a panitumumab-resistant EGFR, or a combination thereof.

105. The peptide complex of claim 104, wherein the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

106. The peptide complex of any one of claims 13-105, wherein an off rate of the cellular receptor-binding peptide from the cellular receptor is slower than a recycling rate of the cellular receptor.

107. The peptide complex of any one of claims 13-106, wherein a half-life of dissociation of the cellular receptor-binding peptide from the cellular receptor is no faster than 1 minute, no faster than 2 minutes, no faster than 3 minutes, no faster than 4 minutes, no faster than 5 minutes, no faster than 7 minutes, no faster than 10 minutes, no faster than 15 minutes, no faster than 20 minutes, no faster than 30 minutes, no faster than 45 minutes, no faster than 60 minutes, no faster than 90 minutes, or no faster than 120 minutes.

108. The peptide complex of any one of claims 13-107, wherein a rate of dissociation of the EGFR-binding peptide from the target molecule is faster than a recycling rate of the cellular receptor.

109. The peptide complex of any one of claims 13-108, wherein a half-life of dissociation of a target binding-binding peptide from the target molecule is less than 10 seconds, less than 20seconds, less than 30 seconds, less than 1 minute, less than 2 minutes, less than 5 minutes, less than 10 minutes, less than 20 minutes, less than 30 minutes, less than 45 minutes, or less than 60 minutes in endosomal conditions.

110. The peptide complex of any one of claims 13-109, wherein the peptide complex is capable of being endocytosed via receptor-mediated endocytosis.

111. The peptide complex of claim 110, wherein the receptor-mediated endocytosis is transferrin receptor-mediated endocytosis.

112. The peptide complex of claim 110, wherein the receptor-mediated endocytosis is PD-L1- mediated endocytosis.

113. The peptide complex of any one of claims 13-112, wherein the cellular receptor-binding peptide remains bound to the cellular receptor inside an endocytic vesicle.

114. The peptide complex of any one of claims 13-113, wherein the peptide complex is recycled to the cell surface when the cellular receptor-binding peptide is bound to the cellular receptor and the cellular receptor is recycled.

115. The peptide complex of any one of claims 13-114, wherein the target molecule is released or dissociated from the EGFR-binding peptide after the peptide complex is endocytosed via receptor-mediated endocytosis.

116. The peptide complex of any one of claims 13-115, wherein the target molecule is an extracellular protein, a circulating protein, or a soluble protein.

117. The peptide complex of any one of claims 13-116, wherein the target molecule is a cell surface protein.

118. The peptide complex of any one of claims 13-117, wherein the target molecule is a transmembrane protein.

119. The peptide complex of any one of claims 13-118, further comprising a half-life modifying agent coupled to the cellular receptor-binding peptide, the EGFR-binding peptide, or both.

120. The peptide complex of claim 119, wherein the half-life modifying agent is a polymer, a polyethylene glycol (PEG), a hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a zwitterionic water soluble polymer, a water soluble poly(amino acid), a water soluble polymer of proline, alanine and serine, a water soluble polymer containing glycine, glutamic acid, and serine, an Fc region, a fatty acid, palmitic acid, an albumin, or a molecule that binds to albumin.

121. The peptide complex of claim 120, wherein the molecule that binds to albumin is a serum albumin-binding peptide.

122. The peptide complex of claim 121, wherein the serum albumin-binding peptide comprises a sequence of any one of SEQ ID NO: 178, SEQ ID NO: 179, or SEQ ID NO: 193.

123. The peptide complex of any one of claims 13-122, wherein the cellular receptor-binding peptide, the EGFR-binding peptide, or both is recombinantly expressed.

124. The peptide complex of any one of claims 13-123, wherein the EGFR-binding peptide is configured to dissociate from the target molecule at pH 6.5, pH 6.0, pH 5.8, pH 5.5, pH 5.0, or pH 4.5.

125. The peptide complex of any one of claims 13-124, wherein the cellular receptor-binding peptide is configured to dissociate from the cellular receptor at pH 6.5, pH 6.0, pH 5.5, pH 5.0, or pH 4.5.

126. The peptide complex of any one of claims 13-125, wherein the peptide complex comprises a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 499 - SEQ ID NO: 501, SEQ ID NO: 504, SEQ ID NO: 505, SEQ ID NO: 507, SEQ ID NO: 508, or SEQ ID NO: 511 - SEQ ID NO: 526.

127. The peptide complex of any one of claims 13-126, wherein the peptide complex comprises a sequence having at least 80%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with any one of SEQ ID NO: 495 - SEQ ID NO: 497, SEQ ID NO: 503, or SEQ ID NO: 506.

128. A peptide-active agent complex comprising a peptide complexed to an active agent, wherein the peptide comprises the EGFR-binding peptide of any one of claims 1-12 or the peptide complex of any one of claims 13-127.

129. The peptide-active agent complex of claim 128, wherein the active agent comprises a peptide, a peptidomimetic, an oligonucleotide, a DNA, an RNA, an antibody, a single chain variable fragment (scFv), an antibody fragment, an aptamer, or a small molecule.

130. The peptide-active agent complex of claim 129, wherein the DNA comprises cDNA, ssDNA, or dsDNA.

131. The peptide-active agent complex of claim 129, wherein the RNA comprises RNAi, microRNA, snRNA, dsRNA, or an antisense oligonucleotide.

132. The peptide-active agent complex of any one of claims 128-131, wherein the active agent is a therapeutic agent or a detectable agent.

133. The peptide-active agent complex of claim 132, wherein the detectable agent comprises a dye, a fluorophore, a fluorescent biotin compound, a luminescent compound, a chemiluminescent compound, a radioisotope, nanoparticle, a paramagnetic metal ion, or a combination thereof.

134. The peptide-active agent complex of claim 132, wherein the therapeutic agent comprises a chemical agent, a small molecule, a therapeutic, a drug, a peptide, an antibody protein, any fragment thereof, or any combination thereof.

135. The peptide-active agent complex of any one of claims 132-134, wherein the therapeutic agent comprises an oncology agent, an autoimmune disease agent, an acute and chronic neurodegeneration agent, a pain management agent, or an anti-cancer agent.

136. The peptide-active agent complex of claim 135, wherein the anti-cancer agent comprises a radionuclide, radioisotope, a chemotherapeutic agent, a platinum therapeutic, a toxin, an enzyme, a sensitizing drug, an anti-angiogenic agent, cisplatin, an anti-metabolite, an anti- metabolic therapeutic, a mitotic inhibitor, a growth factor inhibitor, paclitaxel, temozolomide, topotecan, fluorouracil, vincristine, vinblastine, procarbazine, decarbazine, altretamine, methotrexate, mercaptopurine, thioguanine, fludarabine phosphate, cladribine, pentostatin,cytarabine, azacitidine, etoposide, teniposide, irinotecan, docetaxel, doxorubicin, daunorubicin, dactinomycin, idarubicin, plicamycin, mitomycin, bleomycin, tamoxifen, flutamide, leuprolide, goserelin, aminogluthimide, anastrozole, amsacrine, asparaginase, mitoxantrone, mitotane, or amifostine.

137. The peptide-active agent complex of claim 135 or claim 136, wherein the anti-cancer agent targets other oncogenic signaling pathways, targets immune response pathways, directly drives an immune response to cancer cells, or targets disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells.

138. A pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent.

139. The pharmaceutical composition of claim 138, wherein the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof.

140. The pharmaceutical composition of claim 138, wherein the EGFR-binding peptide is the EGFR-binding peptide of any one of claims 1-12.

141. A pharmaceutical composition comprising: a peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; andthe EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent.

142. The pharmaceutical composition of claim 141, wherein the peptide complex is the peptide complex of any one of claims 13-127 or the peptide-active agent complex of any one of claims 128-137.

143. A method of administering a pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent, to a subject.

144. The method of claim 143, wherein the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof.

145. The method of claim 146, wherein the EGFR-binding peptide is the EGFR-binding peptide of any one of claims 1-12.

46. A method of administering a pharmaceutical composition comprising: a peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; and the EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent, a subject.

147. The method of claim 146, wherein the peptide complex is the peptide complex of any one of claims 13-127 or the peptide-active agent complex of any one of claims 128-137.

148. The method of any one of claims 143-147, wherein the pharmaceutical composition is the pharmaceutical composition of any one of claims 138-142.

149. A method of inhibiting EGFR in a subject, the method comprising: administering to the subject a pharmaceutical composition, wherein the pharmaceutical composition comprises: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide binds to EGFR on the cell of the subject and inhibits activation of the EGFR.

150. The method of claim 149, wherein the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof.

151. The method of claim 149, wherein the EGFR-binding peptide is the EGFR-binding peptide of any one of claims 1-12.

152. A method of inhibiting EGFR in a subject, the method comprising: administering to the subject a pharmaceutical composition, wherein the pharmaceutical composition comprises: a peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; andat least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, 138D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; and the EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide binds to EGFR on the cell of the subject and inhibits activation of the EGFR.

153. The method of claim 152, wherein the peptide complex is the peptide complex of any one of claims 13-127 or the peptide-active agent complex of any one of claims 128-137.

154. The method of any one of claims 149-153, wherein the pharmaceutical composition is the pharmaceutical composition of any one of claims 142-145.

155. The method of any one of claims 149-154, wherein the EGFR-binding peptide inhibits activation of the EGFR by disrupting multimerization, dimerization, or heterodimerization of the EGFR on the cell of the subject that expresses EGFR.

156. A method of selectively depleting a target molecule, the method comprising:(a) contacting a peptide complex to a cell expressing a cellular receptor, wherein the peptide complex comprises:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein:there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, 138D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; and the EGFR-binding peptide has affinity for a target molecule;(b) binding the EGFR-binding peptide to the target molecule under extracellular conditions;(c) binding the cellular receptor-binding peptide to the cellular receptor under extracellular conditions; and(d) endocytosing the peptide complex, the target molecule, and the cellular receptor into an endocytic or lysosomal compartment, thereby depleting the target molecule.

157. The method of claim 156, wherein the peptide complex of any one of claims 13-127 or the peptide-active agent complex of any one of claims 128-137.

158. The method of claim 156 or claim 157, further comprising:(e) dissociating the EGFR-binding peptide from the target molecule, the cellular- receptor-binding peptide from the cellular receptor, or both, under endosomal or lysosomal conditions.

159. The method of any one of claims 156-158, further comprising:(f) degrading the target molecule, thereby further depleting the target molecule.

160. The method of any one of claims 156-159, further comprising recycling the peptide complex and the cellular receptor to the cell surface.

161. A method of treating a disease or condition in a subject in need thereof, the method comprising administering a pharmaceutical composition comprising: an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent, to the subject, thereby treating the disease or condition.

162. A method of treating a disease or condition in a subject in need thereof, the method comprising: administering a pharmaceutical composition comprising:an EGFR-binding peptide comprising a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; and a pharmaceutically acceptable excipient or diluent; and delivering the EGFR-binding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-binding peptide inhibits the EGFR on the cell of the subject, thereby treating the disease or condition.

163. The method of claim 161 or claim 162, wherein the EGFR-binding peptide comprises a sequence having at least 85%, at least 90%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO: 494 or a fragment thereof.

164. The method of claim 161 or claim 162, wherein the EGFR-binding peptide is the EGFR- binding peptide of any one of claims 1-12.

165. A method of treating a disease or condition in a subject in need thereof, the method comprising administering a pharmaceutical composition comprising: a peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; and the EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent, to the subject, thereby treating the disease or condition.

166. A method of treating a disease or condition in a subject in need thereof, the method comprising:administering a pharmaceutical composition comprising: a peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acid residue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, 138D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; and the EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent; anddelivering the EGFR-b inding peptide to a cell of the subject that expresses EGFR, wherein the EGFR-b inding peptide inhibits the EGFR on the cell of the subject, thereby treating the disease or condition.

167. The method of claim 165 or claim 166, wherein the peptide complex of any one of claims 13-127 or the peptide-active agent complex of any one of claims 128-137.

168. The method of any one of claims 161-167, wherein the pharmaceutical composition is the pharmaceutical composition of any one of claims 138-142.

169. The method of any one of claims 161-168, wherein the EGFR comprises wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximabresistant EGFR, or a panitumumab-resistant EGFR.

170. The method of claim 169, wherein the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

171. A method of treating a disease or condition in a subject in need thereof, the method comprising:(a) administering a pharmaceutical composition to the subject, wherein the pharmaceutical composition comprises: a peptide complex comprising:(i) a cellular receptor-binding peptide; and(ii) an EGFR-binding peptide, wherein: the EGFR-binding peptide comprises a sequence having at least 80% sequence identity to SEQ ID NO: 494 or a fragment thereof, wherein the sequence comprises: a first cysteine amino acid residue, a second cysteine amino acid residue, a third cysteine amino acid residue, a fourth cysteine amino acidresidue, a fifth cysteine amino acid residue, and a sixth cysteine amino acid residue, wherein: there are seven amino acid residues between the first cysteine amino acid residue and the second cysteine amino acid residue, there are five amino acid residues between the second cysteine amino acid residue and the third cysteine amino acid residue, there are ten amino acid residues between the third cysteine amino acid residue and the fourth cysteine amino acid residue, there is one amino acid residue between the fourth cysteine amino acid residue and the fifth cysteine amino acid residue, and there are eight amino acid residues between the fifth cysteine amino acid residue and the sixth cysteine amino acid residue; and at least six mutations relative to SEQ ID NO: 317 comprising amino acid substitutions of M21R, A30W, V34S, I38D, K48T, and W49R; the EGFR-binding peptide is complexed with the cellular receptorbinding peptide; and the EGFR-binding peptide has affinity for a target molecule; and a pharmaceutically acceptable excipient or diluent;(b) binding the EGFR-binding peptide under extracellular conditions to a target molecule associated with the disease or condition on a cell of the subject expressing the target molecule and a cellular receptor;(c) binding the cellular receptor-binding peptide under extracellular conditions to the cellular receptor on the cell of the subject; and(d) endocytosing the peptide complex, the target molecule, and the cellular receptor, thereby treating the disease or condition.

172. The method of claim 171, wherein the peptide complex is the peptide complex of any one of claims 13-127 or the peptide-active agent complex of any one of claims 128-137.

173. The method of claim 171 or claim 172, wherein the pharmaceutical composition is the pharmaceutical composition of any one of claims 138-142.

174. The method of any one of claims 171-173, further comprising:(e) dissociating the EGFR-binding peptide from the target molecule, the cellular- receptor-binding peptide from the cellular receptor, or both under endosomal conditions.

175. The method of any one of claims 171-174, further comprising:(f) degrading the target molecule.

176. The method of any one of claims 165-175, wherein the target molecule comprises an EGFR.

177. The method of claim 176, wherein the EGFR comprises wild-type EGFR, EGFRvIII, tyrosine kinase inhibitor-resistant EGFR, EGFR containing an exon 19 deletion, EGFR containing an exon21 L858R mutation, EGFR mutant T790M, a cetuximab-resistant EGFR, or a panitumumab-resistant EGFR.

178. The method of claim 177, wherein the tyrosine kinase inhibitor-resistant EGFR comprises a EGFR L692V mutant, EGFR E709K mutant, EGFR L718Q mutant, EGFR L718V mutant, EGFR G719A mutant, EGFR G724S mutant, EGFR L747S mutant, EGFR D761Y mutant, EGFR S768I mutant, EGFR SV768IL mutant, EGFR G769X mutant, EGFR T790M mutant, EGFR L792X mutant, EGFR G796R mutant, EGFR G796S mutant, EGFR G796D mutant, EGFR C797X mutant, EGFR L798I mutant, EGFR V834I mutant, EGFR V834L mutant, EGFR V843I mutant, EGFR T854I mutant, or EGFR H870R mutant.

179. The method of any one of claims 161-178, wherein the disease or condition is a cancer.

180. The method of claim 179, wherein the cancer expresses EGFR, overexpresses EGFR, or contains mutant EGFR.

181. The method of claim 179 or claim 180, wherein the cancer is breast cancer, liver cancer, colon cancer, brain cancer, leukemia, lymphoma, non-Hodgkin lymphoma, myeloma, blood- cell-derived cancer, lung cancer, sarcoma, stomach cancer, a gastrointestinal cancer, glioblastoma, head and neck cancer, squamous head and neck cancer, non-small-cell lungcancer, squamous non-small cell lung cancer, pancreatic cancer, ovarian cancer, endometrial cancer, blood cancer, skin cancer, liver cancer, kidney cancer, or colorectal cancer.

182. The method of any one of claims 179-181, wherein the cancer is TKI -resistant, cetuximab-resistant, necitumumab-resistant, or panitumumab-resistant.

183. The method of any one of claims 179-182, wherein the cancer has one or more of the following: overexpresses EGFR, KRAS mutation, KRAS G12S mutation, KRAS G12C mutation, PTEN loss, EGFR exonl9 deletion, EGFR L858R mutation, EGFR T790M mutation, PIK3CA mutation, TP53 R273H mutation, PIK3CA amplification, PIK3CA G118D, TP53 R273H, EGFR C797X mutation, EGFR G724S mutation, EGFR L718Q mutation, EGFR S768I mutation, an EGFR mutation, a cetuximab-resistant EGFR, a panitumumab-resistant EGFR, or a combination thereof.

184. The method of claim any one of claims 179-183, wherein the cancer expresses or has upregulated c-MET, Her2, Her3 that heterodimerizes with EGFR.

185. The method of any one of claims 179-184, wherein the cancer is a primary cancer, an advanced cancer, a metastatic cancer, a metastatic cancer in the central nervous system, a primary cancer in the central nervous system, metastatic colorectal cancer, metastatic head and neck cancer, metastatic non-small-cell lung cancer, metastatic breast cancer, metastatic skin cancer, a refractory cancer, a KRAS wild type cancer, a KRAS mutant cancer, or an exon20 mutant non-small-cell lung cancer.

186. The method of any one of claims 161-185, further comprising administering an additional therapy to the subject.

187. The method of claim 186, where the additional therapy is adjuvant, first-line, or combination therapy.

188. The method of claim 186 or claim 187, wherein the additional therapy targets other oncogenic signaling pathways, targets immune response pathways, directly drives an immune response to cancer cells, or targets disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells.

189. The method of claim 188, wherein targeting of other oncogenic signaling pathways comprises administration of inhibitors of MEK / ERK pathway signaling, PI3K / AKT pathway signaling, JAK / STAT pathway signaling, or WNT / p-catenin pathway signaling.

190. The method of claim 188, wherein targeting of immune response pathways comprises PD-1 / PD-L1 checkpoint inhibition.

191. The method of claim 188, wherein directly driving an immune response to cancer cells comprises bispecific T cell engagers or chimeric antigen receptor expressing T cells.

192. The method of claim 188, wherein the targeting disrupting the growth, metabolism, or oncogenic signaling capabilities of senescent cells comprises administering senolytic agents to a subject.

193. The method of any one of claims 186-192, wherein the additional therapy comprises administering fluorouracil, FOLFIRI, irinotecan, FOLFOX, gemcitabine, or cisplatin, irinotecan, oxiplatin, fluoropyrimidine to the subject.

194. The method of any one of claims 146-148, 152-160, 165-193, further comprising forming a ternary complex between the peptide complex, the target molecule, and the cellular receptor.

195. The method of claim 194, wherein formation of the ternary complex increases, facilitates, or stabilizes recycling or turnover of the cellular receptor, the target molecule, or both.

196. The method of claim 194 or claim 195, wherein formation of the ternary complex increases, facilitates, or stabilizes binding of the target molecule to the cellular receptor.

197. The method of any one of claims 146-148, 152-160, 165-196, wherein the peptide complex binds at higher levels to cells that overexpress the target molecule and the cellular receptor than to cells that have lower levels of the target molecule or the cellular receptor or both.

198. The method of any one of claims 146-148, 152-160, 165-197, wherein the peptide complex has a larger, longer, or wider therapeutic window as compared to an alternative therapy.

199. The method of claim 198, wherein the alternative therapy is not recycled to the cell surface.

200. The method of claim 198 or claim 199, wherein the alternative therapy is a lysosomal targeting therapy, a ubiquitin-proteosome system (UPS) targeting therapy, a non-selective therapeutic agent, an existing biologic, or a lysosomal delivery molecule.

201. The method of any one of claims 143-200, wherein the peptide complex or the EGFR- binding peptide is administered at lower molar dosage than alternative therapies.

202. The method of claim 143-201, wherein the peptide complex or the EGFR-binding peptide binds at higher levels to cancer cells than to normal cells.

203. The method of claim 143-202, wherein the peptide complex or the EGFR-binding peptide has a higher antiproliferative effect, a higher target molecule depletion effect, or a higher viability effect on cancer cells than on normal cells in vitro or in vivo.

204. The method of claim 143-203, wherein the peptide complex or the EGFR-binding peptide has a larger, longer, or wider therapeutic window than an anti-EGFR antibody or a TKI.

205. The method of claim 143-204, wherein the peptide complex or the EGFR-binding peptide has lower toxicity on skin or on keratinocytes than an anti-EGFR antibody or a TKI.

206. The method of any one of claims 161-205, further comprising causing remission in, reducing, ameliorating, or ablating the disease or condition.

207. The method of any one of claims 179-206, further comprising causing remission in, reducing, ameliorating, or ablating the cancer.

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