Protein-based delivery system

Abstract

Provided herein are elastin-like polypeptides, endosomal escape peptides, and compounds comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides. Also provided herein are compositions comprising a compound provided herein and an agent, methods of delivering an agent, and methods of treating or preventing a disease (e.g., a cardiovascular disease, a lung or respiratory disease, a musculoskeletal disease, a hematological disease, an immune disorder, an infectious disease, a genetic disease, cancer, a neurological disease, a psychiatric disorder, a metabolic disorder, or an inflammatory disease).

Description

PROTEIN-BASED DELIVERY SYSTEM CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority under 35 U.S.C. § 119(e) to U.S. Provisional Application No. 63 / 514,222 filed July 18, 2023, the contents of which are incorporated herein by reference in their entirety. GOVERNMENTSUPPORT

[0002] This invention was made with government support under grant AI150551 awarded by the National Institutes of Health. The government has certain rights in the invention. REFERENCE TO ANELECTRONICSEQUENCELISTING

[0003] The contents of the electronic sequence listing (B066270120WO00-SEQ-CLL.xml; Size: 462,142 bytes; and Date of Creation: July 18, 2024) is herein incorporated by reference in its entirety. BACKGROUND

[0004] There is growing interest in the use of nucleic acids and protein-based drugs to modulate intracellular targets or enable cytosolic protein production. The past decade has witnessed the emergence of highly effective therapies for vaccination,1,2gene knockdown,3gene editing,4,5protein replacement,6and cell reprogramming7,8for the treatment of a variety of infectious diseases, genetic diseases, and cancer. Nonetheless, the full potential of these therapeutic modalities has yet to be realized due to a number of challenges associated with the delivery of these molecules. In particular, achieving efficient cytosolic entry is a key impediment shared across all classes of therapeutic macromolecules, as the large size of these molecules prevents their ability to easily traverse the cell membrane. Several classes of viral and non-viral delivery vehicles have been explored to facilitate either direct cell membrane penetration or endosomal escape.9,10Among these, cationic membrane disruptive peptides, including cell penetrating peptides such as Tat11, oligoarginine12, and penetratin13or other endosomolytic peptides, as exemplified by S1014have been examined as vectors in the form of conjugates or as untethered peptide shuttles.15,16However, these peptides remain limited by low potency and cytotoxicity. Moreover, in vivo biodistribution has been difficult to control due to the cationic nature of many of these peptides and the inability to precisely control the size and morphology of the resulting macromolecular complex. Altogether, these factors have1 / 179B0662.70120WO00limited the efficacy of peptide-based macromolecule delivery systems for therapeutic applications. SUMMARY OF THEINVENTION

[0005] The present disclosure relates in part to elastin-like polypeptides, endosomal escape peptides, and compounds comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides. The compounds provided herein may be useful in compositions comprising an agent, such as a micelle or particle, for delivery of the agent to a subject or cell. The present disclosure also provides pharmaceutical compositions and kits comprising the compounds, agents, particles, or compositions provided herein. The present disclosure also provides methods of treating or preventing a disease, as well as methods of delivery.

[0006] In one aspect, provided herein is a compound comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides.

[0007] In another aspect, provided herein is an elastin-like polypeptide comprising a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 1) wherein: X3is V or A; and X4is E or G.

[0008] In another aspect, provided herein is an endosomal escape peptide comprising a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL; WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K; TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK; MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR; LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF; LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR; RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K; KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK; KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK;2 / 179B0662.70120WO00KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF; KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG; KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK; KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK; KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL; and ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NOs: 2-16, from top to bottom).

[0009] In another aspect, provided herein is a composition comprising a compound provided herein and an agent.

[0010] In another aspect, provided herein is a kit comprising a compound or composition provided herein and instructions for using the compound or composition.

[0011] In another aspect, provided herein is a method of delivering an agent to a subject, cell, or biological sample, comprising administering to the subject or contacting the cell or biological sample with an effective amount of a composition provided herein.

[0012] In another aspect, provided herein is a method of treating or preventing a disease in a subject, comprising administering to the subject a composition provided herein.

[0013] The details of certain embodiments of the invention are set forth in the Detailed Description of Certain Embodiments, as described below. Other features, objects, and advantages of the invention will be apparent from the Definitions, Examples, Figures, and Claims. It should be understood that the aspects described herein are not limited to specific embodiments, methods, or configurations, and as such can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and, unless specifically defined herein, is not intended to be limiting. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] The accompanying drawings, which constitute a part of this specification, illustrate several embodiments of the invention and together with the description, provide non-limiting examples of the invention.

[0015] Throughout the drawings, the following sequence abbreviations are used:3 / 179B0662.70120WO00

[0016] FIG. 1 shows elastin-like polypeptide (ELP) nanoparticles for multimodal macromolecule delivery. Micelle-forming tetrablock ELPs containing (from N- to C- terminus) a hydrophilic block, cysteine-based crosslinking block, hydrophobic block, and endosomolytic block were designed for siRNA, mRNA, and protein cargo delivery. From top to bottom, the sequences are V1: SEQ ID NO: 117, V2: SEQ ID NO: 115, V3: SEQ ID NO: 115, and V4: SEQ ID NO: 113.

[0017] FIGs. 2A-2M show formation of ELP-protein nanoparticles. a) Schematic representation of Cre-V1-ELP nanoparticles following evolved sortase-mediated conjugation and b) PAGE gel image depicting result of sortase-mediated Cre- LPETG conjugation to V1- ELP at varying molar ratios. The reaction solution underwent centrifugal filtration against a 100 kDa filter to remove unreacted Cre. c) Quantification of Cre-V1-ELP conjugation efficiency via densitometry analysis, measured as percentage of total ELP conjugated to Cre. d) Diameter of Cre-V1-ELP nanoparticles (n = 3 technical replicates). e) Schematic representation of SpCas9 RNP-V1-ELP nanoparticles. f) PAGE gel image depicting result of sortase-mediated SpCas9-LPETG conjugation to V1-ELP at varying ratios. g) Diameter and h) Zeta potential of SpCas9-V1-ELP nanoparticles (n = 3 technical replicates). i) Click and sortase conjugation reaction scheme for anti-CD117 mAb conjugation to V1-ELP. The sequence depicted is KLPETGG (SEQ ID NO: 118). j) PAGE gel image and k) Densitometry analysis of mAb-ELP conjugation efficiency. l) MFI and m) Fluorescent images of CD117+ P815 mast cells treated with CD117 mAb- functionalized V1-ELP nanoparticles compared to unfunctionalized controls (n = 3S). *p < 0.05, **p < 0.01, ***p < 0.005, ****p < 0.001, two- tailed Student’s t-test. Data represented by mean ± SEM.

[0018] FIGs. 3A-3G show multigenerational screen to identify ELPs for effective intracellular protein delivery. a) ELP block designs of each tetrablock ELP version assessed.4 / 179B0662.70120WO00Block designs are described as YmZn, where Y and Z are the identity of the guest residues X in that block (e.g., VPGXG (SEQ ID NO: 124)), and m and n are the number of those residue present in each 5-monomer repeat (e.g., [VPGXG]5 (SEQ ID NO: 212), m = 3, n = 2). b) Measured diameter and particle size distribution by mass of V2-ELP at varying pHs ([ELP] = 10 µM). c) % RFP+ of HEK293-RFP following treated with Cre conjugated to V1-ELP compared to histidine-containing V2-ELP, with and without chloroquine supplementation (n = 3). d) % RFP+ of treated HEK293-RFP following treatment with V3 ELPs conjugated to Cre (Cre-EEP-V2-ELPs) at escalating doses (n = 3). e) % RFP+ of treated HEK293-RFP following co-treatment with V4-ELP-EEPs and Cre protein (n = 3). f) % RFP+ and g) Cell viability following co-treatment with V4- ELP-S10 or S10 peptide with Cre recombinase at 2 µM. Data represented by mean ± SEM.

[0019] FIGs. 4A-4I show discovery of enhanced endosomolytic peptides via in silico screening. a) Comparison of the observed and predicted % GFP+ of HeLa cells treated with GFP proteins and a library of membrane disruptive peptides. Data from Ref 55. b) In silico alpha-helix library screening approach. c) Activity of V4-ELP-Gen2 and d) V4-ELP-Gen3 EEP constructs in Cre protein delivery to HEK293-RFP (average efficacies 10.1% and 27.1% respectively). e) Predicted structure (via AlphaFold2) of EEP13 along with sequence labeled composition. SEQ ID NOs: 119 (LL-III), 120 (Linker), and 121 (Mast18) are shown. f) Helical wheel diagrams of alpha helical peptides composing EEP13. g) Diameter of V4-ELP- EEP13 series micelle ([ELP] = 10 µM). h) ELP- and i) Cre treatment dose titration comparing activity of V4-ELP-EEP13 monomer, dimer, and trimer in HEK293-RFP. *p < 0.05, **p < 0.01, ***p < 0.005, two-tailed Student’s t-test. Data represented by mean ± SEM.

[0020] FIGs. 5A-5F show ELP-mediated protein delivery to primary cells. a-b) % tdTomato+ in Ai9-derived mouse lung fibroblasts, c-d) peritoneal macrophages, e) splenic CD4+ T cells, and f) Lin+Sca-1+c-Kit- hematopoietic stem cells following Cre and ELP co-treatment. [Cre], [ELP], ELP identity, and conjugation format are labeled on each plot. n = 3, data represented by mean ± SEM.

[0021] FIGs. 6A-6H show complexation and delivery of siRNA by tetrablock ELP nanoparticles. a) Schematic of ELP-siRNA nanoparticle formation via combined electrostatic and heat-induced hydrophobic interactions. b) Gel shift and Ribogreen assay evaluating V4- ELP-EEP13, V4-ELP-[EEP13]2, and V4-ELP-[EEP13]3 siRNA encapsulation efficiency (n = 3 for Ribogreen). c) Diameter of V4-ELP-EEP13 series-siRNA nanoparticles (n=3). d) Zeta potential of V4-ELP-EEP13 -siRNA nanoparticles (n = 3). e) MFI of HeLa treated with Alexa Fluor 647-labeled siRNA complexed with V4-ELP-EEP13 series compared to RNAiMAX5 / 179B0662.70120WO00after 1, 2, 4, and 24 hours of treatment. f) % GFP+ and g) viability of HeLa-d2eGFP cells following treatment with siGFP complexed with V4-ELP-EEP13 series at varying doses (n = 3). h) MFI (bars) and viability (points) of HeLa cells treated with Alexa Fluor 647-labeled siRNA complexed with V4- ELP-EEP13 series following treatment with chemical endocytosis inhibitors. Cells were pretreated with inhibitors for 1 hour then treated with ELP- siRNA nanoparticles for an additional 2 hours (n = 3). All data represented by mean ± SEM.

[0022] FIGs. 7A-7G show complexation and delivery of mRNA-encoded Cre recombinase by tetrablock ELP nanoparticles. a) Comparison of V4-ELP-EEP13 monomer, dimer, and trimer Cre mRNA complexation efficiency via gel shift and RiboGreen assays. b) Diameter and c) zeta potential of V4-ELP-EEP13 nanoparticles encapsulating Cre mRNA (n = 3). PDI = 0.112 and 0.17, -mRNA and +mRNA. d) % RFP+ of HEK293-RFP following treatment with Cre mRNA complexed with V4-ELP-EEP13 at varying ELP and Cre mRNA concentrations (n = 3). e) Dose titration of V4-ELP- EEP13 monomer, dimer, and trimer for Cre mRNA delivery (n = 3). f) % tdTomato+ of Ai9 lung fibroblasts following treatment with Cre mRNA encapsulated by V4-ELP-EEP13 (n = 3). g) Correlation between protein and mRNA-form Cre delivery efficacy by V4-ELP EEPs (30 constructs assessed). All data represented by mean ± SEM.

[0023] FIGs. 8A-8E show ELP-mediated delivery of therapeutic gene editor and transcription factor proteins. a) Schematic of V4-ELP-S10 + SpCas9 treatment solution. sgRNA targeting HPRT1 was encapsulated into nanoparticles prior to treatment. b) Editing efficiency in HEK29348 hours after treatment with SpCas9 and HPRT1 sgRNA encapsulated by V4-ELP-S10, assessed by Sanger sequencing (n = 1). c) Schematic of pro-inflammatory reprogramming in treated macrophages following IRF5 intracellular delivery mediated by ELPs. d) IFNg and e) IL12b expression assessed via qPCR 72 hours after treatment with IRF5 alone, V4-ELP-EEP13 alone, and both agents (n = 6). All data represented by mean ± SEM.

[0024] FIGs. 9A-9C show ELP-mediated intradermal delivery of Cre protein in Ai9 mice. a) Images of intradermal administration procedures. A patch of flank skin was shaved to facilitate precise injection. b) % tdTomato+ in skin T cells and innate lymphoid cells, c) fibroblasts, and d) macrophages and neutrophils 72 hours following administration. Mice were treated with PBS (n = 1), Cre alone (n = 1), Cre conjugated S10-V2-ELP (n = 3), and S10-V2-ELP and unconjugated Cre (n = 3). All data represented by mean ± SEM.

[0025] FIGs. 10A-10D show ELP micelle size and cellular internalization. a) Size and polydispersity index (PDI) of V1-ELP micelles formed with varying concentrations of ELP.6 / 179B0662.70120WO00b) Representative light scattering intensity plot of V1-ELP ([ELP] = 8 µM). c) Uptake of PS800-labeled V1-ELP micelles in HeLa cells over the course of 24 hours. d) Viability of HeLa and HEK293 cells following treatment with V1-ELPs at varying concentrations.

[0026] FIGs. 11A-11B show protease-mediated protein cargo release from ELP micelles. a) Schematic depicting Cathepsin B cleavage sites in Cre-V1-ELP. The sequence (GGS)9 (SEQ ID NO: 91) is depicted. b) Anti-Histag western blot depicting result of digestion of GFP-Cre conjugate and Cre-V1-ELP with Cathepsin B.

[0027] FIG. 12 shows quantification of ELP conjugation efficiency to SpCas9.

[0028] FIGs. 13A-13B show CD117-targeted ELP micelle uptake via anti-CD117 mAb functionalization. a) Histograms depicting distribution of Alexa Fluor 633 MFI in CD117+ P815 cells treated with CD117-functionalized V1-ELP-AF633 compared to unmodified control. b) Z-stack confocal images of CD117-V1-ELP-AF633 treated P815 cells.

[0029] FIGs. 14A-14C show the influence of pH on the size of ELP micelles. a) Diameter of V2-ELPs at varying pHs ([ELP] = 10 µM). b) Diameter and c) Size distribution of V1-ELPs at varying pHs ([ELP] = 10 µM).

[0030] FIGs. 15A-15D show formation and characterization of Cre-V2-ELP micelles. a) PAGE depicting Cre conjugation to V2-ELP. b-d) Size distribution of V1-, V2- Cre-V1-, and Cre-V2-ELPs.

[0031] FIGs. 16A-16E show formation and characterization of Cre-V3-ELP micelles. a-d) Diameter of V3-ELPs after 10 min and 72 hr incubation at 25 °C. e) PAGE depicting conjugation of Cre to V3-ELPs.

[0032] FIG. 17 shows HEK293 viability following Cre-V3-ELP treatment for 48 hours.

[0033] FIGs. 18A-18B show optimization of protease cleavable linker sequences to improve cargo release from ELP micelles. a) Linker sequences on the C-terminal end of Cre protein variants. From top to bottom, SEQ ID NOs are 91-97. b) Ratio of cis (conjugated) to trans (unconjugated) Cre protein delivery efficacy in HEK293-loxP-RFP via S10-V3, quantified by RFP+ percentage.

[0034] FIG. 19 shows diameter and PDI of V4-ELP micelles.

[0035] FIG. 20 shows HEK293-RFP viability following Cre recombinase and V4-ELP-EEP co-treatment for 48 hours.

[0036] FIGs. 21A-21B show an assessment of Lipofectamine 3000 for Cre protein delivery to HEK293-RFP. a) Viability and b) RFP+ percentage following transfection.

[0037] FIGs. 22A-22B show the predicted structure and helical wheel depiction of S10. a) Predicted structure of S10 (via AlphaFold2). SEQ ID NOs: 122 (CM18 Variant), 1207 / 179B0662.70120WO00(Linker), and 123 (PTD4 Variant) are shown. b) Helical wheel diagrams of alpha helices composing S10.

[0038] FIG. 23 shows quantification of peptide feature importance for protein delivery efficacy.

[0039] FIG. 24 shows multiple linear regression model for prediction of V4-ELP-EEP construct efficiency in siGFP delivery.

[0040] FIG. 25 shows multi-stage α-helical database screen for the discovery of Gen3 EEPs.

[0041] FIGs. 26A-26B show a comparison of EEP13 and S10 in standalone format in Cre protein delivery. a) RFP+ percentage and b) viability of HEK293-RFP cells co-treated with S10 or EEP13 peptide with Cre protein.

[0042] FIGs. 27A-27B show HEK293 RFP expression and viability following treatment with protein-form Cre recombinase and V4-ELP-EEP13 multimer series. a) RFP+ percentage and b) viability of HEK293-RFP following treatment with varying doses of Cre protein and V4- EEP13 ELPs ([ELP] = 8 µM) for 48 hours.

[0043] FIGs. 28A-28F show primary cell viability following co-treatment with protein-form Cre recombinase and ELP micelles. a-b) Viability of Ai9 mouse-isolated lung fibroblasts, c-d) peritoneal macrophages, e) splenic CD4+ T cells, and f) Lin-Sca-1+ hematopoietic stem cells following ELP and Cre protein co-treatment.

[0044] FIGs. 29A-29B show an assessment of Lipofectamine 3000 for Cre protein delivery to primary mouse fibroblasts. a) Viability and b) tdTomato expression of Ai9 fibroblasts following treated with Lipofectamine 3000 and Cre protein.

[0045] FIG. 30 shows gating strategy for isolation of Lin-Sca-1+c-Kit+ hematopoietic stem cells from Ai9-SauSpyCas9 mice.

[0046] FIGs. 31A-31B show delivery of fluorescently labeled Cre recombinase to LS HSPCs via CD117-targeted ELP micelles. a) Schematic of mAb- and Alexa Fluor 633 labeled Cre- functionalized V2-ELP. b) Cre+ LS cell percentage following treatment with CD117 mAb- functionalized Cre-V2-ELP nanoparticles compared to unmodified Cre-V2-ELP nanoparticles. n = 3.

[0047] FIG. 32 shows flow cytometry plots of HeLa-d2eGFP following V4-ELP-[EEP13]2- mediated siGFP delivery.

[0048] FIGs. 33A-33B show HeLa-d2eGFP GFP expression and viability following treatment with siGFP formulated with V4-ELP-EEP13 multimer series. a) GFP+ percentage and b) viability of HeLa-d2eGFP following treatment with varying doses of siGFP and V4- EEP13 ELPs ([ELP] = 10 µM) for 24 hours.8 / 179B0662.70120WO00

[0049] FIGs. 34A-34B show HEK293-RFP RFP expression and viability following treatment with mRNA-encoded Cre recombinase via V4-ELP-EEP13 multimeric variants. a) RFP+ percentage and b) viability of HEK293-RFP following treatment with varying doses of mRNA-encoded Cre recombinase and V4-EEP13 ELPs ([ELP] = 10 µM) for 48 hours.

[0050] FIG. 35 shows Ai9-SauSpyCas9 lung fibroblast viability following treatment with mRNA-encoded Cre recombinase via V4-ELP-EEP13.

[0051] FIGs. 36A-36B show V4-ELP-EEP screen for a) Cre recombinase protein and b) mRNA delivery to HEK293-GFP.

[0052] FIG. 37 shows gating strategy for skin cell isolation.

[0053] FIG. 38 shows representative flow cytometry plots from isolated Ai9 skin cell populations following intradermal Cre-ELP injections.

[0054] FIGs. 39A-39L show ELP-mediated protein delivery to primary cells. a-b) % tdTomato+ in Ai9-derived mouse lung fibroblasts, c-d) peritoneal macrophages, e-f) splenic CD4+ T cells, and g-h) Lin-Sca-1+c-kit+ (LSK) hematopoietic stem cells following Cre and ELP co-treatment (n = 3). In a, c, e - [Cre] = 8 μM. b, d – [ELP] = 8 μM. Data represented by mean ± SEM. i) Fluorescence images with nuclear staining (blue) taken 2 hours after treatment of murine peritoneal macrophages with 100 nM AF488-labeled IRF5 alone (top) and in combination with 4 μM V4-ELP-EEP13. Scalebars = 25 μm. j) IFNg and k) IL12b, and l) IL1b expression in murine macrophages assessed via qPCR 72 hours after treatment with IRF5 alone (500 nM), V4-ELP-EEP13 alone (8 μM), and both agents (n = 3).

[0055] FIGs. 40A-40B show complexation and delivery of siRNA by tetrablock ELP nanoparticles. a) GAPDH mRNA expression in Ai9-derived murine lung fibroblasts following 48 hours of treatment with siGAPDH, assessed via qPCR. Expression levels are normalized to cells treated with siRNA alone. [ELP] = 10 μM, [siRNA] = 500 nM, n = 3. b) MFI (bars) and viability (points) of HeLa cells treated with Alexa Fluor 647-labeled siRNA (50 nM) complexed with V4-ELP-EEP13 (5 μM) following treatment with chemical endocytosis inhibitors. Cells were pretreated with inhibitors for 1 hour then treated with ELP- siRNA nanoparticles for an additional 2 hours (n = 3). *p < 0.05, **p < 0.01, ***p < 0.005, ****p < 0.001, two-way ANOVA with Dunnett’s or Šídák correction for multiple comparisons as appropriate. All data represented by mean ± SEM.

[0056] FIGs. 41A-41E show complexation and delivery of mRNA by tetrablock ELP nanoparticles. a) % RFP+ of HEK293-RFP following treatment with Cre pDNA at escalation [pDNA] (n = 3, [ELP] = 10 μM). b) Delivery of Cas9 mRNA and c) pDNA with a9 / 179B0662.70120WO00chemically modified sgRNA targeting HPRT1 to HEK293 at varying mRNA / pDNA:sgRNA molar ratios. [mRNA] = 6 ng / μL in mRNA study, [ELP] = 20 μM in pDNA study, n = 3. d) % tdTomato+ of Ai9 lung fibroblasts and e) peritoneal macrophages following treatment with Cre mRNA encapsulated by V4-ELP-EEP13 (n = 3). All data represented by mean ± SEM.

[0057] FIGs. 42A-42E show ELP-mediated delivery of Cre protein to the bronchial epithelium in vivo. a) Schematic of V4-ELP-EEP13-mediated Cre protein delivery study to the bronchial epithelium via intranasal instillation in Ai9 mice. b) Editing efficiency observed in small and c) large airway epithelial cells following administration of Cre protein ± V4- ELP-EEP13 (n = 3, ≥ 6 large airway images / mouse, ≥ 20 small airway images / mouse). d) Representative images of large airway and e) small airways following co-treatment with Cre protein and V4-ELP-EEP13 nanoparticles. Scalebars: 200 µm (left, d), 50 µm (right, d), 100 µm (e). *p < 0.05, **p < 0.01, ***p < 0.005, ****p < 0.001, unpaired two-tailed t-test.

[0058] FIG. 43 shows a negative stain TEM image of V4-ELP nanoparticles. Scalebar = 50 nm, inset scalebar = 10 nm. f) % RFP+ HEK293-RFP cells following treatment with V4- ELP-EEPs and free Cre protein. [ELP] = 8 μM, [Cre protein] = 2 μM, n = 3.

[0059] FIG. 44 shows Editing efficiency in HEK293 cells 72 hours after treatment with SpCas9 RNP containing HPRT1 sgRNA and V4-ELP-EEP13 assessed by Sanger sequencing. [ELP] = 8 μM, n = 3. Data represented by mean ± SEM.

[0060] FIGs. 45A-45I show physicochemical and functional characterization of V1-ELPs. a) Size and polydispersity index (PDI) of V1-ELP micelles formed with varying concentrations of ELP. b) Representative light scattering intensity plot of V1-ELP ([ELP] = 8 µM). c) Uptake of PS800-labeled V1-ELP micelles in HeLa cells over the course of 24 hours. d) Viability of HeLa and HEK293 cells following treatment with V1-ELPs at varying concentrations. e) Schematic depicting Cathepsin B cleavage sites in Cre-V1-ELP. The sequence (GGS)9 (SEQ ID NO: 91) is depicted. f) Anti-Histag western blot depicting result of digestion of a GFP-Cre conjugate and Cre-V1-ELP with Cathepsin B. g) Quantification of ELP conjugation efficiency to SpCas9 via densitometry analysis (see gel in FIG. 2F). h) Histograms depicting distribution of Alexa Fluor 633 MFI in CD117+ P815 cells treated with CD117-functionalized V1-ELP-AF633 compared to unmodified control. i) Z-stack confocal images of CD117-V1-ELP-AF633 treated P815 cells.

[0061] FIGs. 46A-46J show formation and characterization of Cre-V2-ELP and Cre-V3-ELP nanoparticles. a) PAGE depicting Cre conjugation to V2-ELP. b-d) Size distribution of V1-, V2- Cre-V1-, and Cre-V2-ELPs. e-h) Diameter of V3-ELPs after a 10 minute and 72 hour10 / 179B0662.70120WO00incubation at 25 °C. i) PAGE depicting conjugation of Cre to V3-ELPs. j) HEK293 viability following Cre-V3-ELP treatment for 48 hours (n = 3).

[0062] FIGs. 47A-47B show optimization of protease cleavable linker sequences to improve cargo release from ELP nanoparticles. a) Linker sequences on the C-terminal end of Cre protein variants. From top to bottom, SEQ ID NOs: 91-97 are shown. b) Ratio of conjugated to unconjugated Cre protein delivery efficacy in HEK293-loxP-RFP cells via S10-V3, quantified by percentage RFP+ cells.

[0063] FIGs. 48A-48B show characterization and functional assessment of V4-ELP nanoparticles. a) Diameter and PDI of V4-ELP nanoparticles at varying ELP concentrations. b) HEK293-RFP cell viability following Cre recombinase and V4-ELP-EEP co-treatment for 48 hours (n = 3, delivery efficiency data in FIG. 3E).

[0064] FIGs. 49A-49B show assessment of Lipofectamine 3000 for Cre protein delivery to HEK293-RFP cells. a) Cell viability and b) percentage RFP+ cells 48 hours following transfection (n = 3).

[0065] FIGs. 50A-50C show alpha helical peptide database screening for discovery of endosomal escape peptides. a) Quantification of the importance of peptide features for effective protein delivery for the Gen1 EEP predictive model. The five features highlighted in green were incorporated into the model parameters. b) Quantification of peptide feature importance for protein delivery efficacy for Gen2 EEP predictive model. The three features highlighted in green were incorporated into the model parameters. c) Multi-stage α-helical database screen for the discovery of Gen3 EEPs.

[0066] FIGs. 51A-51B show a comparison of free EEP13 and S10 peptides in Cre protein delivery. a) Percentage of RFP+ cells and b) the viability of HEK293-RFP cells co-treated with either free S10 or EEP13 peptides and Cre protein (*p < 0.05, **p < 0.01, ***p < 0.005, ****p < 0.001, unpaired two-tailed t-test with Bonferroni’s correction for multiple comparisons).

[0067] FIGs. 52A-52B show % RFP+ HEK293-RFP cells and cell viability following treatment with protein-form Cre recombinase and V4-ELP-[EEP13]n nanoparticles. a) The percentage of RFP+ cells and b) viability of HEK293-RFP cells following treatment with varying doses of Cre protein and V4-EEP13 ELPs ([ELP] = 8 µM) for 48 hours.

[0068] FIG. 53 shows ELP-EEP-mediated endosomal escape of FITC-labeled dextran. HeLa cells were co-treated with V4-ELP-EEP13 or V4-ELP-[EEP13]2 and FITC-dextran with11 / 179B0662.70120WO00varying molecular weights of FITC-labeled dextran for 6 hours, fixed, and imaged using a fluorescence microscopy (scale bar = 10 µm).

[0069] FIG. 54A-54H show viability of primary cells following co-treatment with protein- form Cre recombinase and ELP nanoparticles. a-b) Viability of Ai9 mouse-isolated lung fibroblasts, c-d) peritoneal macrophages, e-f) splenic CD4+ T cells, and g-h) Lin-Sca-1+ckit+ hematopoietic stem cells following treatment with V4-ELP-EEP13 and Cre protein. Constant [Cre] = 8 μM or constant [ELP] = 8 μM as applicable, n = 3.

[0070] FIGs. 55A-55B show assessment of Lipofectamine 3000 for Cre protein delivery to primary mouse fibroblasts. a) Cell viability and b) tdTomato expression of Ai9 fibroblasts following treated with Lipofectamine 3000 and Cre protein.

[0071] FIGs. 56A-56C show cell viability and GFP expression of HeLa-d2eGFP cells after V4-ELP-EEP-mediated siGFP delivery. a) GFP expression of HeLa-d2eGFP cells following treatment with siGFP alone or after delivery via RNAiMAX ([siRNA] = 100 nM, n = 3). b) Viability of HeLA-d2eGFP cells following treatment with 100 nM siGFP via V4-ELP-EEP13 and V4-ELP-[EEP13]2 nanoparticles for 24 hours. c) GFP expression of HeLa-d2eGFP cells treated with varying concentrations of V4-ELP-EEP13 nanoparticles carrying 100 nM siGFP.

[0072] FIGs. 57A-57E show cell viability following treatment with Cre mRNA formulated with V4-ELP-[EEP13]n nanoparticles. a) HEK293-loxP-RFP cell viability following treatment with V4-ELP-EEP13-Cre mRNA nanoparticles ([mRNA] = 1 ng / μL, varying ELP concentration) and b) [ELP] = 10 μM, varying mRNA concentration for 48 hours. c) Viability of HEK293-loxP-RFP cells treated with Cre recombinase pDNA via Lipofectamine or V4- ELP-EEP13 nanoparticles. d) Viability of Ai9-derived fibroblasts and e) peritoneal macrophages following treatment with V4-ELP-EEP13-Cre mRNA nanoparticles for 48 hours. (n = 3 for all studies, mean ± SEM).

[0073] FIG. 58 shows sequences of ELP hydrophilic blocks, as well as their charge and hydrophilicity. From top to bottom, SEQ ID NOs: 17, 18, 204, 205, and 206 are shown.

[0074] FIG. 59A shows PAGE analysis of V2-, V4-, and V5-ELP-EEP13 conjugates to Cre recombinase. FIG. 59B shows percent conjugation of V2-, V4-, and V5-ELP-EEP13 conjugates determined by relative intensity of Cre-ELP versus ELP bands.

[0075] FIG. 60 shows delivery efficiency of Cre protein (FIG. 60A) and Cre mRNA (FIG. 60B) by V5-ELP particles to HEK293 Cre reporter cells.12 / 179B0662.70120WO00

[0076] FIG. 61 shows size and dispersity of V5-ELP particles with and without mRNA: V5[S]-ELP-EEP13 (FIG. 61A), V5[G] -ELP-EEP13 (FIG. 61B), V5[S]-ELP-EEP13 (FIG. 61C), V5[G]-ELP-EEP13 (FIG. 61D).

[0077] FIG. 62 shows PAGE depicting result of V5(S)-ELP-EEP13 reacted with LPETG- linker functionalized anti-CD117 mAb via sortase reaction.

[0078] FIG. 63 shows size exclusion chromatography of ELP-EEP13 + varying amounts of AF647-labeled siRNA.

[0079] FIG. 64 shows the SEC protocol for mAb-ELP-siRNA nanoparticles. FIG. 64A shows PAGE analysis of ELPs purified by SEC. FIG. 64B shows SEC fractions of mAb-ELP siRNA nanoparticles, free mAb, free ELP, and free sortase.

[0080] FIG. 65A shows improved uptake of CD117-V5(S)-EEP13 over V5(S)-EEP13 formulated with AF647-labeled siRNA. FIG. 65B shows cells untreated, with V5(S)-EEP13, or with CD117-V5(S)-EEP13 at 30 minutes (top) versus 24 hours (bottom).

[0081] FIG. 66 shows uptake of AF647-labeled siRNA formulated ELP nanoparticles in Lineage-Sca-1+c-kit+ cells (LSK, FIG. 66A) and CD117+ cells (FIG. 66B) and overally (FIG. 66C) using CD117-V5(S)-EEP13 versus V5(S)-EEP13 nanoparticles.

[0082] FIGs. 67A and 67B show conjugation of an anti-CD-5 antibody to V5(S)-ELP- EEP13.

[0083] FIG. 68 shows a schematic for in silico discovery of EEPs.

[0084] FIG. 69 shows a schematic of a variational autoencoder to generate EEP sequences. From top to bottom, SEQ ID NOs: 207-210 are shown.

[0085] FIG. 70 shows a schematic of a transfer learning predictive model.

[0086] FIG. 71 shows a table of first generation ELP-E3P sequences and predicted efficacies for in vitro Cre protein delivery (normalized to V4-ELP-EEP13). From top to bottom, SEQ ID NOs: 133-164 are shown.

[0087] FIG. 72 shows Cre protein delivery activity of first generation ELP-E3Ps to HEK293 Cre reporter cells.

[0088] FIG. 73 shows a second generation predictive model to generate EEP sequences. FIG. 73A shows predictive model performance on Gen0 and Gen1 ELP-EEP efficacy dataof second generation peptides. FIG. 73B shows a synthetic minority oversampling with Gaussian noise technique.

[0089] FIG. 74 shows computational and empirical evaluation of Gen0 and Gen1EEP sequences.13 / 179B0662.70120WO00

[0090] FIG. 75 shows a table of second generation ELP-E3P sequences, fitness function scores, and predicted efficacies. From top to bottom, SEQ ID NOs: 165-180 are shown.

[0091] FIG. 76 shows Cre protein delivery activity of second generation ELP-E3Ps to HEK293 Cre reporter cells.

[0092] FIG. 77 shows delivery efficacy of V4-ELP-EEP13 and V4-ELP-E3P2.5 for Cre protein (FIG. 77A) and Cre mRNA (FIG. 77B) to HEK293 Cre reporter cells. FIG. 77C shows the structure of E3P2.5.

[0093] FIG. 78 shows a table of third generation ELP-E3Ps. From top to bottom, SEQ ID NOs: 181-196 are shown.

[0094] FIG. 79A shows delivery activity of third generation ELP-E3Ps for Cre protein delivery to HEK293 Cre reporter cells (normalized to EEP13). FIG. 79B shows a comparison of EEPs, E3P first generation, E3P second generation, and E3P third generation.

[0095] FIG. 80A shows strategy #1: Prevention of intramolecular crosslinking via rigid linkers. In the table, (GGGGS)ncorresponds to SEQ ID NO: 213 and (EAAAK)ncorresponds to SEQ ID NO: 214. Under the table, from left to right, SEQ ID NOs: 19, 125, and 126 are shown. FIG. 80B shows strategy #2: Spacing cysteines via placement in hydrophobic block. SEQ ID NO: 132 is shown.

[0096] FIG. 81 shows crosslinking efficiency at 4 hours (left) and 24 hours (right) for ELP crosslinked nanoparticles.

[0097] FIG. 82 shows crosslinking of V4-ELP-Cys7-EEP13 and V4-ELP-[CEAAAK]5over time with and without siRNA.

[0098] FIG. 83 shows degree of crosslinking over time for V4-ELP-Cys7-EEP13 and V4- ELP-[CEAAAK]5 both with and without siRNA cargo.

[0099] FIG. 84 shows size analysis of Cys-containing ELP-EEP constructs with and without Cre-mRNA and over time.

[0100] FIG. 85 shows size analysis of Cys-containing ELPs (non-EEP13-functionalized) without mRNA.

[0101] FIG. 86A shows crosslinking of V4-ELP-Cys7-EEP13 particles for up to 24 hours. FIG. 86B shows delivery efficacy for Cre protein and Cre mRNA after crosslinking for up to 24 hours.

[0102] FIG. 87A shows an image of gel band densitometry of V4-C4G3-EEP13 formulated with Cre mRNA. FIG. 87B shows ELP-EEP13-mRNA nanoparticle crosslinking efficiencies.

[0103] FIG. 88 shows mRNA delivery efficiency of V4-ELP-EEP13 nanoparticles.14 / 179B0662.70120WO00

[0104] FIG. 89A shows V4-ELP-EEP13 + / - Cys7 at 10 uM w / 500 nM siRNA, crosslinking at RT (rotating) for 24 hrs. 24% (-Cys7) and 66% (+Cys7) crosslinked per densitometry analysis. FIG. 89B shows agarose gel shift assay of crosslinked and uncrosslinked ELP constructs with free and bound siRNA.

[0105] FIG. 90 shows biodistribution of siRNA cargo. FIG. 90A shows quantification of radiant efficiency of crosslinked and uncrosslinked particles in the liver, lungs, spleen, kidneys, heart, femurs, and quads. FIG. 90B shows images of distribution of crosslinked and uncrosslinked particles to the liver, lungs, spleen, kidneys, heart, femurs, and quads.

[0106] FIG. 91 shows crosslinking efficiency assessed via densitometry for Cys-containing ELP-EEP13 constructs ( - = not crosslinked; + = crosslinked).

[0107] FIG. 92A shows peptide placements in ELP-EEP13 variants modified with OTI peptide. FIG. 92B shows ELP-EEP13 variants modified with OTI peptide.

[0108] FIG. 93A shows viability of untreated cells and cells treated with OTI-functionalized ELPs. FIG. 93B shows delivery of ovalbumin protein using OTI-functionalized ELPs.

[0109] FIG. 94A shows viability of untreated cells and cells treated with varying ratios of N-terminal and middle OTI-modified ELPs. FIG. 94B shows delivery of ovalbumin protein using varying ratios of N-terminal and middle OTI-modified ELPs. DEFINITIONS

[0110] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art. The following definitions are provided to help interpret the disclosure and claims of this application. In the event a definition in this section is not consistent with definitions elsewhere, the definition set forth in this section shall control.

[0111] Unless otherwise required by context, singular terms shall include pluralities, and plural terms shall include the singular. The singular terms “a,” “an,” and “the” include plural references unless the context clearly indicates otherwise. Similarly, the word “or” is intended to include “and” unless the context clearly indicates otherwise.

[0112] The language “in some embodiments” and the language “in certain embodiments” are used interchangeably.

[0113] Other than in the examples, or where otherwise indicated, all numbers expressing quantities of ingredients or reaction conditions used herein should be understood as modified in all instances by the term “about.” “About” and “approximately” shall generally mean an acceptable degree of error for the quantity measured given the nature or precision of the15 / 179B0662.70120WO00measurements. Exemplary degrees of error are within 20 percent (%), typically, within 10%, or more typically, within 5%, 4%, 3%, 2%, or 1% of a given value or range of values.

[0114] The term “amino acid” refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids. Every amino acid contains an amine (-NH2) and a carboxylic acid (-COOH) functional group. Each amino acid contains a unique side chain, designated by the “R” substituent shown below.

[0115] Exemplary amino acids include, without limitation, alpha-amino acids such as D– and L–isomers of the 20 common naturally occurring alpha amino acids found in peptides, natural amino acids which are not the 20 common naturally occurring amino acids, and unnatural alpha-amino acids. Amino acids used in the construction of peptides of the present disclosure may be prepared by organic synthesis, or obtained by other routes, such as, for example, degradation of or isolation from a natural source. Amino acids may be commercially available or may be synthesized. Amino acids may be referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission.

[0116] In a particular embodiment, the term “amino acid” refers to a naturally occurring amino acid. Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, γ-carboxyglutamate, and O-phosphoserine. In certain embodiments, the amino acid is an N-alkyl amino acid, where the hydrogen on any non-proline amine (N) is replaced with an alkyl (e.g., methyl (-CH3)) group. In certain embodiments, the N-alkyl amino acid is sarcosine (Sar). Amino acid analogs refer to compounds that have the same basic chemical structure as a naturally occurring amino acid, i.e., a carbon that is bound to a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid. Amino acid mimetics refers to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid. Unnatural (or non-natural) amino acids refer to those not naturally incorporated16 / 179B0662.70120WO00into proteins during translation. Examples of unnatural amino acids include, but are not limited to, β-amino acids (e.g., β2and β3), homo-amino acids, proline derivatives, pyruvic acid derivatives, alanine derivatives (e.g., 1'- and 2'-naphthylalanine), glycine derivatives, ring-substituted phenylalanine and tyrosine derivatives, linear core amino acids, and N- methyl amino acids. In certain embodiments, a compound provided herein comprises an amino acid side chain selected from the 20 proteinogenic amino acids (i.e., an amino acid incorporated into proteins during translation) shown in Table A. The term amino acid may also refer to non-proteinogenic amino acids, such as, for example, selenocysteine (-CH2SeH). Table A. Amino acids and side chains.

[0117] Amino acids with hydrophobic side chains include Gly, Pro, Ala, Ile, Leu, Val, Phe, Met, Trp, and Tyr. In certain embodiments, amino acids with hydrophobic side chains include Gly, Pro, Ala, Ile, Leu, Val, and Phe. In certain embodiments, amino acids with hydrophobic side chains include Ala, Ile, Leu, and Val. Amino acids with polar side chains include Gln, Asn, His, Ser, Thr , Tyr, Cys, Met, Trp. In certain embodiments, amino acids17 / 179B0662.70120WO00with polar side chains include Asn, Cys, Gln, Met, Ser, and Thr. Amino acids with aromatic side chains include Phe, Trp, Tyr, and His. Amino acids with hydrophobic aromatic side chains include Phe, Typ, and Tyr. Amino acids with charged side chains include Asp, Glu, Arg, His, and Lys. Negatively charged side chains include Asp and Glu. Positively charged side chains include Arg, His, and Lys. Neutral amino acids are selected from the group consisting of Ala, Ser, Val, Leu, Ile, Pro, Phe, Trp, Met, Gly, Thr, Cys, Tyr, Asn, and Gln.

[0118] A “protein,” “peptide,” or “polypeptide” comprises a polymer of amino acid residues linked together by peptide bonds. The term refers to proteins, polypeptides, and peptides of any size, structure, or function. Typically, a protein will be at least three amino acids long. A protein may refer to an individual protein or a collection of proteins. Inventive proteins preferably contain only natural amino acids, although non-natural amino acids (i.e., compounds that do not occur in nature but that can be incorporated into a polypeptide chain) and / or amino acid analogs as are known in the art may alternatively be employed. Also, one or more of the amino acids in a protein may be modified, for example, by the addition of a chemical entity such as a carbohydrate group, a hydroxyl group, a phosphate group, a farnesyl group, an isofarnesyl group, a fatty acid group, a linker for conjugation or functionalization, or other modification. A protein may also be a single molecule or may be a multi-molecular complex. A protein may be a fragment of a naturally occurring protein or peptide. A protein may be naturally occurring, recombinant, synthetic, or any combination of these.

[0119] As to amino acid sequences, one of skill will recognize that individual substitutions to a peptide, polypeptide, or protein sequence which alters a single amino acid or a small percentage of amino acids in the encoded sequence is a "conservatively modified variant" where the alteration results in the substitution of an amino acid with a chemically similar amino acid. Conservative substitution tables providing functionally similar amino acids are well known in the art.

[0120] The term “homologous,” as used herein is an art-understood term that refers to nucleic acids or proteins that are highly related at the level of nucleotide or amino acid sequence. Nucleic acids or proteins that are homologous to each other are termed homologues. Homologous may refer to the degree of sequence similarity between two sequences (i.e., nucleotide sequence or amino acid). The homology percentage figures referred to herein reflect the maximal homology possible between two sequences, i.e., the percent homology when the two sequences are so aligned as to have the greatest number of matched (homologous) positions. Homology can be readily calculated by known methods18 / 179B0662.70120WO00such as those described in: Computational Molecular Biology, Lesk, A. M., ed., Oxford University Press, New York, 1988; Biocomputing: Informatics and Genome Projects, Smith, D. W., ed., Academic Press, New York, 1993; Sequence Analysis in Molecular Biology, von Heinje, G., Academic Press, 1987; Computer Analysis of Sequence Data, Part I, Griffin, A. M., and Griffin, H. G., eds., Humana Press, New Jersey, 1994; and Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York, 1991; each of which is incorporated herein by reference. Methods commonly employed to determine homology between sequences include, but are not limited to those disclosed in Carillo, H., and Lipman, D., SIAM J Applied Math., 48:1073 (1988); incorporated herein by reference. Techniques for determining homology are codified in publicly available computer programs. Exemplary computer software to determine homology between two sequences include, but are not limited to, GCG program package, Devereux, J., et al., Nucleic Acids Research, 12(1), 387 (1984)), BLASTP, BLASTN, and PASTA Atschul, S. F. et al., J Molec. Biol., 215, 403 (1990)).

[0121] As used herein, the term “identity” refers to the overall relatedness between polymeric molecules, e.g., between nucleic acid molecules (e.g. DNA molecules and / or RNA molecules) and / or between polypeptide molecules. Calculation of the percent identity of two amino acid sequences, for example, can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second amino acid sequences for optimal alignment and non-identical sequences can be disregarded for comparison purposes). In certain embodiments, the length of a sequence aligned for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or 100% of the length of the reference sequence. The amino acids at corresponding positions are then compared. When a position in the first sequence is occupied by the same amino acid as the corresponding position in the second sequence, then the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. For example, the percent identity can be calculated by optimal alignment of the sequences using a similarity-scoring matrix such as the Blosum62 matrix described in Henikoff S. and Henikoff J.G., P.N.A.S. USA 1992, 89: 10915-10919. Calculation of the percentage identity and optimal alignment of two sequences using the19 / 179B0662.70120WO00Blosum62 similarity matrix and the algorithm of Needleman and Wunsch (J. Mol. Biol. 1970, 48: 443-453) can be performed using the GAP program of the Genetics Computer Group (GCG, Madison, Wl, USA) using the default parameters of the program. Specific parameters for calculating percentage identity for protein sequences and nucleic acid sequences in respect of the present invention are described below.

[0122] As used herein, to “modulate” means to act as an antagonist, i.e., partially or fully inhibit, reduce, alleviate, block or prevent; or to increase or stimulate, i.e., to act as an agonist, partial agonist or inverse agonist. The modulation may be direct or indirect or allosteric.

[0123] As used herein the term “inhibit” or “inhibition,” for example, in the context of enzymes refers to a reduction in the activity of the enzyme. In some embodiments, the term refers to a reduction of the level of enzyme activity to a level that is statistically significantly lower than an initial level, which may, for example, be a baseline level of enzyme activity. In some embodiments, the term refers to a reduction of the level of enzyme activity to a level that is less than 75%, less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.5%, less than 0.1%, less than 0.01%, less than 0.001%, or less than 0.0001% of an initial level, which may, for example, be a baseline level of enzyme activity.

[0124] The terms “composition” and “formulation” are used interchangeably.

[0125] A “subject” to which administration is contemplated refers to a human (i.e., male or female of any age group, e.g., pediatric subject (e.g., infant, child, or adolescent) or adult subject (e.g., young adult, middle-aged adult, or senior adult)) or non-human animal. In certain embodiments, the non-human animal is a mammal (e.g., primate (e.g., cynomolgus monkey or rhesus monkey), commercially relevant mammal (e.g., cattle, pig, horse, sheep, goat, cat, or dog), or bird (e.g., commercially relevant bird, such as chicken, duck, goose, or turkey)). In certain embodiments, the non-human animal is a fish, reptile, or amphibian. The non-human animal may be a male or female at any stage of development. The non-human animal may be a transgenic animal or genetically engineered animal. The term “patient” refers to a human subject in need of treatment of a disease.

[0126] The term “biological sample” refers to any sample including tissue samples (such as tissue sections and needle biopsies of a tissue); cell samples (e.g., cytological smears (such as Pap or blood smears) or samples of cells obtained by microdissection); samples of whole organisms (such as samples of yeasts or bacteria); or cell fractions, fragments or organelles20 / 179B0662.70120WO00(such as obtained by lysing cells and separating the components thereof by centrifugation or otherwise). Other examples of biological samples include blood, serum, urine, semen, fecal matter, cerebrospinal fluid, interstitial fluid, mucous, tears, sweat, pus, biopsied tissue (e.g., obtained by a surgical biopsy or needle biopsy), nipple aspirates, milk, vaginal fluid, saliva, swabs (such as buccal swabs), or any material containing biomolecules that is derived from a first biological sample.

[0127] The term “target tissue” refers to any biological tissue of a subject (including a group of cells, a body part, or an organ) or a part thereof, including blood and / or lymph vessels, which is the object to which a compound, particle, and / or composition of the present disclosure is delivered. A target tissue may be an abnormal or unhealthy tissue, which may need to be treated. A target tissue may also be a normal or healthy tissue that is under a higher than normal risk of becoming abnormal or unhealthy, which may need to be prevented. In certain embodiments, the target tissue is the liver. In certain embodiments, the target tissue is the lung.

[0128] A “non-target tissue” is any biological tissue of a subject (including a group of cells, a body part, or an organ) or a part thereof, including blood and / or lymph vessels, which is not a target tissue

[0129] The term “administer,” “administering,” or “administration” refers to implanting, absorbing, ingesting, injecting, inhaling, or otherwise introducing a compound described herein, or a composition thereof, in or on a subject.

[0130] The terms “condition,” “disease,” and “disorder” are used interchangeably.

[0131] The terms “treatment,” “treat,” and “treating” refer to reversing, alleviating, delaying the onset of, or inhibiting the progress of a disease described herein. In some embodiments, treatment may be administered after one or more signs or symptoms of the disease have developed or have been observed. In other embodiments, treatment may be administered in the absence of signs or symptoms of the disease. For example, treatment may be administered to a susceptible subject prior to the onset of symptoms (e.g., in light of a history of symptoms and / or in light of exposure to a pathogen). Treatment may also be continued after symptoms have resolved, for example, to delay or prevent recurrence.

[0132] The term “prevent,” “preventing,” or “prevention” refers to a prophylactic treatment of a subject who is not and was not with a disease but is at risk of developing the disease or who was with a disease, is not with the disease, but is at risk of regression of the disease. In certain embodiments, the subject is at a higher risk of developing the disease or at a higher risk of regression of the disease than an average healthy member of a population.21 / 179B0662.70120WO00

[0133] An “effective amount” of a compound or agent described herein refers to an amount sufficient to elicit the desired biological response. An effective amount of a compound or agent described herein may vary depending on such factors as the desired biological endpoint, severity of side effects, disease, or disorder, the identity, pharmacokinetics, and pharmacodynamics of the particular compound, the condition being treated, the mode, route, and desired or required frequency of administration, the species, age and health or general condition of the subject. In certain embodiments, an effective amount is a therapeutically effective amount. In certain embodiments, an effective amount is a prophylactic treatment. In certain embodiments, an effective amount is the amount of a compound or agent described herein in a single dose. In certain embodiments, an effective amount is the combined amounts of a compound or agent described herein in multiple doses. In certain embodiments, the desired dosage is delivered three times a day, two times a day, once a day, every other day, every third day, every week, every two weeks, every three weeks, or every four weeks. In certain embodiments, the desired dosage is delivered using multiple administrations (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, or more administrations).

[0134] In certain embodiments, an effective amount of a compound or agent for administration one or more times a day to a 70 kg adult human comprises about 0.0001 mg to about 3000 mg, about 0.0001 mg to about 2000 mg, about 0.0001 mg to about 1000 mg, about 0.001 mg to about 1000 mg, about 0.01 mg to about 1000 mg, about 0.1 mg to about 1000 mg, about 1 mg to about 1000 mg, about 1 mg to about 100 mg, about 10 mg to about 1000 mg, or about 100 mg to about 1000 mg, of a compound per unit dosage form.

[0135] In certain embodiments, the compounds or agents of the present disclosure are administered orally or parenterally at dosage levels sufficient to deliver from about 0.001 mg / kg to about 100 mg / kg, from about 0.01 mg / kg to about 50 mg / kg, preferably from about 0.1 mg / kg to about 40 mg / kg, preferably from about 0.5 mg / kg to about 30 mg / kg, from about 0.01 mg / kg to about 10 mg / kg, from about 0.1 mg / kg to about 10 mg / kg, and more preferably from about 1 mg / kg to about 25 mg / kg, of subject body weight per day, one or more times a day, to obtain the desired therapeutic effect.

[0136] It will be appreciated that dose ranges as described herein provide guidance for the administration of provided pharmaceutical compositions to an adult. The amount to be administered to, for example, a child or an adolescent can be determined by a medical practitioner or person skilled in the art and can be lower or the same as that administered to an adult.22 / 179B0662.70120WO00

[0137] A “therapeutically effective amount” of a compound or agent described herein is an amount sufficient to provide a therapeutic benefit in the treatment of a condition or to delay or minimize one or more symptoms associated with the condition. A therapeutically effective amount of a compound or agent means an amount of therapeutic agent, alone or in combination with other therapies, which provides a therapeutic benefit in the treatment of the condition. The term “therapeutically effective amount” can encompass an amount that improves overall therapy, reduces or avoids symptoms, signs, or causes of the condition, and / or enhances the therapeutic efficacy of another therapeutic agent. In some embodiments, a therapeutically effective amount of a compound provided herein is an amount sufficient to deliver an agent to a subject, cell, or biological sample.

[0138] A “prophylactically effective amount” of a compound or agent described herein is an amount sufficient to prevent a condition, or one or more symptoms associated with the condition or prevent its recurrence. A prophylactically effective amount of a compound or agent means an amount of a therapeutic agent, alone or in combination with other agents, which provides a prophylactic benefit in the prevention of the condition. The term “prophylactically effective amount” can encompass an amount that improves overall prophylaxis or enhances the prophylactic efficacy of another prophylactic agent. In some embodiments, a prophylactically effective amount of a compound provided herein is an amount sufficient to deliver an agent to a subject, cell, or biological sample.

[0139] The term “infectious disease” refers to diseases or disorders associated with infection, presence, or growth of a pathogenic agent in a host subject. In some embodiments, infectious disease is spread directly or indirectly from one host subject to another. Infectious pathogenic agents include, but are not limited to, viruses, bacteria, fungi, protozoa, parasites, and aberrant proteins. Viruses associated with infectious disease include but are not limited to, herpes simplex viruses, cytomegalovirus, Epstein-Barr virus, Varicella-zoster virus, herpes viruses, Vesicular stomatitis virus, Hepatitis viruses, Rhinovints, Coronavirus, Influenza viruses, Measles virus, Polyomavirus, Human Papilomavirus, Respiratory syncytial virus, Adenovirus, Coxsackie virus, Dengue virus, Mumps virus, Poliovirus, Rabies virus, Rous sarcoma virus, Yellow fever virus, Ebola virus, Simian Immunodeficiency viruses, Human Immunodeficiency viruses. Protozoa associated with infectious disease include, but are not limited to, Entamoeba histolytica, Toxoplasma gondii, Schistosoma mansoni, Cryptosporidium sp., Leishmania donovani, Neospora caninum, and Plasmodium spp. Bacteria associated with infectious disease include, but are not limited to, M. tuberculosis,23 / 179B0662.70120WO00Salmonella species, E. coli, Chlamydia species, Staphylococcus species, Bacillus species, and Pseudomonas species.

[0140] The term “genetic disease” refers to a disease caused by one or more abnormalities in the genome of a subject, such as a disease that is present from birth of the subject. Genetic diseases may be heritable and may be passed down from the parents’ genes. A genetic disease may also be caused by mutations or changes of the DNAs and / or RNAs of the subject. In such cases, the genetic disease will be heritable if it occurs in the germline. Exemplary genetic diseases include Aarskog-Scott syndrome, Aase syndrome, achondroplasia, acrodysostosis, addiction, adreno-leukodystrophy, albinism, ablepharon-macrostomia syndrome, alagille syndrome, alkaptonuria, alpha-1 antitrypsin deficiency, Alport’s syndrome, Alzheimer’s disease, asthma, autoimmune polyglandular syndrome, androgen insensitivity syndrome, Angelman syndrome, ataxia, ataxia telangiectasia, atherosclerosis, attention deficit hyperactivity disorder (ADHD), autism, baldness, Batten disease, Beckwith- Wiedemann syndrome, Best disease, bipolar disorder, brachydactyl, breast cancer, Burkitt lymphoma, chronic myeloid leukemia, Charcot-Marie-Tooth disease, Crohn’s disease, cleft lip, Cockayne syndrome, Coffin Lowry syndrome, colon cancer, congenital adrenal hyperplasia, Cornelia de Lange syndrome, Costello syndrome, Cowden syndrome, craniofrontonasal dysplasia, Crigler-Najjar syndrome, Creutzfeldt-Jakob disease, cystic fibrosis, deafness, depression, diabetes, diastrophic dysplasia, DiGeorge syndrome, Down’s syndrome, dyslexia, Duchenne muscular dystrophy, Dubowitz syndrome, ectodermal dysplasia Ellis-van Creveld syndrome, Ehlers-Danlos, epidermolysis bullosa, epilepsy, essential tremor, familial hypercholesterolemia, familial Mediterranean fever, fragile X syndrome, Friedreich’s ataxia, Gaucher’s disease, glaucoma, glucose galactose malabsorption, glutaricaciduria, gyrate atrophy, Goldberg Shprintzen syndrome (velocardiofacial syndrome), Gorlin syndrome, Hailey-Hailey disease, hemihypertrophy, hemochromatosis, hemophilia, hereditary motor and sensory neuropathy (HMSN), hereditary non polyposis colorectal cancer (HNPCC), Huntington’s disease, immunodeficiency with hyper-IgM, juvenile onset diabetes, Klinefelter’s syndrome, Kabuki syndrome, Leigh’s disease, long QT syndrome, lung cancer, malignant melanoma, manic depression, Marfan syndrome, Menkes syndrome, miscarriage, mucopolysaccharide disease, multiple endocrine neoplasia, multiple sclerosis, muscular dystrophy, myotrophic lateral sclerosis, myotonic dystrophy, neurofibromatosis, Niemann-Pick disease, Noonan syndrome, obesity, ovarian cancer, pancreatic cancer, Parkinson’s disease, paroxysmal nocturnal hemoglobinuria, Pendred syndrome, peroneal muscular atrophy, phenylketonuria (PKU), polycystic kidney24 / 179B0662.70120WO00disease, Prader-Willi syndrome, primary biliary cirrhosis, prostate cancer, REAR syndrome, Refsum disease, retinitis pigmentosa, retinoblastoma, Rett syndrome, Sanfilippo syndrome, schizophrenia, severe combined immunodeficiency, sickle cell anemia, spina bifida, spinal muscular atrophy, spinocerebellar atrophy, sudden adult death syndrome, Tangier disease, Tay-Sachs disease, thrombocytopenia absent radius syndrome, Townes-Brocks syndrome, tuberous sclerosis, Turner syndrome, Usher syndrome, von Hippel-Lindau syndrome, Waardenburg syndrome, Weaver syndrome, Werner syndrome, Williams syndrome, Wilson’s disease, xeroderma piginentosum, and Zellweger syndrome.

[0141] A “proliferative disease” refers to a disease that occurs due to abnormal growth or extension by the multiplication of cells (Walker, Cambridge Dictionary of Biology; Cambridge University Press: Cambridge, UK, 1990). A proliferative disease may be associated with: 1) the pathological proliferation of normally quiescent cells; 2) the pathological migration of cells from their normal location (e.g., metastasis of neoplastic cells); 3) the pathological expression of proteolytic enzymes such as the matrix metalloproteinases (e.g., collagenases, gelatinases, and elastases); or 4) the pathological angiogenesis as in proliferative retinopathy and tumor metastasis. Exemplary proliferative diseases include cancers (i.e., “malignant neoplasms”), benign neoplasms, angiogenesis, inflammatory diseases, and autoimmune diseases.

[0142] The term “angiogenesis” refers to the physiological process through which new blood vessels form from pre-existing vessels. Angiogenesis is distinct from vasculogenesis, which is the de novo formation of endothelial cells from mesoderm cell precursors. The first vessels in a developing embryo form through vasculogenesis, after which angiogenesis is responsible for most blood vessel growth during normal or abnormal development. Angiogenesis is a vital process in growth and development, as well as in wound healing and in the formation of granulation tissue. However, angiogenesis is also a fundamental step in the transition of tumors from a benign state to a malignant one, leading to the use of angiogenesis inhibitors in the treatment of cancer. Angiogenesis may be chemically stimulated by angiogenic proteins, such as growth factors (e.g., VEGF). “Pathological angiogenesis” refers to abnormal (e.g., excessive or insufficient) angiogenesis that amounts to and / or is associated with a disease.

[0143] The terms “neoplasm” and “tumor” are used herein interchangeably and refer to an abnormal mass of tissue wherein the growth of the mass surpasses and is not coordinated with the growth of a normal tissue. A neoplasm or tumor may be “benign” or “malignant,” depending on the following characteristics: degree of cellular differentiation (including25 / 179B0662.70120WO00morphology and functionality), rate of growth, local invasion, and metastasis. A “benign neoplasm” is generally well differentiated, has characteristically slower growth than a malignant neoplasm, and remains localized to the site of origin. In addition, a benign neoplasm does not have the capacity to infiltrate, invade, or metastasize to distant sites. Exemplary benign neoplasms include lipoma, chondroma, adenomas, acrochordon, senile angiomas, seborrheic keratoses, lentigos, and sebaceous hyperplasias. In some cases, certain “benign” tumors may later give rise to malignant neoplasms, which may result from additional genetic changes in a subpopulation of the tumor’s neoplastic cells, and these tumors are referred to as “pre-malignant neoplasms.” An exemplary pre-malignant neoplasm is a teratoma. In contrast, a “malignant neoplasm” is generally poorly differentiated (anaplasia) and has characteristically rapid growth accompanied by progressive infiltration, invasion, and destruction of the surrounding tissue. Furthermore, a malignant neoplasm generally has the capacity to metastasize to distant sites. The term “metastasis,” “metastatic,” or “metastasize” refers to the spread or migration of cancerous cells from a primary or original tumor to another organ or tissue and is typically identifiable by the presence of a “secondary tumor” or “secondary cell mass” of the tissue type of the primary or original tumor and not of that of the organ or tissue in which the secondary (metastatic) tumor is located. For example, a prostate cancer that has migrated to bone is said to be metastasized prostate cancer and includes cancerous prostate cancer cells growing in bone tissue.

[0144] The term “cancer” refers to a class of diseases characterized by the development of abnormal cells that proliferate uncontrollably and have the ability to infiltrate and destroy normal body tissues. See, e.g., Stedman’s Medical Dictionary, 25thed.; Hensyl ed.; Williams & Wilkins: Philadelphia, 1990. The cancer may be a solid tumor. The cancer may be a hematological malignancy. Exemplary cancers include acoustic neuroma; adenocarcinoma; adrenal gland cancer; anal cancer; angiosarcoma (e.g., lymphangiosarcoma, lymphangioendotheliosarcoma, hemangiosarcoma); appendix cancer; benign monoclonal gammopathy; biliary cancer (e.g., cholangiocarcinoma); bladder cancer; breast cancer (e.g., adenocarcinoma of the breast, papillary carcinoma of the breast, mammary cancer, medullary carcinoma of the breast); brain cancer (e.g., meningioma, glioblastomas, glioma (e.g., astrocytoma, oligodendroglioma), medulloblastoma); bronchus cancer; carcinoid tumor; cervical cancer (e.g., cervical adenocarcinoma); choriocarcinoma; chordoma; craniopharyngioma; colorectal cancer (e.g., colon cancer, rectal cancer, colorectal adenocarcinoma); connective tissue cancer; epithelial carcinoma; ependymoma; endotheliosarcoma (e.g., Kaposi’s sarcoma, multiple idiopathic hemorrhagic sarcoma);26 / 179B0662.70120WO00endometrial cancer (e.g., uterine cancer, uterine sarcoma); esophageal cancer (e.g., adenocarcinoma of the esophagus, Barrett’s adenocarcinoma); Ewing’s sarcoma; ocular cancer (e.g., intraocular melanoma, retinoblastoma); familiar hypereosinophilia; gall bladder cancer; gastric cancer (e.g., stomach adenocarcinoma); gastrointestinal stromal tumor (GIST); germ cell cancer; head and neck cancer (e.g., head and neck squamous cell carcinoma, oral cancer (e.g., oral squamous cell carcinoma), throat cancer (e.g., laryngeal cancer, pharyngeal cancer, nasopharyngeal cancer, oropharyngeal cancer)); hematopoietic cancers (e.g., leukemia such as acute lymphocytic leukemia (ALL) (e.g., B-cell ALL, T-cell ALL), acute myelocytic leukemia (AML) (e.g., B-cell AML, T-cell AML), chronic myelocytic leukemia (CML) (e.g., B-cell CML, T-cell CML), and chronic lymphocytic leukemia (CLL) (e.g., B- cell CLL, T-cell CLL)); lymphoma such as Hodgkin lymphoma (HL) (e.g., B-cell HL, T-cell HL) and non-Hodgkin lymphoma (NHL) (e.g., B-cell NHL such as diffuse large cell lymphoma (DLCL) (e.g., diffuse large B-cell lymphoma), follicular lymphoma, chronic lymphocytic leukemia / small lymphocytic lymphoma (CLL / SLL), mantle cell lymphoma (MCL), marginal zone B-cell lymphomas (e.g., mucosa-associated lymphoid tissue (MALT) lymphomas, nodal marginal zone B-cell lymphoma, splenic marginal zone B-cell lymphoma), primary mediastinal B-cell lymphoma, Burkitt lymphoma, lymphoplasmacytic lymphoma (i.e., Waldenström’s macroglobulinemia), hairy cell leukemia (HCL), immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma and primary central nervous system (CNS) lymphoma; and T-cell NHL such as precursor T-lymphoblastic lymphoma / leukemia, peripheral T-cell lymphoma (PTCL) (e.g., cutaneous T-cell lymphoma (CTCL) (e.g., mycosis fungoides, Sezary syndrome), angioimmunoblastic T-cell lymphoma, extranodal natural killer T-cell lymphoma, enteropathy type T-cell lymphoma, subcutaneous panniculitis-like T-cell lymphoma, and anaplastic large cell lymphoma); a mixture of one or more leukemia / lymphoma as described above; and multiple myeloma (MM)), heavy chain disease (e.g., alpha chain disease, gamma chain disease, mu chain disease); hemangioblastoma; hypopharynx cancer; inflammatory myofibroblastic tumors; immunocytic amyloidosis; kidney cancer (e.g., nephroblastoma a.k.a. Wilms’ tumor, renal cell carcinoma); liver cancer (e.g., hepatocellular cancer (HCC), malignant hepatoma); lung cancer (e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), adenocarcinoma of the lung); leiomyosarcoma (LMS); mastocytosis (e.g., systemic mastocytosis); muscle cancer; myelodysplastic syndrome (MDS); mesothelioma; myeloproliferative disorder (MPD) (e.g., polycythemia vera (PV), essential thrombocytosis (ET), agnogenic myeloid metaplasia (AMM) a.k.a. myelofibrosis (MF), chronic idiopathic27 / 179B0662.70120WO00myelofibrosis, chronic myelocytic leukemia (CML), chronic neutrophilic leukemia (CNL), hypereosinophilic syndrome (HES)); neuroblastoma; neurofibroma (e.g., neurofibromatosis (NF) type 1 or type 2, schwannomatosis); neuroendocrine cancer (e.g., gastroenteropancreatic neuroendoctrine tumor (GEP-NET), carcinoid tumor); osteosarcoma (e.g.,bone cancer); ovarian cancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarian adenocarcinoma); papillary adenocarcinoma; pancreatic cancer (e.g., pancreatic andenocarcinoma, intraductal papillary mucinous neoplasm (IPMN), Islet cell tumors); penile cancer (e.g., Paget’s disease of the penis and scrotum); pinealoma; primitive neuroectodermal tumor (PNT); plasma cell neoplasia; paraneoplastic syndromes; intraepithelial neoplasms; prostate cancer (e.g., prostate adenocarcinoma); rectal cancer; rhabdomyosarcoma; salivary gland cancer; skin cancer (e.g., squamous cell carcinoma (SCC), keratoacanthoma (KA), melanoma, basal cell carcinoma (BCC)); small bowel cancer (e.g., appendix cancer); soft tissue sarcoma (e.g., malignant fibrous histiocytoma (MFH), liposarcoma, malignant peripheral nerve sheath tumor (MPNST), chondrosarcoma, fibrosarcoma, myxosarcoma); sebaceous gland carcinoma; small intestine cancer; sweat gland carcinoma; synovioma; testicular cancer (e.g., seminoma, testicular embryonal carcinoma); thyroid cancer (e.g., papillary carcinoma of the thyroid, papillary thyroid carcinoma (PTC), medullary thyroid cancer); urethral cancer; vaginal cancer; and vulvar cancer (e.g., Paget’s disease of the vulva).

[0145] The term “neurological disease” refers to any disease of the nervous system, including diseases that involve the central nervous system (brain, brainstem and cerebellum), the peripheral nervous system (including cranial nerves), and the autonomic nervous system (parts of which are located in both central and peripheral nervous system). Neurodegenerative diseases refer to a type of neurological disease marked by the loss of nerve cells, including, but not limited to, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, tauopathies (including frontotemporal dementia), and Huntington’s disease. Examples of neurological diseases include, but are not limited to, headache, stupor and coma, dementia, seizure, sleep disorders, trauma, infections, neoplasms, neuro-ophthalmology, movement disorders, demyelinating diseases, spinal cord disorders, and disorders of peripheral nerves, muscle and neuromuscular junctions. Addiction and mental illness, include, but are not limited to, bipolar disorder and schizophrenia, are also included in the definition of neurological diseases. Further examples of neurological diseases include acquired epileptiform aphasia; acute disseminated encephalomyelitis; adrenoleukodystrophy; agenesis of the corpus callosum; agnosia; Aicardi syndrome; Alexander disease; Alpers’ disease;28 / 179B0662.70120WO00alternating hemiplegia; Alzheimer’s disease; amyotrophic lateral sclerosis; anencephaly; Angelman syndrome; angiomatosis; anoxia; aphasia; apraxia; arachnoid cysts; arachnoiditis; Arnold-Chiari malformation; arteriovenous malformation; Asperger syndrome; ataxia telangiectasia; attention deficit hyperactivity disorder; autism; autonomic dysfunction; back pain; Batten disease; Behcet’s disease; Bell’s palsy; benign essential blepharospasm; benign focal; amyotrophy; benign intracranial hypertension; Binswanger’s disease; blepharospasm; Bloch Sulzberger syndrome; brachial plexus injury; brain abscess; bbrain injury; brain tumors (including glioblastoma multiforme); spinal tumor; Brown-Sequard syndrome; Canavan disease; carpal tunnel syndrome (CTS); causalgia; central pain syndrome; central pontine myelinolysis; cephalic disorder; cerebral aneurysm; cerebral arteriosclerosis; cerebral atrophy; cerebral gigantism; cerebral palsy; Charcot-Marie-Tooth disease; chemotherapy- induced neuropathy and neuropathic pain; Chiari malformation; chorea; chronic inflammatory demyelinating polyneuropathy (CIDP); chronic pain; chronic regional pain syndrome; Coffin Lowry syndrome; coma, including persistent vegetative state; congenital facial diplegia; corticobasal degeneration; cranial arteritis; craniosynostosis; Creutzfeldt- Jakob disease; cumulative trauma disorders; Cushing’s syndrome; cytomegalic inclusion body disease (CIBD); cytomegalovirus infection; dancing eyes-dancing feet syndrome; Dandy-Walker syndrome; Dawson disease; De Morsier’s syndrome; Dejerine-Klumpke palsy; dementia; dermatomyositis; diabetic neuropathy; diffuse sclerosis; dysautonomia; dysgraphia; dyslexia; dystonias; early infantile epileptic encephalopathy; empty sella syndrome; encephalitis; encephaloceles; encephalotrigeminal angiomatosis; epilepsy; Erb’s palsy; essential tremor; Fabry’s disease; Fahr’s syndrome; fainting; familial spastic paralysis; febrile seizures; Fisher syndrome; Friedreich’s ataxia; frontotemporal dementia and other “tauopathies”; Gaucher’s disease; Gerstmann’s syndrome; giant cell arteritis; giant cell inclusion disease; globoid cell leukodystrophy; Guillain-Barre syndrome; HTLV-1 associated myelopathy; Hallervorden-Spatz disease; head injury; headache; hemifacial spasm; hereditary spastic paraplegia; heredopathia atactica polyneuritiformis; herpes zoster oticus; herpes zoster; Hirayama syndrome; HIV-associated dementia and neuropathy (see also neurological manifestations of AIDS); holoprosencephaly; Huntington’s disease and other polyglutamine repeat diseases; hydranencephaly; hydrocephalus; hypercortisolism; hypoxia; immune- mediated encephalomyelitis; inclusion body myositis; incontinentia pigmenti; infantile; phytanic acid storage disease; Infantile Refsum disease; infantile spasms; inflammatory myopathy; intracranial cyst; intracranial hypertension; Joubert syndrome; Kearns-Sayre syndrome; Kennedy disease; Kinsbourne syndrome; Klippel Feil syndrome; Krabbe disease;29 / 179B0662.70120WO00Kugelberg-Welander disease; kuru; Lafora disease; Lambert-Eaton myasthenic syndrome; Landau-Kleffner syndrome; lateral medullary (Wallenberg) syndrome; learning disabilities; Leigh’s disease; Lennox-Gastaut syndrome; Lesch-Nyhan syndrome; leukodystrophy; Lewy body dementia; lissencephaly; locked-in syndrome; Lou Gehrig’s disease (aka motor neuron disease or amyotrophic lateral sclerosis); lumbar disc disease; lyme disease-neurological sequelae; Machado-Joseph disease; macrencephaly; megalencephaly; Melkersson-Rosenthal syndrome; Menieres disease; meningitis; Menkes disease; metachromatic leukodystrophy; microcephaly; migraine; Miller Fisher syndrome; mini-strokes; mitochondrial myopathies; Mobius syndrome; monomelic amyotrophy; motor neurone disease; moyamoya disease; mucopolysaccharidoses; multi-infarct dementia; multifocal motor neuropathy; multiple sclerosis and other demyelinating disorders; multiple system atrophy with postural hypotension; muscular dystrophy; myasthenia gravis; myelinoclastic diffuse sclerosis; myoclonic encephalopathy of infants; myoclonus; myopathy; myotonia congenital; narcolepsy; neurofibromatosis; neuroleptic malignant syndrome; neurological manifestations of AIDS; neurological sequelae of lupus; neuromyotonia; neuronal ceroid lipofuscinosis; neuronal migration disorders; Niemann-Pick disease; O’Sullivan-McLeod syndrome; occipital neuralgia; occult spinal dysraphism sequence; Ohtahara syndrome; olivopontocerebellar atrophy; opsoclonus myoclonus; optic neuritis; orthostatic hypotension; overuse syndrome; paresthesia; Parkinson’s disease; paramyotonia congenita; paraneoplastic diseases; paroxysmal attacks; Parry Romberg syndrome; Pelizaeus-Merzbacher disease; periodic paralyses; peripheral neuropathy; painful neuropathy and neuropathic pain; persistent vegetative state; pervasive developmental disorders; photic sneeze reflex; phytanic acid storage disease; Pick’s disease; pinched nerve; pituitary tumors; polymyositis; porencephaly; Post-Polio syndrome; postherpetic neuralgia (PHN); postinfectious encephalomyelitis; postural hypotension; Prader-Willi syndrome; primary lateral sclerosis; prion diseases; progressive; hemifacial atrophy; progressive multifocal leukoencephalopathy; progressive sclerosing poliodystrophy; progressive supranuclear palsy; pseudotumor cerebri; Ramsay-Hunt syndrome (Type I and Type II); Rasmussen’s Encephalitis; reflex sympathetic dystrophy syndrome; Refsum disease; repetitive motion disorders; repetitive stress injuries; restless legs syndrome; retrovirus-associated myelopathy; Rett syndrome; Reye’s syndrome; Saint Vitus Dance; Sandhoff disease; Schilder’s disease; schizencephaly; septo-optic dysplasia; shaken baby syndrome; shingles; Shy-Drager syndrome; Sjogren’s syndrome; sleep apnea; Soto’s syndrome; spasticity; spina bifida; spinal cord injury; spinal cord tumors; spinal muscular atrophy; stiff-person syndrome; stroke; Sturge-Weber syndrome; subacute30 / 179B0662.70120WO00sclerosing panencephalitis; subarachnoid hemorrhage; subcortical arteriosclerotic encephalopathy; sydenham chorea; syncope; syringomyelia; tardive dyskinesia; Tay-Sachs disease; temporal arteritis; tethered spinal cord syndrome; Thomsen disease; thoracic outlet syndrome; tic douloureux; Todd’s paralysis; Tourette syndrome; transient ischemic attack; transmissible spongiform encephalopathies; transverse myelitis; traumatic brain injury; tremor; trigeminal neuralgia; tropical spastic paraparesis; tuberous sclerosis; vascular dementia (multi-infarct dementia); vasculitis including temporal arteritis; Von Hippel-Lindau Disease (VHL); Wallenberg’s syndrome; Werdnig-Hoffman disease; West syndrome; whiplash; Williams syndrome; Wilson’s disease; and Zellweger syndrome.

[0146] The term “psychiatric disorder” refers to a disease of the mind and includes diseases and disorders listed in the Diagnostic and Statistical Manual of Mental Disorders - Fourth Edition (DSM-IV), published by the American Psychiatric Association, Washington D. C. (1994). Psychiatric disorders include, but are not limited to, anxiety disorders (e.g., acute stress disorder agoraphobia, generalized anxiety disorder, obsessive-compulsive disorder, panic disorder, posttraumatic stress disorder, separation anxiety disorder, social phobia, and specific phobia), childhood disorders, (e.g., attention-deficit / hyperactivity disorder, conduct disorder, and oppositional defiant disorder), eating disorders (e.g., anorexia nervosa and bulimia nervosa), mood disorders (e.g., depression, bipolar disorder, cyclothymic disorder, dysthymic disorder, and major depressive disorder), personality disorders (e.g., antisocial personality disorder, avoidant personality disorder, borderline personality disorder, dependent personality disorder, histrionic personality disorder, narcissistic personality disorder, obsessive-compulsive personality disorder, paranoid personality disorder, schizoid personality disorder, and schizotypal personality disorder), psychotic disorders (e.g., brief psychotic disorder, delusional disorder, schizoaffective disorder, schizophreniform disorder, schizophrenia, and shared psychotic disorder), substance-related disorders (e.g., alcohol dependence, amphetamine dependence, cannabis dependence, cocaine dependence, hallucinogen dependence, inhalant dependence, nicotine dependence, opioid dependence, phencyclidine dependence, and sedative dependence), adjustment disorder, autism, delirium, dementia, multi-infarct dementia, learning and memory disorders (e.g., amnesia and age- related memory loss), and Tourette’s disorder.

[0147] The term "metabolic disorder" refers to any disorder that involves an alteration in the normal metabolism of carbohydrates, lipids, proteins, nucleic acids, or a combination thereof. A metabolic disorder is associated with either a deficiency or excess in a metabolic pathway resulting in an imbalance in metabolism of nucleic acids, proteins, lipids, and / or31 / 179B0662.70120WO00carbohydrates. Factors affecting metabolism include, and are not limited to, the endocrine (hormonal) control system (e.g., the insulin pathway, the enteroendocrine hormones including GLP-1, PYY or the like), the neural control system (e.g., GLP-1 in the brain), or the like. Examples of metabolic disorders include, but are not limited to, diabetes (e.g., Type I diabetes, Type II diabetes, gestational diabetes), hyperglycemia, hyperinsulinemia, insulin resistance, and obesity.

[0148] The terms “inflammatory disease” and “inflammatory condition” are used interchangeably herein, and refer to a disease or condition caused by, resulting from, or resulting in inflammation. Inflammatory diseases and conditions include those diseases, disorders or conditions that are characterized by signs of pain (dolor, from the generation of noxious substances and the stimulation of nerves), heat (calor, from vasodilatation), redness (rubor, from vasodilatation and increased blood flow), swelling (tumor, from excessive inflow or restricted outflow of fluid), and / or loss of function (functio laesa, which can be partial or complete, temporary or permanent. Inflammation takes on many forms and includes, but is not limited to, acute, adhesive, atrophic, catarrhal, chronic, cirrhotic, diffuse, disseminated, exudative, fibrinous, fibrosing, focal, granulomatous, hyperplastic, hypertrophic, interstitial, metastatic, necrotic, obliterative, parenchymatous, plastic, productive, proliferous, pseudomembranous, purulent, sclerosing, seroplastic, serous, simple, specific, subacute, suppurative, toxic, traumatic, and / or ulcerative inflammation. The term “inflammatory disease” may also refer to a dysregulated inflammatory reaction that causes an exaggerated response by macrophages, granulocytes, and / or T-lymphocytes leading to abnormal tissue damage and / or cell death. An inflammatory disease can be either an acute or chronic inflammatory condition and can result from infections or non-infectious causes. Inflammatory diseases include, without limitation, atherosclerosis, arteriosclerosis, autoimmune disorders, multiple sclerosis, systemic lupus erythematosus, polymyalgia rheumatica (PMR), gouty arthritis, degenerative arthritis, tendonitis, bursitis, psoriasis, cystic fibrosis, arthrosteitis, rheumatoid arthritis, inflammatory arthritis, Sjogren’s syndrome, giant cell arteritis, progressive systemic sclerosis (scleroderma), ankylosing spondylitis, polymyositis, dermatomyositis, pemphigus, pemphigoid, diabetes (e.g., Type I), myasthenia gravis, Hashimoto’s thyroiditis, Graves’ disease, Goodpasture’s disease, mixed connective tissue disease, sclerosing cholangitis, inflammatory bowel disease, Crohn’s disease, ulcerative colitis, pernicious anemia, inflammatory dermatoses, usual interstitial pneumonitis (UIP), asbestosis, silicosis, bronchiectasis, berylliosis, talcosis, pneumoconiosis, sarcoidosis, desquamative interstitial pneumonia, lymphoid interstitial pneumonia, giant cell interstitial32 / 179B0662.70120WO00pneumonia, cellular interstitial pneumonia, extrinsic allergic alveolitis, Wegener’s granulomatosis and related forms of angiitis (temporal arteritis and polyarteritis nodosa), inflammatory dermatoses, hepatitis, delayed-type hypersensitivity reactions (e.g., poison ivy dermatitis), pneumonia, respiratory tract inflammation, Adult Respiratory Distress Syndrome (ARDS), encephalitis, immediate hypersensitivity reactions, asthma, hayfever, allergies, acute anaphylaxis, rheumatic fever, glomerulonephritis, pyelonephritis, cellulitis, cystitis, chronic cholecystitis, ischemia (ischemic injury), reperfusion injury, allograft rejection, host- versus-graft rejection, appendicitis, arteritis, blepharitis, bronchiolitis, bronchitis, cervicitis, cholangitis, chorioamnionitis, conjunctivitis, dacryoadenitis, dermatomyositis, endocarditis, endometritis, enteritis, enterocolitis, epicondylitis, epididymitis, fasciitis, fibrositis, gastritis, gastroenteritis, gingivitis, ileitis, iritis, laryngitis, myelitis, myocarditis, nephritis, omphalitis, oophoritis, orchitis, osteitis, otitis, pancreatitis, parotitis, pericarditis, pharyngitis, pleuritis, phlebitis, pneumonitis, proctitis, prostatitis, rhinitis, salpingitis, sinusitis, stomatitis, synovitis, testitis, tonsillitis, urethritis, urocystitis, uveitis, vaginitis, vasculitis, vulvitis, vulvovaginitis, angitis, chronic bronchitis, osteomyelitis, optic neuritis, temporal arteritis, transverse myelitis, necrotizing fasciitis, and necrotizing enterocolitis. An ocular inflammatory disease includes, but is not limited to, post-surgical inflammation.

[0149] Additional exemplary inflammatory conditions include, but are not limited to, inflammation associated with acne, anemia (e.g., aplastic anemia, haemolytic autoimmune anaemia), asthma, arteritis (e.g., polyarteritis, temporal arteritis, periarteritis nodosa, Takayasu's arteritis), arthritis (e.g., crystalline arthritis, osteoarthritis, psoriatic arthritis, gouty arthritis, reactive arthritis, rheumatoid arthritis and Reiter's arthritis), ankylosing spondylitis, amylosis, amyotrophic lateral sclerosis, autoimmune diseases, allergies or allergic reactions, atherosclerosis, bronchitis, bursitis, chronic prostatitis, conjunctivitis, Chagas disease, chronic obstructive pulmonary disease, cermatomyositis, diverticulitis, diabetes (e.g., type I diabetes mellitus, Type II diabetes mellitus), a skin condition (e.g., psoriasis, eczema, burns, dermatitis, pruritus (itch)), endometriosis, Guillain-Barre syndrome, infection, ischaemic heart disease, Kawasaki disease, glomerulonephritis, gingivitis, hypersensitivity, headaches (e.g., migraine headaches, tension headaches), ileus (e.g., postoperative ileus and ileus during sepsis), idiopathic thrombocytopenic purpura, interstitial cystitis (painful bladder syndrome), gastrointestinal disorder (e.g., selected from peptic ulcers, regional enteritis, diverticulitis, gastrointestinal bleeding, eosinophilic gastrointestinal disorders (e.g., eosinophilic esophagitis, eosinophilic gastritis, eosinophilic gastroenteritis, eosinophilic colitis), gastritis, diarrhea, gastroesophageal reflux disease (GORD, or its synonym GERD), inflammatory33 / 179B0662.70120WO00bowel disease (IBD) (e.g., Crohn's disease, ulcerative colitis, collagenous colitis, lymphocytic colitis, ischaemic colitis, diversion colitis, Behcet's syndrome, indeterminate colitis) and inflammatory bowel syndrome (IBS)), lupus, multiple sclerosis, morphea, myeasthenia gravis, myocardial ischemia, nephrotic syndrome, pemphigus vulgaris, pernicious aneaemia, peptic ulcers, polymyositis, primary biliary cirrhosis, neuroinflammation associated with brain disorders (e.g., Parkinson's disease, Huntington's disease, and Alzheimer's disease), prostatitis, chronic inflammation associated with cranial radiation injury, pelvic inflammatory disease, reperfusion injury, regional enteritis, rheumatic fever, systemic lupus erythematosus, schleroderma, scierodoma, sarcoidosis, spondyloarthopathies, Sjogren's syndrome, thyroiditis, transplantation rejection, tendonitis, trauma or injury (e.g., frostbite, chemical irritants, toxins, scarring, burns, physical injury), vasculitis, vitiligo and Wegener's granulomatosis. In certain embodiments, the inflammatory disorder is selected from arthritis (e.g., rheumatoid arthritis), inflammatory bowel disease, inflammatory bowel syndrome, asthma, psoriasis, endometriosis, interstitial cystitis and prostatistis. In certain embodiments, the inflammatory condition is an acute inflammatory condition (e.g., for example, inflammation resulting from infection). In certain embodiments, the inflammatory condition is a chronic inflammatory condition (e.g., conditions resulting from asthma, arthritis and inflammatory bowel disease). The compounds may also be useful in treating inflammation associated with trauma and non-inflammatory myalgia. The compounds disclosed herein may also be useful in treating inflammation associated with cancer.

[0150] The term “musculoskeletal disease” or “MSD” refers to an injury and / or pain in a subject’s joints, ligaments, muscles, nerves, tendons, and structures that support limbs, neck, and back. In certain embodiments, an MSD is a degenerative disease. In certain embodiments, an MSD includes an inflammatory condition. Body parts of a subject that may be associated with MSDs include upper and lower back, neck, shoulders, and extremities (arms, legs, feet, and hands). In certain embodiments, an MSD is a bone disease, such as achondroplasia, acromegaly, bone callus, bone demineralization, bone fracture, bone marrow disease, bone marrow neoplasm, dyskeratosis congenita, leukemia (e.g., hairy cell leukemia, lymphocytic leukemia, myeloid leukemia, Philadelphia chromosome-positive leukemia, plasma cell leukemia, stem cell leukemia), systemic mastocytosis, myelodysplastic syndromes, paroxysmal nocturnal hemoglobinuria, myeloid sarcoma, myeloproliferative disorders, multiple myeloma, polycythemia vera, pearson marrow-pancreas syndrome, bone neoplasm, bone marrow neoplasm, Ewing sarcoma, osteochondroma, osteoclastoma, osteosarcoma, brachydactyly, Camurati-Engelmann syndrome, Craniosynostosis, Crouzon34 / 179B0662.70120WO00craniofacial dysostosis, dwarfism, achondroplasia, bloom syndrome, Cockayne syndrome, Ellis-van Creveld syndrome, Seckel syndrome, spondyloepiphyseal dysplasia, spondyloepiphyseal dysplasia congenita, Werner syndrome, hyperostosis, osteophyte, Klippel-Trenaunay-Weber syndrome, Marfan syndrome, McCune-Albright syndrome, osteitis, osteoarthritis, osteochondritis, osteochondrodysplasia, Kashin-Beck disease, Leri- Weill dyschondrosteosis, osteochondrosis, osteodystrophy, osteogenesis imperfecta, osteolysis, Gorham-Stout syndrome, osteomalacia, osteomyelitis, osteonecrosis, osteopenia, osteopetrosis, osteoporosis, osteosclerosis, otospondylomegaepiphyseal dysplasia, pachydermoperiostosis, Paget disease of bone, Polydactyly, Meckel syndrome, rickets, Rothmund-Thomson syndrome, Sotos syndrome, spondyloepiphyseal dysplasia, spondyloepiphyseal dysplasia congenita, syndactyly, Apert syndrome, syndactyly type II, or Werner syndrome. In certain embodiments, an MSD is a cartilage disease, such as cartilage neoplasm, osteochondritis, osteochondrodysplasia, Kashin-Beck disease, or Leri-Weill dyschondrosteosis. In certain embodiments, an MSD is hernia, such as intervertebral disk hernia. In certain embodiments, an MSD is a joint disease, such as arthralgia, arthritis (e.g., gout (e.g., Kelley-Seegmiller syndrome, Lesch-Nyhan syndrome), Lyme disease, osteoarthritis, psoriatic arthritis, reactive arthritis, rheumatic fever, rheumatoid arthritis, Felty syndrome, synovitis, Blau syndrome, nail-patella syndrome, spondyloarthropathy, reactive arthritis, Stickler syndrome, synovial membrane disease, synovitis, or Blau syndrome. In certain embodiments, an MSD is Langer-Giedion syndrome. In certain embodiments, an MSD is a muscle disease, such as Barth syndrome, mitochondrial encephalomyopathy, MELAS syndrome, MERRF syndrome, MNGIE syndrome, mitochondrial myopathy, Kearns-Sayre syndrome, myalgia, fibromyalgia, polymyalgia rheumatica, myoma, myositis, dermatomyositis, neuromuscular disease, Kearns-Sayre syndrome, muscular dystrophy, myasthenia, congenital myasthenic syndrome, Lambert-Eaton myasthenic syndrome, myasthenia gravis, myotonia, myotonia congenita, spinal muscular atrophy, tetany, ophthalmoplegia, or rhabdomyolysis. In certain embodiments, an MSD is Proteus syndrome. In certain embodiments, an MSD is a rheumatic diseases, such as arthritis (e.g., gout (e.g., Kelley-Seegmiller syndrome, Lesch-Nyhan lyme disease)), osteoarthritis, psoriatic arthritis, reactive arthritis, rheumatic fever, rheumatoid arthritis, Felty syndrome, synovitis, Blau syndrome, gout (e.g., Kelley-Seegmiller syndrome, Lesch-Nyhan syndrome), polymyalgia rheumatica, rheumatic fever, rheumatic heart disease, or Sjogren syndrome. In certain embodiments, an MSD is Schwartz-Jampel syndrome. In certain embodiments, an MSD is a skeleton disease, such as Leri-Weill dyschondrosteosis, skeleton malformations, Melnick-35 / 179B0662.70120WO00Needles syndrome, pachydermoperiostosis, Rieger syndrome, spinal column disease, intervertebral disk hernia, scoliosis, spina bifida, spondylitis, ankylosing spondylitis, spondyloarthropathy, reactive arthritis, spondyloepiphyseal dysplasia, spondyloepiphyseal dysplasia congenita, or spondylosis.

[0151] A “hematological disease” includes a disease which affects a hematopoietic cell or tissue. Hematological diseases include diseases associated with aberrant hematological content and / or function. Examples of hematological diseases include diseases resulting from bone marrow irradiation or chemotherapy treatments for cancer, diseases such as pernicious anemia, hemorrhagic anemia, hemolytic anemia, aplastic anemia, sickle cell anemia, sideroblastic anemia, anemia associated with chronic infections such as malaria, trypanosomiasis, HTV, hepatitis virus or other viruses, myelophthisic anemias caused by marrow deficiencies, renal failure resulting from anemia, anemia, polycythemia, infectious mononucleosis (EVI), acute non-lymphocytic leukemia (ANLL), acute myeloid leukemia (AML), acute promyelocytic leukemia (APL), acute myelomonocytic leukemia (AMMoL), polycythemia vera, lymphoma, acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia, Wilm’s tumor, Ewing’s sarcoma, retinoblastoma, hemophilia, disorders associated with an increased risk of thrombosis, herpes, thalassemia, antibody-mediated disorders such as transfusion reactions and erythroblastosis, mechanical trauma to red blood cells such as micro-angiopathic hemolytic anemias, thrombotic thrombocytopenic purpura and disseminated intravascular coagulation, infections by parasites such as Plasmodium, chemical injuries from, e.g., lead poisoning, and hypersplenism.

[0152] Immune disorders, such as auto-immune disorders, include, but are not limited to, arthritis (including rheumatoid arthritis, spondyloarthopathies, gouty arthritis, degenerative joint diseases such as osteoarthritis, systemic lupus erythematosus, Sjogren's syndrome, ankylosing spondylitis, undifferentiated spondylitis, Behcet's disease, haemolytic autoimmune anaemias, multiple sclerosis, amyotrophic lateral sclerosis, amylosis, acute painful shoulder, psoriatic, and juvenile arthritis), asthma, atherosclerosis, osteoporosis, bronchitis, tendonitis, bursitis, skin condition (e.g., psoriasis, eczema, burns, dermatitis, pruritus (itch)), enuresis, eosinophilic disease, gastrointestinal disorder (e.g., selected from peptic ulcers, regional enteritis, diverticulitis, gastrointestinal bleeding, eosinophilic gastrointestinal disorders (e.g., eosinophilic esophagitis, eosinophilic gastritis, eosinophilic gastroenteritis, eosinophilic colitis), gastritis, diarrhea, gastroesophageal reflux disease (GORD, or its synonym GERD), inflammatory bowel disease (IBD) (e.g., Crohn's disease, ulcerative colitis, collagenous colitis, lymphocytic colitis, ischaemic colitis, diversion colitis,36 / 179B0662.70120WO00Behcet's syndrome, indeterminate colitis) and inflammatory bowel syndrome (IBS)), and disorders ameliorated by a gastroprokinetic agent (e.g., ileus, postoperative ileus and ileus during sepsis; gastroesophageal reflux disease (GORD, or its synonym GERD); eosinophilic esophagitis, gastroparesis such as diabetic gastroparesis; food intolerances and food allergies and other functional bowel disorders, such as non-ulcerative dyspepsia (NUD) and non- cardiac chest pain (NCCP, including costo-chondritis)).

[0153] An “autoimmune disease” or “autoimmune disorder” refers to a disease arising from an inappropriate immune response of the body of a subject against substances and tissues normally present in the body. In other words, the immune system mistakes some part of the body as a pathogen and attacks its own cells. This may be restricted to certain organs (e.g., in autoimmune thyroiditis) or involve a particular tissue in different places (e.g., Goodpasture’s disease which may affect the basement membrane in both the lung and kidney). The treatment of autoimmune diseases is typically with immunosuppression, e.g., medications which decrease the immune response. Exemplary autoimmune diseases include, but are not limited to, glomerulonephritis, Goodpasture’s syndrome, necrotizing vasculitis, lymphadenitis, peri-arteritis nodosa, systemic lupus erythematosis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosis, psoriasis, ulcerative colitis, systemic sclerosis, dermatomyositis / polymyositis, anti-phospholipid antibody syndrome, scleroderma, pemphigus vulgaris, ANCA-associated vasculitis (e.g., Wegener’s granulomatosis, microscopic polyangiitis), uveitis, Sjogren’s syndrome, Crohn’s disease, Reiter’s syndrome, ankylosing spondylitis, Lyme disease, Guillain-Barré syndrome, Hashimoto’s thyroiditis, and cardiomyopathy.

[0154] In certain embodiments, the inflammatory disorder and / or the immune disorder is a gastrointestinal disorder. In some embodiments, the gastrointestinal disorder is selected from gastrointestinal disorder (e.g., selected from peptic ulcers, regional enteritis, diverticulitis, gastrointestinal bleeding, eosinophilic gastrointestinal disorders (e.g., eosinophilic esophagitis, eosinophilic gastritis, eosinophilic gastroenteritis, eosinophilic colitis), gastritis, diarrhea, gastroesophageal reflux disease (GORD, or its synonym GERD), inflammatory bowel disease (IBD) (e.g., Crohn's disease, ulcerative colitis, collagenous colitis, lymphocytic colitis, ischaemic colitis, diversion colitis, Behcet's syndrome, indeterminate colitis) and inflammatory bowel syndrome (IBS)). In certain embodiments, the gastrointestinal disorder is inflammatory bowel disease (IBD). In certain embodiments, the inflammatory condition and / or immune disorder is a skin condition. In some embodiments, the skin condition is37 / 179B0662.70120WO00pruritus (itch), psoriasis, eczema, burns or dermatitis. In certain embodiments, the skin condition is psoriasis. In certain embodiments, the skin condition is pruritis.

[0155] The term “cardiovascular disease” refers to diseases and disorders of the heart and circulatory system. Exemplary cardiovascular diseases, including cholesterol- or lipid-related disorders, include, but are not limited to acute coronary syndrome, angina, arrhythmia, arteriosclerosis, atherosclerosis, atherosclerotic lesions, carotid atherosclerosis, cerebrovascular disease, cerebral infarction, congestive heart failure, congenital heart disease, coronary heart disease, coronary artery disease, coronary plaque stabilization, dyslipidemias, dyslipoproteinemias, endothelium dysfunctions, familial hypercholeasterolemia, familial combined hyperlipidemia, hypoalphalipoproteinemia, hypertriglyceridemia, hyperbetalipoproteinemia, hypercholesterolemia, hypertension, hyperlipidemia, intermittent claudication, ischemia, ischemia reperfusion injury, ischemic heart diseases, cardiac ischemia, metabolic syndrome, multi-infarct dementia, myocardial infarction, obesity, peripheral vascular disease, reperfusion injury, restenosis, renal artery atherosclerosis, rheumatic heart disease, stroke, thrombotic disorder, thrombus formation, thromboembolism, primary or recurrent venous thromboembolism (VTE), deep vein thrombosis, pulmonary embolism, non-occlusive venous thrombosis, transitory ischemic attacks, and lipoprotein abnormalities associated with Alzheimer's disease, obesity, diabetes mellitus, syndrome X, impotence, multiple sclerosis, Parkinson's diseases, and inflammatory diseases.

[0156] The term “lung disease” or “pulmonary disease” refers to a disease of the lung. Examples of lung diseases include, but are not limited to, bronchiectasis, bronchitis, bronchopulmonary dysplasia, interstitial lung disease, occupational lung disease, emphysema, cystic fibrosis, acute respiratory distress syndrome (ARDS), severe acute respiratory syndrome (SARS), asthma (e.g., intermittent asthma, mild persistent asthma, moderate persistent asthma, severe persistent asthma), chronic bronchitis, chronic obstructive pulmonary disease (COPD), emphysema, interstitial lung disease, sarcoidosis, asbestosis, aspergilloma, aspergillosis, pneumonia (e.g., lobar pneumonia, multilobar pneumonia, bronchial pneumonia, interstitial pneumonia), pulmonary fibrosis, pulmonary tuberculosis, rheumatoid lung disease, pulmonary embolism, and lung cancer (e.g., non-small-cell lung carcinoma (e.g., adenocarcinoma, squamous-cell lung carcinoma, large-cell lung carcinoma), small-cell lung carcinoma). In some embodiments, a lung disease is a respiratory disease.38 / 179B0662.70120WO00DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS

[0157] Provided herein are elastin-like polypeptides, endosomal escape peptides, and compounds comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides. Also provided herein are compositions comprising a compound provided herein and an agent. In addition, provided herein are methods of delivering an agent and methods of treating or preventing a disease. Compounds

[0158] In one aspect, provided herein is a compound comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides.

[0159] In some embodiments, the compound comprises an elastin-like polypeptide chemically conjugated to one or more endosomal escape peptides. In some embodiments, the elastin-like polypeptide is chemically conjugated to one or more endosomal escape peptides via a chemical reaction that creates a covalent bond. In some embodiments, the elastin-like polypeptide is chemically conjugated to one or more endosomal escape peptides via a chemical reaction with one or more additional reagents. In some embodiments, the elastin- like polypeptide is chemically conjugated to one or more endosomal escape peptides via reaction at a lysine residue, a cysteine residue, or a tyrosine residue. In some embodiments, the elastin-like polypeptide is chemically conjugated to one or more endosomal escape peptides via reaction with one or more of N-hydroxysuccinimide esters, isocyanates, isothiocyanates, benzoyl fluorides, maleimides, iodoacetamides, 2-thiopyridines, 3- arylpropiolonitriles, diazonium salts, or cyclic diazodicarboxyamide derivatives. In some embodiments, the elastin-like polypeptide is chemically conjugated to one or more endosomal escape peptides via acylation, disulfide bond formation, amide bond formation, electrophilic aromatic substitution, or Mannich reaction. In some embodiments, the elastin- like polypeptide is chemically conjugated to one or more endosomal escape peptides via reaction at the N-terminus. In some embodiments, the elastin-like polypeptide is chemically conjugated to one or more endosomal escape peptides via reaction at the C-terminus. In some embodiments, the elastin-like polypeptide is chemically conjugated to one or more endosomal escape peptides via functionalization of the N- or C-terminus amino acid.

[0160] In some embodiments, the compound comprises an elastin-like polypeptide recombinantly conjugated to one or more endosomal escape peptides. In some embodiments, the recombinant conjugation comprises producing the compound via an oligonucleotide sequence encoding both the elastin-like polypeptide and one or more endosomal escape39 / 179B0662.70120WO00peptides. In some embodiments, the compound comprises an elastin-like polypeptide recombinantly conjugated to one or more endosomal escape peptides via a method provided herein.

[0161] In some embodiments, the elastin-like polypeptide comprises a hydrophilic block fused to a hydrophobic block. In some embodiments, the elastin-like polypeptide comprises a hydrophilic block fused to a hydrophobic block separated by a crosslinking block.

[0162] In some embodiments, the elastin-like polypeptide is conjugated via an intervening agent to one or more endosomal escape peptides. In some embodiments, the elastin-like polypeptide is conjugated at its C-terminus via an intervening agent to one or more endosomal escape peptides. In some embodiments, the elastin-like polypeptide is conjugated at its N-terminus via an intervening agent to one or more endosomal escape peptides. In some embodiments, the intervening agent is any agent provided herein. In some embodiments, the intervening agent is a peptide. In some embodiments, the peptide is ovalbumin-derived MHC class I epitope or ovalbumin-derived MHC class II epitope. In some embodiments, the peptide is ovalbumin-derived MHC class I epitope. In some embodiments, the peptide is ovalbumin-derived MHC class II epitope.

[0163] In some embodiments, the hydrophilic block has the sequence: [VPGXG]n (SEQ ID NO: 114) wherein: each instance of X is independently any amino acid except proline; and n is at least 5.

[0164] In some embodiments, the hydrophilic block has the sequence: [VPGXG]n (SEQ ID NO: 114) wherein: each instance of X is any amino acid except proline; and n is at least 5.

[0165] In some embodiments, n is at least 5. In some embodiments, n is at least 10. In some embodiments, n is at least 20. In some embodiments, n is at least 30. In some embodiments, n is 5-200. In some embodiments, n is 5-150. In some embodiments, n is 5-100. In some embodiments, n is 5-75. In some embodiments, n is 5-50. In some embodiments, n is 10-200. In some embodiments, n is 10-150. In some embodiments, n is 10-100. In some embodiments, n is 10-75. In some embodiments, n is 10-50. In some embodiments, n is 25- 200. In some embodiments, n is 25-150. In some embodiments, n is 25-100. In some40 / 179B0662.70120WO00embodiments, n is 25-75. In some embodiments, n is 25-50. In some embodiments, n is 30- 70.

[0166] In some embodiments, each instance of X is independently A, R, N, D, C, E, Q, G, H, I, L, K, M, F, S, T, W, Y, or V. In some embodiments, each instance of X is independently E, V, A, G, S, or Q. In some embodiments, each instance of X is independently G, S, or Q. In some embodiments, each instance of X is independently E, V, A, or G. In some embodiments, each instance of X is E, V, A, or G. In some embodiments, each instance of X is independently E or V. In some embodiments, each instance of X is E or V. In some embodiments, each instance of X is independently E or A. In some embodiments, each instance of X is independently E or G. In some embodiments, each instance of X is independently A or G. In some embodiments, each instance of X is A or G. In some embodiments, each instance of X is independently A or V. In some embodiments, each instance of X is independently V or G. In some embodiments, each instance of X is independently S or G. In some embodiments, each instance of X is independently Q or G. In some embodiments, each instance of X is G.

[0167] In some embodiments, each instance of X is independently X1or X2. In some embodiments, X is X1or X2.

[0168] In some embodiments, the hydrophilic block has the sequence: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]n1 (SEQ ID NO: 22) wherein: X1and X2are any amino acid except proline; each instance of X1is the same; each instance of X2is the same; X1and X2are different amino acids; and n1 = n / 5.

[0169] In some embodiments, each instance of X1is A, R, N, D, C, E, Q, G, H, I, L, K, M, F, S, T, W, Y, or V. In some embodiments, each instance of X1is S, Q, E, V, A, or G. In some embodiments, each instance of X1is S, Q, V, A, or G. In some embodiments, each instance of X1is E, V, A, or G. In some embodiments, each instance of X1is V or A. In some embodiments, each instance of X1is V or each instance of X1is A. In some embodiments, each instance of X1is G, S, or Q. In some embodiments, each instance of X1is G, each instance of X1is S, or each instance of X1is Q. In some embodiments, each instance of X1is A. In some embodiments, each instance of X1is V. In some embodiments, each instance of41 / 179B0662.70120WO00X1is G. In some embodiments, each instance of X1is S. In some embodiments, each instance of X1is Q.

[0170] In some embodiments, each instance of X2is A, R, N, D, C, E, Q, G, H, I, L, K, M, F, S, T, W, Y, or V. In some embodiments, each instance of X2is E, V, A, or G. In some embodiments, each instance of X2is E or G. In some embodiments, each instance of X2is E or each instance of X2is G. In some embodiments, each instance of X2is E. In some embodiments, each instance of X2is G.

[0171] In some embodiments, each instance of X1is V, and each instance of X2is E. In some embodiments, each instance of X1is V, and each instance of X2is G. In some embodiments, each instance of X1is A, and each instance of X2is E. In some embodiments, each instance of X1is A, and each instance of X2is G. In some embodiments, each instance of X1is G, and each instance of X2is G. In some embodiments, each instance of X1is G, and each instance of X2is E. In some embodiments, each instance of X1is S, and each instance of X2is G. In some embodiments, each instance of X1is S, and each instance of X2is E. In some embodiments, each instance of X1is Q, and each instance of X2is G. In some embodiments, each instance of X1is Q, and each instance of X2is E.

[0172] In some embodiments, the hydrophilic block has the sequence: [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10 (SEQ ID NO: 23).

[0173] In some embodiments, the hydrophilic block has the sequence: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10(SEQ ID NO: 24).

[0174] In some embodiments, the hydrophilic block has the sequence: [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10 (SEQ ID NO: 202).

[0175] In some embodiments, the hydrophilic block has the sequence: [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10(SEQ ID NO: 215).

[0176] In some embodiments, the hydrophilic block has the sequence: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10 (SEQ ID NO: 216).

[0177] In some embodiments, the crosslinking block comprises one or more cysteine residues. In some embodiments, the crosslinking block has the sequence –CCCCGGG– (SEQ ID NO: 19). In some embodiments, the crosslinking block has the sequence –[CAP]5– (SEQ ID NO: 125). In some embodiments, the crosslinking block has the sequence –[CEAAAK]5– (SEQ ID NO: 126). In some embodiments, the crosslinking block has the sequence –C5G3– (SEQ ID NO: 127).

[0178] In some embodiments, the crosslinking block is 4-25 amino acids in length. In some embodiments, the crosslinking block is 4-15 amino acids in length. In some embodiments, the42 / 179B0662.70120WO00crosslinking block is 4-10 amino acids in length. In some embodiments, the crosslinking block is 6-25 amino acids in length. In some embodiments, the crosslinking block is 6-15 amino acids in length. In some embodiments, the crosslinking block is 6-10 amino acids in length. In some embodiments, the crosslinking block is 10-25 amino acids in length. In some embodiments, the crosslinking block is 10-15 amino acids in length. In some embodiments, the crosslinking block is 10-20 amino acids in length. In some embodiments, the crosslinking block is 15-25 amino acids in length. In some embodiments, the crosslinking block is 20-25 amino acids.

[0179] In some embodiments, the hydrophobic block comprises one or more instances of G, H, P, A, I, L, M, F, W, Y, V, or C. In some embodiments, the hydrophobic block comprises one or more instances of G, H, P, A, I, L, M, F, W, Y, or V. In some embodiments, the hydrophobic block comprises one or more instances of G, P, H, Y, V, I, L, F, or W. In some embodiments, the hydrophobic block comprises one or more instances of G, P, H, Y, V, I, L, F, W, or C. In some embodiments, the hydrophobic block comprises one or more tyrosine residues. In some embodiments, the hydrophobic block comprises one or more cysteine residues.

[0180] In some embodiments, the hydrophobic block has the sequence: (Z1PGZ2G)m (SEQ ID NO: 25) wherein: Z1and Z2are each independently a hydrophobic amino acid; and m is an integer from 5 to 400.

[0181] In some embodiments, each instance of Z1is independently a hydrophobic amino acid. In some embodiments, each instance of Z1is independently P, A, I, L, M, F, W, Y, or V. In some embodiments, each instance of Z1is independently I, V, A, or L. In some embodiments, each instance of Z1is independently I, V, or L. In some embodiments, each instance of Z1is independently I, V, or A. In some embodiments, each instance of Z1is independently I, A, or L. In some embodiments, each instance of Z1is independently V, A, or L. In some embodiments, each instance of Z1is independently I or V. In some embodiments, each instance of Z1is independently I or L. In some embodiments, each instance of Z1is independently I or A. In some embodiments, each instance of Z1is independently V or A. In some embodiments, each instance of Z1is independently V or L. In some embodiments, each instance of Z1is independently A or L. In some embodiments, Z1is I. In some embodiments, Z1is L. In some embodiments, Z1is V. In some embodiments, Z1is A.43 / 179B0662.70120WO00

[0182] In some embodiments, each instance of Z2is independently a hydrophobic amino acid. In some embodiments, each instance of Z2is independently C, P, A, I, L, M, F, W, Y, or V. In some embodiments, each instance of Z2is independently C, P, H, A, I, L, M, F, W, Y, or V. In some embodiments, each instance of Z2is independently C, H, Y, V, I, L, F, or W. In some embodiments, each instance of Z2is independently P, A, I, L, M, F, W, Y, or V. In some embodiments, each instance of Z2is independently P, H, A, I, L, M, F, W, Y, or V. In some embodiments, each instance of Z2is independently H, Y, V, I, L, F, or W. In some embodiments, each instance of Z2is independently C, H, or Y. In some embodiments, Z2is an aromatic amino acid. In some embodiments, Z2is H, F, Y, or W. In some embodiments, Z2is H, F, or Y. In some embodiments, Z2is H, F, or W. In some embodiments, Z2is H, Y, or W. In some embodiments, Z2is F, Y, or W. In some embodiments, Z2is H or F. In some embodiments, Z2is H or Y. In some embodiments, Z2is H or W. In some embodiments, Z2is F, or Y. In some embodiments, Z2is F or W. In some embodiments, Z2is Y or W. In some embodiments, at least one instance of Z2is H. In some embodiments, Z2is H. In some embodiments, Z2is F. In some embodiments, at least one instance of Z2is Y. In some embodiments, Z2is Y. In some embodiments, Z2is W. In some embodiments, at least one instance of Z2is C.

[0183] In some embodiments, m is an integer from 5 to 400. In some embodiments, m is an integer from 5 to 300. In some embodiments, m is an integer from 5 to 200. In some embodiments, m is an integer from 5 to 100. In some embodiments, m is an integer from 15 to 400. In some embodiments, m is an integer from 15 to 300. In some embodiments, m is an integer from 15 to 200. In some embodiments, m is an integer from 15 to 100. In some embodiments, m is an integer from 25 to 400. In some embodiments, m is an integer from 25 to 300. In some embodiments, m is an integer from 25 to 200. In some embodiments, m is an integer from 25 to 100. In some embodiments, m is an integer from 50 to 400. In some embodiments, m is an integer from 50 to 300. In some embodiments, m is an integer from 50 to 200. In some embodiments, m is an integer from 50 to 100.

[0184] In some embodiments, the hydrophobic block has the sequence: [(IPGZ2G)2(VPGZ2G)(IPGZ2G)2]m1 (SEQ ID NO: 26), wherein m1 is an integer from 1 to 80.

[0185] In some embodiments, m1 is an integer from 1 to 70. In some embodiments, m1 is an integer from 1 to 60. In some embodiments, m1 is an integer from 1 to 50. In some embodiments, m1 is an integer from 1 to 40. In some embodiments, m1 is an integer from 1 to 30. In some embodiments, m1 is an integer from 1 to 20. In some embodiments, m1 is an44 / 179B0662.70120WO00integer from 5 to 20. In some embodiments, m1 is an integer from 5 to 30. In some embodiments, m1 is an integer from 5 to 40. In some embodiments, m1 is an integer from 5 to 50. In some embodiments, m1 is an integer from 5 to 60. In some embodiments, m1 is an integer from 5 to 70. In some embodiments, m1 is an integer from 5 to 80. In some embodiments, m1 is an integer from 1 to 20. In some embodiments, m1 is an integer from 20 to 40. In some embodiments, m1 is an integer from 40 to 60. In some embodiments, m1 is an integer from 60 to 80.

[0186] In some embodiments, the hydrophobic block has the sequence: [(IPGVG)2(VPGYG)(IPGVG)2]15 (SEQ ID NO: 27).

[0187] In some embodiments, the hydrophobic block has the sequence: [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 28).

[0188] In some embodiments, the hydrophobic block has the sequence: [(IPGZ2G)2(VPGYG)(IPGZ2G)2]15 (SEQ ID NO: 128); wherein: each instance of Z2is independently C or H.

[0189] In some embodiments, three instances of Z2are C. In some embodiments, five instances of Z2are C. In some embodiments, seven instances of Z2are C. In some embodiments, thirteen instances of Z2are C. In some embodiments, three instances of Z2are C and the remaining instances of Z2are H. In some embodiments, five instances of Z2are C and the remaining instances of Z2are H. In some embodiments, seven instances of Z2are C and the remaining instances of Z2are H. In some embodiments, thirteen instances of Z2are C and the remaining instances of Z2are H.

[0190] In some embodiments, the hydrophobic block has the sequence: [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGH GIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPG HGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIP GHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 230); [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGH GIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPG45 / 179B0662.70120WO00HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPG HGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGCGIPGHGVPGYGIP GHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 231); [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGH GIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPG CGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIP GHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 232); or [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGH GIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPG CGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIP GHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 233).

[0191] In some embodiments, the elastin-like polypeptide comprises the sequence of: [VPGXG]n–CCCCGGG–(Z1PGZ2G)m(SEQ ID NO: 29) wherein: each instance of X is any amino acid except proline; n is at least 5; Z1and Z2are each independently a hydrophobic amino acid; and m is an integer from 5 to 400.

[0192] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]n1–CCCCGGG–(Z1PGZ2G)m(SEQ ID NO: 129) wherein: X1and X2are any amino acid except proline; each instance of X1is the same; each instance of X2is the same;46 / 179B0662.70120WO00n1 = n / 5; Z1and Z2are each independently a hydrophobic amino acid; and m is an integer from 5 to 400.

[0193] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]n1–CCCCGGG–(Z1PGZ2G)m (SEQ ID NO: 30) wherein: X1and X2are any amino acid except proline; each instance of X1is the same; each instance of X2is the same; X1and X2are different amino acids; n1 = n / 5; Z1and Z2are each independently a hydrophobic amino acid; and m is an integer from 5 to 400.

[0194] In some embodiments, the elastin-like polypeptide comprises the sequence of: [VPGXG]n–CCCCGGG–[(IPGZ2G)2(VPGZ2G)(IPGZ2G)2]m1 (SEQ ID NO: 31) wherein: each instance of X is any amino acid except proline; n is at least 5; m1 is an integer from 1 to 80; and each instance of Z2is independently a hydrophobic amino acid.

[0195] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]n1–CCCCGGG– [(IPGZ2G)2(VPGZ2G)(IPGZ2G)2]m1(SEQ ID NO: 130) wherein: X1and X2are any amino acid except proline; each instance of X1is the same; each instance of X2is the same; n1 = n / 5; m1 is an integer from 1 to 80; and each instance of Z2is independently a hydrophobic amino acid.

[0196] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]n1–CCCCGGG– [(IPGZ2G)2(VPGZ2G)(IPGZ2G)2]m1(SEQ ID NO: 32)47 / 179B0662.70120WO00wherein: X1and X2are any amino acid except proline; each instance of X1is the same; each instance of X2is the same; X1and X2are different amino acids; n1 = n / 5; m1 is an integer from 1 to 80; and each instance of Z2is independently a hydrophobic amino acid.

[0197] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 217), wherein: each instance of X1is independently G, S, Q, V, or A; and each instance of X2is independently E or G.

[0198] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 33), wherein: each instance of X1is independently V or A; and each instance of X2is independently E or G.

[0199] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 33), wherein: X1is V or A; and X2is E or G.

[0200] In some embodiments, the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 218), wherein: each instance of X1is independently G, S, or Q; and each instance of X2is G.

[0201] In some embodiments, the elastin-like polypeptide comprises a sequence of:48 / 179B0662.70120WO00[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 34).

[0202] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 113).

[0203] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 131).

[0204] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 219).

[0205] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 220).

[0206] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CAP]5– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 199).

[0207] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CEAAAK]5– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 200).

[0208] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–C5G3– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 201).

[0209] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[(IPGZ2G)2(VPGYG)(IPG Z2G)2]15 (SEQ ID NO: 132); wherein: each instance of Z2is independently C or H.

[0210] In some embodiments, the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP49 / 179B0662.70120WO00GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 234); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 235); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 236); or [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 237).

[0211] In some embodiments, the elastin-like polypeptide is a polypeptide disclosed in Kim, W. et al. Recombinant Amphiphilic Protein Micelles for Drug Delivery. Langmuir, 2011, 27, 14329-14334, in Kim, W. et al. Targeted Antithrombotic Protein Micelles. Angew. Chem. Int. Ed., 2015, 54, 1461-1465, in Abdelghani, M. et al. Applications of elastin-like polypeptides50 / 179B0662.70120WO00in nanomedicine. [Phd Thesis 1 (Research TU / e / Graduation TU / e), Biomedical Engineering], Technische Universiteit Eindhoven, September 2, 2021, or in Yi, A. Development of elastin-like polypeptide for targeted specific gene delivery in vivo. J. Nanobiotechnol., 2020, 18:15, the contents of which are incorporated herein by reference in their entireties.

[0212] Endosomal escape peptides (EEPs) disrupt endosomal membranes, acting after endocytosis to trigger cargo release into the cytosol. In some embodiments, the endosomal escape peptide is a membrane active peptide. In some embodiments, the endosomal escape peptide facilitates entry of macromolecular cargo into the cytosol following endocytosis. In some embodiments, a compound provided herein (e.g., an ELP-EEP compound) contains a core-facing membrane active peptide. In some embodiments, a compound provided herein (e.g., an ELP-EEP compound) enables endosomolytic function separate from any cell penetrating capacity. Without wishing to be bound by any particular theory, this effect is achieved because the peptide is hidden from the particle surface at the time of endocytosis.

[0213] In some embodiments, the endosomal escape peptide is not a cell penetrating peptide (CPP). CPPs function to drive nanoparticle uptake into cells. CPPs, e.g. Tat,11penetratin,41transportan-10,50Aurein1.2, or oligoarginine49demonstrate non-specific membrane disruptive capacity that facilitates cargo internalization at the level of the plasma membrane via translocation or endocytosis, but can also induce significant cellular toxicity. CPPs are generally not sufficient to enable functional cytosolic delivery. In some embodiments, the endosomal escape peptide is not one or more of S10, Penetratin, Tat, R9, Transportan-10, L17E M-Lycotoxin, or Aurein1.2. In some embodiments, the endosomal escape peptide is one or more of S10, Penetratin, Tat, R9, Transportan-10, L17E M-Lycotoxin, or Aurein1.2.

[0214] In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 200 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 190 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 180 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 170 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 160 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 150 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 140 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 130 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 1051 / 179B0662.70120WO00to about 120 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 110 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 100 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 90 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 80 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 70 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 10 to about 60 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 20 to about 60 amino acids in length.

[0215] In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 200 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 190 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 180 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 170 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 160 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 150 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 140 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 130 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 120 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 110 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 100 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 60 to about 90 amino acids in length.

[0216] In some embodiments, the endosomal escape peptide is a peptide of about 100 to about 200 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 100 to about 190 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 100 to about 180 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 100 to about 170 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 100 to about 160 amino acids in length. In some embodiments, the endosomal escape52 / 179B0662.70120WO00peptide is a peptide of about 100 to about 150 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of about 100 to about 140 amino acids in length.

[0217] In some embodiments, the endosomal escape peptide is a peptide of at least 10 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of at least 20 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of at least 30 amino acids in length.

[0218] In some embodiments, the endosomal escape peptide is a peptide of at most 200 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of at most 170 amino acids in length. In some embodiments, the endosomal escape peptide is a peptide of at most 140 amino acids in length.

[0219] In some embodiments, the endosomal escape peptide is a peptide comprising one or more amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising one to ten amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising two to eight amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising two to six amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising one amphipathic alpha helical motif. In some embodiments, the endosomal escape peptide is a peptide comprising one or more amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising one or two amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising one amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising two amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising two or more amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising three amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising four amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising five amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising six amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising seven amphipathic alpha helical motifs. In some embodiments, the endosomal escape peptide is a peptide comprising eight amphipathic alpha helical motifs.

[0220] In some embodiments, the amphipathic alpha helical motifs comprise a positively charged hydrophilic outer face and a hydrophobic outer face. In some embodiments, the53 / 179B0662.70120WO00amphipathic alpha helical motifs are separated by a flexible peptide linker. In some embodiments, an amphipathic alpha helix comprises a sequence in which a hydrophilic amino acid is present at every second or third position in the sequence. In some embodiments, an amphipathic alpha helix comprises a sequence in which a hydrophilic amino acid is present at every second position in the sequence. In some embodiments, an amphipathic alpha helix comprises a sequence in which a hydrophilic amino acid is present at every third position in the sequence. In some embodiments, an amphipathic alpha helix comprises a sequence in which a hydrophobic amino acid is present at every third or fourth position in the sequence. In some embodiments, an amphipathic alpha helix comprises a sequence in which a hydrophobic amino acid is present at every third position in the sequence. In some embodiments, an amphipathic alpha helix comprises a sequence in which a hydrophobic amino acid is present at every fourth position in the sequence.

[0221] In some embodiments, the endosomal escape peptide comprises about 10% to about 85% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V (i.e., about 10% to about 85% of the amino acids of the endosomal escape peptide are selected from A, C, G, I, L, M, F, P, W, Y, R, and V). In some embodiments, the endosomal escape peptide comprises about 20% to about 75% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 30% to about 70% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 30% to about 65% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 25% to about 65% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 40% to about 65% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 40% to about 55% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 30% to about 55% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 20% to about 30% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 30% to about 40% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 40% to about 50% of any54 / 179B0662.70120WO00combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 50% to about 60% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 55% to about 65% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V. In some embodiments, the endosomal escape peptide comprises about 65% to about 75% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V.

[0222] In some embodiments, the endosomal escape peptide comprises about 5% to about 65% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 15% to about 55% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 15% to about 45% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 25% to about 55% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 25% to about 45% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 35% to about 45% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 25% to about 35% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 15% to about 25% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 25% to about 35% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 35% to about 45% of any combination of the amino acids K and R. In some embodiments, the endosomal escape peptide comprises about 45% to about 55% of any combination of the amino acids K and R.

[0223] In some embodiments, the endosomal escape peptide has a net charge between about +6 and about +20. In some embodiments, the endosomal escape peptide has a net charge between about +8 and about +18. In some embodiments, the endosomal escape peptide has a net charge between about +10 and about +16. In some embodiments, the endosomal escape peptide has a net charge between about +10 and about +18. In some embodiments, the endosomal escape peptide has a net charge between about +8 and about +16. In some embodiments, the endosomal escape peptide has a net charge between about +12 and about +16. In some embodiments, the endosomal escape peptide has a net charge between about +10 and about +14. In some embodiments, the endosomal escape peptide has a net charge between about +6 and about +8. In some embodiments, the endosomal escape peptide has a55 / 179B0662.70120WO00net charge between about +8 and about +10. In some embodiments, the endosomal escape peptide has a net charge between about +10 and about +12. In some embodiments, the endosomal escape peptide has a net charge between about +12 and about +14. In some embodiments, the endosomal escape peptide has a net charge between about +14 and about +16. In some embodiments, the endosomal escape peptide has a net charge between about +16 and about +18. In some embodiments, the endosomal escape peptide has a net charge between about +18 and about +20.

[0224] In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 2 and about 20. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 4 and about 18. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 6 and about 16. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 4 and about 16. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 6 and about 18. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 6 and about 14. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 8 and about 16. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 2 to about 6. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 6 to about 10. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 10 to about 12. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 12 to about 16. In some embodiments, the endosomal escape peptide has a hydrophobic moment between about 16 to about 20.

[0225] In some embodiments, the endosomal escape peptide has a helical content between about 5% and about 80%. In some embodiments, the endosomal escape peptide has a helical content between about 10% and about 75%. In some embodiments, the endosomal escape peptide has a helical content between about 14% and about 72%. In some embodiments, the endosomal escape peptide has a helical content between about 10% and about 80%. In some embodiments, the endosomal escape peptide has a helical content between about 5% and about 75%. In some embodiments, the endosomal escape peptide has a helical content between about 10% and about 70%. In some embodiments, the endosomal escape peptide has a helical content between about 15% and about 75%. In some embodiments, the endosomal escape peptide has a helical content between about 5% to about 25%. In some embodiments, the endosomal escape peptide has a helical content between about 25% to about 45%. In56 / 179B0662.70120WO00some embodiments, the endosomal escape peptide has a helical content between about 45% to about 65%. In some embodiments, the endosomal escape peptide has a helical content between about 65% to about 85%.

[0226] In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 90% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 80% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 70% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 60% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 50% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 40% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 30% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 20% and about 100% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 40% and about 90% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 40% and about 80% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 40% and about 70% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 40% and about 60% K. In some embodiments, wherein positively charged residues of the endosomal escape peptide comprise between about 60% and about 80% K.

[0227] In some embodiments, the endosomal escape peptide is conjugated to the N-terminus of the elastin-like polypeptide. In some embodiments, the endosomal escape peptide is conjugated to the C-terminus of the elastin-like polypeptide.

[0228] In some embodiments, the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 2); WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K (SEQ ID NO: 3); TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK (SEQ ID NO: 4);57 / 179B0662.70120WO00MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 5); LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 6); LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 7); RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K (SEQ ID NO: 8); KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 9); KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 10); KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 11); KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 12); KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 13); KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 14); KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 15); ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 16); KWKLARAFARAIKKLGGSGGGSYARALRRQARTG (SEQ ID NO: 36); RQIKIWFQNRRMKWKK (SEQ ID NO: 37); CKRKKRRQRRRG (SEQ ID NO: 38); RRRRRRRRR (SEQ ID NO: 39); AGYLLGKINLKALAALAKKIL (SEQ ID NO: 40); IWLTALKFLGKHAAKHEAKQQLSKL (SEQ ID NO: 41); and GLFDIIKKIAESF (SEQ ID NO: 42).

[0229] In some embodiments, the endosomal escape peptide comprises a sequence of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 2).

[0230] In some embodiments, the endosomal escape peptide comprises a sequence of: WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K (SEQ ID NO: 3).

[0231] In some embodiments, the endosomal escape peptide comprises a sequence of: TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK (SEQ ID NO: 4).

[0232] In some embodiments, the endosomal escape peptide comprises a sequence of:58 / 179B0662.70120WO00MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 5).

[0233] In some embodiments, the endosomal escape peptide comprises a sequence of: LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 6).

[0234] In some embodiments, the endosomal escape peptide comprises a sequence of: LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 7).

[0235] In some embodiments, the endosomal escape peptide comprises a sequence of: RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K (SEQ ID NO: 8).

[0236] In some embodiments, the endosomal escape peptide comprises a sequence of: KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 9).

[0237] In some embodiments, the endosomal escape peptide comprises a sequence of: KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 10).

[0238] In some embodiments, the endosomal escape peptide comprises a sequence of: KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 11).

[0239] In some embodiments, the endosomal escape peptide comprises a sequence of: KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 12).

[0240] In some embodiments, the endosomal escape peptide comprises a sequence of: KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 13).

[0241] In some embodiments, the endosomal escape peptide comprises a sequence of: KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 14).

[0242] In some embodiments, the endosomal escape peptide comprises a sequence of: KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 15).

[0243] In some embodiments, the endosomal escape peptide comprises a sequence of: ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 16).

[0244] In some embodiments, the endosomal escape peptide comprises a sequence of: KWKLARAFARAIKKLGGSGGGSYARALRRQARTG (SEQ ID NO: 36).

[0245] In some embodiments, the endosomal escape peptide comprises a sequence of: RQIKIWFQNRRMKWKK (SEQ ID NO: 37).

[0246] In some embodiments, the endosomal escape peptide comprises a sequence of: CKRKKRRQRRRG (SEQ ID NO: 38).59 / 179B0662.70120WO00

[0247] In some embodiments, the endosomal escape peptide comprises a sequence of: RRRRRRRRR (SEQ ID NO: 39).

[0248] In some embodiments, the endosomal escape peptide comprises a sequence of: AGYLLGKINLKALAALAKKIL (SEQ ID NO: 40).

[0249] In some embodiments, the endosomal escape peptide comprises a sequence of: IWLTALKFLGKHAAKHEAKQQLSKL(SEQ ID NO: 41).

[0250] In some embodiments, the endosomal escape peptide comprises a sequence of:

[0251] GLFDIIKKIAESF (SEQ ID NO: 42).In some embodiments, the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: RMKFNVWAWICVVKKKKFFPHWKIRYIIGRAWLL (SEQ ID NO: 133); DVHRRIWIILLPGHRIKFNWKQWKWRFEFEFKAVRK(SEQ ID NO: 134); KLVRYISYLKLRGGVKILLAKIKPFYEVKLKLKRLSLII(SEQ ID NO: 135); YFLARARARAILIKFFGKLKPFKAKKLAKQLKWVART(SEQ ID NO: 136); ANHKFGVYFGGSGGGSGVKWQVKFGVNHANHVKF(SEQ ID NO: 137); ANHGHKFGVKLNHGGSGGGSYFKFGKAPGVKW(SEQ ID NO: 138); GVGVREWKFGGSGGGSYYFGVKGHKKGVGV(SEQ ID NO: 139); GVKFGVGVKKYFKYFASGVKGVKYF(SEQ ID NO: 140); GVGVKYFKVAVKNHKPKWFSYKAAWKKFG(SEQ ID NO: 141); GNHKAYKWVRAPRTNHKHKKAVKWKHKPG(SEQ ID NO: 142); LKFWGASAPAPHKFAWKFGVGARASGHKLL(SEQ ID NO: 143); VKWKWQWGVKGAPGYFYLGFGVKTAWAPAWAW(SEQ ID NO: 144); FLKFGGSGGGSHKVKFGVVGNHKKAPVHKW(SEQ ID NO: 145); GVGASLGKQWKFGGSGGGSNHVKWFGVKPSKGHKGAWKWS(SEQ ID NO: 146); KPIQWGVGGSGGGSYLAPRTKKHVKFGVKRTHKFAP(SEQ ID NO: 147); WKWLKAPKPILKILGKPVKHKWKPAWKGKWQ(SEQ ID NO: 148); KSILKWGAKPKKALILASLGVKKLLAPSGAPW(SEQ ID NO: 149); GVKPAWRASLGGKHVKRLSGFHKPRTKAPSL(SEQ ID NO: 150); KGVKWLLKFGGSGGGSLILKFGKFGFKAPGAPGIL(SEQ ID NO: 151); AWQGVKGFKLAPKKGKPLGKPTKKKAHVRKL(SEQ ID NO: 152); FGAPSKHVKKILEWQKGGSGGGSWKILKFGGKPGVGF(SEQ ID NO: 153); EWKFGFGAGKSLKLGVVHVFGGSGGGSKILILKPVGAP(SEQ ID NO: 154); GFKQAPGVILAPRGFKWLIKLGKAKQKVKPFK(SEQ ID NO: 155);60 / 179B0662.70120WO00GVKPLKKCKPAILVAPKKGKQEWLKW(SEQ ID NO: 156); GVGKKVAWWQAQAKAPAWKGFGKWKKAKAAP(SEQ ID NO: 157); GIWKLGKAPKILGGSGGGSKWGVKKGALAPI(SEQ ID NO: 158); TGVKIWKVAPKKPGKFGGSGGGSLKKAPGKPAKI(SEQ ID NO: 159); AKAPKVHWKYYGFVHKFSKGAITWLWFLAKAW(SEQ ID NO: 160); IKARLKKKKCFAILLVAKKKKKAIG(SEQ ID NO: 161); WSGKAGSLLAKKKKPFTALLAKKGAKLKKKKG(SEQ ID NO: 162); RVKGKALWLGGSGGGSKWILGAKVGSLR(SEQ ID NO: 163); and KLILLILAKARGKVGGSGGGSWKAYLLKKKAKLAVNWKEWKKLGKAWKA GGKF (SEQ ID NO: 164).

[0252] In some embodiments, the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: KKGIGKWGKKHKKGIGGIKVVWGKVVGIWGIL(SEQ ID NO: 165); WGKLAFGKIKGIYKWGKKGWGKLAGIIGIGIKGI(SEQ ID NO: 166); GKKKLGKGIKGIIGKWGIIGIIGIKGIGIW(SEQ ID NO: 167); KKGIRRIGIGIGKGIGIIGKIFLSKLGWWGIW(SEQ ID NO: 168); WGIKLWGKVKGIKAIGYKAFGKIANKVINKYKGIG(SEQ ID NO: 169); KKGIGKIGKAFGKWGIKAIGIGKGALAILKVVIG(SEQ ID NO: 170); KKGIKAFIGIGKKWGGSGGGSKIWGIIGWGIAFGIAFN(SEQ ID NO: 171); WGKKSKKVWGIWKVLSGWGIKVWGK(SEQ ID NO: 172); PKMKGIGIKVAKGIKKLGKKLKNGIGKVKVY(SEQ ID NO: 173); KFWIGIGAGIIKVVKIGGSGGGSKNWGKWGKGWGI(SEQ ID NO: 174); RKWWGKLHKAIGGSGGGSKKKLSKVVKVVMAGI(SEQ ID NO: 175); KVVRKWGGIGGSGGGSKKWGHHKAINVLAFIKAF(SEQ ID NO: 176); RKIGGIKKWGGSGGGSVVKVVKGIWGAVVAIWGI(SEQ ID NO: 177); WGKKVKIKVAGIGKVWKKAIGNQAKVIGLG(SEQ ID NO: 178); KMHGIGAFGKWGKGIIGIIKKIGIWGIIGWG(SEQ ID NO: 179); and WKVWRKIGKGGSGGGSYKVKLGIIIGIGIG (SEQ ID NO: 180).

[0253] In some embodiments, the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: RILSVNRIKGGSGGGSVLLKRISVLSLKGWVNW(SEQ ID NO: 181); RIKKITGLSKKVLIRISVLGKRIATGWRTGSRIRI(SEQ ID NO: 182); RIWKKVLLATGGSGGGSVNILSRIFKRISVL(SEQ ID NO: 183); RIKKFLKVNKLFSVNWAKTGQLGVLRVNRIGKLSV(SEQ ID NO: 184);61 / 179B0662.70120WO00WKRILSKVNTGGSGGGSKNKRIIRRIKRIIRI(SEQ ID NO: 185); KRIRRIAKVGGSGGGSVNKRILSVLVNW(SEQ ID NO: 186); RRITGWARIGGSGGGSVNWKRIWVLSVL(SEQ ID NO: 187); WSVLRIRITGVLSVLGGSGGGSVRIIRIWRI(SEQ ID NO: 188); RIRIWKRISVLGGSGGGSVRIISVLWKKRLRIRRIL(SEQ ID NO: 189); SVLRRISKRIGGSGGGSVLSVNRIMKRIVNW(SEQ ID NO: 190); TGWGAIKGWKGTLTGWLSVFRRKLRITGKKVN(SEQ ID NO: 191); SVLRIHRIRRVNWARIRVLSVRSV(SEQ ID NO: 192); YGLSIKRISVLSKKLSVNKLG(SEQ ID NO: 193); GWKVLRILLSQVNKLSVLLSKRTGNRIGRI(SEQ ID NO: 194); WMTGWSVLGGGSGGGSVLSKKVLKKVLK(SEQ ID NO: 195); and GLGKVSVLSVRIFVRIGLTGWKRIG (SEQ ID NO: 196).

[0254] In some embodiments, the endosomal escape peptide comprises a multimer. In some embodiments, the endosomal escape peptide comprises a dimer. In some embodiments, the dimer comprises two iterations of an endosomal escape peptide sequence provided herein. In some embodiments, the endosomal escape peptide comprises a trimer. In some embodiments, the trimer comprises three iterations of an endosomal escape peptide sequence provided herein.

[0255] In some embodiments, the endosomal escape peptide comprises a sequence of: [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 43).

[0256] In some embodiments, the endosomal escape peptide comprises a sequence of: [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 44).

[0257] In some embodiments, the endosomal escape peptide is one or more of S10, Penetratin, Tat, R9, Transportan-10, L17E M-Lycotoxin, or Aurein1.2. In some embodiments, the endosomal escape peptide is S10, Penetratin, Tat, R9, Transportan-10, L17E M-Lycotoxin, or Aurein1.2. In some embodiments, the endosomal escape peptide is S10. In some embodiments, the endosomal escape peptide is Penetratin. In some embodiments, the endosomal escape peptide is Tat. In some embodiments, the endosomal escape peptide is R9. In some embodiments, the endosomal escape peptide is Transportan-10. In some embodiments, the endosomal escape peptide is L17E M-Lycotoxin. In some embodiments, the endosomal escape peptide is Aurein1.2.62 / 179B0662.70120WO00

[0258] In some embodiments, the endosomal escape peptide is a peptide disclosed in Machida, S. et al. Design of a Novel Membrane-Destabilizing Peptide Selectively Acting on Acidic Liposomes. Biosci. Biotechnol. Biochem. 2000, 64, 985-994, in Krishnamurthy, S. et al. Engineered amphiphilic peptides enable delivery of proteins and CRISPR-associated nucleases. Nat. Commun. 2019, 10, 4906, or in Cerovsky, V. et al. Lasioglossins: Three Novel Antimicrobial Peptides from the Venom of the Eusocial Bee Lasioglossum laticeps (Hymenoptera: Halictidae). Chem. Bio. Chem. 2009, 10, 2089-2099, the contents of which are incorporated herein by reference in their entireties.

[0259] In some embodiments, the compound has a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 45); WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 46); TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 47); MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 48); LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 49); LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 50); RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 51); KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 52);63 / 179B0662.70120WO00KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 53); KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 54); KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 55); KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 56); KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 57); KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 58); ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 59); KWKLARAFARAIKKLGGSGGGSYARALRRQARTG– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 60); RQIKIWFQNRRMKWKK–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 61); CKRKKRRQRRRG–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 62); RRRRRRRRR–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 63); AGYLLGKINLKALAALAKKIL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 64); IWLTALKFLGKHAAKHEAKQQLSKL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 65); and64 / 179B0662.70120WO00GLFDIIKKIAESF–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 66).

[0260] In some embodiments, the compound has a sequence selected from the group consisting of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 67); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWKK (SEQ ID NO: 68); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK (SEQ ID NO: 69); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 70); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 71); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 72); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKLK (SEQ ID NO: 73); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 74); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–65 / 179B0662.70120WO00KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 75); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 76); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 77); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 78); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 79); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 80); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 81); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 82); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 83); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–KWKLARAFARAIKKLGGSGGGSYARALRRQARTG (SEQ ID NO: 84); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–RQIKIWFQNRRMKWKK (SEQ ID NO: 85); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG–66 / 179B0662.70120WO00[(IPGHG)2(VPGYG)(IPGHG)2]15–CKRKKRRQRRRG (SEQ ID NO: 86); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–RRRRRRRRR (SEQ ID NO: 87); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–AGYLLGKINLKALAALAKKIL (SEQ ID NO: 88); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–IWLTALKFLGKHAAKHEAKQQLSKL (SEQ ID NO: 89); and [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GLFDIIKKIAESF (SEQ ID NO: 90).

[0261] In some embodiments, the compound has a sequence selected from the group consisting of: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 – KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 221); [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 – KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 222); and [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 – KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 223).

[0262] In some embodiments, the compound has a sequence selected from the group consisting of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CAP]5– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 197); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CEAAAK]5– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 224); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–C5G3– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 225); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH67 / 179B0662.70120WO00GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 238); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 239); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 240); and [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 241).68 / 179B0662.70120WO00

[0263] In some embodiments, the compound has a sequence: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[(IPG(C / H)G)2(VPGYG)(IPG (C / H)G)2]15–KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 226).

[0264] In some embodiments, the compound further comprises a protease cleavable linker at the C-terminus.

[0265] In some embodiments, the compound further comprises a protease cleavable linker at the N-terminus. In some embodiments, the protease cleavable linker sequence is selected from the group consisting of: (GGS)9(SEQ ID NO: 91); GGSGGSEVRFGGSGGS (SEQ ID NO: 92); GGSGGSGGVGFLGGGS (SEQ ID NO: 93); GGSGGSGTQFFRLGGS (SEQ ID NO: 94); KKESTFEERSYWQSQV (SEQ ID NO: 95); GGSGGSRVRRGGSGGS (SEQ ID NO: 96); and NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 97).

[0266] In some embodiments, the protease cleavable linker is (GGS)9 (SEQ ID NO: 91). In some embodiments, the protease cleavable linker is GGSGGSEVRFGGSGGS (SEQ ID NO: 92). In some embodiments, the protease cleavable linker is GGSGGSGGVGFLGGGS (SEQ ID NO: 93). In some embodiments, the protease cleavable linker is GGSGGSGTQFFRLGGS (SEQ ID NO: 94). In some embodiments, the protease cleavable linker is KKESTFEERSYWQSQV (SEQ ID NO: 95). In some embodiments, the protease cleavable linker is GGSGGSRVRRGGSGGS (SEQ ID NO: 96). In some embodiments, the protease cleavable linker is NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 97).

[0267] In some embodiments, the compound comprises a protease cleavable linker conjugated to the elastin-like polypeptide. In some embodiments, compound comprises a protease cleavable linker conjugated to the elastin-like polypeptide and comprises a sequence selected from the group consisting of: (GGS)9–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 98); GGSGGSEVRFGGSGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 99);69 / 179B0662.70120WO00GGSGGSGGVGFLGGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 100); GGSGGSGTQFFRLGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 101); KKESTFEERSYWQSQV–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 102); NNTHDLVGDVRLAGVQSVASSRRHKRFAGV– [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 103);and GGSGGSRVRRGGSGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 104).

[0268] In some embodiments, compound comprises a protease cleavable linker conjugated to the elastin-like polypeptide and comprises a sequence selected from the group consisting of: [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–(GGS)9 (SEQ ID NO: 105); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSEVRFGGSGGS (SEQ ID NO: 106); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSGGVGFLGGGS (SEQ ID NO: 107); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSGTQFFRLGGS (SEQ ID NO: 108); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–KKESTFEERSYWQSQV (SEQ ID NO: 109); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSRVRRGGSGGS (SEQ ID NO: 110); and [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 111).

[0269] In another aspect, provided herein is an elastin-like polypeptide comprising a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–X*– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 198), wherein: each instance of X3is independently G, S, Q, V, or A; each instance of X4is independently E or G; and70 / 179B0662.70120WO00X*is -CCCCGGG- (SEQ ID NO: 19), -[CAP]5- (SEQ ID NO: 125), [CEAAAK]5(SEQ ID NO: 126), or -C5G3- (SEQ ID NO: 127).

[0270] In some embodiments, X* is -CCCCGGG- (SEQ ID NO: 19) or -C5G3- (SEQ ID NO: 127). In some embodiments, X* is -[CAP]5- (SEQ ID NO: 125), [CEAAAK]5(SEQ ID NO: 126), or -C5G3- (SEQ ID NO: 127). In some embodiments, X* is -CCCCGGG- (SEQ ID NO: 19). In some embodiments, X* is -[CAP]5- (SEQ ID NO: 125). In some embodiments, X* is [CEAAAK]5(SEQ ID NO: 126). In some embodiments, X* is -C5G3- (SEQ ID NO: 127).

[0271] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 227), wherein: X3is G, S, Q, V, or A; and X4is E or G.

[0272] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 112) wherein: X3is V or A; and X4is E or G.

[0273] In some embodiments, each instance of X3is V, each instance of X3is A, each instance of X3is G, each instance of X3is S, or each instance of X3is Q. In some embodiments, each instance of X3is V or each instance of X3is A. In some embodiments, each instance of X3is G, each instance of X3is S, or each instance of X3is Q. In some embodiments, X3is V. In some embodiments, X3is A. In some embodiments, each instance of X3is the same. In some embodiments, each instance of X3is A. In some embodiments, each instance of X3is V.

[0274] In some embodiments, each instance of X4is E or each instance of X4is G. In some embodiments, X4is E. In some embodiments, X4is G. In some embodiments, each instance of X4is the same. In some embodiments, each instance of X4is E. In some embodiments, each instance of X4is G.

[0275] In some embodiments, X3is V, and X4is E. In some embodiments, X3is V, and X4is G. In some embodiments, X3is A, and X4is G. In some embodiments, X3is A, and X4is E.

[0276] In some embodiments, the elastin-like polypeptide comprises a sequence of:71 / 179B0662.70120WO00[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 115).

[0277] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 113).

[0278] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 131); [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 228); or [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 229).

[0279] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CAP]5– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 199); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CEAAAK]5– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 200); or [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–C5G3– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 201).

[0280] In some embodiments, the elastin-like polypeptide comprises a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10– [(IPGZ4G)2(VPGYG)(IPGZ4G)2]15 (SEQ ID NO: 203), wherein: each instance of X3is independently G, S, Q, V, or A; each instance of X4is independently E or G; and each instance of Z3is independently C or H.

[0281] In some embodiments, three instances of Z3are C and the remaining instances of Z3are H. In some embodiments, five instances of Z3are C and the remaining instances of Z3are H. In some embodiments, seven instances of Z3are C and the remaining instances of Z3are H. In some embodiments, thirteen instances of Z3are C and the remaining instances of Z3are H.

[0282] In another aspect, provided herein is an endosomal escape peptide comprising a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 2);72 / 179B0662.70120WO00WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K (SEQ ID NO: 3); TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK (SEQ ID NO: 4); MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 5); LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 6); LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 7); RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K (SEQ ID NO: 8); KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 9); KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 10); KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 11); KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 12); KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 13); KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 14); KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 15); and ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 16).

[0283] In another aspect, provided herein is an endosomal escape peptide comprising a sequence selected from the group consisting of: RMKFNVWAWICVVKKKKFFPHWKIRYIIGRAWLL(SEQ ID NO: 133); DVHRRIWIILLPGHRIKFNWKQWKWRFEFEFKAVRK(SEQ ID NO: 134); KLVRYISYLKLRGGVKILLAKIKPFYEVKLKLKRLSLII(SEQ ID NO: 135); YFLARARARAILIKFFGKLKPFKAKKLAKQLKWVART(SEQ ID NO: 136); ANHKFGVYFGGSGGGSGVKWQVKFGVNHANHVKF(SEQ ID NO: 137); ANHGHKFGVKLNHGGSGGGSYFKFGKAPGVKW(SEQ ID NO: 138); GVGVREWKFGGSGGGSYYFGVKGHKKGVGV(SEQ ID NO: 139); GVKFGVGVKKYFKYFASGVKGVKYF(SEQ ID NO: 140); GVGVKYFKVAVKNHKPKWFSYKAAWKKFG(SEQ ID NO: 141); GNHKAYKWVRAPRTNHKHKKAVKWKHKPG(SEQ ID NO: 142); LKFWGASAPAPHKFAWKFGVGARASGHKLL(SEQ ID NO: 143);73 / 179B0662.70120WO00VKWKWQWGVKGAPGYFYLGFGVKTAWAPAWAW(SEQ ID NO: 144); FLKFGGSGGGSHKVKFGVVGNHKKAPVHKW(SEQ ID NO: 145); GVGASLGKQWKFGGSGGGSNHVKWFGVKPSKGHKGAWKWS(SEQ ID NO: 146); KPIQWGVGGSGGGSYLAPRTKKHVKFGVKRTHKFAP(SEQ ID NO: 147); WKWLKAPKPILKILGKPVKHKWKPAWKGKWQ(SEQ ID NO: 148); KSILKWGAKPKKALILASLGVKKLLAPSGAPW(SEQ ID NO: 149); GVKPAWRASLGGKHVKRLSGFHKPRTKAPSL(SEQ ID NO: 150); KGVKWLLKFGGSGGGSLILKFGKFGFKAPGAPGIL(SEQ ID NO: 151); AWQGVKGFKLAPKKGKPLGKPTKKKAHVRKL(SEQ ID NO: 152); FGAPSKHVKKILEWQKGGSGGGSWKILKFGGKPGVGF(SEQ ID NO: 153); EWKFGFGAGKSLKLGVVHVFGGSGGGSKILILKPVGAP(SEQ ID NO: 154); GFKQAPGVILAPRGFKWLIKLGKAKQKVKPFK(SEQ ID NO: 155); GVKPLKKCKPAILVAPKKGKQEWLKW(SEQ ID NO: 156); GVGKKVAWWQAQAKAPAWKGFGKWKKAKAAP(SEQ ID NO: 157); GIWKLGKAPKILGGSGGGSKWGVKKGALAPI(SEQ ID NO: 158); TGVKIWKVAPKKPGKFGGSGGGSLKKAPGKPAKI(SEQ ID NO: 159); AKAPKVHWKYYGFVHKFSKGAITWLWFLAKAW(SEQ ID NO: 160); IKARLKKKKCFAILLVAKKKKKAIG(SEQ ID NO: 161); WSGKAGSLLAKKKKPFTALLAKKGAKLKKKKG(SEQ ID NO: 162); RVKGKALWLGGSGGGSKWILGAKVGSLR(SEQ ID NO: 163); and KLILLILAKARGKVGGSGGGSWKAYLLKKKAKLAVNWKEWKKLGKAWKA GGKF (SEQ ID NO: 164).

[0284] In another aspect, provided herein is an endosomal escape peptide comprising a sequence selected from the group consisting of: KKGIGKWGKKHKKGIGGIKVVWGKVVGIWGIL(SEQ ID NO: 165); WGKLAFGKIKGIYKWGKKGWGKLAGIIGIGIKGI(SEQ ID NO: 166); GKKKLGKGIKGIIGKWGIIGIIGIKGIGIW(SEQ ID NO: 167); KKGIRRIGIGIGKGIGIIGKIFLSKLGWWGIW(SEQ ID NO: 168); WGIKLWGKVKGIKAIGYKAFGKIANKVINKYKGIG(SEQ ID NO: 169); KKGIGKIGKAFGKWGIKAIGIGKGALAILKVVIG(SEQ ID NO: 170); KKGIKAFIGIGKKWGGSGGGSKIWGIIGWGIAFGIAFN(SEQ ID NO: 171); WGKKSKKVWGIWKVLSGWGIKVWGK(SEQ ID NO: 172); PKMKGIGIKVAKGIKKLGKKLKNGIGKVKVY(SEQ ID NO: 173);74 / 179B0662.70120WO00KFWIGIGAGIIKVVKIGGSGGGSKNWGKWGKGWGI(SEQ ID NO: 174); RKWWGKLHKAIGGSGGGSKKKLSKVVKVVMAGI(SEQ ID NO: 175); KVVRKWGGIGGSGGGSKKWGHHKAINVLAFIKAF(SEQ ID NO: 176); RKIGGIKKWGGSGGGSVVKVVKGIWGAVVAIWGI(SEQ ID NO: 177); WGKKVKIKVAGIGKVWKKAIGNQAKVIGLG(SEQ ID NO: 178); KMHGIGAFGKWGKGIIGIIKKIGIWGIIGWG(SEQ ID NO: 179); and WKVWRKIGKGGSGGGSYKVKLGIIIGIGIG (SEQ ID NO: 180).

[0285] In another aspect, provided herein is an endosomal escape peptide comprising a sequence selected from the group consisting of: RILSVNRIKGGSGGGSVLLKRISVLSLKGWVNW(SEQ ID NO: 181); RIKKITGLSKKVLIRISVLGKRIATGWRTGSRIRI(SEQ ID NO: 182); RIWKKVLLATGGSGGGSVNILSRIFKRISVL(SEQ ID NO: 183); RIKKFLKVNKLFSVNWAKTGQLGVLRVNRIGKLSV(SEQ ID NO: 184); WKRILSKVNTGGSGGGSKNKRIIRRIKRIIRI(SEQ ID NO: 185); KRIRRIAKVGGSGGGSVNKRILSVLVNW(SEQ ID NO: 186); RRITGWARIGGSGGGSVNWKRIWVLSVL(SEQ ID NO: 187); WSVLRIRITGVLSVLGGSGGGSVRIIRIWRI(SEQ ID NO: 188); RIRIWKRISVLGGSGGGSVRIISVLWKKRLRIRRIL(SEQ ID NO: 189); SVLRRISKRIGGSGGGSVLSVNRIMKRIVNW(SEQ ID NO: 190); TGWGAIKGWKGTLTGWLSVFRRKLRITGKKVN(SEQ ID NO: 191); SVLRIHRIRRVNWARIRVLSVRSV(SEQ ID NO: 192); YGLSIKRISVLSKKLSVNKLG(SEQ ID NO: 193); GWKVLRILLSQVNKLSVLLSKRTGNRIGRI(SEQ ID NO: 194); WMTGWSVLGGGSGGGSVLSKKVLKKVLK(SEQ ID NO: 195); and GLGKVSVLSVRIFVRIGLTGWKRIG (SEQ ID NO: 196).

[0286] In some embodiments, the term “compound provided herein” refers to a compound comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides; to an elastin-like polypeptide; or to an endosomal escape peptide provided herein. Pharmaceutical Compositions, Kits, and Administration

[0287] In another aspect, provided herein is a composition comprising a compound provided herein and an agent.75 / 179B0662.70120WO00

[0288] In some embodiments, the composition is in the form of a particle. In some embodiments, the particle is a nanoparticle. In certain embodiments, the particle is a microparticle. In some embodiments, the composition is in the form of a micelle.

[0289] In certain embodiments, the average diameter of the particle is at least about 10 nm, at least about 50 nm, at least about 100 nm, at least about 500 nm, at least about 1 μm, at least about 5 μm, at least about 10 μm, at least about 50 μm, at least about 100 μm, at least about 500 μm, or at least about 1 mm. In certain embodiments, the average diameter of the particle is less than about 1 mm, less than about 500 μm, less than about 100 μm, less than about 50 μm less than about 10 μm, less than about 5 μm, less than about 1 μm, less than about 500 nm, less than about 100 nm, less than about 50 nm, or less than about 10 nm.

[0290] In some embodiments, the average diameter of the particle is about 15 nm to about 800 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 700 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 600 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 500 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 400 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 300 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 250 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 200 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 150 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 100 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 90 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 80 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 70 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 60 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 50 nm. In some embodiments, the average diameter of the particle is about 15 nm to about 25 nm.

[0291] In some embodiments, the average diameter of the particle is about 25 nm to about 300 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 250 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 200 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 150 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 100 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 90 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 8076 / 179B0662.70120WO00nm. In some embodiments, the average diameter of the particle is about 25 nm to about 70 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 60 nm. In some embodiments, the average diameter of the particle is about 25 nm to about 50 nm.

[0292] In some embodiments, the average diameter of the particle is about 50 nm to about 300 nm. In some embodiments, the average diameter of the particle is about 50 nm to about 250 nm. In some embodiments, the average diameter of the particle is about 50 nm to about 200 nm. In some embodiments, the average diameter of the particle is about 50 nm to about 150 nm. In some embodiments, the average diameter of the particle is about 50 nm to about 100 nm.

[0293] In some embodiments, the average dimeter of the particle is about 20 nm to about 200 nm. In some embodiments, the average dimeter of the particle is about 30 nm to about 200 nm. In some embodiments, the average dimeter of the particle is about 40 nm to about 200 nm. In some embodiments, the average dimeter of the particle is about 60 nm to about 200 nm. In some embodiments, the average dimeter of the particle is about 75 nm to about 200 nm.

[0294] In some embodiments, the average diameter of the particle is about 100 nm to about 200 nm. In some embodiments, the average diameter of the particle is about 150 nm to about 250 nm. In some embodiments, the average diameter of the particle is about 250 nm to about 350 nm. In some embodiments, the average diameter of the particle is about 350 nm to about 450 nm. In some embodiments, the average diameter of the particle is about 450 nm to about 550 nm. In some embodiments, the average diameter of the particle is about 550 nm to about 650 nm.

[0295] In some embodiments, the polydispersity index is about 0.05 to about 0.8. In some embodiments, the polydispersity index is about 0.1 to about 0.6. In some embodiments, the polydispersity index is about 0.1 to about 0.5. In some embodiments, the polydispersity index is about 0.2 to about 0.4. In some embodiments, the polydispersity index is about 0.1 to about 0.3. In some embodiments, the polydispersity index is about 0.3 to about 0.5. In some embodiments, the polydispersity index is about 0.4 to about 0.6. In some embodiments, the polydispersity index is determined by light scattering.

[0296] In some embodiments, the particle or micelle encapsulates the agent.

[0297] In some embodiments, the compound and the agent are not covalently attached. In some embodiments, the compound and the agent are electrostatically complexed. In some embodiments, the compound and the agent are electrostatically complexed at the N-terminus77 / 179B0662.70120WO00of the compound. In some embodiments, the compound and the agent are electrostatically complexed at the C-terminus of the compound.

[0298] In some embodiments, the compound and the agent are covalently attached. In some embodiments, the compound and the agent are covalently attached via the N-terminus of the compound. In some embodiments, the compound and the agent are covalently attached via the C-terminus of the compound.

[0299] In some embodiments, the compound and the agent are covalently attached via a protease cleavable linker. In some embodiments, the protease cleavable linker is selected from the group consisting of: (GGS)9(SEQ ID NO: 91); GGSGGSEVRFGGSGGS (SEQ ID NO: 92); GGSGGSGGVGFLGGGS (SEQ ID NO: 93); GGSGGSGTQFFRLGGS (SEQ ID NO: 94); KKESTFEERSYWQSQV (SEQ ID NO: 95); GGSGGSRVRRGGSGGS (SEQ ID NO: 96); and NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 97).

[0300] In some embodiments, the protease cleavable linker is (GGS)9(SEQ ID NO: 91). In some embodiments, the protease cleavable linker is GGSGGSEVRFGGSGGS (SEQ ID NO: 92). In some embodiments, the protease cleavable linker is GGSGGSGGVGFLGGGS (SEQ ID NO: 93). In some embodiments, the protease cleavable linker is GGSGGSGTQFFRLGGS (SEQ ID NO: 94). In some embodiments, the protease cleavable linker is KKESTFEERSYWQSQV (SEQ ID NO: 95). In some embodiments, the protease cleavable linker is GGSGGSRVRRGGSGGS (SEQ ID NO: 96). In some embodiments, the protease cleavable linker is NNTHDLVGDVRLAGVQSVASSRRHKRFAGV(SEQ ID NO: 97).

[0301] In some embodiments, the agent is an organic molecule, protein, peptide, polypeptide, polynucleotide, an isotopically labeled chemical compound, vaccine, antibody, or an immunological agent. In some embodiments, the agent is an organic molecule, protein, polypeptide, polynucleotide, an isotopically labeled chemical compound, vaccine, or an immunological agent. In some embodiments, the agent is an organic molecule or an isotopically labeled chemical compound. In some embodiments, the agent is a protein, polypeptide, or polynucleotide. In some embodiments, the agent is a polynucleotide or polypeptide. In some embodiments, the agent is a peptide or polypeptide. In some embodiments, the agent is a vaccine or an immunological agent. In some embodiments, the78 / 179B0662.70120WO00agent is an organic molecule. In some embodiments, the agent is a protein. In some embodiments, the agent is a peptide. In some embodiments, the agent is a polypeptide. In some embodiments, the agent is a polynucleotide. In some embodiments, the agent is an isotopically labeled chemical compound. In some embodiments, the agent is a vaccine. In some embodiments, the agent is an antibody. In some embodiments, the agent is an immunological agent.

[0302] In some embodiments, the agent is an RNA. In some embodiments, the RNA is messenger RNA (mRNA), pre-messenger RNA (pre-mRNA), ribosomal RNA (rRNA), transfer RNA (tRNA), small nuclear RNA (snRNA), small nucleolar RNA (snoRNA), Piwi- interacting RNA (piRNA), microRNA (miRNA), small interfering RNA (siRNA), antisense RNA (asRNA or aRNA), transfer-messenger RNA (tmRNA), single-stranded RNA (ssRNA), double-stranded RNA (dsRNA), precursor messenger RNA (pre-mRNA), small hairpin RNA or short hairpin RNA (shRNA), guide RNA (gRNA), heterogeneous nuclear RNA (hnRNA), coding RNA, non-coding RNA (ncRNA), long non-coding RNA (long ncRNA or lncRNA), satellite RNA, viral satellite RNA, signal recognition particle RNA, small cytoplasmic RNA, polyinosinic acid, ribozyme, flexizyme, spliced leader RNA, viral RNA, or viral satellite RNA. In some embodiments, the RNA is messenger RNA (mRNA). In some embodiments, the RNA is pre-messenger RNA (pre-mRNA). In some embodiments, the RNA is ribosomal RNA (rRNA). In some embodiments, the RNA is transfer RNA (tRNA). In some embodiments, the RNA is small nuclear RNA (snRNA). In some embodiments, the RNA is small nucleolar RNA (snoRNA). In some embodiments, the RNA is Piwi-interacting RNA (piRNA). In some embodiments, the RNA is microRNA (miRNA). In some embodiments, the RNA is small interfering RNA (siRNA). In some embodiments, the RNA is antisense RNA (asRNA or aRNA). In some embodiments, the RNA is transfer-messenger RNA (tmRNA). In some embodiments, the RNA is single-stranded RNA (ssRNA). In some embodiments, the RNA is double-stranded RNA (dsRNA). In some embodiments, the RNA is precursor messenger RNA (pre-mRNA). In some embodiments, the RNA is small hairpin RNA or short hairpin RNA (shRNA). In some embodiments, the RNA is guide RNA (gRNA). In some embodiments, the RNA is heterogeneous nuclear RNA (hnRNA). In some embodiments, the RNA is coding RNA. In some embodiments, the RNA is non-coding RNA (ncRNA). In some embodiments, the RNA is long non-coding RNA (long ncRNA or lncRNA). In some embodiments, the RNA is satellite RNA. In some embodiments, the RNA is viral satellite RNA. In some embodiments, the RNA is signal recognition particle RNA. In some embodiments, the RNA is small cytoplasmic RNA. In some embodiments, the RNA is79 / 179B0662.70120WO00polyinosinic acid. In some embodiments, the RNA is ribozyme. In some embodiments, the RNA is flexizyme. In some embodiments, the RNA is spliced leader RNA. In some embodiments, the RNA is viral RNA. In some embodiments, the RNA is viral satellite RNA.

[0303] In some embodiments, the polynucleotide is a DNA. In some embodiments, the DNA is a plasmid DNA (pDNA).

[0304] In some embodiments, the peptide is ovalbumin-derived MHC class I epitope or ovalbumin-derived MHC class II epitope. In some embodiments, the peptide is ovalbumin- derived MHC class I epitope. In some embodiments, the peptide is ovalbumin-derived MHC class II epitope.

[0305] In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the monoclonal antibody is anti-CD117. In some embodiments, the monoclonal antibody is anti-CD5.

[0306] In some embodiments, the molar ratio of the agent to the compound is from about 1:0.05 to about 1:5000. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:5000. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:3000. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:1500. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:750. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:300. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:150. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:60. In some embodiments, the molar ratio of the agent to the compound is from about 1:1 to about 1:30. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:5000. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:3000. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:1500. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:750. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:300. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:150. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:60. In some embodiments, the molar ratio of the agent to the compound is from about 1:10 to about 1:30.

[0307] In some embodiments, the molar ratio of the agent to the compound is about 1:0.05 to about 1:30. In some embodiments, the molar ratio of the agent to the compound is about80 / 179B0662.70120WO001:30 to about 1:60. In some embodiments, the molar ratio of the agent to the compound is about 1:60 to about 1:150. In some embodiments, the molar ratio of the agent to the compound is about 1:150 to about 1:300. In some embodiments, the molar ratio of the agent to the compound is about 1:300 to about 1:750. In some embodiments, the molar ratio of the agent to the compound is about 1:750 to about 1:1500. In some embodiments, the molar ratio of the agent to the compound is about 1:1500 to about 1:3000.

[0308] Pharmaceutical compositions described herein can be prepared by any method known in the art of pharmaceutics. In general, such preparatory methods include bringing a compound, agent, particle, or composition described herein (e.g., the “active ingredient”) into association with a carrier or excipient, and / or one or more other accessory ingredients, and then, if necessary and / or desirable, shaping, and / or packaging the product into a desired single- or multi-dose unit.

[0309] Pharmaceutical compositions can be prepared, packaged, and / or sold in bulk, as a single unit dose, and / or as a plurality of single unit doses. A “unit dose” is a discrete amount of the pharmaceutical composition comprising a predetermined amount of the active ingredient. The amount of the active ingredient is generally equal to the dosage of the active ingredient which would be administered to a subject and / or a convenient fraction of such a dosage, such as one-half or one-third of such a dosage.

[0310] Relative amounts of the active ingredient, the pharmaceutically acceptable excipient, and / or any additional ingredients in a pharmaceutical composition described herein will vary, depending upon the identity, size, and / or condition of the subject treated and further depending upon the route by which the composition is to be administered. The composition may comprise between 0.1% and 100% (w / w) active ingredient.

[0311] Pharmaceutically acceptable excipients used in the manufacture of provided pharmaceutical compositions include inert diluents, dispersing and / or granulating agents, surface active agents and / or emulsifiers, disintegrating agents, binding agents, preservatives, buffering agents, lubricating agents, and / or oils. Excipients such as cocoa butter and suppository waxes, coloring agents, coating agents, sweetening, flavoring, and perfuming agents may also be present in the composition.

[0312] Exemplary diluents include calcium carbonate, sodium carbonate, calcium phosphate, dicalcium phosphate, calcium sulfate, calcium hydrogen phosphate, sodium phosphate lactose, sucrose, cellulose, microcrystalline cellulose, kaolin, mannitol, sorbitol, inositol, sodium chloride, dry starch, cornstarch, powdered sugar, and mixtures thereof.81 / 179B0662.70120WO00

[0313] Exemplary granulating and / or dispersing agents include potato starch, corn starch, tapioca starch, sodium starch glycolate, clays, alginic acid, guar gum, citrus pulp, agar, bentonite, cellulose, and wood products, natural sponge, cation-exchange resins, calcium carbonate, silicates, sodium carbonate, cross-linked poly(vinyl-pyrrolidone) (crospovidone), sodium carboxymethyl starch (sodium starch glycolate), carboxymethyl cellulose, cross- linked sodium carboxymethyl cellulose (croscarmellose), methylcellulose, pregelatinized starch (starch 1500), microcrystalline starch, water insoluble starch, calcium carboxymethyl cellulose, magnesium aluminum silicate (Veegum), sodium lauryl sulfate, quaternary ammonium compounds, and mixtures thereof.

[0314] Exemplary surface active agents and / or emulsifiers include natural emulsifiers (e.g., acacia, agar, alginic acid, sodium alginate, tragacanth, chondrux, cholesterol, xanthan, pectin, gelatin, egg yolk, casein, wool fat, cholesterol, wax, and lecithin), colloidal clays (e.g., bentonite (aluminum silicate) and Veegum (magnesium aluminum silicate)), long chain amino acid derivatives, high molecular weight alcohols (e.g., stearyl alcohol, cetyl alcohol, oleyl alcohol, triacetin monostearate, ethylene glycol distearate, glyceryl monostearate, and propylene glycol monostearate, polyvinyl alcohol), carbomers (e.g., carboxy polymethylene, polyacrylic acid, acrylic acid polymer, and carboxyvinyl polymer), carrageenan, cellulosic derivatives (e.g., carboxymethylcellulose sodium, powdered cellulose, hydroxymethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, methylcellulose), sorbitan fatty acid esters (e.g., polyoxyethylene sorbitan monolaurate (Tween®20), polyoxyethylene sorbitan (Tween®60), polyoxyethylene sorbitan monooleate (Tween®80), sorbitan monopalmitate (Span®40), sorbitan monostearate (Span®60), sorbitan tristearate (Span®65), glyceryl monooleate, sorbitan monooleate (Span®80), polyoxyethylene esters (e.g., polyoxyethylene monostearate (Myrj®45), polyoxyethylene hydrogenated castor oil, polyethoxylated castor oil, polyoxymethylene stearate, and Solutol®), sucrose fatty acid esters, polyethylene glycol fatty acid esters (e.g., Cremophor®), polyoxyethylene ethers, (e.g., polyoxyethylene lauryl ether (Brij®30)), poly(vinyl-pyrrolidone), diethylene glycol monolaurate, triethanolamine oleate, sodium oleate, potassium oleate, ethyl oleate, oleic acid, ethyl laurate, sodium lauryl sulfate, Pluronic®F-68, poloxamer P-188, cetrimonium bromide, cetylpyridinium chloride, benzalkonium chloride, docusate sodium, and / or mixtures thereof.

[0315] Exemplary binding agents include starch (e.g., cornstarch and starch paste), gelatin, sugars (e.g., sucrose, glucose, dextrose, dextrin, molasses, lactose, lactitol, mannitol, etc.), natural and synthetic gums (e.g., acacia, sodium alginate, extract of Irish moss, panwar gum, ghatti gum, mucilage of isapol husks, carboxymethylcellulose, methylcellulose,82 / 179B0662.70120WO00ethylcellulose, hydroxyethylcellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, microcrystalline cellulose, cellulose acetate, poly(vinyl-pyrrolidone), magnesium aluminum silicate (Veegum®), and larch arabogalactan), alginates, polyethylene oxide, polyethylene glycol, inorganic calcium salts, silicic acid, polymethacrylates, waxes, water, alcohol, and / or mixtures thereof.

[0316] Exemplary preservatives include antioxidants, chelating agents, antimicrobial preservatives, antifungal preservatives, antiprotozoan preservatives, alcohol preservatives, acidic preservatives, and other preservatives. In certain embodiments, the preservative is an antioxidant. In other embodiments, the preservative is a chelating agent.

[0317] Exemplary antioxidants include alpha tocopherol, ascorbic acid, ascorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene, monothioglycerol, potassium metabisulfite, propionic acid, propyl gallate, sodium ascorbate, sodium bisulfite, sodium metabisulfite, and sodium sulfite.

[0318] Exemplary chelating agents include ethylenediaminetetraacetic acid (EDTA) and salts and hydrates thereof (e.g., sodium edetate, disodium edetate, trisodium edetate, calcium disodium edetate, dipotassium edetate, and the like), citric acid and salts and hydrates thereof (e.g., citric acid monohydrate), fumaric acid and salts and hydrates thereof, malic acid and salts and hydrates thereof, phosphoric acid and salts and hydrates thereof, and tartaric acid and salts and hydrates thereof. Exemplary antimicrobial preservatives include benzalkonium chloride, benzethonium chloride, benzyl alcohol, bronopol, cetrimide, cetylpyridinium chloride, chlorhexidine, chlorobutanol, chlorocresol, chloroxylenol, cresol, ethyl alcohol, glycerin, hexetidine, imidurea, phenol, phenoxyethanol, phenylethyl alcohol, phenylmercuric nitrate, propylene glycol, and thimerosal.

[0319] Exemplary antifungal preservatives include butyl paraben, methyl paraben, ethyl paraben, propyl paraben, benzoic acid, hydroxybenzoic acid, potassium benzoate, potassium sorbate, sodium benzoate, sodium propionate, and sorbic acid.

[0320] Exemplary alcohol preservatives include ethanol, polyethylene glycol, phenol, phenolic compounds, bisphenol, chlorobutanol, hydroxybenzoate, and phenylethyl alcohol.

[0321] Exemplary acidic preservatives include vitamin A, vitamin C, vitamin E, beta- carotene, citric acid, acetic acid, dehydroacetic acid, ascorbic acid, sorbic acid, and phytic acid.

[0322] Other preservatives include tocopherol, tocopherol acetate, deteroxime mesylate, cetrimide, butylated hydroxyanisol (BHA), butylated hydroxytoluened (BHT), ethylenediamine, sodium lauryl sulfate (SLS), sodium lauryl ether sulfate (SLES), sodium83 / 179B0662.70120WO00bisulfite, sodium metabisulfite, potassium sulfite, potassium metabisulfite, Glydant®Plus, Phenonip®, methylparaben, Germall®115, Germaben®II, Neolone®, Kathon®, and Euxyl®.

[0323] Exemplary buffering agents include citrate buffer solutions, acetate buffer solutions, phosphate buffer solutions, ammonium chloride, calcium carbonate, calcium chloride, calcium citrate, calcium glubionate, calcium gluceptate, calcium gluconate, D-gluconic acid, calcium glycerophosphate, calcium lactate, propanoic acid, calcium levulinate, pentanoic acid, dibasic calcium phosphate, phosphoric acid, tribasic calcium phosphate, calcium hydroxide phosphate, potassium acetate, potassium chloride, potassium gluconate, potassium mixtures, dibasic potassium phosphate, monobasic potassium phosphate, potassium phosphate mixtures, sodium acetate, sodium bicarbonate, sodium chloride, sodium citrate, sodium lactate, dibasic sodium phosphate, monobasic sodium phosphate, sodium phosphate mixtures, tromethamine, magnesium hydroxide, aluminum hydroxide, alginic acid, pyrogen- free water, isotonic saline, Ringer’s solution, ethyl alcohol, and mixtures thereof.

[0324] Exemplary lubricating agents include magnesium stearate, calcium stearate, stearic acid, silica, talc, malt, glyceryl behanate, hydrogenated vegetable oils, polyethylene glycol, sodium benzoate, sodium acetate, sodium chloride, leucine, magnesium lauryl sulfate, sodium lauryl sulfate, and mixtures thereof.

[0325] Exemplary natural oils include almond, apricot kernel, avocado, babassu, bergamot, black current seed, borage, cade, camomile, canola, caraway, carnauba, castor, cinnamon, cocoa butter, coconut, cod liver, coffee, corn, cotton seed, emu, eucalyptus, evening primrose, fish, flaxseed, geraniol, gourd, grape seed, hazel nut, hyssop, isopropyl myristate, jojoba, kukui nut, lavandin, lavender, lemon, litsea cubeba, macademia nut, mallow, mango seed, meadowfoam seed, mink, nutmeg, olive, orange, orange roughy, palm, palm kernel, peach kernel, peanut, poppy seed, pumpkin seed, rapeseed, rice bran, rosemary, safflower, sandalwood, sasquana, savoury, sea buckthorn, sesame, shea butter, silicone, soybean, sunflower, tea tree, thistle, tsubaki, vetiver, walnut, and wheat germ oils. Exemplary synthetic oils include, but are not limited to, butyl stearate, caprylic triglyceride, capric triglyceride, cyclomethicone, diethyl sebacate, dimethicone 360, isopropyl myristate, mineral oil, octyldodecanol, oleyl alcohol, silicone oil, and mixtures thereof.

[0326] Liquid dosage forms for oral and parenteral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredients, the liquid dosage forms may comprise inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl84 / 179B0662.70120WO00benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (e.g., cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents. In certain embodiments for parenteral administration, the conjugates described herein are mixed with solubilizing agents such as Cremophor®, alcohols, oils, modified oils, glycols, polysorbates, cyclodextrins, polymers, and mixtures thereof.

[0327] Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions can be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation can be a sterile injectable solution, suspension, or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that can be employed are water, Ringer’s solution, U.S.P., and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or di-glycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.

[0328] The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

[0329] In order to prolong the effect of a drug, it is often desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This can be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution, which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form may be accomplished by dissolving or suspending the drug in an oil vehicle.

[0330] Compositions for rectal or vaginal administration are typically suppositories which can be prepared by mixing the conjugates described herein with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol, or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active ingredient.85 / 179B0662.70120WO00

[0331] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active ingredient is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and / or (a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, (b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, (c) humectants such as glycerol, (d) disintegrating agents such as agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, I solution retarding agents such as paraffin, (f) absorption accelerators such as quaternary ammonium compounds, (g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, (h) absorbents such as kaolin and bentonite clay, and (i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may include a buffering agent.

[0332] Solid compositions of a similar type can be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the art of pharmacology. They may optionally comprise opacifying agents and can be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of encapsulating compositions which can be used include polymeric substances and waxes. Solid compositions of a similar type can be employed as fillers in soft and hard- filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.

[0333] The active ingredient can be in a micro-encapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings, and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active ingredient can be admixed with at least one inert diluent such as sucrose, lactose, or starch. Such dosage forms may comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may comprise buffering agents. They may optionally comprise opacifying agents and can be of a composition that they release the active ingredient(s) only, or86 / 179B0662.70120WO00preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of encapsulating agents which can be used include polymeric substances and waxes.

[0334] Dosage forms for topical and / or transdermal administration of a compound, agent, particle, or composition described herein may include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants, and / or patches. Generally, the active ingredient is admixed under sterile conditions with a pharmaceutically acceptable carrier or excipient and / or any needed preservatives and / or buffers as can be required. Additionally, the present disclosure contemplates the use of transdermal patches, which often have the added advantage of providing controlled delivery of an active ingredient to the body. Such dosage forms can be prepared, for example, by dissolving and / or dispensing the active ingredient in the proper medium. Alternatively or additionally, the rate can be controlled by either providing a rate controlling membrane and / or by dispersing the active ingredient in a polymer matrix and / or gel.

[0335] Suitable devices for use in delivering intradermal pharmaceutical compositions described herein include short needle devices. Intradermal compositions can be administered by devices which limit the effective penetration length of a needle into the skin. Alternatively or additionally, conventional syringes can be used in the classical mantoux method of intradermal administration. Jet injection devices which deliver liquid formulations to the dermis via a liquid jet injector and / or via a needle which pierces the stratum corneum and produces a jet which reaches the dermis are suitable. Ballistic powder / particle delivery devices which use compressed gas to accelerate the compound, agent, particle, or composition in powder form through the outer layers of the skin to the dermis are suitable.

[0336] Formulations suitable for topical administration include, but are not limited to, liquid and / or semi-liquid preparations such as liniments, lotions, oil-in-water and / or water-in-oil emulsions such as creams, ointments, and / or pastes, and / or solutions and / or suspensions. Topically administrable formulations may, for example, comprise from about 1% to about 10% (w / w) active ingredient, although the concentration of the active ingredient can be as high as the solubility limit of the active ingredient in the solvent. Formulations for topical administration may further comprise one or more of the additional ingredients described herein.

[0337] A pharmaceutical composition described herein can be prepared, packaged, and / or sold in a formulation suitable for pulmonary administration via the buccal cavity. Such a formulation may comprise dry particles which comprise the active ingredient and which have87 / 179B0662.70120WO00a diameter in the range from about 0.5 to about 7 nanometers, or from about 1 to about 6 nanometers. Such compositions are conveniently in the form of dry powders for administration using a device comprising a dry powder reservoir to which a stream of propellant can be directed to disperse the powder and / or using a self-propelling solvent / powder dispensing container such as a device comprising the active ingredient dissolved and / or suspended in a low-boiling propellant in a sealed container. Such powders comprise particles wherein at least 98% of the particles by weight have a diameter greater than 0.5 nanometers and at least 95% of the particles by number have a diameter less than 7 nanometers. Alternatively, at least 95% of the particles by weight have a diameter greater than 1 nanometer and at least 90% of the particles by number have a diameter less than 6 nanometers. Dry powder compositions may include a solid fine powder diluent such as sugar and are conveniently provided in a unit dose form.

[0338] Low boiling propellants generally include liquid propellants having a boiling point of below 65 °F at atmospheric pressure. Generally the propellant may constitute 50 to 99.9% (w / w) of the composition, and the active ingredient may constitute 0.1 to 20% (w / w) of the composition. The propellant may further comprise additional ingredients such as a liquid non-ionic and / or solid anionic surfactant and / or a solid diluent (which may have a particle size of the same order as particles comprising the active ingredient).

[0339] Pharmaceutical compositions described herein formulated for pulmonary delivery may provide the active ingredient in the form of droplets of a solution and / or suspension. Such formulations can be prepared, packaged, and / or sold as aqueous and / or dilute alcoholic solutions and / or suspensions, optionally sterile, comprising the active ingredient, and may conveniently be administered using any nebulization and / or atomization device. Such formulations may further comprise one or more additional ingredients including, but not limited to, a flavoring agent such as saccharin sodium, a volatile oil, a buffering agent, a surface active agent, and / or a preservative such as methylhydroxybenzoate. The droplets provided by this route of administration may have an average diameter in the range from about 0.1 to about 200 nanometers.

[0340] Formulations described herein as being useful for pulmonary delivery are useful for intranasal delivery of a pharmaceutical composition described herein. Another formulation suitable for intranasal administration is a coarse powder comprising the active ingredient and having an average particle from about 0.2 to 500 micrometers. Such a formulation is administered by rapid inhalation through the nasal passage from a container of the powder held close to the nares.88 / 179B0662.70120WO00

[0341] Formulations for nasal administration may, for example, comprise from about as little as 0.1% (w / w) to as much as 100% (w / w) of the active ingredient, and may comprise one or more of the additional ingredients described herein. A pharmaceutical composition described herein can be prepared, packaged, and / or sold in a formulation for buccal administration. Such formulations may, for example, be in the form of tablets and / or lozenges made using conventional methods, and may contain, for example, 0.1 to 20% (w / w) active ingredient, the balance comprising an orally dissolvable and / or degradable composition and, optionally, one or more of the additional ingredients described herein. Alternately, formulations for buccal administration may comprise a powder and / or an aerosolized and / or atomized solution and / or suspension comprising the active ingredient. Such powdered, aerosolized, and / or aerosolized formulations, when dispersed, may have an average particle and / or droplet size in the range from about 0.1 to about 200 nanometers, and may further comprise one or more of the additional ingredients described herein.

[0342] A pharmaceutical composition described herein can be prepared, packaged, and / or sold in a formulation for ophthalmic administration. Such formulations may, for example, be in the form of eye drops including, for example, a 0.1-1.0% (w / w) solution and / or suspension of the active ingredient in an aqueous or oily liquid carrier or excipient. Such drops may further comprise buffering agents, salts, and / or one or more other of the additional ingredients described herein. Other opthalmically-administrable formulations which are useful include those which comprise the active ingredient in microcrystalline form and / or in a liposomal preparation. Ear drops and / or eye drops are also contemplated as being within the scope of this disclosure.

[0343] Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and / or perform such modification with ordinary experimentation.

[0344] Compounds, agents, particles, or compositions provided herein are typically formulated in dosage unit form for ease of administration and uniformity of dosage. It will be understood, however, that the total daily usage of the compositions described herein will be decided by a physician within the scope of sound medical judgment. The specific89 / 179B0662.70120WO00therapeutically effective dose level for any particular subject or organism will depend upon a variety of factors including the disease being treated and the severity of the disorder; the activity of the specific active ingredient employed; the specific composition employed; the age, body weight, general health, sex, and diet of the subject; the time of administration, route of administration, and rate of excretion of the specific active ingredient employed; the duration of the treatment; drugs used in combination or coincidental with the specific active ingredient employed; and like factors well known in the medical arts.

[0345] The compounds, agents, particles, and compositions provided herein can be administered by any route, including enteral (e.g., oral), parenteral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (as by powders, ointments, creams, and / or drops), mucosal, nasal, buccal, sublingual; by intratracheal instillation, bronchial instillation, and / or inhalation; and / or as an oral spray, nasal spray, and / or aerosol. Specifically contemplated routes are oral administration, intravenous administration (e.g., systemic intravenous injection), regional administration via blood and / or lymph supply, and / or direct administration to an affected site. In general, the most appropriate route of administration will depend upon a variety of factors including the nature of the agent (e.g., its stability in the environment of the gastrointestinal tract), and / or the condition of the subject (e.g., whether the subject is able to tolerate oral administration). In certain embodiments, the compound, agent, particle, or composition described herein is suitable for topical administration to the eye of a subject.

[0346] The exact amount of a compound, agent, particle, or composition required to achieve an effective amount will vary from subject to subject, depending, for example, on species, age, and general condition of a subject, severity of the side effects or disorder, identity of the particular compound, agent, particle, or composition, mode of administration, and the like. An effective amount may be included in a single dose (e.g., single oral dose) or multiple doses (e.g., multiple oral doses). In certain embodiments, when multiple doses are administered to a subject or applied to a tissue or cell, any two doses of the multiple doses include different or substantially the same amounts of a compound, agent, particle, or composition described herein. In certain embodiments, when multiple doses are administered to a subject or applied to a tissue or cell, the frequency of administering the multiple doses to the subject or applying the multiple doses to the tissue or cell is three doses a day, two doses a day, one dose a day, one dose every other day, one dose every third day, one dose every week, one dose every two weeks, one dose every three weeks, or one dose every four weeks.90 / 179B0662.70120WO00In certain embodiments, the frequency of administering the multiple doses to the subject or applying the multiple doses to the tissue or cell is one dose per day. In certain embodiments, the frequency of administering the multiple doses to the subject or applying the multiple doses to the tissue or cell is two doses per day. In certain embodiments, the frequency of administering the multiple doses to the subject or applying the multiple doses to the tissue or cell is three doses per day. In certain embodiments, when multiple doses are administered to a subject or applied to a tissue or cell, the duration between the first dose and last dose of the multiple doses is one day, two days, four days, one week, two weeks, three weeks, one month, two months, three months, four months, six months, nine months, one year, two years, three years, four years, five years, seven years, ten years, fifteen years, twenty years, or the lifetime of the subject, tissue, or cell. In certain embodiments, the duration between the first dose and last dose of the multiple doses is three months, six months, or one year. In certain embodiments, the duration between the first dose and last dose of the multiple doses is the lifetime of the subject, tissue, or cell. In certain embodiments, a dose (e.g., a single dose, or any dose of multiple doses) described herein includes independently between 0.1 µg and 1 µg, between 0.001 mg and 0.01 mg, between 0.01 mg and 0.1 mg, between 0.1 mg and 1 mg, between 1 mg and 3 mg, between 3 mg and 10 mg, between 10 mg and 30 mg, between 30 mg and 100 mg, between 100 mg and 300 mg, between 300 mg and 1,000 mg, or between 1 g and 10 g, inclusive, of a compound, agent, particle, or composition described herein. In certain embodiments, a dose described herein includes independently between 1 mg and 3 mg, inclusive, of a compound, agent, particle, or composition described herein. In certain embodiments, a dose described herein includes independently between 3 mg and 10 mg, inclusive, of a compound, agent, particle, or composition described herein. In certain embodiments, a dose described herein includes independently between 10 mg and 30 mg, inclusive, of a compound, agent, particle, or composition described herein. In certain embodiments, a dose described herein includes independently between 30 mg and 100 mg, inclusive, of a compound, agent, particle, or composition described herein.

[0347] Dose ranges as described herein provide guidance for the administration of provided pharmaceutical compositions to an adult. The amount to be administered to, for example, a child or an adolescent can be determined by a medical practitioner or person skilled in the art and can be lower or the same as that administered to an adult.

[0348] A compound, agent, particle, or composition, as described herein, can be administered in combination with one or more additional pharmaceutical agents (e.g., therapeutically and / or prophylactically active agents). The compounds, agents, particles, or91 / 179B0662.70120WO00compositions can be administered in combination with additional pharmaceutical agents that improve their activity (e.g., activity (e.g., potency and / or efficacy) in treating a disease in a subject in need thereof, in preventing a disease in a subject in need thereof, in reducing the risk to develop a disease in a subject in need thereof, and / or in inhibiting the activity of a protein kinase in a subject or cell), improve bioavailability, improve safety, reduce drug resistance, reduce and / or modify metabolism, inhibit excretion, and / or modify distribution in a subject or cell. It will also be appreciated that the therapy employed may achieve a desired effect for the same disorder, and / or it may achieve different effects. In certain embodiments, a pharmaceutical composition described herein including a compound, agent, particle, or composition described herein and an additional pharmaceutical agent shows a synergistic effect that is absent in a pharmaceutical composition including one of the compound, agent, particle, or composition and the additional pharmaceutical agent, but not both. In some embodiments, the additional pharmaceutical agent achieves a desired effect for the same disorder. In some embodiments, the additional pharmaceutical agent achieves different effects.

[0349] The compound, agent, particle, or composition can be administered concurrently with, prior to, or subsequent to one or more additional pharmaceutical agents, which may be useful as, e.g., combination therapies. Pharmaceutical agents include therapeutically active agents. Pharmaceutical agents also include prophylactically active agents. Pharmaceutical agents include small organic molecules such as drug compounds (e.g., compounds approved for human or veterinary use by the U.S. Food and Drug Administration as provided in the Code of Federal Regulations (CFR)), peptides, proteins, carbohydrates, monosaccharides, oligosaccharides, polysaccharides, nucleoproteins, mucoproteins, lipoproteins, synthetic polypeptides or proteins, small molecules linked to proteins, glycoproteins, steroids, nucleic acids, DNAs, RNAs, nucleotides, nucleosides, oligonucleotides, antisense oligonucleotides, lipids, hormones, vitamins, and cells. In certain embodiments, the additional pharmaceutical agent is a pharmaceutical agent useful for treating and / or preventing a disease (e.g., proliferative disease, hematological disease, neurological disease, painful condition, psychiatric disorder, or metabolic disorder). Each additional pharmaceutical agent may be administered at a dose and / or on a time schedule determined for that pharmaceutical agent. The additional pharmaceutical agents may also be administered together with each other and / or with the compound, agent, particle, or composition described herein in a single dose or composition or administered separately in different doses or compositions. The particular combination to employ in a regimen will take into account compatibility of the compound,92 / 179B0662.70120WO00agent, particle, or composition described herein with the additional pharmaceutical agent(s) and / or the desired therapeutic and / or prophylactic effect to be achieved. In general, it is expected that the additional pharmaceutical agent(s) in combination be utilized at levels that do not exceed the levels at which they are utilized individually. In some embodiments, the levels utilized in combination will be lower than those utilized individually.

[0350] The additional pharmaceutical agents include, but are not limited to, anti- proliferative agents, anti-cancer agents, anti-angiogenesis agents, steroidal or non-steroidal anti-inflammatory agents, immunosuppressants, anti-bacterial agents, anti-viral agents, cardiovascular agents, cholesterol-lowering agents, anti-diabetic agents, anti-allergic agents, contraceptive agents, pain-relieving agents, anesthetics, anti–coagulants, inhibitors of an enzyme, steroidal agents, steroidal or antihistamine, antigens, vaccines, antibodies, decongestant, sedatives, opioids, analgesics, anti–pyretics, hormones, and prostaglandins. In certain embodiments, the additional pharmaceutical agent is an anti-proliferative agent. In certain embodiments, the additional pharmaceutical agent is an anti-cancer agent. In certain embodiments, the additional pharmaceutical agent is an anti-viral agent. In certain embodiments, the additional pharmaceutical agent is an binder or inhibitor of a protein kinase. In certain embodiments, the additional pharmaceutical agent is selected from the group consisting of epigenetic or transcriptional modulators (e.g., DNA methyltransferase inhibitors, histone deacetylase inhibitors (HDAC inhibitors), lysine methyltransferase inhibitors), antimitotic drugs (e.g., taxanes and vinca alkaloids), hormone receptor modulators (e.g., estrogen receptor modulators and androgen receptor modulators), cell signaling pathway inhibitors (e.g., tyrosine protein kinase inhibitors), modulators of protein stability (e.g., proteasome inhibitors), Hsp90 inhibitors, glucocorticoids, all-trans retinoic acids, and other agents that promote differentiation. In certain embodiments, the compounds, agents, particles, or compositions described herein or pharmaceutical compositions can be administered in combination with an anti-cancer therapy including, but not limited to, surgery, radiation therapy, transplantation (e.g., stem cell transplantation, bone marrow transplantation), immunotherapy, and chemotherapy. Additional pharmaceutical agents include small organic molecules such as drug compounds (e.g., compounds approved by the US Food and Drug Administration as provided in the Code of Federal Regulations (CFR)), peptides, proteins, carbohydrates, monosaccharides, oligosaccharides, polysaccharides, nucleoproteins, mucoproteins, lipoproteins, synthetic polypeptides or proteins, small molecules linked to proteins, glycoproteins, steroids, nucleic acids, DNAs, RNAs,93 / 179B0662.70120WO00nucleotides, nucleosides, oligonucleotides, antisense oligonucleotides, lipids, hormones, vitamins and cells.

[0351] Also encompassed by the disclosure are kits (e.g., pharmaceutical packs). The kits provided may comprise a pharmaceutical composition or compound, agent, particle, or composition described herein and a container (e.g., a vial, ampule, bottle, syringe, and / or dispenser package, or other suitable container). In some embodiments, provided kits may optionally further include a second container comprising a pharmaceutical excipient for dilution or suspension of a pharmaceutical composition or compound, agent, particle, or composition described herein. In some embodiments, the pharmaceutical composition or compound, agent, particle, or composition described herein provided in the first container and the second container are combined to form one unit dosage form.

[0352] Thus, in one aspect, provided are kits comprising a compound provided herein or a composition provided herein and instructions for using the compound or composition. In some embodiments, the kit comprises including a first container comprising a compound, agent, particle, or composition or pharmaceutical composition described herein. In certain embodiments, the kits are useful for treating a disease (e.g., an infectious disease, genetic disease, or cancer) in a subject in need thereof. In certain embodiments, the kits are useful for preventing a disease (e.g., an infectious disease, genetic disease, or cancer) in a subject in need thereof. In certain embodiments, the kits are useful for reducing the risk of developing a disease (e.g., an infectious disease, genetic disease, or cancer) in a subject in need thereof. In certain embodiments, the kits are useful for delivering an agent to a cell.

[0353] In certain embodiments, a kit described herein further includes instructions for using the kit. A kit described herein may also include information as required by a regulatory agency such as the U.S. Food and Drug Administration (FDA). In certain embodiments, the information included in the kits is prescribing information. In certain embodiments, the kits and instructions provide for treating a disease (e.g., an infectious disease, genetic disease, or cancer) in a subject in need thereof. In certain embodiments, the kits and instructions provide for preventing a disease (e.g., an infectious disease, genetic disease, or cancer) in a subject in need thereof. In certain embodiments, the kits and instructions provide for reducing the risk of developing a disease (e.g., an infectious disease, genetic disease, or cancer) in a subject in need thereof. In certain embodiments, the kits and instructions provide for delivering an agent. A kit described herein may include one or more additional pharmaceutical agents described herein as a separate composition.94 / 179B0662.70120WO00Methods of Treatment and Uses

[0354] In another aspect, provided herein is a method of treating or preventing a disease in a subject, comprising administering to the subject a compound or composition provided herein. In some embodiments, the method comprises administering a compound provided herein. In some embodiments, the method comprises administering a composition provided herein.

[0355] In some embodiments, the disease is a cardiovascular disease, a lung or respiratory disease, a musculoskeletal disease, a hematological disease, an immune disorder, an infectious disease, a genetic disease, cancer, a neurological disease, a psychiatric disorder, a metabolic disorder, or an inflammatory disease. In some embodiments, the disease is an infectious disease, a genetic disease, or cancer. In some embodiments, the disease is a cardiovascular disease. In some embodiments, the disease is a lung disease. In some embodiments, the disease is a respiratory disease. In some embodiments, the disease is a musculoskeletal disease. In some embodiments, the disease is a hematological disease. In some embodiments, the disease is an immune disorder. In some embodiments, the disease is an infectious disease. In some embodiments, the disease is a genetic disease. In some embodiments, the disease is cancer. In some embodiments, the disease is a neurological disease. In some embodiments, the disease is a psychiatric disorder. In some embodiments, the disease is a metabolic disorder. In some embodiments, the disease is an inflammatory disease.

[0356] In certain embodiments, the subject is an animal. The animal may be of either sex and may be at any stage of development. In certain embodiments, the subject described herein is a human. In certain embodiments, the subject is a non-human animal. In certain embodiments, the subject is a mammal. In certain embodiments, the subject is a non-human mammal. In certain embodiments, the subject is a domesticated animal, such as a dog, cat, cow, pig, horse, sheep, or goat. In certain embodiments, the subject is a companion animal, such as a dog or cat. In certain embodiments, the subject is a livestock animal, such as a cow, pig, horse, sheep, or goat. In certain embodiments, the subject is a zoo animal. In another embodiment, the subject is a research animal, such as a rodent (e.g., mouse, rat), dog, pig, or non-human primate. In certain embodiments, the animal is a genetically engineered animal. In certain embodiments, the animal is a transgenic animal (e.g., transgenic mice and transgenic pigs). In certain embodiments, the subject is a fish or reptile.95 / 179B0662.70120WO00Additional Methods and Uses

[0357] In another aspect, provided herein is a method of delivering an agent to a subject, cell, or biological sample, comprising administering to the subject or contacting the cell or biological sample with an effective amount of a compound or composition provided herein. In some embodiments, the method comprises administering to the subject or contacting the cell or biological sample with a compound provided herein. In some embodiments, the method comprises administering to the subject or contacting the cell or biological sample with a composition provided herein.

[0358] In some embodiments, the agent is any agent provided herein. In some embodiments, the agent is an organic molecule, protein, polypeptide, polynucleotide, an isotopically labeled chemical compound, vaccine, or an immunological agent. In some embodiments, the agent is a protein, polypeptide, or polynucleotide. In some embodiments, the agent is a protein. In some embodiments, the agent is a polypeptide. In some embodiments, the agent is a polynucleotide. In some embodiments, the agent is an RNA. In some embodiments, the RNA is messenger RNA (mRNA).In some embodiments, the RNA is small interfering RNA (siRNA). In some embodiments, the polynucleotide is a DNA. In some embodiments, the DNA is a plasmid DNA (pDNA).

[0359] In some embodiments, the agent is delivered to a subject. In some embodiments, the agent is delivered to a target tissue of the subject. In some embodiments, the agent is delivered to a cell. In some embodiments, the cell is in vivo, e.g., in an organism. In certain embodiments, the cell is in vitro, e.g., in cell culture. In some embodiments, the cell is ex vivo, meaning the cell is removed from an organism prior to the delivery. In some embodiments, the cell is a eukaryotic cell. In some embodiments, the cell is a HeLa cell, HEK293 cell, a fibroblast, a macrophage, a T cell, or a hematopoietic stem cell. In some embodiments, the cell is a HEK293 cell. In some embodiments, the cell is a fibroblast. In some embodiments, the cell is a macrophage. In some embodiments, the cell is a T cell. In some embodiments, the cell is a hematopoietic stem cell. In some embodiments, the cell is a HeLa cell.

[0360] In some embodiments, the agent is delivered from an extracellular space to the cytosol of a cell. In some embodiments, the agent is delivered from an extracellular space to the nucleus of the cell.

[0361] In some embodiments, the composition is administered by any method provided herein.96 / 179B0662.70120WO00

[0362] In some embodiments, the effective amount is sufficient to increase the transduction efficiency of the polynucleotide or polypeptide. In some embodiments, the effective amount is sufficient to increase the transduction efficiency of the polynucleotide. In some embodiments, the effective amount is sufficient to increase the transduction efficiency of the polypeptide. EXAMPLES

[0363] In order that the present disclosure may be more fully understood, the following examples are set forth. The synthetic and biological examples described in this application are offered to illustrate the compounds, pharmaceutical compositions, and methods provided herein and are not to be construed in any way as limiting in their scope. EXAMPLE 1

[0364] Effective intracellular delivery of biomacromolecules such as small interfering RNA (siRNA), protein-encoding messenger RNA (mRNA), and proteins can facilitate treatment of a variety of clinical disorders. A recombinant elastin-like polypeptide (ELP)-based delivery platform was developed for effective siRNA, mRNA, and protein cargo delivery in vitro and in vivo. Through an iterative design process, cationic endosomal escape peptide (EEP)-fused ELPs were developed. These EEP-fused ELPs demonstrated micellar self-assembly, enabling EEP shielding from the particle surface, effective nucleic acid encapsulation, as well as acidic pH-responsive micelle disassembly permitting EEP activity and intracellular cargo delivery. Leveraging a machine learning-based predictive model of peptide membrane disruptive activity, in silico screening of α-helical peptide libraries led to the discovery of a various EEPs (including EEP13 (SEQ ID NO: 14)) with dramatically improved efficiency in both mRNA and protein delivery compared to previously reported EEPs. Multimerization of EEPs further improved potency. Due to the multivalent nature and micellar architecture facilitating EEP shielding from the nanoparticle surface, ELP-EEP nanoparticles also demonstrated greatly improved potency and reduced cytotoxicity as compared to standalone EEPs. ELP-EEP delivery vehicles outperformed lipid-based reagents in the delivery of mRNA-encoded and Cre recombinase protein, as well as GFP-targeted siRNA to multiple cell lines and primary cells. The ability of this system to mediate in vivo delivery of Cre protein was confirmed after intradermal administration. The results establish ELP nanoparticles as a first-in-class non- viral, recombinant protein-based delivery platform for diverse macromolecular cargo of therapeutic interest.97 / 179B0662.70120WO00Introduction

[0365] Elastin-like polypeptides (ELPs) are recombinant protein polymers that exhibit thermally responsive self-assembly, enabling the formation of micelles that have been utilized as drug carriers for various cargo.17–19ELPs are composed of a short repeating peptide motif of Val-Pro- Gly-X-Gly derived from the hydrophobic domain of tropoelastin, where X represents a guest residue that can be any amino acid except proline. ELPs exhibit lower critical solution temperature (LCST) phase behavior. Specifically, ELPs are soluble below a transition temperature (Tt), inversely correlated to guest residue (X) hydrophobicity and protein size20,21, but above the transition temperature reversibly phase separate to form insoluble coacervates.22ELPs can be synthesized with high precision and programmability, are biodegradable, and exhibit minimal immunogenicity.23Previously designed amphiphilic diblock ELPs can self-assemble into micellular nanoparticles with a hydrophilic surface and hydrophobic core for targeted extracellular drug delivery, including proteins and small molecules.24–26Fusion constructs between elastin-like polypeptides and therapeutic proteins have also been developed, including vascular endothelial growth factor A (VEGF-A)27,28, glucagon-like peptide-1 (GLP-1)29, and chemotherapeutic agents.30,31Attempts to utilize ELPs for the intracellular delivery of proteins, gene editor ribonucleoproteins (RNPs), and mRNA have not been reported and efforts to deliver nucleic acids, such as siRNA and pDNA, have been associated with low potency, presumably due to an inability to mediate effective endosomal escape.32,33

[0366] An ELP-based delivery system (Elastin-based Nanoparticles for Therapeutic delivery (ENTER)) that achieves efficient intracellular delivery of major therapeutic macromolecule modalities, including proteins, RNP, siRNA, mRNA, and plasmid DNA to a variety of cell types both in vitro and in vivo was designed. Over four generations of designs, engineered ELP multiblock protein polymers were developed that self-assemble as crosslinked nanoparticles displaying a hydrophilic corona and a histidine-containing hydrophobic core, which includes cationic endosomal escape peptides (EEPs) (FIG. 1). Significantly, iterative machine learning-based in silico design and in vitro screening led to the discovery of novel EEP sequences (e.g., EEP13 (SEQ ID NO: 14)) that greatly enhanced intracellular delivery of both proteins and nucleic acids. ELPs enabled efficient delivery of Cre recombinase protein and mRNA to HEK293 cells, as well as fibroblasts, macrophages, T cells, and hematopoietic stem cells (HSCs) and were easily modified for cell specific delivery. Of note, EEPs as a component of ELP nanoparticles displayed dramatically enhanced potency with minimal cytotoxicity as compared to free EEP peptide shuttles. Moreover, this system provided an98 / 179B0662.70120WO00effective means for intracellular delivery of siRNA, mRNA, plasmid DNA, proteins, as well as Cas9 RNP and IRF5, as a prototypical transcription factor, in cell lines as well as primary fibroblasts, macrophages, T cells, and hematopoietic stem cells. Delivery of interferon regulatory factor 5 (IRF5), an M1 macrophage lineage-defining transcription, to macrophages in vitro significantly upregulated the expression of key pro-inflammatory cytokines. In vivo delivery of Cre protein was confirmed in Ai9 mice. Intranasal administration of ELP-EEP13 combined with Cre recombinase protein achieved effective editing of lung epithelial cells in reporter mice. These studies establish ELPs and the ENTER system as a platform for intracellular delivery of proteins and nucleic acids for a variety of therapeutic applications. Formation and Characterization of ELP Nanoparticles

[0367] A first-generation micelle-forming ELP (V1-ELP) bearing an N-terminal triglycine motif for chemical or sortase-mediated conjugation was previously reported.26V1-ELPs are diblock copolymers composed of a glutamic acid-containing hydrophilic block, a tyrosine- containing hydrophobic block, and a cysteine midblock domain enabling disulfide crosslinking (FIG. 1).34Micelle self-assembly was observed above the transition temperature of the hydrophobic block (10 °C)34with dynamic light scattering (DLS) demonstrating an average particle diameter of ~60 nm above a critical micelle concentration (CMC) of ~500 nM (FIGs. 10A-10B).34V1- ELPs were effectively internalized by HeLa cells and HEK293 cells without cytotoxicity (FIGs. 10C-10D).

[0368] The ability to conjugate Cre recombinase and Streptococcus pyogenes Cas9 (SpCas9) modified with a C-terminal LPETG motif and a protease cleavable (GGS)9linker35to ELP nanoparticles by sortase transpeptidation was assessed.36SDS-PAGE confirmed the ability to conjugate Cre to about 50% of accessible GGG motifs (FIGs. 2A-2C) with an increase in nanoparticle diameter to 70-80 nm (FIG. 2D). Cathepsin B, an endosomal protease, effectively liberated Cre and induced degradation of ELP protein polymer (FIGs. 11A-11B). Similarly, Cas9 was easily conjugated to V1-ELPs (FIGs. 2E-2F, FIG. 12) with an increase in particle diameter, further augmented by the introduction of single guide RNA (sgRNA) with a commensurate reduction in zeta potential to generate ELP bound Cas9 ribonucleoproteins (RNP) (FIGs. 2G-2H). An anti-CD117 antibody was able to be conjugated to V1-ELPs (FIG. 2I). CD117 (c-kit) is upregulated on human and murine hematopoietic stem cells (HSC) and previously used for selective HSC targeting.37,38SDS- PAGE and band densitometry analysis confirmed antibody conjugation (FIGs. 2J-2K). Antibody mediated ELP targeting to P815 cells, a CD117+ murine mast cell line, was99 / 179B0662.70120WO00evaluated by conjugating Alexa Fluor 633 to a V1-ELP variant containing a C-terminal cysteine domain. On target binding of ELPs was confirmed (FIGs. 2L-2M, 13A-13B). ELP-mediated intracellular protein delivery

[0369] After confirming that V1-ELP is a suitable backbone for straightforward functionalization with protein cargo, an iterative design strategy was pursued to optimize ELP-mediated protein and nucleic acid intracellular delivery (FIG. 3A). The valine residues present in the hydrophobic block of V1-ELP were replaced with histidine (V2-ELP) to increase the hydrophilicity of the hydrophobic block in order to facilitate micelle disassembly within acidic endosomes following histidine protonation, and promote endosomal swelling and osmolysis through a proton sponge effect39. Measurement of V2-ELP micelle size at varying pH demonstrated effective micelle disassembly at pH below the pKa of the histidine side chain (~6.8), while no such trend was observed for V1-ELPs (FIG. 3B, 14A-14C). HEK293 Cre reporter cells (HEK293-RFP) were treated with Cre bearing V2-ELP nanoparticles (Cre-V2-ELP) (FIGs. 15A-15D), but RFP expression was only observed in the presence of chloroquine, a cationic amphipathic drug with known endosomolytic capacity40(FIG. 3D). Notably, RFP expression was greater when Cre was delivered by V2-ELPs as compared to V1-ELPs (~90% vs. 60%), consistent with the ability of histidine residues to enhance endosomolysis.

[0370] A third generation ELP was designed to eliminate the need for an exogenous endosomolytic agent. To endow ELPs with membrane disruption capabilities, V2-ELPs were modified with endosomal escape peptides (EEPs), known to enhance intracellular delivery of macromolecular cargo (V3-ELPs).11,14,41–43V3-ELPs were generated containing one of five EEPs positioned at the N-terminus of the protein polymer (FIGs. 16A-16E); each construct demonstrated stable size at room temperature over the course of 72 hours and was amenable to sortase-mediated functionalization with Cre recombinase. HEK293-RFP cells treated with Cre-V3-ELPs for 48 hours displayed dose-dependent RFP expression, up to an order of magnitude greater than those treated with unmodified V2-ELPs (FIG. 3D). Effective ELPs contained Tat15or or S1014, enabling RFP expression in up to 30% of cells at a concentration of 16 µM, though some cytotoxicity was observed at this dose (FIG. 17). To further optimize Cre release from ELP nanoparticles, a linker library (SEQ ID NOs: 91-97) was screened with known susceptibility to a variety of endolysosomal proteases (FIG. 18A).44–47HEK293- RFP cells were treated with V3-ELPs bearing an S10 EEP (SEQ ID NO: 60) for 48 hrs. A furin cleavable linker flanked by GGS repeats (GGSGGSRVRRGGSGGS)(SEQ ID NO: 96)100 / 179B0662.70120WO00demonstrated the highest release efficiency, as evident by the ratio of RFP+ cells after treatment with Cre in ELP-conjugated or unconjugated formats (FIG. 18B). This linker was used in subsequent studies.

[0371] Cytotoxicity remains a shortcoming of all existing cell penetrating and membrane disruptive peptides, which has been attributed to excessive disruption of the plasma cell membrane.48To address this limitation, V4-ELPs were designed with the EEP positioned at the C-terminus of the protein polymer, so as to be localized within the nanoparticle core. Without wishing to be bound by any particular theory, it was postulated that once concentrated within an endosome, protonation of histidines would lead to micelle disassembly, exposing EEPs and enabling endosomal disruption. It was further hypothesized that these cationic EEPs would also facilitate encapsulation of nucleic acids. To circumvent electrostatic interactions between the C-terminal cationic EEP and the N-terminal anionic hydrophilic block, glutamic acid residues in hydrophilic block were replaced by alanine and glycine at a 3:2 ratio (FIG. 3A). V4-ELPs form monodisperse spherical micelles with a diameter of ~70 nm as determined by DLS (FIG. 19) and TEM imaging and were further modified with the EEPs noted above, as well as nonaarginine (R9)49, transportan-1050, and L17E M-lycotoxin51. Treatment of HEK293-RFP cells with V4-ELP-EEPs (8 µM) and free, unbound Cre (2 µM) revealed that V4-ELPs containing S10 (SEQ ID NO: 84) enabled RFP expression in nearly all cells without cytotoxicity (FIGs. 3E, 20). Notably, Cre delivery by V4-ELP-S10 (SEQ ID NO: 84) was superior to that achieved using S10 (SEQ ID NO: 36) shuttle peptides alone (i.e., as an untethered peptide), with over 20-fold less ELP being required to achieving 50% RFP+ cells (~1.5 µM vs 32 µM, FIG. 3F). This demonstrates that ELP- mediated EEP multivalency greatly improves the ability to achieve effective intracellular target protein delivery. Remarkably, cell viability was unaffected by treatment with V4-ELPs at all doses tested, in contrast to free S10 (SEQ ID NO: 36) peptide, which was associated with substantial toxicity at its EC50 (FIG. 3G). V4-ELPs also outperformed Lipofectamine 3000 in the delivery of Cre protein, with RFP expression in only 40% cells under optimal Lipofectamine conditions in contrast to 100% expression when delivered by V4-ELPs (FIGs. 21A-21B). These findings confirm that V4-ELPs are effective vectors for intracellular delivery of protein cargo. Discovery of enhanced EEPs via in silico screening

[0372] To further improve the efficacy of V4-ELPs, EEPs were designed with enhanced activity. Among EEPs initially screened for Cre protein delivery in HEK293-RFP cells, the101 / 179B0662.70120WO00most effective peptide was S10 (SEQ ID NO: 36). S10 is comprised of amphipathic α-helical peptide variants of CM1852and PTD453, separated by a Gly-Ser linker, in contrast to other polycationic EEPs that are either unstructured or contain a single α-helical domain (FIGs. 22A-22B). The amphipathic α-helical structure is thought to facilitate intercalation within lipid bilayers, particularly negatively charged bilayers, characteristic of late endosomes.54,55

[0373] Motivated by this insight, an α-helix peptide database was compiled and screened to identify next generation EEPs with enhanced functionality. To enable prediction of EEP efficacy, a multiple linear regression model was trained on a published dataset of 73 membrane disruptive peptides encoded by a set of 15 physicochemical, structural, and compositional features, labeled with efficacy values derived from an in vitro GFP protein delivery assay (Table 1).56F-score screening of each feature with respect to peptide efficacy revealed specific physicochemical properties, including hydrophobic moment, net charge, and amphipathic α- helical structure or lack thereof, which were highly correlated with delivery efficacy (FIG. 23). Regression models using the top k features (k = 1 to 15) were trained and evaluated by leave-one-out cross validation (FIG. 4A). The model with the top performance on held-out data points (root mean squared error (RMSE) = 12.4) is described by the equation: Predicted efficacy (%) = 5.9 × hydrophobic moment + 3.2 × net charge – 3.2 × pI – 4.3 × % negative residues + 20.8 × is amphipathic helix Table 1. Physicochemical and compositional descriptors used to encode peptide library for predictive model training.102 / 179B0662.70120WO00

[0374] This model was used in a multistage screen of a library of peptides compiled from the Antimicrobial Peptide Database (APD)57, the Database of Antimicrobial Activity and Structure of Peptides (DBAASP)58, and the Therapeutic Peptide Design database (TP-DB)59, a database containing all α-helical segments extracted from protein structures compiled in the Protein Data Bank (FIG. 4B). Filters were applied to select peptides labeled as helical from the APD (491 peptides), without chemical modifications from the DBAASP (5,836 peptides), and those peptides likely to remain as stable as helices when isolated from a parent protein in the TP-DB (8,956 peptides). After removal of duplicates, a total of 11,084 peptides were evaluated using the model. Peptides with predicted efficacies greater than 10% were combined into dimers separated by a GGSGGGS (SEQ ID NO: 117) linker and then reassessed. Seven lead peptides were selected (Table 2), with priority given to sequences with predicted efficacies exceeding that of S10 in the GFP protein delivery assay (73.2%) or of human protein origin. Table 2. Endosomal escape peptides (EEPs) designed via in silico ⍺-helical peptide library screening.103 / 179B0662.70120WO00104 / 179B0662.70120WO00

[0375] V4-ELPs containing each of these derived EEPs were generated and assessed for their ability to deliver free Cre protein at a dose of 100 nM to avoid effect saturation (near 100% RFP positivity) observed in prior screening studies at 2 μM. The highest performing V4-ELP-EEP (V4-ELP-EEP5 (SEQ ID NO: 71)) led to RFP expression in 25.7% of cells, but underperformed S10 (44.1%, FIG. 4C). The two highest performing V4-ELP-EEPs (V4- ELP-EEP1 and -EEP5 (SEQ ID NOs: 67 and 71)) were enriched in lysine over arginine among positively charged residues, with 87% and 85% of positive residues comprised of lysine, respectively. Motivated by this insight, a second generation of EEPs was identified by rescreening the peptide library described above with filters applied to select for105 / 179B0662.70120WO00peptides enriched in lysine (FIGs. 24-25). The top 8 peptides derived from this in silico screen were evaluated for their capacity to deliver Cre protein to HEK293-RFP cells as V4- ELPs. Two of these constructs, V4-ELP-EEP13 and V4-ELP-EEP14 (SEQ ID NOs: 79 and 82), significantly outperformed S10 (FIG. 4D). EEP13 (SEQ ID NO: 79) is a dimeric amphipathic α-helical peptide composed of two synthetic antimicrobial peptides (FIGs. 4E- 4F).60,61contrast, as a free shuttle peptide, S10 (SEQ ID NO: 36) significantly outperformed EEP13 (SEQ ID NO: 14) in Cre protein delivery and demonstrated less cytotoxicity in a peptide dose escalation study (FIGs. 26A-26B). The separation in performance between the two peptides emerges at doses where EEP13 (SEQ ID NO: 14) displayed increased cytotoxicity, suggesting that elevated cytotoxicity may contribute to reduced potency. As a consequence, nanoparticles designed to shield highly potent EEPs until localized within the endosome achieve higher levels of endosomal escape efficiency that would otherwise not be achievable with the use of free peptides due to inherent cytotoxicity.

[0376] Multimerization of cell-penetrating peptides enhances membrane disruption by increasing binding affinity of multimeric cationic peptides for anionic proteoglycans in the plasma membrane.62,63Given that endosomal membranes are enriched in anionic phospholipid bis(monoacylglycero)phosphate (BMP)64, it was hypothesized that EEP13 multimerization would further increase potency. V4-ELPs containing EEP13 as a monomer, dimer, or trimer (SEQ ID NOs: 79, 80, or 81, respectively) were evaluated for their ability to deliver free Cre protein. All three constructs formed uniform micelles between 60 and 80 nm (e.g., ~75 nm) in diameter (FIG. 4G), with the micelle size increasing as additional copies of EEP13 were incorporated into the ELP. V4-ELPs containing an EEP13 dimer (SEQ ID NO: 80) or trimer (SEQ ID NO: 81) outperformed those containing an EEP13 monomer (SEQ ID NO: 79) at doses between 500 nM (50.8% vs.7.02% RFP+) and 2 µM with limited cytotoxicity noted at the highest dose (FIGs. 4H-4I). However, some increased cytotoxicity associated with the EEP13 dimer and trimer relative to the monomer was observed at and above 1 µM (FIGs. 4H-4I). At a constant concentration of V4-ELP, differences in efficiency or cell viability were not observed with increasing amounts of Cre protein. V4-ELP-[EEP13]2 displayed an EC50 of 500 nM in the presence of 2 μM Cre, while all three variants of V4-ELP-EEP13 led to RFP expression in 50% of cells at a Cre dose of 30 nM (FIGs. 27A-27B). To examine the ability of V4-ELP-EEPs to deliver cargo larger than Cre recombinase (38 kDa), FITC-labeled dextrans with molecular weight up to 2 megadaltons were tested as cargo in a co-formulation study. Both V4-ELP-EEP13 and V4-ELP-[EEP13]2 effectively facilitated cytoplasmic entry of dextrans of106 / 179B0662.70120WO00all tested sizes (FIG. 53). Collectively, these findings support the ability of V4-ELP-EEP13 nanoparticles to deliver large macromolecules.

[0377] Gene disruption or repair via intracellular delivery of gene editing agents, such as Cas nuclease, base editors, and prime editors are promising as a therapeutic strategy for many diseases.72Delivery of gene editors in protein form rather than mRNA or DNA has the potential to reduce off-target editing by limiting the lifetime of the editing agent within treated cells.73However, the large size and complex charge of Cas9 RNP presents multiple delivery challenges. To test the ability of our ELP delivery system to overcome these challenges, we used V4-ELP-EEP13 to deliver free SpCas9 RNP formulated with a sgRNA targeting HPRT1 (hypoxanthine-guanine phosphoribosyltransferase 1) to HEK293 cells. At an ELP concentration of 8 µM, V4-ELP-EEP13 facilitated efficient gene editing at a level (~65%) comparable to the positive control CRISPRMAX, a commercial lipid-based system optimized for RNP delivery (FIG. 44), establishing the potential of ENTER to serve as a delivery vehicle for gene editors. In summary, in silico screening led to the discovery of a novel EEP that dramatically enhanced the ability of ELPs to mediate intracellular protein and RNP delivery with high potency. ELP-mediated protein delivery to primary cells ex vivo

[0378] Efficient delivery to primary cells presents another hurdle for delivery systems. The ability of ELP nanoparticles to deliver Cre protein was evaluated in a variety of primary cells, including lung fibroblasts, peritoneal macrophages, splenic CD4+ T cells, and hematopoietic stem cells from reporter mice (e.g., from Ai9-SauSpyCas9 mice). Delivery of Cre recombinase leads to the excision of a stop cassette flanked by loxP recombination sites with induction of tdTomato expression.

[0379] In fibroblasts, V4- ELP-[EEP13]2(SEQ ID NO: 80) significantly outperformed V4- ELP-EEP13 (SEQ ID NO: 79) particularly at low ELP concentrations, achieving nearly 90% tdTomato+ cells at 4 µM ELP with minimal cytotoxicity (FIGs. 5A-5B, 28A-28B). Notably, Lipofectamine 3000 was much less effective in delivering Cre protein with tdTomato+ expression observed in fewer than 15% of cells at the highest tested dose (FIGs.29A-29B). Optimal delivery efficiency to macrophages (~70% tdTomato+) was observed following co- treatment with 4 or 8 µM V4-ELP-EEP13 (SEQ ID NO: 79) and 8 or 16 µM Cre, respectively (FIGs. 5C-5D). Viability at these effective doses was generally high (~80% from ~90% viability when untreated), though some ELP- and Cre-dose dependent toxicity was observed (FIGs. 28C-28D). Up to 80% of CD4+ T cells were tdTomato+ after treatment with Cre-107 / 179B0662.70120WO00bearing V4-ELPs for 48 hours, with improved efficacy observed for co-delivery of V4-ELPs and free Cre protein at the lowest Cre and ELP concentrations tested (FIG. 5E). Unlike treated fibroblasts and macrophages, treatment of CD4+ T cells result in notable dose- dependent cytotoxicity (FIG.28E).

[0380] Due to concerns about cytotoxicity associated with the multimerized EEP13 in primary cells, monomeric V4-ELP-EEP13 was further evaluated. In fibroblasts, co-treatment with V4-ELP-EEP13 and Cre resulted in a dose-dependent increase in tdTomato+ cells of up to 70% with minimal cytotoxicity (FIGs. 39A-39B, FIGs. 54A-54B). In contrast, Lipofectamine 3000 exhibited lower effectiveness, achieving tdTomato+ expression in fewer than 15% of cells at the highest tested dose (FIGs. 55A-55B). In macrophages, an optimal dose of 8 μM V4-ELP-EEP13 with 8 μM Cre resulted in editing in over 80% of treated cells, showing a near-linear relationship with the treatment concentration of Cre (FIGs. 39C-39D). A modest decrease (~10%) in cell viability was noted at this dose of V4-ELP-EEP13 (FIGs. 54C-54D). CD4+ T cells treated with Cre and V4-ELP-EEP13 exhibited up to 50% tdTomato+ cells with both ELP and Cre dose-dependent editing efficiency (FIGs. 39E-39F). Notable cytotoxicity was observed at the highest tested concentration of ELP, while Cre dose escalation did not result in significant changes in cell viability (FIGs. 54E-54F). Finally, V4- ELP-EEP13 facilitated effective delivery of Cre into Lin-Sca-1+c-kit+ hematopoietic stem cells, enabling ~20% tdTomato+ cells at optimal dosing with minimal cytotoxicity (FIGs. 39G-39H, FIGs. 54G-54H). Altogether, V4-ELP-EEP13 demonstrates robust delivery efficacy across various primary cell types, including cells within hematological lineages.

[0381] The ability to facilitate Cre protein delivery was also examined in Sca-1+ primary bone marrow cells isolated from Ai9 mice. V2-ELPs were functionalized with AF633-labeled Cre and an anti-CD117 mAb via sortase conjugation (FIG. 31A). CD117 targeting enhanced Cre delivery to Lin-Sca1+ HSPCs populations (45% vs.18%) (FIGs. 30, 31B). V3-ELPs bearing Cre and containing penetratin enabled expression of tdTomato+ in ~2% in Lin- Sca1+c- Kit+ (LSK) HSPCs (FIG. 5F) with some dose-dependent cytotoxicity observed (FIG. 28F).

[0382] The ability of ENTER to facilitate delivery of transcription factors was assessed; transcription factors represent another class of proteins of therapeutic interest due to their ability to reprogram cell state.74For example, tumor-associated macrophages have been reprogrammed towards an antitumor phenotype by delivery of mRNA-encoded interferon regulatory factor 5 (IRF5), a proposed master transcription factor for pro-inflammatory M1 macrophages.75,97However, the delivery of protein-form IRF-5 has not been previously108 / 179B0662.70120WO00achieved and could be advantageous compared to mRNA due to improved cargo stability, dosing accuracy and its insensitive nature to the translational activity of targeted cells. Motivated by this, V4-ELP-EEP13 was evaluated for its ability to deliver IRF5 protein to primary mouse macrophages in vitro. V4-ELP-EEP13 effectively enabled the internalization of AF488-labeled IRF5 into murine peritoneal macrophages after 2 hours (FIG. 39I), supporting a synergistic uptake of IRF-5 stimulated by ELP nanoparticles. Macrophages were then treated with V4-ELP-EEP13 alone, IRF5 alone, or both for 48 hours. All recombinant proteins went through endotoxin removal to avoid endotoxin-stimulated gene expression. Expression of Ifng, Il12b, and Il1b, all genes under IRF5 regulation, significantly increased after co-treatment with IRF5 and V4-ELP-EEP13 (FIGs. 39J-39K). Limited increases in cytokine expression were observed following treatment with V4-ELP-EEP13 or recombinant IRF5 alone, suggesting enhanced production is related to IRF5 transcriptional activity enabled by ELP-mediated cytosolic entry. Evaluation of ELP micelles / nanoparticles for in vitro siRNA delivery

[0383] Cationic polymeric micelles have been widely explored as delivery vectors for nucleic acids65, including siRNA and pDNA cargo.66–69However, achieving high delivery efficacy remains a major barrier to clinical translation. It was hypothesized that V4-ELPs containing a cationic C-terminal EEP module would effectively encapsulate siRNA through combining electrostatic and physical complexation (FIG. 6A). Importantly, the combination of both electrostatic interactions and phase separation driven micelle assembly has been previously demonstrated to stabilize micelles in ion-rich environments.70It was postulated that the EEP domain would remain effective as an endosomolytic agent following pH- induced ELP disassembly within the endosome, enabling potent RNA delivery.

[0384] V4-ELP-EEP13 (SEQ ID NO: 79), V4-ELP-[EEP13]2(SEQ ID NO: 80), and V4- ELP-[EEP13]3 (SEQ ID NO: 81) were assessed for their ability to complex siRNA. All three ELPs displayed effective dose-dependent complexation of siRNA via gel shift and RiboGreen RNA binding dye exclusion assays (FIG. 6B). Near 100% encapsulation efficiency was observed at a V4-ELP protein polymer:siRNA molar ratio above 10, with the trimeric EEP13 variant displaying more efficient complexation than the monomeric or dimeric variants, as depicted in the gel shift assay. After siRNA complexation, V4-ELP- EEP13 and V4-ELP-[EEP13]2 nanoparticles showed a modest (~10-20 nm) increase in size (FIG. 6C). This increase in size was accompanied by a decrease in zeta potential (FIG. 6D).109 / 179B0662.70120WO00

[0385] The ability of V4-ELPs to enable intracellular siRNA delivery to HeLa cells was assessed. V4-ELP-EEP13 (SEQ ID NO: 79), V4-ELP-[EEP13]2 (SEQ ID NO: 80), and V4- ELP-[EEP13]3 (SEQ ID NO: 81) enabled rapid delivery, with nearly an order of magnitude greater uptake as compared to a lipid- based transfection reagent (RNAiMAX) (FIG. 6E). To evaluate functional siRNA delivery, V4-ELP-EEP13 series constructs were used to deliver an siRNA mediating GFP knockdown in HeLa cells expressing a destabilized GFP (HeLa- d2eGFP). Both the dimeric and trimeric constructs displayed high potency relative to the monomeric construct. The dimeric and trimeric constructs enabled 80-85% GFP knockdown at an ELP concentration of 1 µM with 100 nM siGFP, following treatment for 24 hours with minimal cytotoxicity (FIG. 6F), and the dimeric construct displayed mediated up to 60% GFP knockdown at an ELP concentration of 5 μM with 100 nM siGFP following treatment for 24 hours, with minimal cytotoxicity (FIG. 6F-6G). Interestingly, the efficacy decreased significantly with increasing ELP treatment concentration with the dimeric construct, while a positive dose-response relationship was observed with the monomeric construct. This effect may be related to the effect of these constructs on cell viability; both V4-ELP-[EEP13]2 (SEQ ID NO: 80) and V4-ELP-[EEP13]3 (SEQ ID NO: 81) displayed notable cytotoxicity at the highest tested concentrations of 10 and 20 µM (~20-40% loss in cell viability, FIG. 6G). All three constructs showed dose-dependent efficacy in an siGFP dose titration study, with V4- ELP-EEP13 (SEQ ID NO: 79) displaying the highest efficacy at the tested ELP concentration (10 µM) across all doses (FIGs. 33A-33B). Alternatively or additionally, this effect may be related to the efficiency of siRNA release from the particles. At high ELP concentrations, an excess of EEP-associated positive charge in V4-ELP-[EEP13]2 relative to the V4-ELP-EEP13 may be detrimental for intracellular decomplexation, reducing delivery efficiency. Notably, V4-ELP-EEP13 (SEQ ID NO: 79) also effectively facilitated delivery of a GAPDH-targeted siRNA into Ai9-derived murine lung fibroblasts, inducing ~95% knockdown of mRNA expression comparable to RNAiMAX (FIG. 40A). Collectively, these data demonstrate that ELPs serve as an effective vehicle for in vitro siRNA delivery.

[0386]

[0387] To interrogate the endocytic pathways facilitating ELP nanoparticle internalization, HEK293 cells were pretreated for 1 hour with chemical inhibitors known to primarily affect specific pathways: chlorpromazine (clathrin-mediated endocytosis, CME), nystatin (caveolin and clathrin independent carrier pathways), Dynasore (dynamin-dependent pathways, including CME), and amiloride (macropinocytosis)82, and were then treated with Alexa Fluor 647-labeled siRNA complexed with V4-ELP-EEP13 (SEQ ID NO: 79) series110 / 179B0662.70120WO00constructs for 2 hours. Of the four inhibitors assessed, both chlorpromazine and Dynasore resulted in a clear reduction in nanoparticle uptake (FIG. 40B). This finding suggests that a clathrin- and dynamin-dependent endocytic pathway is central to ELP nanoparticle internalization in HEK293 cells. In vitro and ex vivo mRNA delivery

[0388] To further establish ELPs as a platform for therapeutic macromolecule delivery, their ability to deliver protein-encoding mRNA was investigated. Given that V4-ELPs were effective in encapsulating siRNA through electrostatic and physical entrapment, it was hypothesized that mRNA would be complexed in a similar manner. The mRNA binding capacity of V4-ELP- EEP13 (SEQ ID NO: 79), V4-ELP-[EEP13]2(SEQ ID NO: 80), and V4- ELP-[EEP13]3(SEQ ID NO: 81) was initially assessed through gel shift and Ribogreen assays. Cre mRNA (1335 nt) was added to V4-ELPs at varying molar ratios. As expected, higher ELP:mRNA molar ratios were required for effective complexation as compared to ELP- siRNA complexation, given the larger size of the mRNA cargo (150 vs 25 molar ratios for V4- ELP-EEP13 complexation of Cre mRNA and siRNA, respectively) (FIG. 7A). The size and zeta potential of ELP-mRNA complexes were assessed to determine the ability of ELPs to effectively encapsulate mRNA. The size of V4-ELP-EEP13 series nanoparticles increased between 5-20 nm following Cre mRNA complexation, with the largest size increase being observe with the monomeric construct (FIG.7B). The zeta potential of V4-ELP-EEP13 micelles following mRNA complexation was unchanged (FIG.7C), indicative of efficient encapsulation and supporting that the majority of mRNA is shielded in the core.

[0389] The ability of V4-ELP-EEP13 series constructs to facilitate mRNA delivery was investigated through the delivery of mRNA-encoded Cre recombinase to HEK293-RFP cells. To assess whether the efficacy of ELP-EEPs is cargo dependent, ELPs containing EEPs generated through three generations of iterative design were assessed for their ability to deliver Cre mRNA to HEK293-RFP cells and compared to their Cre protein delivery efficiency (FIG. 36B and FIG. 7G). Consistently, both V4-ELP-EEP13 (SEQ ID NO: 79) and -EEP14 (SEQ ID NO: 82) significantly outperformed V4-ELP-S10 (SEQ ID NO: 84). A strong correlation was noted for protein and mRNA delivery (R = 0.9062, FIG. 7G) with EEP13 identified as most effective, despite the disparate nature of the two cargos. This suggests the existence of a common mechanistic pathway by which macromolecule endosomal escape is triggered, independent of the cargo’s physiochemical properties.111 / 179B0662.70120WO00

[0390] In a follow-up assessment of V4-ELP-EEP13 (SEQ ID NO: 79), a maximum % RFP+ of 86% was observed following treatment with 10 µM V4-ELP-EEP13 (SEQ ID NO: 79) (FIG. 7D). Consistent with the results observed with siRNA cargo, V4-ELP-[EEP13]2 (SEQ ID NO: 80) outperformed V4- ELP-EEP13 (SEQ ID NO: 79) at lower ELP concentrations (1 and 2 µM), but underperformed at higher concentrations. This effect may be due to less efficient release of RNA cargo as a result of higher binding affinity exhibited by the multimeric constructs. An increase in cytotoxicity was also observed with increased EEP13 repeat length, though all constructs displayed limited toxicity at the most effective treatment concentrations (< 10 µM, FIG. 7E) Notably, V4-ELP-EEP13 (SEQ ID NO: 79) enabled RFP+ expression at a level comparable to or greater than that achieved by MessengerMAX, a commercial lipid-based transfection agent, with significantly less observed cytotoxicity (FIGs. 34A-34B). Similarly, V4-ELP-EEP13 (SEQ ID NO: 79) performed comparably to Lipofectamine in delivering plasmid DNA-encoded Cre recombinase to HEK293-RFP with minimal cytotoxicity (FIG. 41A, FIG. 57C).

[0391] V4-ELP-EEP13 (SEQ ID NO: 79) also demonstrated effective Cre mRNA delivery to primary Ai9 lung fibroblasts in a dose-dependent manner, outperforming MessengerMAX at ELP concentrations greater than 10 µM (57.1% vs.39.9%, FIG. 7F). Interestingly, while cytotoxicity was observed on increasing ELP dose, cell viability was rescued by increasing mRNA concentration (FIG. 35). This suggests that increasing mRNA concentration reduces the presence of free, non- micelle associated, cell penetrating, albeit cytotoxic, ELP-EEP protein polymers.

[0392] In demonstrating that V4-ELP-EEP13 (SEQ ID NO: 79) effectively delivered both Cre protein and mRNA, 30 V4-ELPs containing EEPs generated through three generations of iterative design were assessed for their ability to deliver Cre mRNA and protein to HEK293- RFP cells (FIGs. 36A-36B). A strong correlation was noted for protein and mRNA delivery (R = 0.82, FIG. 7G) with EEP13 identified as the most effective, despite the disparate nature of the two cargos. This suggests the existence of a common mechanistic pathway by which macromolecule endosomal escape is triggered, irrespective of the cargo’s physicochemical properties. Altogether, these results indicate that without modification, the same ELP constructs can be effectively used for protein, siRNA, and mRNA delivery in vitro.

[0393] Next, the ability of V4-ELP-EEP13 (SEQ ID NO: 79) to mediate mRNA- and pDNA-encoded Cas9 delivery as a representative therapeutic gene editor cargo was assessed. SpCas9 mRNA or pDNA were co-complexed with a chemically modified sgRNA targeting HPRT1 at varying mRNA / pDNA:sgRNA weight ratios. In both formats, a weight ratio of 2:1112 / 179B0662.70120WO00facilitated improved delivery efficiency, resulting in 26% and 29% editing efficiency for mRNA and pDNA cargo respectively (FIGs. 41B-41C). Editing efficiency improved with both increased ELP concentration, noted in the mRNA delivery study, as well as nucleic acid concentration, noted in the pDNA delivery study. V4-ELP-EEP13 (SEQ ID NO: 79) also demonstrated effective Cre mRNA delivery to both primary Ai9 lung fibroblasts and macrophages in a dose-dependent manner, facilitating 54% and 56.3% tdTomato expression respectively at optimal dosing. While some dose-dependent cytotoxicity was observed in treated fibroblasts (~20% reduction in viability at the most efficacious dose), minimal toxicity was observed in treated macrophages (FIGs. 57D-57E). Altogether, these experiments demonstrate the ability of V4-ELP-EEP13 (SEQ ID NO: 79) to deliver therapeutically relevant genetically encoded cargo to diverse cell types, including primary cells. ELP delivery of genome editors and transcription factors

[0394] Gene disruption or repair via intracellular delivery of gene editing agents, such as Cas nuclease, base editors, and prime editors holds great promise as a therapeutic strategy for many diseases.72Delivery of gene editors in protein form rather than mRNA or DNA has the potential to reduce off-target editing by limiting the lifetime of the editing agent within treated cells.73The ability of V4-ELP-S10 (SEQ ID NO: 84) to deliver free SpCas9 along with an encapsulated sgRNA targeting HPRT1 (hypoxanthine-guanine phosphoribosyltransferase 1) to HEK293 cells was assessed (FIG. 8A). Modest editing efficiency (~20%) was observed at the highest doses tested (100 nM sgRNA / Cas9, 8 µM ELP, FIG. 8B) establishing the potential of ELPs to serve as a delivery vehicle for gene editors.

[0395] Transcription factors represent another class of proteins of therapeutic interest due to their cell polarizing activity.74For example, tumor-associated macrophages have been reprogramed towards an antitumor phenotype by delivery of mRNA-encoded interferon regulatory factor 5 (IRF5), a transcription factor that regulates the pro-inflammatory M1 polarization of macrophages.75V4-ELP-EEP13 (SEQ ID NO: 79) was evaluated for its ability to deliver IRF5 to primary mouse macrophages ex vivo (FIG. 8C). Peritoneal Ai9 macrophages were treated with V4-ELP-EEP13, soluble IRF5, or both for 48 hours. Expression of Ifng and Il1b, both under the control of IRF5-bound promoters, increased in a dose-dependent fashion after co-treatment with IRF5 and V4-ELP-EEP13. (FIGs. 8D-8E). Very limited increases in cytokine expression were observed following treatment with V4- ELP-EEP13 (SEQ ID NO: 79) or recombinant IRF5 alone, suggesting enhanced production113 / 179B0662.70120WO00is related to IRF5 transcriptional activity enabled by ELP-mediated cytosolic entry and nuclear localization. Collectively, these data support that ELP micelles can serve as an effective delivery vector for therapeutic gene editors and transcription factors. In vivo delivery of Cre recombinase via ELP nanoparticles

[0396] S10-V3-ELP (SEQ ID NO: 60) was administered with Cre protein, either free or micelle bound, via a single intradermal injection to Ai9 mice. Skin was harvested 72 hours later and lymphoid cells (CD45+CD90.2+), fibroblasts (CD45-CD90.2+), and myeloid cells (CD45+CD90.2- CD11b+) were isolated for analysis via flow cytometry (FIG. 37). tdTomato expression was observed in 10% of lymphoid cells, 2.5% of fibroblasts, and 6% of myeloid cells (FIGs. 9A-9C, 38) with comparable editing efficiency for the delivery of free or micelle bound Cre in association with S10-V3-ELP (SEQ ID NO: 60). ENTER-mediated delivery of Cre recombinase to lung epithelium in vivo

[0397] To validate the utility of ENTER for therapeutic delivery in vivo, V4-ELP-EEP13- mediated delivery of Cre protein to the airway epithelium lining was assessed in mice. Ai9 mice received three daily doses of V4-ELP-EEP13 (SEQ ID NO: 79) nanoparticles (100 μM, ~24 mg / kg) co-formulated with Cre recombinase protein (50 μM, ~8 mg / kg) or Cre protein alone via intranasal instillation; lungs were harvested for image-based evaluation of tdTomato expression 5 days following the final treatment (FIG. 42A). Compared to the group treated with Cre protein alone, mice co-treated with Cre protein and V4-ELP-EEP13 (SEQ ID NO: 79) displayed significantly higher editing of the bronchial epithelium in both large airways (FIG. 42B and FIG. 42D) and small airways (FIG. 42C and FIG. 42E). 25.4 ± 3.7% of epithelial cells in the large airways of co-treated mice were tdTomato+ (8.9 ± 1.8% in mice treated with Cre protein alone), while 9.2 ± 0.8% of epithelial cells in the small airways of co-treated mice were tdTomato+ (1.8 ± 0.3% in mice treated with Cre protein alone). ENTER can thus be leveraged for intracellular delivery of therapeutic protein cargo in vivo. Discussion

[0398] A non-viral, recombinant protein nanoparticle system (Elastin-based Nanoparticles for Therapeutic delivERy (ENTER)) was developed as a versatile platform for effective cytosolic delivery of multiple types of biomacromolecules of therapeutic interest, including siRNA, mRNA, pDNA, RNPs, and protein cargo. Prior success in cytosolic delivery of mRNAs, proteins, and RNPs have largely been achieved via lipid nanoparticles and virus-like particles.98,99114 / 179B0662.70120WO00This ELP-based platform is believed to be the first use of non-viral recombinant protein nanoparticles for intracellular mRNA and protein delivery. Notably, the ability to delivery diverse biomacromolecules is achieved without the need for cargo-specific vehicle modifications, evidence of the versatility of the platform.

[0399] The fourth-generation multiblock micelle copolymer design differs in multiple ways from previous ELP systems utilized for cytosolic delivery of nucleic acids, such as pDNA38,85,86, siRNA87, and CpG88. Unlike the amorphous polyion complexes (PICs) formed by these systems, ENTER displays an organized micellar structure. The hydrophilic shell ensures well-controlled surface chemistry, allowing for flexibility in modification with functional modalities like targeting moieties through sortase-mediated conjugation or genetic fusion. Secondly, the core-facing cationic EEP remains shielded, facilitating the encapsulation of nucleic acid cargo in the core. Micelle assembly is driven by a combination of hydrophobic and electrostatic interactions, strengthened by cysteine crosslinking. Furthermore, the presence of histidines in the ELP hydrophobic block facilitates micelle disassembly in acidic environments, enabling the exposure of EEP and cargo release only in acidic late endosomes. This feature contributes significantly to low cytotoxicity and high delivery efficiency. Notably, ENTER shares similarities with other state-of-the-art delivery systems, such as lipid nanoparticles, viruses, and virus-like particles, each exhibiting organized structures with surface modules separated from core-encapsulated cargo, accompanied by a triggered release mechanism. In contrast, many existing polymer, peptide, and protein based delivery systems lack such organized structure and release mechanisms, limiting their potency and clinical translation. Accordingly, ENTER represents an advancement in engineering the structural complexity and responsiveness required for potent macromolecule delivery.

[0400] In addition, the effectiveness of ENTER is significantly enhanced by the identification of novel and potent EEPs. Through an iterative process, ELP nanoparticles were designed to facilitate nucleic acid encapsulation and effective intracellular delivery. By in silico screening of peptide databases using a predictive model of membrane disruptive activity, a novel EEP (EEP13, SEQ ID NO: 14) was discovered, which outperformed S10 (SEQ ID NO: 36) when incorporated as a component of ELP nanoparticles14, a high performing cell penetrating, endosomolytic, shuttle peptide, in both mRNA and protein delivery, while maintaining high cell viability. These findings establish ELPs as a versatile delivery platform for diverse macromolecular cargo of therapeutic interest.115 / 179B0662.70120WO00

[0401] Several important benefits are derived from the recombinant fusion of membrane disruptive EEPs to ELPs, when compared to cell penetrating or endosomolytic peptides used in isolation as shuttle peptides for macromolecular delivery. One benefit is the effect of EEP multivalency on delivery potency. ELP-S10 displayed nearly 20-fold higher potency in Cre protein delivery as compared to S10 alone (SEQ ID NO: 36) (FIG. 2F), with potency further enhanced by the incorporation of EEP dimers at the C-terminus of the ELP protein polymer (FIG. 4G). A dimer ([EEP13]2(SEQ ID NO: 80), 87 AAs) has a length that would make its manufacturing by solid phase peptide synthesis challenging. Given that the mechanism of action of such amphipathic α-helical peptides is related to their ability to intercalate, disrupt, and induce fusion of lipid membranes54, increased local endosomal EEP concentration afforded by ELP micelle multivalency is presumed to directly enhance this activity.

[0402] Secondly, ELP-EEP fusion dramatically reduced EEP-associated cytotoxicity. Standalone EEP-mediated transfection often requires peptide removal from cell culture media after a brief incubation due to high cytotoxicity, limiting the in vivo applicability of this approach.76In the ELP-EEP system, prolonged peptide incubation was conducted in various cell types without significant cytotoxicity. Without wishing to be bound by any particular theory, several factors may contribute to this phenomenon. First, unlike free cell penetrating or endosomolytic peptides, which directly interact with the plasma membrane, ELP nanoparticles require endocytosis and acidification- induced disassembly to expose the encapsulated EEP. The kinetics of such exposure and the nature of endosomal membrane disruption likely contributes to substantially less cytotoxicity irrespective of the efficiency of cargo release into the cytosol.

[0403] In addition, segregation of the EEP from the surface of ELP nanoparticle has established a unique system which can be used to evaluate endosomolytic potential of membrane disruptive peptides in isolation from cell uptake or cytotoxicity. This feature enabled the discovery of more potent EEPs that would not emerge from a screen of free peptides due to their cytotoxic potential. Overall, significant advantages in potency and cytotoxicity, in addition to the capacity to easily incorporate targeting specificity, establishes the promise of ELP nanoparticles for effective intracellular delivery of a variety of macromolecules.

[0404] V4-ELP-EEP13, demonstrated an EC50in the submicromolar to low single-digit micromolar range for delivering protein and nucleic acid cargos into various cell lines and primary cells. This represents a significant improvement in efficacy compared to previous reports, in which standalone endosomolytic peptides were functional in the 10 to 40 μM116 / 179B0662.70120WO00range.14,100Notably, overcoming the high molar concentration requirement for protein and peptide-based delivery systems addresses a significant bottleneck that limits in vivo and clinical translation of such technologies.

[0405] Moreover, V4-ELPs are comparable or even outcompete commonly used lipid-based transfection reagents like Lipofectamine 3000, RNAiMAX, and MessengerMax under a variety of conditions in the delivery of protein, siRNA and mRNA, respectively. V4-ELP- EEP13 in particular exhibited comparable or superior performance in delivering proteins and nucleic acids. The enhanced efficiency of V4-ELP is especially evident when compared to Lipofectamine for Cre protein delivery. This is likely due, in part, to the positive surface charge of Cre, creating repulsive forces between the protein and positively charged lipids leading to inefficient encapsulation. Unlike the charge-dependent nature of Lipofectamine, V4-ELP appears to potentiate cellular uptake of both positively charged Cre and negatively charged IRF-5 as a consequence of nanoparticle stimulated collaborative uptake or macropinocytosis.15,77This highlights the charge-insensitive nature of the V4-ELP-EEP system and its ability to deliver positively charged cargo, which remains a challenge for lipid nanoparticles and other nonviral delivery systems. V4-ELPs were also effective as a delivery system for a variety of cell lines and primary cells, including hematopoietic stem cells, macrophages, and primary T cells. This is significant as transfecting these cells has historically posed a challenge for non-viral delivery systems. These observations suggest that the mechanisms responsible for V4-ELP uptake and endosomal disruption are conserved across different cell types.

[0406] Multiblock micelle copolymer designs in which the cationic domain is shielded from the surface offer a number of key advantages.66,67Such designs reduce cytotoxicity, enable nucleic acid encapsulation, improve particle stability through combining hydrophobic and electrostatic assembly, and provide great flexibility for engineering the nanoparticle surface. ELP nanoparticles were designed for efficient delivery of siRNA and mRNA (FIG. 6C and FIG. 7B). While the ability of ELPs to mediate intracellular protein delivery has not been previously reported, prior efforts directed at the delivery of nucleic acids, including pDNA33,78,79, siRNA80, CpG81have been in the form of amorphous polyion complexes (PICs). Such complexes do not display organized structure or the stability of the V4-ELP nanoparticles.

[0407] The uniqueness of ELPs as a recombinant protein offers several advantages over other nanoparticle systems, such as LNPs or synthetic polymeric nanoparticles. The genetically defined composition of ELPs enables greater manufacturing precision when117 / 179B0662.70120WO00compared to more dispersed chemically synthesized materials. This genetically defined composition also creates an opportunity for precise incorporation of additional peptide domains of interest, such as EEPs. In contrast to lipid and synthetic polymer formulations, which often require re-optimization with new cargo types82, the data suggest that V4-ELPs use a common endosomolytic mechanism with high efficacy for both RNA and protein cargo (FIG. 7G).

[0408] Outside of ELPs and other recombinant protein polymers, such as silk-like peptides83, computationally designed protein nanoparticles have emerged as a related class of delivery vehicle.84While designs have been reported which enable RNA encapsulation85and pH- induced disassembly86, both functions useful for a macromolecule drug carrier, such systems have largely been used for extracellular protein delivery or presentation such as vaccination, largely due to the challenges associated with endosomal escape.87,88This investigation demonstrates that membrane disruptive amphipathic α-helical peptides remain highly functional even when appropriately incorporated within an ELP suggesting their ability to enable endosomal escape of a variety of other nanoparticles.

[0409] In summary, these studies introduced a novel recombinant protein-based delivery system for diverse therapeutic macromolecules, including siRNA, mRNA, plasmid DNA, RNPs, and proteins. ELPs and ENTER represent a promising in vivo and ex vivo delivery platform for various cargos across numerous therapeutic indications. Methods Plasmid Cloning

[0410] DNA sequences coding ELP were codon optimized for E. Coli expression, synthesized, and cloned into previously reported pQE-80L backbone using commercial services from GenScript.34Cre and Cas9 with LPETG tag and GGS linker were initially subcloned in house using previously reported genome editor coding plasmids.35Cre with variable cleavable linkers were cloned using GenScript. A constitutively active mutant of human IRF5 splice variant 5 carrying five serine to aspartic acid mutation (IRF-5 / 4D) with furin cleavable linker and LPETG tag was codon optimized for E. Coli expression, synthesized, and cloned into pET backbone by GenScript.92Protein expression and purification

[0411] All plasmids were transformed into E. coli strain BL21(DE3) (Invitrogen, Thermo Fisher Scientific; C601003) for expression. E. coli were grown in Luria-Bertani medium supplemented with 100 ug / ml of ampicillin at 37 °C with continuous shaking at 225 rpm till118 / 179B0662.70120WO00OD600reaching 0.6 - 0.8. For all ELP expression, 1mM of Isopropyl ß-D-1- thiogalactopyranoside (IPTG) (UBPBio; P1010100) were added to induce expression for another 4 hours at 37C. Harvested E. coli pellets were resuspended in denaturing buffer containing 8 M Urea, 1 M NaCl, and 10mM TCEP in PBS at pH=7, (GoldBio; 51805459) with Protease Inhibitor Cocktail (Sigma-Aldrich; P8849). Cells were lysed by sonication on ice and cell lysates were incubated at 4C overnight to ensure complete solubilization of ELPs from inclusion body. The lysates were then centrifuged at 20,000 g for 30 min and 4 °C and soluble supernatant were collected. His-tag carrying ELP were purified by Immobilized Talon Metal Affinity chromatography (IMAC, Takara Bio; 635503) under denaturing condition following commercial protocol. This fraction was used in high throughput screening of large batches of ELP-EEPs. For thorough characterization of lead ELPs, an additional round of inverse thermal cycling was carried as previously described to improve purity.17Briefly, ELPs were precipitated in hot cycle by incubating with 3M NaCl at 30 °C for 50 minutes. Following centrifugation, ELP pellets were resolubilized in PBS with 10mM DTT at 4C during in the cold cycle. Un-resoluble impurities were removed by centrifugation at 20,000g at 4 °C for 30 minutes. For final peptides in the ELP-EEP13 series, cationic exchange chromatography (HiTrap SP HP, Cytiva) was utilized as the second step of purification following manufacturer’s protocol under denaturing condition. ELP-EEP13 and ELP-(EEP13)2 were eluted at about 200mM and 300 mM NaCl concentrations respectively. All final ELP elutes were thoroughly dialyzed in deionized water, lyophilized, and stored at -20 °C till usage.

[0412] For the expression of all variants of CRE and Cas9, 0.5 mM IPTG was added at OD 0.6 to 0.8. For IRF-5 expression, 0.1 mM IPTG and 3% ethanol were added at OD 0.6 to 0.8 to improve the solubility of the recombinant protein. Expression was carried overnight at 20 °C. Cell pellets were resuspended in lysis buffer containing 1M NaCl, 20% glycerol, 10mM TCEP in 50 mM Tris-HCl at PH-8.0 followed by similar sonication and IMAC purification protocol as described above under non-denaturing condition. CRE and Cas9 were further purified by SP column and IRF-5 by Q column following commercial protocols. Protein products were concentrated using centrifugal filter to above 100 μM. Small aliquots were snapped frozen in liquid nitrogen to avoid freeze and thaw cycles and stored in -80 °C till usage.

[0413] Before primary cell treatment and in vivo applications, Tris based buffer in protein cargo samples was exchanged to HBSS to reduce potential buffer associated toxicity. Both ELP and protein cargo samples were further purified by Pierce High-Capacity Endotoxin Removal Resin (Fisher) and sterilized by 0.22 μM filter. Final endotoxin level in a typical 8119 / 179B0662.70120WO00µM sample was quantified by Pyrogent-5000 kinetic turbidimetric LAL assay (Lonza) to be less than 5 EU / ml. Sortase-mediated protein conjugation to ELP and purification

[0414] Cre, Cas9 and antibody carrying C-terminus LPETG tag were incubated with ELP carrying N-terminus tri-glycines at indicated concentrations in figures together with 1 µM evolved sortase A in HBSS buffer with calcium and magnesium and incubated at room temperature for 1 hour with agitation.36Unconjugated Cre and antibody were removed by centrifugal filter with 100KD molecule cut off (Amicon, Sigma). Six filtration cycles were carried at 37 °C which allows the stable formation of micelle nanoparticles and large size difference compared to unconjugated proteins. Unconjugated Cas9 was removed by glycerol gradient ultracentrifugation. Briefly, solution was layered on top of a glycerol gradience from 5 to 55% and centrifuged at 39,000RPM in a Beckman XPN-80 ultracentrifuge for 4 hours at room temperature (about 22 °C). Unconjugated Cas9 presented in 5 to 15% glycerol fractions whereas formed Cas9-ELP nanoparticles resided in 35 to 45% fractions. Antibody modification, conjugation, and targeted delivery

[0415] Anti-CD117 antibody was azide-modified using the SiteClick Antibody Azido Modification kit (Invitrogen, Thermo Fisher Scientific; S10900) according to the manufacturer’s protocol. Azide modified antibody was successfully conjugated to ELP via both sortase reaction and chemical conjugation. For sortase mediated conjugation, KLPETGG (SEQ ID NO: 118) peptide with N-terminus amine acetylated was commercially synthesized (GenScript) and the only primary amine on the side chain of lysine was coupled with DBCO-NHCO-PEG13-NHS ester linker (BroadPharm). Unreacted linkers were removed by HPLC and correct molecular weight of DBCO-NHCO- PEG13-amide-KLPETGG was confirmed by MALDI mass spectrum. DBCO modified KLPETGG (SEQ ID NO: 118) linker was then clicked onto azide-modified Anti-CD117 antibody and unreacted excessive linkers were removed by 10KD centrifugal filter. The molecular weight shift of linker modified antibody heavy chain was confirmed using SDS-PAGE. Ab carrying LPETG tag was further conjugated to ELP via sortase reaction as described above. For chemical conjugation, DBCO- NHCO-PEG13-NHS ester linker (BroadPharm) was incubated with V1-ELP-A633, which only carries one free amine at N-terminus for bio-orthogonal conjugation. Unreacted linkers were removed via 10KD centrifugal filter. Purified DBCO-Diblcok-cysteine-A633 was then incubated with azide modified CD117(2B8) antibody at room temperature overnight with constant agitation. Unreacted Ab was removed via 100KD centrifugal filter at 37 °C.120 / 179B0662.70120WO00

[0416] For targeting study in P815 cells, 4uM of V1-ELP-A633 with or without CD117 antibody conjugation were added to 200,000 P815 cells in full medium (DMEM with 10%FBS) at 37 °C for 0.5, 1, 2, 4, 24 hours. Medium were removed and cells were washed with PBS containing 1 unit of heparin. Part of samples were immediately examined by flow cytometry for cell associated A633 signals. Part of samples were fixed in PBS containing 4% formaldehyde and 2ug / ml Hoechst 33342 (Fisher) for 15mins at RT, washed and aliquoted to slides, mounted in ProLong Gold antifade reagent (Cell Signaling) and examined by Zeiss TIRF / LSM 710 confocal with 63 oil lens. Diameter and zeta potential measurement

[0417] The diameter and zeta potential of ELP micelles was evaluated using a ZetaSizer NanoSeries (Nano ZS, Malvern Instruments, UK). ELP micelles were formed in 1x PBS. Dynamic light scattering measurements were recorded at a scattering angle of 173°. Samples were incubated at 37 °C for five minutes prior to measurement to ensure micelle formation and recapitulate cell treatment conditions. Three measurements with at least 10 runs were recorded for each sample. ELP-RNA nanoparticle formation

[0418] To form ELP-RNA NPs, siRNA or mRNAs were added to ELPs at varying molar ratios. An ELP stock concentration of 200 µM was generally used. For siRNA particle formation, the NPs were incubated at room temperature with continuous shaking at 500 rpm for 30 minutes. siRNAs u...

Claims

CLAIMS What is claimed is:

1. A compound comprising an elastin-like polypeptide chemically or recombinantly conjugated to one or more endosomal escape peptides.

2. The compound of claim 1, wherein the elastin-like polypeptide comprises a hydrophilic block fused to a hydrophobic block.

3. The compound of claim 1, wherein the elastin-like polypeptide comprises a hydrophilic block fused to a hydrophobic block separated by a crosslinking block.

4. The compound of any one of claims 1-3, wherein the hydrophilic block has the sequence: [VPGXG]n(SEQ ID NO: 114) wherein: each instance of X is independently any amino acid except proline; and n is at least 5.

5. The compound of any one of claims 1-4, wherein the hydrophilic block has the sequence: [VPGXG]n (SEQ ID NO: 114) wherein: each instance of X is any amino acid except proline; and n is at least 5.

6. The compound of claim 4 or 5, wherein n is 30-70.

7. The compound of any one of claims 2-6, wherein the hydrophilic block has the sequence: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]n1(SEQ ID NO: 22) wherein: X1and X2are any amino acid except proline;145 / 179B0662.70120WO00each instance of X1is the same; each instance of X2is the same; X1and X2are different amino acids; and n1 = n / 5.

8. The compound of any one of claims 4-7, wherein each instance of X is independently E, V, A, G, S, or Q.

9. The compound of any one of claims 4-8, wherein each instance of X is independently G, S, or Q.

10. The compound of any one of claims 4-8, wherein each instance of X is independently E, V, A, or G.

11. The compound of any one of claims 4-8 or 10, wherein each instance of X is E, V, A, or G.

12. The compound of any one of claims 4-8, 10, or 11, wherein each instance of X is independently E or V.

13. The compound of any one of claims 4-8 or 10-12, wherein each instance of X is E or V.

14. The compound of any one of claims 2-8 or 10-13, wherein the hydrophilic block has the sequence: [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10 (SEQ ID NO: 23).

15. The compound of any one of claims 4-8, 10, or 11, wherein each instance of X is independently A or G.

16. The compound of any one of claims 4-8, 10, 11, or 15, wherein each instance of X is A or G.146 / 179B0662.70120WO0017. The compound of any one of claims 2-8, 10, 11, 15, or 16, wherein the hydrophilic block has the sequence: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10 (SEQ ID NO: 24).

18. The compound of any one of claims 4-9, wherein each instance of X is independently S or G.

19. The compound of any one of claims 2-9 or 18, wherein the hydrophilic block has the sequence: [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10(SEQ ID NO: 202).

20. The compound of any one of claims 4-9, wherein each instance of X is independently Q or G.

21. The compound of any one of claims 2-9 or 20, wherein the hydrophilic block has the sequence: [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10(SEQ ID NO: 215).

22. The compound of any one of claims 4-11, 15, 16, 18, or 20, wherein each instance of X is G.

23. The compound of any one of claims 2-11, 18, 20, or 22, wherein the hydrophilic block has the sequence: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10(SEQ ID NO: 216).

24. The compound of any one of claims 3-23, wherein the crosslinking block comprises one or more cysteine (C) residues.

25. The compound of any one of claims 3-24, wherein the crosslinking block has the sequence –CCCCGGG– (SEQ ID NO: 19).

26. The compound of any one of claims 3-24, wherein the crosslinking block has the sequence –[CAP]5– (SEQ ID NO: 125).147 / 179B0662.70120WO0027. The compound of any one of claims 3-24, wherein the crosslinking block has the sequence –[CEAAAK]5– (SEQ ID NO: 126).

28. The compound of any one of claims 3-24, wherein the crosslinking block has the sequence –C5G3– (SEQ ID NO: 127).

29. The compound of any one of claims 2-28, wherein the hydrophobic block comprises one or more tyrosine (Y) residues.

30. The compound of any one of claims 2-29, wherein the hydrophobic block comprises one or more cysteine (C) residues.

31. The compound of any one of claims 2-30, wherein the hydrophobic block has the sequence: (Z1PGZ2G)m (SEQ ID NO: 25) wherein: Z1and Z2are each independently a hydrophobic amino acid; and m is an integer from 5 to 400.

32. The compound of claim 31, wherein each instance of Z1is independently I, V, A, or L.

33. The compound of claim 31 or 32, wherein each instance of Z1is independently I or V.

34. The compound of any one of claims 2-33, wherein the hydrophobic block has the sequence: [(IPGZ2G)2(VPGZ2G)(IPGZ2G)2]m1 (SEQ ID NO: 26) wherein m1 is an integer from 1 to 80.

35. The compound of any one of claims 31-34, wherein each instance of Z2is independently C, H, Y, V, I, L, F, or W.148 / 179B0662.70120WO0036. The compound of any one of claims 31-35, wherein each instance of Z2is independently H, Y, V, I, L, F, or W.

37. The compound of any one of claims 31-35, wherein each instance of Z2is independently C, H, or Y.

38. The compound of any one of claims 31-37, wherein at least one instance of Z2is H.

39. The compound of any one of claims 31-38, wherein Z2is H.

40. The compound of any one of claims 31-38, wherein at least one instance of Z2is C.

41. The compound of any one of claims 31-39, wherein Z2is an aromatic amino acid.

42. The compound of any one of claims 31-38, 40, or 41, wherein at least one instance of Z2is Y.

43. The compound of any one of claims 31-37, 41, or 42, wherein Z2is Y.

44. The compound of any one of claims 2-36 or 41-43, wherein the hydrophobic block has the sequence: [(IPGVG)2(VPGYG)(IPGVG)2]15 (SEQ ID NO: 27) 45. The compound of any one of claims 2-43, wherein the hydrophobic block has the sequence: [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 28).

46. The compound of any one of claims 2-43, wherein the hydrophobic block has the sequence: [(IPGZ2G)2(VPGYG)(IPGZ2G)2]15 (SEQ ID NO: 128); wherein: each instance of Z2is independently C or H.149 / 179B0662.70120WO0047. The compound of claim 46, wherein three instances of Z2are C and the remaining instances of Z2are H.

48. The compound of claim 46, wherein five instances of Z2are C and the remaining instances of Z2are H.

49. The compound of claim 46, wherein seven instances of Z2are C and the remaining instances of Z2are H.

50. The compound of claim 46, wherein thirteen instances of Z2are C and the remaining instances of Z2are H.

51. The compound of any one of claims 1-43 or 46-50, wherein the hydrophobic block has the sequence: [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGH GIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPG HGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIP GHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 230); [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGH GIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPG HGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGCGIPGHGVPGYGIP GHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 231); [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGH GIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPG150 / 179B0662.70120WO00CGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIP GHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 232); or [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGH GIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPG YGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPG HGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPG CGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIP GHGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIP GHGIPGHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 233).

52. The compound of claim 1 or 3, wherein the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 217), wherein: each instance of X1is independently G, S, Q, V, or A; and each instance of X2is independently E or G.

53. The compound of claim 1, 3, or 52, wherein the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 33), wherein: each instance of X1is independently V or A; and each instance of X2is independently E or G.

54. The compound of claim 1, 3, or 52, wherein the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 33), wherein: X1is V or A; and151 / 179B0662.70120WO00X2is E or G.

55. The compound of any one of claims 52-54, wherein each instance of X1is V or each instance of X1is A.

56. The compound of any one of claims 52-55, wherein each instance of X2is E or each instance of X2is G.

57. The compound of claim 1, 3, or 52, wherein the elastin-like polypeptide comprises the sequence of: [(VPGX1G)(VPGX2G)(VPGX1G)(VPGX2G)(VPGX1G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 218), wherein: each instance of X1is independently G, S, or Q; and each instance of X2is G.

58. The compound of claim 57, wherein each instance of X1is G, each instance of X1is S, or each instance of X1is Q.

59. The compound of any one of claims 1, 3, or 52-56, wherein the elastin-like polypeptide comprises the sequence of: [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 34).

60. The compound of any one of claims 1, 3, or 52-56, wherein the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 113).

61. The compound of any one of claims 1, 3, 52, 57, or 58, wherein the elastin-like polypeptide has the sequence of: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 131).152 / 179B0662.70120WO0062. The compound of any one of claims 1, 3, 52, 57, or 58, wherein the elastin-like polypeptide has the sequence of: [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 219).

63. The compound of any one of claims 1, 3, 52, 57, or 58, wherein the elastin-like polypeptide has the sequence of: [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 220).

64. The compound of claim 1 or 3, wherein the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CAP]5– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 199).

65. The compound of claim 1 or 3, wherein the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CEAAAK]5– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 200).

66. The compound of claim 1 or 3, wherein the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–C5G3– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 201).

67. The compound of any one of claims 1, 2, 4-8, 10, 11, 15-17, 29-35, 37-43, or 46, wherein the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[(IPGZ2G)2(VPGYG)(IPG Z2G)2]15 (SEQ ID NO: 132); wherein: each instance of Z2is independently C or H.153 / 179B0662.70120WO0068. The compound of claim 67, wherein three instances of Z2are C and the remaining instances of Z2are H.

69. The compound of claim 67, wherein five instances of Z2are C and the remaining instances of Z2are H.

70. The compound of claim 67, wherein seven instances of Z2are C and the remaining instances of Z2are H.

71. The compound of claim 67, wherein thirteen instances of Z2are C and the remaining instances of Z2are H.

72. The compound of claim 67, wherein the elastin-like polypeptide has the sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 234); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 235); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG154 / 179B0662.70120WO00HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 236); or [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG] (SEQ ID NO: 237).

73. The compound of any one of claims 1-72, wherein the endosomal escape peptide is a peptide of about 10 to about 170 amino acids in length.

74. The compound of any one of claims 1-73, wherein the endosomal escape peptide is a peptide of about 10 to about 140 amino acids in length.

75. The compound of any one of claims 1-74, wherein the endosomal escape peptide is a peptide of about 10 to about 60 amino acids in length.

76. The compound of any one of claims 1-75, wherein the endosomal escape peptide is a peptide of at least 10 amino acids in length.

77. The compound of any one of claims 1-76, wherein the endosomal escape is a peptide of at least 30 amino acids in length.

78. The compound of any one of claims 1-77, wherein the endosomal escape peptide is a peptide comprising one or more amphipathic alpha helical motifs.155 / 179B0662.70120WO0079. The compound of any one of claims 1-78, wherein the endosomal escape peptide is a peptide comprising two amphipathic alpha helical motifs.

80. The compound of claim 78 or 79, wherein the amphipathic alpha helical motifs comprise a positively charged hydrophilic outer face and a hydrophobic outer face.

81. The compound of any one of claims 78-80, wherein the amphipathic alpha helical motifs are separated by a flexible peptide linker.

82. The compound of any one of claims 1-81, wherein the endosomal escape peptide comprises about 30% to about 65% of any combination of the amino acids A, C, G, I, L, M, F, P, W, Y, R, and V.

83. The compound of any one of claims 1-82, wherein the endosomal escape peptide comprises about 25% to about 45% of any combination of the amino acids K and R.

84. The compound of any one of claims 1-83, wherein the endosomal escape peptide has a net charge between about +10 and about +16.

85. The compound of any one of claims 1-84, wherein the endosomal escape peptide has a hydrophobic moment between about 6 and about 16.

86. The compound of any one of claims 1-85, wherein the endosomal escape peptide has a helical content between about 14% and about 72%.

87. The compound of any one of claims 1-86, wherein positively charged residues of the endosomal escape peptide comprise between about 40% and about 100% K.

88. The compound of any one of claims 1-87, wherein the endosomal escape peptide is conjugated to the N-terminus of the elastin-like polypeptide.

89. The compound of any one of claims 1-87, wherein the endosomal escape peptide is conjugated to the C-terminus of the elastin-like polypeptide.156 / 179B0662.70120WO0090. The compound of any one of claims 1-89, wherein the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 2); WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K (SEQ ID NO: 3); TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK (SEQ ID NO: 4); MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 5); LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 6); LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 7); RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K (SEQ ID NO: 8); KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 9); KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 10); KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 11); KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 12); KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 13); KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 14); KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 15); ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 16); KWKLARAFARAIKKLGGSGGGSYARALRRQARTG (SEQ ID NO: 36); RQIKIWFQNRRMKWKK (SEQ ID NO: 37); CKRKKRRQRRRG (SEQ ID NO: 38); RRRRRRRRR (SEQ ID NO: 39); AGYLLGKINLKALAALAKKIL (SEQ ID NO: 40); IWLTALKFLGKHAAKHEAKQQLSKL (SEQ ID NO: 41); and GLFDIIKKIAESF (SEQ ID NO: 42).157 / 179B0662.70120WO0091. The compound of any one of claims 1-89, wherein the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: RMKFNVWAWICVVKKKKFFPHWKIRYIIGRAWLL (SEQ ID NO: 133); DVHRRIWIILLPGHRIKFNWKQWKWRFEFEFKAVRK(SEQ ID NO: 134); KLVRYISYLKLRGGVKILLAKIKPFYEVKLKLKRLSLII(SEQ ID NO: 135); YFLARARARAILIKFFGKLKPFKAKKLAKQLKWVART(SEQ ID NO: 136); ANHKFGVYFGGSGGGSGVKWQVKFGVNHANHVKF(SEQ ID NO: 137); ANHGHKFGVKLNHGGSGGGSYFKFGKAPGVKW(SEQ ID NO: 138); GVGVREWKFGGSGGGSYYFGVKGHKKGVGV(SEQ ID NO: 139); GVKFGVGVKKYFKYFASGVKGVKYF(SEQ ID NO: 140); GVGVKYFKVAVKNHKPKWFSYKAAWKKFG(SEQ ID NO: 141); GNHKAYKWVRAPRTNHKHKKAVKWKHKPG(SEQ ID NO: 142); LKFWGASAPAPHKFAWKFGVGARASGHKLL(SEQ ID NO: 143); VKWKWQWGVKGAPGYFYLGFGVKTAWAPAWAW(SEQ ID NO: 144); FLKFGGSGGGSHKVKFGVVGNHKKAPVHKW(SEQ ID NO: 145); GVGASLGKQWKFGGSGGGSNHVKWFGVKPSKGHKGAWKWS(SEQ ID NO: 146); KPIQWGVGGSGGGSYLAPRTKKHVKFGVKRTHKFAP(SEQ ID NO: 147); WKWLKAPKPILKILGKPVKHKWKPAWKGKWQ(SEQ ID NO: 148); KSILKWGAKPKKALILASLGVKKLLAPSGAPW(SEQ ID NO: 149); GVKPAWRASLGGKHVKRLSGFHKPRTKAPSL(SEQ ID NO: 150); KGVKWLLKFGGSGGGSLILKFGKFGFKAPGAPGIL(SEQ ID NO: 151); AWQGVKGFKLAPKKGKPLGKPTKKKAHVRKL(SEQ ID NO: 152); FGAPSKHVKKILEWQKGGSGGGSWKILKFGGKPGVGF(SEQ ID NO: 153); EWKFGFGAGKSLKLGVVHVFGGSGGGSKILILKPVGAP(SEQ ID NO: 154); GFKQAPGVILAPRGFKWLIKLGKAKQKVKPFK(SEQ ID NO: 155); GVKPLKKCKPAILVAPKKGKQEWLKW(SEQ ID NO: 156); GVGKKVAWWQAQAKAPAWKGFGKWKKAKAAP(SEQ ID NO: 157); GIWKLGKAPKILGGSGGGSKWGVKKGALAPI(SEQ ID NO: 158); TGVKIWKVAPKKPGKFGGSGGGSLKKAPGKPAKI(SEQ ID NO: 159); AKAPKVHWKYYGFVHKFSKGAITWLWFLAKAW(SEQ ID NO: 160); IKARLKKKKCFAILLVAKKKKKAIG(SEQ ID NO: 161); WSGKAGSLLAKKKKPFTALLAKKGAKLKKKKG(SEQ ID NO: 162); RVKGKALWLGGSGGGSKWILGAKVGSLR(SEQ ID NO: 163); and158 / 179B0662.70120WO00KLILLILAKARGKVGGSGGGSWKAYLLKKKAKLAVNWKEWKKLGKAWKA GGKF (SEQ ID NO: 164).

92. The compound of any one of claims 1-89, wherein the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: KKGIGKWGKKHKKGIGGIKVVWGKVVGIWGIL(SEQ ID NO: 165); WGKLAFGKIKGIYKWGKKGWGKLAGIIGIGIKGI(SEQ ID NO: 166); GKKKLGKGIKGIIGKWGIIGIIGIKGIGIW(SEQ ID NO: 167); KKGIRRIGIGIGKGIGIIGKIFLSKLGWWGIW(SEQ ID NO: 168); WGIKLWGKVKGIKAIGYKAFGKIANKVINKYKGIG(SEQ ID NO: 169); KKGIGKIGKAFGKWGIKAIGIGKGALAILKVVIG(SEQ ID NO: 170); KKGIKAFIGIGKKWGGSGGGSKIWGIIGWGIAFGIAFN(SEQ ID NO: 171); WGKKSKKVWGIWKVLSGWGIKVWGK(SEQ ID NO: 172); PKMKGIGIKVAKGIKKLGKKLKNGIGKVKVY(SEQ ID NO: 173); KFWIGIGAGIIKVVKIGGSGGGSKNWGKWGKGWGI(SEQ ID NO: 174); RKWWGKLHKAIGGSGGGSKKKLSKVVKVVMAGI(SEQ ID NO: 175); KVVRKWGGIGGSGGGSKKWGHHKAINVLAFIKAF(SEQ ID NO: 176); RKIGGIKKWGGSGGGSVVKVVKGIWGAVVAIWGI(SEQ ID NO: 177); WGKKVKIKVAGIGKVWKKAIGNQAKVIGLG(SEQ ID NO: 178); KMHGIGAFGKWGKGIIGIIKKIGIWGIIGWG(SEQ ID NO: 179); and WKVWRKIGKGGSGGGSYKVKLGIIIGIGIG (SEQ ID NO: 180).

93. The compound of any one of claims 1-89, wherein the endosomal escape peptide is an endosomal escape peptide comprising a sequence selected from the group consisting of: RILSVNRIKGGSGGGSVLLKRISVLSLKGWVNW(SEQ ID NO: 181); RIKKITGLSKKVLIRISVLGKRIATGWRTGSRIRI(SEQ ID NO: 182); RIWKKVLLATGGSGGGSVNILSRIFKRISVL(SEQ ID NO: 183); RIKKFLKVNKLFSVNWAKTGQLGVLRVNRIGKLSV(SEQ ID NO: 184); WKRILSKVNTGGSGGGSKNKRIIRRIKRIIRI(SEQ ID NO: 185); KRIRRIAKVGGSGGGSVNKRILSVLVNW(SEQ ID NO: 186); RRITGWARIGGSGGGSVNWKRIWVLSVL(SEQ ID NO: 187); WSVLRIRITGVLSVLGGSGGGSVRIIRIWRI(SEQ ID NO: 188); RIRIWKRISVLGGSGGGSVRIISVLWKKRLRIRRIL(SEQ ID NO: 189); SVLRRISKRIGGSGGGSVLSVNRIMKRIVNW(SEQ ID NO: 190);159 / 179B0662.70120WO00TGWGAIKGWKGTLTGWLSVFRRKLRITGKKVN(SEQ ID NO: 191); SVLRIHRIRRVNWARIRVLSVRSV(SEQ ID NO: 192); YGLSIKRISVLSKKLSVNKLG(SEQ ID NO: 193); GWKVLRILLSQVNKLSVLLSKRTGNRIGRI(SEQ ID NO: 194); WMTGWSVLGGGSGGGSVLSKKVLKKVLK(SEQ ID NO: 195); and GLGKVSVLSVRIFVRIGLTGWKRIG (SEQ ID NO: 196).

94. The compound of any one of claims 1-93, wherein the endosomal escape peptide comprises a multimer.

95. The compound of claim 94, wherein the endosomal escape peptide comprises a dimer.

96. The compound of claim 95, wherein the dimer comprises two iterations of a sequence of any one of claims 87-90.

97. The compound of claim 94, wherein the endosomal escape peptide comprises a trimer.

98. The compound of claim 97, wherein the trimer comprises three iterations of a sequence of any one of claims 87-90.

99. The compound of claim 90, wherein the endosomal escape peptide comprises a sequence of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 2).

100. The compound of claim 90, wherein the endosomal escape peptide comprises a sequence of: LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 6).

101. The compound of claim 90, wherein the endosomal escape peptide comprises a sequence of: KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 11).160 / 179B0662.70120WO00102. The compound of claim 90, wherein the endosomal escape peptide comprises a sequence of: KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 13).

103. The compound of claim 90, wherein the endosomal escape peptide comprises a sequence of: KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 14).

104. The compound of any one of claims 94-96, wherein the endosomal escape peptide comprises a sequence of: [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 43).

105. The compound of any one of claims 94, 97, or 98, wherein the endosomal escape peptide comprises a sequence of: [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 44).

106. The compound of claim 90, wherein the endosomal escape peptide comprises a sequence of: KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 15).

107. The compound of any one of claims 1-70, wherein the endosomal escape peptide is not one or more of S10, Penetratin, Tat, R9, Transportan-10, L17E M-Lycotoxin, or Aurein1.

2.

108. The compound of claim 1, having a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 45);161 / 179B0662.70120WO00WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 46); TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 47); MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 48); LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 49); LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 50); RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 51); KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 52); KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 53); KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 54); KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 55); KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 56); KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK–162 / 179B0662.70120WO00[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 57); KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 58); ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 59); KWKLARAFARAIKKLGGSGGGSYARALRRQARTG– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 60); RQIKIWFQNRRMKWKK–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 61); CKRKKRRQRRRG–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 62); RRRRRRRRR–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 63); AGYLLGKINLKALAALAKKIL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 64); IWLTALKFLGKHAAKHEAKQQLSKL– [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 65); and GLFDIIKKIAESF–[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 66).

109. The compound of claim 1, having a sequence selected from the group consisting of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 67); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWKK (SEQ ID NO: 68); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–163 / 179B0662.70120WO00TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK (SEQ ID NO: 69); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 70); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 71); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 72); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKLK (SEQ ID NO: 73); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 74); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 75); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 76); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 77); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 78); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 79);164 / 179B0662.70120WO00[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 80); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK]–[GGSGGGS]– [KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK] (SEQ ID NO: 81); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 82); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 83); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–KWKLARAFARAIKKLGGSGGGSYARALRRQARTG (SEQ ID NO: 84); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–RQIKIWFQNRRMKWKK (SEQ ID NO: 85); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–CKRKKRRQRRRG (SEQ ID NO: 86); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–RRRRRRRRR (SEQ ID NO: 87); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–AGYLLGKINLKALAALAKKIL (SEQ ID NO: 88); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–IWLTALKFLGKHAAKHEAKQQLSKL (SEQ ID NO: 89); and [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GLFDIIKKIAESF (SEQ ID NO: 90).

110. The compound of claim 1, having a sequence selected from the group consisting of: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 – KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 221);165 / 179B0662.70120WO00[(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 222); and [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10– CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 – KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 223).

111. The compound of claim 1, having a sequence selected from the group consisting of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CAP]5– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 197); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CEAAAK]5– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 224); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–C5G3– [(IPGHG)2(VPGYG)(IPGHG)2]15– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 225); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 238); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGH GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGCGIPGHGVPGYGIPGHGIP GHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 239);166 / 179B0662.70120WO00[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 240); and [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– [IPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGC GVPGYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPG HGIPGHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPG HGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIPGCGVP GYGIPGHGIPGHGIPGHGIPGHGVPGYGIPGCGIPGHGIPGHGIPGHGVPGYGIPGHGIP GHGIPGCGIPGHGVPGYGIPGHGIPGHGIPGHGIPGCGVPGYGIPGHGIPGHGIPGHGIP GHGVPGYGIPGHGIPGCGIPGYG]– KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 241).

112. The compound of any one of claims 1-111, wherein the compound further comprises a protease cleavable linker sequence at the C-terminus.

113. The compound of any one of claims 1-111, wherein the compound further comprises a protease cleavable linker sequence at the N-terminus.

114. The compound of claim 112 or 113, wherein the protease cleavable linker sequence is selected from the group consisting of: (GGS)9 (SEQ ID NO: 91); GGSGGSEVRFGGSGGS (SEQ ID NO: 92); GGSGGSGGVGFLGGGS (SEQ ID NO: 93); GGSGGSGTQFFRLGGS (SEQ ID NO: 94); KKESTFEERSYWQSQV (SEQ ID NO: 95);167 / 179B0662.70120WO00GGSGGSRVRRGGSGGS (SEQ ID NO: 96); and NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 97).

115. The compound of any one of claims 112-114, comprising a sequence selected from the group consisting of: (GGS)9–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 98); GGSGGSEVRFGGSGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 99); GGSGGSGGVGFLGGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 100); GGSGGSGTQFFRLGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 101); KKESTFEERSYWQSQV–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 102); NNTHDLVGDVRLAGVQSVASSRRHKRFAGV– [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 103); and GGSGGSRVRRGGSGGS–[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10– CCCCGGG–[(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 104).

116. The compound of any one of claims 112-114, comprising a sequence selected from the group consisting of: [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–(GGS)9 (SEQ ID NO: 116); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSEVRFGGSGGS (SEQ ID NO: 106); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSGGVGFLGGGS (SEQ ID NO: 107); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSGTQFFRLGGS (SEQ ID NO: 108); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–KKESTFEERSYWQSQV (SEQ ID NO: 109); [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–GGSGGSRVRRGGSGGS (SEQ ID NO: 110); and168 / 179B0662.70120WO00[(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15–NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 111).

117. An elastin-like polypeptide comprising a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–X*– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 198), wherein: each instance of X3is independently G, S, Q, V, or A; each instance of X4is independently E or G; and X*is -CCCCGGG- (SEQ ID NO: 19), -[CAP]5- (SEQ ID NO: 125), [CEAAAK]5 (SEQ ID NO: 126), or -C5G3 (SEQ ID NO: 127).

118. The elastin-like polypeptide of claim 117, comprising a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 227), wherein: X3is G, S, Q, V, or A; and X4is E or G.

119. The elastin-like polypeptide of claim 117 or 118, comprising a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 112), wherein: X3is V or A; and X4is E or G.

120. The elastin-like polypeptide of claim 117 or 118, wherein each instance of X3is V, each instance of X3is A, each instance of X3is G, each instance of X3is S, or each instance of X3is Q.

121. The elastin-like polypeptide of any one of claims 117-120, wherein each instance of X3is V or each instance of X3is A.169 / 179B0662.70120WO00122. The elastin-like polypeptide of any one of claims 117, 118, or 120, wherein each instance of X3is G, each instance of X3is S, or each instance of X3is Q.

123. The elastin-like polypeptide of any one of claims 117-122, wherein each instance of X4is E or each instance of X4is G.

124. The elastin-like polypeptide of any one of claims 117-123, wherein X*is CCCCGGG (SEQ ID NO: 19).

125. The elastin-like polypeptide of any one of claims 117 or 120-123, wherein X*is - [CAP]5- (SEQ ID NO: 125), [CEAAAK]5(SEQ ID NO: 126), or -C5G3(SEQ ID NO: 127).

126. The elastin-like polypeptide of any one of claims 117-119, comprising a sequence of: [(VPGVG)(VPGEG)(VPGVG)(VPGEG)(VPGVG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 115).

127. The elastin-like polypeptide of any one of claims 117-119, comprising a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 113).

128. The elastin-like polypeptide of claim 117 or 118, comprising a sequence of: [(VPGGG)(VPGGG)(VPGGG)(VPGGG)(VPGGG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 131); [(VPGSG)(VPGGG)(VPGSG)(VPGGG)(VPGSG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 228); or [(VPGQG)(VPGGG)(VPGQG)(VPGGG)(VPGQG)]10–CCCCGGG– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 229).

129. The elastin-like polypeptide of claim 117, comprising a sequence of: [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CAP]5– [(IPGHG)2(VPGYG)(IPGHG)2]15(SEQ ID NO: 199); [(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–[CEAAAK]5– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 200); or170 / 179B0662.70120WO00[(VPGAG)(VPGGG)(VPGAG)(VPGGG)(VPGAG)]10–C5G3– [(IPGHG)2(VPGYG)(IPGHG)2]15 (SEQ ID NO: 201).

130. An elastin-like polypeptide comprising a sequence of: [(VPGX3G)(VPGX4G)(VPGX3G)(VPGX4G)(VPGX3G)]10– [(IPGZ4G)2(VPGYG)(IPGZ4G)2]15 (SEQ ID NO: 203), wherein: each instance of X3is independently G, S, Q, V, or A; each instance of X4is independently E or G; and each instance of Z3is independently C or H.

131. The elastin-like polypeptide of claim 130, wherein three instances of Z3are C and the remaining instances of Z3are H.

132. The elastin-like polypeptide of claim 130, wherein five instances of Z3are C and the remaining instances of Z3are H.

133. The elastin-like polypeptide of claim 130, wherein seven instances of Z3are C and the remaining instances of Z3are H.

134. The elastin-like polypeptide of claim 130, wherein thirteen instances of Z3are C and the remaining instances of Z3are H.

135. The elastin-like polypeptide of any one of claims 130-134, wherein each instance of X3is A.

136. The elastin-like polypeptide of any one of claims 130-135, wherein each instance of X4is G.

137. An endosomal escape peptide comprising a sequence selected from the group consisting of: LYKKFKKKLLKSLKRGGSGGGSLYKKFKKKLLKSLKRL (SEQ ID NO: 2); WKKWWKKWWKWWKKWWKKGGSGGGSWKKWWKKWWKWWKKWWK K (SEQ ID NO: 3); TKRVLQVWFQNARAKFRRNLLRGGSGGGSMLSFILTLKRMLKACLRAWK171 / 179B0662.70120WO00(SEQ ID NO: 4); MLSFLLTLKRMLRACLRAWGGSGGGSLQSTKRFIKWYNAWNEKRR (SEQ ID NO: 5); LYKKFKKKLLKSLKRLGGSGGGSYKWWQKYQKRKF (SEQ ID NO: 6); LYKKFKKKLLKSLKRGGSGGGSTKRVLQVWFQNARAKFRRNLLR (SEQ ID NO: 7); RRWVRRVRRWVRRVVRVVRRWVRRGGSGGGSFLKAIKKFGKEFKKIGAKL K (SEQ ID NO: 8); KHKHKHKHKHGGSGGGSKKTWWKTWWTKWSQPKK (SEQ ID NO: 9); KFAKKFAKKFAKKAAKGGSGGGSFKFFKKFFKKWK (SEQ ID NO: 10); KLLKKLLKLWKKLLKKLKGGSGGGSFVVKKKKKVF (SEQ ID NO: 11); KKVVFKVKFKGGSGGGSAIWKKIIKKIIKSAKKIG (SEQ ID NO: 12); KKPWWKPWWPKWKKGGSGGGSGWKKFFKKAAKVGK (SEQ ID NO: 13); KNWKGIAGMAKKLLGKNWGGSGGGSVNWKKILGKIIKVVK (SEQ ID NO: 14); KWKLFKKIGKVLKVLGGSGGGSKWKKFKKIGAVLKVL (SEQ ID NO: 15); and ALWKTLLKKVLKAAAKGGSGGGSVKFAIKKIGKKAAKKVIVKA (SEQ ID NO: 16).

138. An endosomal escape peptide comprising a sequence selected from the group consisting of: RMKFNVWAWICVVKKKKFFPHWKIRYIIGRAWLL(SEQ ID NO: 133); DVHRRIWIILLPGHRIKFNWKQWKWRFEFEFKAVRK(SEQ ID NO: 134); KLVRYISYLKLRGGVKILLAKIKPFYEVKLKLKRLSLII(SEQ ID NO: 135); YFLARARARAILIKFFGKLKPFKAKKLAKQLKWVART(SEQ ID NO: 136); ANHKFGVYFGGSGGGSGVKWQVKFGVNHANHVKF(SEQ ID NO: 137); ANHGHKFGVKLNHGGSGGGSYFKFGKAPGVKW(SEQ ID NO: 138); GVGVREWKFGGSGGGSYYFGVKGHKKGVGV(SEQ ID NO: 139); GVKFGVGVKKYFKYFASGVKGVKYF(SEQ ID NO: 140); GVGVKYFKVAVKNHKPKWFSYKAAWKKFG(SEQ ID NO: 141); GNHKAYKWVRAPRTNHKHKKAVKWKHKPG(SEQ ID NO: 142); LKFWGASAPAPHKFAWKFGVGARASGHKLL(SEQ ID NO: 143); VKWKWQWGVKGAPGYFYLGFGVKTAWAPAWAW(SEQ ID NO: 144); FLKFGGSGGGSHKVKFGVVGNHKKAPVHKW(SEQ ID NO: 145);172 / 179B0662.70120WO00GVGASLGKQWKFGGSGGGSNHVKWFGVKPSKGHKGAWKWS(SEQ ID NO: 146); KPIQWGVGGSGGGSYLAPRTKKHVKFGVKRTHKFAP(SEQ ID NO: 147); WKWLKAPKPILKILGKPVKHKWKPAWKGKWQ(SEQ ID NO: 148); KSILKWGAKPKKALILASLGVKKLLAPSGAPW(SEQ ID NO: 149); GVKPAWRASLGGKHVKRLSGFHKPRTKAPSL(SEQ ID NO: 150); KGVKWLLKFGGSGGGSLILKFGKFGFKAPGAPGIL(SEQ ID NO: 151); AWQGVKGFKLAPKKGKPLGKPTKKKAHVRKL(SEQ ID NO: 152); FGAPSKHVKKILEWQKGGSGGGSWKILKFGGKPGVGF(SEQ ID NO: 153); EWKFGFGAGKSLKLGVVHVFGGSGGGSKILILKPVGAP(SEQ ID NO: 154); GFKQAPGVILAPRGFKWLIKLGKAKQKVKPFK(SEQ ID NO: 155); GVKPLKKCKPAILVAPKKGKQEWLKW(SEQ ID NO: 156); GVGKKVAWWQAQAKAPAWKGFGKWKKAKAAP(SEQ ID NO: 157); GIWKLGKAPKILGGSGGGSKWGVKKGALAPI(SEQ ID NO: 158); TGVKIWKVAPKKPGKFGGSGGGSLKKAPGKPAKI(SEQ ID NO: 159); AKAPKVHWKYYGFVHKFSKGAITWLWFLAKAW(SEQ ID NO: 160); IKARLKKKKCFAILLVAKKKKKAIG(SEQ ID NO: 161); WSGKAGSLLAKKKKPFTALLAKKGAKLKKKKG(SEQ ID NO: 162); RVKGKALWLGGSGGGSKWILGAKVGSLR(SEQ ID NO: 163); and KLILLILAKARGKVGGSGGGSWKAYLLKKKAKLAVNWKEWKKLGKAWKA GGKF (SEQ ID NO: 164).

139. An endosomal escape peptide comprising a sequence selected from the group consisting of: KKGIGKWGKKHKKGIGGIKVVWGKVVGIWGIL(SEQ ID NO: 165); WGKLAFGKIKGIYKWGKKGWGKLAGIIGIGIKGI(SEQ ID NO: 166); GKKKLGKGIKGIIGKWGIIGIIGIKGIGIW(SEQ ID NO: 167); KKGIRRIGIGIGKGIGIIGKIFLSKLGWWGIW(SEQ ID NO: 168); WGIKLWGKVKGIKAIGYKAFGKIANKVINKYKGIG(SEQ ID NO: 169); KKGIGKIGKAFGKWGIKAIGIGKGALAILKVVIG(SEQ ID NO: 170); KKGIKAFIGIGKKWGGSGGGSKIWGIIGWGIAFGIAFN(SEQ ID NO: 171); WGKKSKKVWGIWKVLSGWGIKVWGK(SEQ ID NO: 172); PKMKGIGIKVAKGIKKLGKKLKNGIGKVKVY(SEQ ID NO: 173); KFWIGIGAGIIKVVKIGGSGGGSKNWGKWGKGWGI(SEQ ID NO: 174);173 / 179B0662.70120WO00RKWWGKLHKAIGGSGGGSKKKLSKVVKVVMAGI(SEQ ID NO: 175); KVVRKWGGIGGSGGGSKKWGHHKAINVLAFIKAF(SEQ ID NO: 176); RKIGGIKKWGGSGGGSVVKVVKGIWGAVVAIWGI(SEQ ID NO: 177); WGKKVKIKVAGIGKVWKKAIGNQAKVIGLG(SEQ ID NO: 178); KMHGIGAFGKWGKGIIGIIKKIGIWGIIGWG(SEQ ID NO: 179); and WKVWRKIGKGGSGGGSYKVKLGIIIGIGIG (SEQ ID NO: 180).

140. An endosomal escape peptide comprising a sequence selected from the group consisting of: RILSVNRIKGGSGGGSVLLKRISVLSLKGWVNW(SEQ ID NO: 181); RIKKITGLSKKVLIRISVLGKRIATGWRTGSRIRI(SEQ ID NO: 182); RIWKKVLLATGGSGGGSVNILSRIFKRISVL(SEQ ID NO: 183); RIKKFLKVNKLFSVNWAKTGQLGVLRVNRIGKLSV(SEQ ID NO: 184); WKRILSKVNTGGSGGGSKNKRIIRRIKRIIRI(SEQ ID NO: 185); KRIRRIAKVGGSGGGSVNKRILSVLVNW(SEQ ID NO: 186); RRITGWARIGGSGGGSVNWKRIWVLSVL(SEQ ID NO: 187); WSVLRIRITGVLSVLGGSGGGSVRIIRIWRI(SEQ ID NO: 188); RIRIWKRISVLGGSGGGSVRIISVLWKKRLRIRRIL(SEQ ID NO: 189); SVLRRISKRIGGSGGGSVLSVNRIMKRIVNW(SEQ ID NO: 190); TGWGAIKGWKGTLTGWLSVFRRKLRITGKKVN(SEQ ID NO: 191); SVLRIHRIRRVNWARIRVLSVRSV(SEQ ID NO: 192); YGLSIKRISVLSKKLSVNKLG(SEQ ID NO: 193); GWKVLRILLSQVNKLSVLLSKRTGNRIGRI(SEQ ID NO: 194); WMTGWSVLGGGSGGGSVLSKKVLKKVLK(SEQ ID NO: 195); and GLGKVSVLSVRIFVRIGLTGWKRIG (SEQ ID NO: 196).

141. A composition comprising a compound of any one of claims 1-140 and an agent.

142. The composition of claim 141, wherein the composition is in the form of a particle.

143. The composition of claim 142, wherein the particle is a nanoparticle.

144. The composition of claim 141, wherein the composition is in the form of a micelle.174 / 179B0662.70120WO00145. The composition of any one of claims 141-144, wherein the compound and the agent are not covalently attached.

146. The composition of any one of claims 141-145, wherein the compound and the agent are electrostatically complexed.

147. The composition of claim any one of claims 141-146, wherein the compound and the agent are electrostatically complexed at the N-terminus of the compound.

148. The composition of any one of claims 141-146, wherein the compound and the agent are electrostatically complexed at the C-terminus of the compound.

149. The composition of any one of claims 141-144, wherein the compound and the agent are covalently attached.

150. The composition of any one of claims 141-144 or 149, wherein the compound and the agent are covalently attached via the N-terminus of the compound.

151. The composition of any one of claims 141-144 or 149, wherein the compound and the agent are covalently attached via the C-terminus of the compound.

152. The composition of any one of claims 141-144 or 149-151, wherein the compound and the agent are covalently attached via a protease cleavable linker.

153. The composition of claim 152, wherein the protease cleavable linker is selected from the group consisting of: (GGS)9 (SEQ ID NO: 91); GGSGGSEVRFGGSGGS (SEQ ID NO: 92); GGSGGSGGVGFLGGGS (SEQ ID NO: 93); GGSGGSGTQFFRLGGS (SEQ ID NO: 94); KKESTFEERSYWQSQV (SEQ ID NO: 95);175 / 179B0662.70120WO00GGSGGSRVRRGGSGGS (SEQ ID NO: 96); and NNTHDLVGDVRLAGVQSVASSRRHKRFAGV (SEQ ID NO: 97).

154. The composition of claim 153, wherein the protease cleavable linker is GGSGGSRVRRGGSGGS (SEQ ID NO: 96).

155. The composition of any one of claims 141-154, wherein the agent is an organic molecule, protein, peptide, polypeptide, polynucleotide, an isotopically labeled chemical compound, vaccine, or an immunological agent.

156. The composition of claim 155, wherein the agent is a polynucleotide.

157. The composition of claim 155, wherein the agent is an RNA.

158. The composition of claim 157, wherein the RNA is RNA is messenger RNA (mRNA), pre-messenger RNA (pre-mRNA), ribosomal RNA (rRNA), transfer RNA (tRNA), small nuclear RNA (snRNA), small nucleolar RNA (snoRNA), Piwi-interacting RNA (piRNA), microRNA (miRNA), small interfering RNA (siRNA), antisense RNA (asRNA or aRNA), transfer-messenger RNA (tmRNA), single-stranded RNA (ssRNA), double-stranded RNA (dsRNA), precursor messenger RNA (pre-mRNA), small hairpin RNA or short hairpin RNA (shRNA), guide RNA (gRNA), heterogeneous nuclear RNA (hnRNA), coding RNA, non-coding RNA (ncRNA), long non-coding RNA (long ncRNA or lncRNA), satellite RNA, viral satellite RNA, signal recognition particle RNA, small cytoplasmic RNA, polyinosinic acid, ribozyme, flexizyme, spliced leader RNA, viral RNA, or viral satellite RNA.

159. The composition of claim 157 or 158, wherein the RNA is siRNA.

160. The composition of claim 157 or 158, wherein the RNA is mRNA.

161. The composition of claim 155, wherein the agent is a protein.

162. The composition of claim 155, wherein the agent is a peptide.176 / 179B0662.70120WO00163. The composition of claim 162, wherein the peptide is ovalbumin-derived MHC class I epitope or ovalbumin-derived MHC class II epitope.

164. The composition of claim 155, wherein the agent is a polypeptide.

165. The composition of claim 164, wherein the agent is a monoclonal antibody.

166. The composition of claim 165, wherein the monoclonal antibody is anti-CD117.

167. The composition of claim 165, wherein the monoclonal antibody is anti-CD5.

168. A kit comprising: the compound of any one of claims 1-140 or the composition of any one of claims 141-167; and instructions for using the compound or composition.

169. A method of delivering an agent to a subject, cell, or biological sample, comprising administering to the subject or contacting the cell or biological sample with an effective amount of a compound of any one of claims 1-140 or a composition of any one of claims 141-168.

170. The method of claim 169, wherein the agent is a polynucleotide or a polypeptide.

171. The method of claim 169 or 170, wherein the agent is delivered from an extracellular space to the cytosol of a cell.

172. The method of claim 169 or 170, wherein the agent is delivered from an extracellular space to the nucleus of the cell.

173. The method of any one of claims 169-172, wherein the cell is a eukaryotic cell.177 / 179B0662.70120WO00174. The method of claim 170, wherein the effective amount is sufficient to increase the transduction efficiency of the polynucleotide or polypeptide.

175. A method of treating or preventing a disease in a subject, comprising administering to the subject a compound of any one of claims 1-140 or a composition of any one of claims 141-168.

176. The method of claim 175, wherein the disease is a cardiovascular disease, a lung or respiratory disease, a musculoskeletal disease, a hematological disease, an immune disorder, an infectious disease, a genetic disease, cancer, a neurological disease, a psychiatric disorder, a metabolic disorder, or an inflammatory disease.

177. The method of claim 175 or 176, wherein the disease is an infectious disease, genetic disease, or cancer.178 / 179B0662.70120WO00

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