Hydrophobically interspaced charged peptides for preventing or treating APOE and / or amyloid beta pathologic interaction in alzheimer's disease and related dementias

Abstract

Disclosed herein are peptides or peptoids including from 8 to 20 amino acids, including a plurality of hydrophobic amino acids and a plurality of charged amino acids, wherein spacing of the plurality of hydrophobic amino acids and the plurality of charged amino acids aligns with the spacing of hydrophobic amino acids and oppositely charged amino acids in an ApoE protein or polypeptide. Also disclosed are pharmaceutical compositions that include a peptide or peptoid as disclosed, as well as various methods of using the peptides or peptoids, and such pharmaceutical compositions. These methods include blocking Aβ / ApoE interaction, inhibiting Aβ protein oligomerization, and inhibiting amyloid plaque and / or tau aggregate formation, as well as the treatment of Alzheimer's Disease and other tauopathies.

Description

HYDROPHOBICALLY INTERSPACED CHARGED PEPTIDES FOR PREVENTING OR TREATING APOE AND / OR AMYLOID BETA PATHOLOGIC INTERACTION IN ALZHEIMER'S DISEASE AND RELATED DEMENTIAS

[0001] This invention was made with government support under P01 AG060882 awarded by the National Institutes of Health. The government has certain rights in the invention.

[0002] The instant application contains a computer readable Sequence Listing which has been submitted electronically in XML format (“Sequence Listing XML”) and is hereby incorporated by reference in its entirety. The Sequence Listing XML, created on August 13, 2024, is named 147462.002471. xml and is 1,670,303 bytes in size.CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This application claims the priority benefit of U.S. Provisional Patent Application Serial No. 63 / 579,743, filed August 30, 2023, which is hereby incorporated by reference in its entirety.FIELD

[0004] Disclosed herein are peptides or peptoids, where feasible, pharmaceutical compositions, and methods of using the peptides for disrupting the ApoE / Ap pathogenic interaction, blocking Ap / ApoE binding, inhibiting Ap protein oligomerization, and inhibiting amyloid plaque and / or tau aggregate formation and progression, as well as the treatment of Alzheimer’s Disease, related neurodegenerative diseases, amyloid diseases, and other tauopathies.BACKGROUND

[0005] The pathological accumulation of Ap peptides as conformationally toxic oligomers, fibrillary amyloid plaques and cerebral amyloid angiopathy (CAA), either from increased production of Ap peptides, their inadequate clearance or their accelerated conformational change to a pathological structure, is critical in the pathogenesis of Alzheimer’s disease (AD) (Scheltens et al. “Alzheimer’s Disease,” Lancet 388:505-517 (2016); Selkoe & Hardy, “The Amyloid Hypothesis of Alzheimer’s Disease at 25 years,” EMBO Mol Med 8:595-608 (2016)). The apolipoprotein E4 (apoE4) allele, a major genetic risk factor for late-onset AD, has been strongly associated with increased amyloid plaques deposition and enlargement in the brain parenchyma and advanced vascular amyloid pathology such as CAA (Potter & Wisniewski, “Apolipoprotein E: Essential Catalyst of the Alzheimer Amyloid Cascade,” Int. J. Alz. Dis 2012:489428 (2012); Huang & Mahley, “Apolipoprotein E: Structure and Function in Lipid Metabolism, Neurobiology, and Alzheimer’s Diseases,” Neurobiol Dis 72(Pt A):3-12 (2014); Huynh et al.,“Apolipoprotein E and Alzheimer Disease: The Influence of ApoE on Amyloid-Beta and Other Amyloidogenic Proteins,” J Lipid Res, 58(5):P824-836 (2017).

[0006] Numerous studies have shown that ApoE binds to residues 12-28 of A ? and this binding modulates Ap accumulation hence affecting disease progression (Potter & Wisniewski, “Apolipoprotein E: Essential Catalyst of the Alzheimer Amyloid Cascade,” Int. J. Alz.Dis 2012:489428 (2012); Huang & Mahley, “Apolipoprotein E: Structure and Function in Lipid Metabolism, Neurobiology, and Alzheimer’s Diseases,” Neurobiol Dis 72(Pt A):3-12 (2014); Liao et al. “Apolipoprotein E Metabolism and Functions in Brain and Its Role in Alzheimer’s Disease,” Curr Opin Lipidol 28:60-67 (2017); Yamazaki et al., “Apolipoprotein E as a Therapeutic Target in Alzheimer’s Disease: A Review of Basic Research and Clinical Evidence,” CNS Drugs 30:773-789 (2016); Wisniewski & Frangione, “Apolipoprotein E: A Pathological Chaperone Protein in Patients with Cerebral and Systemic Amyloid,” Neurosci. Lett 135:235-238 (1992); Ma et al. “Amyloid-associated Proteins Alpha 1- Anti chymotrypsin and Apolipoprotein E Promote Assembly of Alzheimer Beta-protein into Filaments,” Nature 372:92-94 (1994); Ma et al., “Alzheimer Ap Neurotoxicity: Promotion by Antichymotrypsin, ApoE4; Inhibition by AP-related Peptides. Neurobiol. Aging 17:773-780 (1996); Sanan et al., “Apolipoprotein E Associates with Beta Amyloid Peptide of Alzheimer’s Disease to Form Novel Monofibrils. Isoform ApoE4 Associates More Efficiently than ApoE3,” J. Clin. Invest 94:860-869 (1994); Golabek et al. “Amyloid P Binding Proteins in vitro and in Normal Human Cerebrospinal Fluid,” Neurosci. Lett 191 :79-82 (1995); Golabek et al., “The Interaction Between Apolipoprotein E and Alzheimer’s Amyloid P-Peptide Is Dependent on P- peptide Conformation,” J. Biol. Chem 271 : 10602-10606 (1996)).

[0007] However, there is no consensus on how different apoE genotypes contribute to the pathogenesis of AD (Potter & Wisniewski, “Apolipoprotein E: Essential Catalyst of the Alzheimer Amyloid Cascade,” Int. J. Alz. Dis 2012:489428 (2012); Liao et al. “Apolipoprotein E Metabolism and Functions in Brain and Its Role in Alzheimer’s Disease,” Curr Opin Lipidol 28:60-67 (2017); Yamazaki et al., “Apolipoprotein E as a Therapeutic Target in Alzheimer’s Disease: A Review of Basic Research and Clinical Evidence,” CNS Drugs 30:773- 789 (2016). It has been hypothesized that ApoE, and ApoE4 in particular, is an amyloid catalyst or “pathological chaperone” (Potter & Wisniewski, “Apolipoprotein E: Essential Catalyst of the Alzheimer Amyloid Cascade,” Int. J. Alz. Dis 2012:489428 (2012); Wisniewski & Frangione, “Apolipoprotein E: A Pathological Chaperone Protein in Patients with Cerebral and Systemic Amyloid,” Neurosci. Lett 135:235-238 (1992)) and that the interaction of Ap and ApoE4 is associated with neurotoxicity (Wu et al., “Co-inj ection of Abetal-40 and ApoE4 Impaired Spatial Memory and Hippocampal Long-term Potentiation in Rats,” Neurosci Lett 648:47-52(2017)). Alternatively, it has been posited that ApoE is an Ap clearance factor, with ApoE4 being impaired at this function compared to ApoE3 or ApoE2 (Liao et al. “Apolipoprotein E Metabolism and Functions in Brain and Its Role in Alzheimer’s Disease,” Curr Opin Lipidol 28:60-67 (2017); Verghese etal., “ApoE Influences Amyloid-Beta (AP) Clearance Despite Minimal ApoE / Abeta Association in Physiological Conditions,” Proc. Natl. Acad. Sci. USA 110:E1807-E1816 (2013).

[0008] It has been suggested that one therapeutic strategy may be to specifically prevent the interaction of ApoE with Ap, allowing to circumvent complications of altering ApoE levels which may cause detrimental effects on the many other beneficial roles ApoE plays in neurobiology (Potter & Wisniewski, “Apolipoprotein E: Essential Catalyst of the Alzheimer Amyloid Cascade,” Int. J. Alz. Dis 2012:489428 (2012); Yamazaki et al., “Apolipoprotein E as a Therapeutic Target in Alzheimer’s Disease: A Review of Basic Research and Clinical Evidence,” CNS Drugs 30:773-789 (2016). Consistent with this proposal, it has been shown previously that blocking the ApoE / Ap interaction by Ap 12-28P peptides could constitute a novel treatment for AD by reducing brain parenchymal and vascular amyloid burden as well as tau related pathology in several AD transgenic mice lines (Sadowski et al. “A Synthetic Peptide Blocking the Apolipoprotein E / B eta- Amyloid Binding Mitigates Beta-Amyloid Toxicity and Fibril Formation in vitro and Reduces Beta-Amyloid Plaques in Transgenic Mice,” Am J Pathol 165:937-948 (2004); Sadowski etal., “Blocking the ApolipoproteinE / Amyloid P Interaction Reduces the Parenchymal and Vascular Amyloid-P Deposition and Prevents Memory Deficit in AD Transgenic Mice,“ Proc. Nat ’I Acad. Sci USA 103: 18787-18792 (2006); Yang et al., “Blocking the Apolipoprotein E / Amyloid P Interaction Reduces Fibrillar Vascular Amyloid Deposition and Cerebral Microhemorrhages in TgSwDI Mice,” J Alzheimer s. Dis 24:269-285 (2011); Liu et al., “Blocking the Apolipoprotein E / Amyloid P Interaction in Triple Transgenic Mice Ameliorates Alzheimer’s Disease Related Amyloid P and Tau Pathology,” J.Neurochem 128:591 (2014)). More recently, using peptidomimetic technology and the macrocyclization method, a peptoid library derived from Ap 12-28P sequence was used to screen for new ApoE / Ap binding inhibitors with higher efficacy and safety for blocking the apoE / Ap interaction (Pankiewicz et al., “Blocking the ApoE / Abeta Interaction Ameliorates Abeta-Related Pathology in APOE epsilon2 and epsilon4 Targeted Replacement Alzheimer Model Mice,” Acta Neuropathol Commun 2:75 (2104); Cho et al., “Efficient Solid-phase Synthesis of a Series of Cyclic and Linear Peptoid-Dexamethasone Conjugates for the Cell Permeability Studies,” J Comb. Chem 12:321-326 (2010); Khan et al. “Ring-closing Metathesis Approaches for the Solid-phase Synthesis of Cyclic Peptoids,” Org. Lett 13: 1582-1585 (2011);Park & Kwon, “Facile Solid-phase Parallel Synthesis of Linear and Cyclic Peptoids for Comparative Studies of Biological Activity,” ACS Combinatorial Science 17: 196-201 (2015).

[0009] Despite these previous reports, all strategies involve the interaction / binding of Ap to the normal conformation of ApoE; however, there remains a need for an effective strategy for blocking the ApoE4 / Ap pathogenic interaction and thereby treating Ap and tau pathologies. The present invention is directed to overcoming these and other deficiencies in the art.SUMMARY

[0010] A first aspect of the disclosure relates to a peptide or peptoids, when feasible, including from 8 to 20 amino acids, preferably 8 to 15 amino acids and including a plurality of hydrophobic amino acids and a plurality of charged amino acids, wherein spacing of the plurality of hydrophobic amino acids and the plurality of charged amino acids aligns with the spacing of hydrophobic amino acids and oppositely charged amino acids in an ApoE protein or polypeptide. As disclosed herein, these peptides should destabilize the pairing of both alpha-helixes in ApoE, which is responsible for its physiologic conformation and necessary for its pathogenic pro- fibrillogenic function.

[0011] According to one embodiment, the peptide or peptoid includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O (SEQ ID NO: 1), whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue or polar amino acid residue;X3 is a hydrophobic amino acid residue;X4is a hydrophobic amino acid residue;X5 is a positively charged amino acid residue;Xe is a positively charged amino acid residue;X7 is a hydrophobic amino acid residue;Xs is a negatively charged amino acid residue;X9 is a hydrophobic amino acid residue;X10 is a negatively charged amino acid residue; and wherein the peptide or peptoid optionally comprises a C-terminal proline residue.

[0012] According to another embodiment, the peptide or peptoid includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11-X12-X13--X14 (SEQ ID NO: 2) whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue;X3 is a negatively charged amino acid residue;X4 is a positively charged amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a positively charged amino acid residue;X7 is a positively charged amino acid residue;Xs is a hydrophobic amino acid residue;X9 is a hydrophobic amino acid residue or polar amino acid residue;X10 is a negatively charged amino acid residue;Xu is a negatively charged amino acid residue;X12 is a hydrophobic amino acid residue;X13 is a hydrophobic amino acid residue;X14 is a hydrophobic amino acid residue; and wherein the peptide or peptoid optionally comprises a C-terminal proline residue.

[0013] According to a further embodiment, the peptide or peptoid includes the amino acid sequence of:Xi-X2-X3-X4-X5-X6-X7-X8-X9-Xio-Xii (SEQ ID NO: 3) whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue or polar amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a hydrophobic amino acid residue;X5 is a negatively charged amino acid residue;Xe is a negatively charged amino acid residue;X?is a hydrophobic amino acid residue;X8is a negatively charged amino acid residue;X9 is a negatively charged amino acid residue;X10 is a negatively charged amino acid residue;Xu is a hydrophobic amino acid residue; wherein the peptide or peptoid optionally comprises a C-terminal proline residue.

[0014] According to yet another embodiment, the peptide or peptoid includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11-X12-X13-X14 (SEQ ID NO: 4) whereinXi is optional and, when present, is a hydrophobic amino acid residue;X2 is optional and, when present, is a positively charged amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a negatively charged amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a negatively charged amino acid residue;X?is a hydrophobic amino acid residue;Xs is a positively charged amino acid residue;X9 is a positively charged amino acid residue;X10 is a hydrophobic amino acid residue;Xu is optional and, when present, is a hydrophobic amino acid residue;X12 is optional and, when present, is a hydrophobic amino acid residue;X13 is optional and, when present, is a hydrophobic amino acid residue;X14 is optional and, when present, is a hydrophobic amino acid residue; and wherein the peptide or peptoid optionally comprises a C-terminal proline residue.

[0015] According to a further embodiment, the peptide or peptoid includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11-X12-X13-X14 (SEQ ID NO: 1507) whereinXi is a hydrophobic amino acid residue;X2 is a positively charged amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a hydrophobic amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a hydrophobic amino acid residue;X7 is a negatively charged amino acid residue;X8is a hydrophobic amino acid residue;X9 is a negatively charged amino acid residue;X10 is a hydrophobic amino acid residue or polar amino acid residue;Xu is a hydrophobic amino acid residue;X12 is a hydrophobic amino acid residue;X13 is a positively charged amino acid residue; and wherein the peptide or peptoid optionally comprises a C-terminal proline residue.

[0016] A second aspect of the disclosure relates to a pharmaceutical composition that includes a peptide according to the first aspect of the disclosure.

[0017] A third aspect of the disclosure relates to a method of blocking pathogenic Ap / ApoE interaction, the method including the step of contacting an ApoE protein and / or an Ap protein with a peptide or peptoid according to the first aspect of the disclosure, whereby the peptide or peptoid binds to the ApoE protein and / or the Ap protein disrupting its functional conformation, thereby blocking the pathogenic interaction of the ApoE protein with the Ap protein.

[0018] A fourth aspect of the disclosure relates to a method of inhibiting Ap protein oligomerization, the method including the step of contacting an ApoE protein and / or an Ap protein with a peptide according to the first aspect of the disclosure, whereby the peptide binds to the ApoE protein and / or the Ap protein, and thereby blocks the pathogenic interaction of the ApoE protein with the Ap protein or blocks the Ap region from where the P-sheet pathologic structure spreads.

[0019] A fifth aspect of the disclosure relates to a method of inhibiting amyloid plaque and / or tau aggregate formation, the method including the step of contacting an ApoE protein and / or an Ap protein with a peptide according to the first aspect of the disclosure, whereby the peptide binds to the ApoE protein and / or the Ap protein, and inhibits amyloid plaque and / or subsequent tau aggregate formation.

[0020] A sixth aspect of the disclosure relates to a method of treating a condition associated with Ap plaque formations or deposits, the method including the step of administering to a patient having such a condition an effective amount of a peptide according to the first aspect of the disclosure, or a pharmaceutical composition according to the second aspect of the disclosure, wherein said administering is effective to inhibit amyloid plaque and / or tau aggregate formation and thereby treat the patient.

[0021] A seventh aspect of the disclosure relates to a method of treating Alzheimer’s Disease, the method comprising administering to a patient having Alzheimer’s Disease an effective amount of a peptide according to the first aspect of the disclosure, or a pharmaceutical composition according to the second aspect of the disclosure, wherein said administering is effective to inhibit amyloid plaque and / or subsequent tau aggregate formation and thereby treat the patient.

[0022] The accompanying Examples demonstrate the design and testing of several novel peptides that block the pathological function of ApoE. Two distinct representative peptides, Peptide 127 (SEQ ID NO: 133) and Peptide 148 (SEQ ID NO: 518), were shown to block the binding between Ap42 and ApoE4 in vitro. Importantly, each was effective in blocking the pathogenic profibrillogenic function of ApoE4 on Ap and also showed strong anti-self- fibrillogenic effects on Ap alone. In a mouse model of Alzheimer’s Disease expressing different knock-in human apoE genes, treatment of mice with both of these peptides together wasassociated with clear cognitive benefits in different behavioral tests, significant decrease of Ap pathology, in the absence of toxicity.BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figs. 1A-1B are partial sequence alignments of ApoE (SEQ ID NO: 1505) with various peptides. Fig. 1 A is a partial sequence alignment of ApoE residues 111-162 with: (i) peptides CSQNKKPDLDP (SEQ ID NO: 133) and CSQNKKVDLDP (SEQ ID NO: 134), which align with ApoE residues 127 to 137 and / or Ap residues 21 to 24 (of SEQ ID NO: 1506) in terms of hydrophobic residue spacing and charged residue spacing; (ii) peptides IIDRARRPSEDVGLP (SEQ ID NO: 517) and IIDRARRVSEDVGLP (SEQ ID NO: 518), which align with ApoE residues 148 to 162 and / or Ap residues 21 to 24 in terms of hydrophobic residue spacing and charged residue spacing; and (iii) GTNIDEIEDEIP (SEQ ID NO: 861), which aligns with ApoE residues 138 to 149 in terms of hydrophobic residue spacing and charged residue spacing. Fig. IB is a partial sequence alignment of ApoE residues 236-271 with: (i) peptide PDLDVKKNQSC (SEQ ID NO: 962), which aligns with ApoE residues 239 to 249 and / or Ap residues 21 to 24 in terms of hydrophobic residue spacing and charged residue spacing; (ii) peptide CDLDVKKNP (SEQ ID NO: 1141), which aligns with ApoE residues 239 to 247 and / or Ap residues 21 to 24 in terms of hydrophobic residue spacing and charged residue spacing; (iii) peptide CRLDLDVKKNP (SEQ ID NO: 1149), which aligns with ApoE residues 237 to 247 and / or Ap residues 21 to 24 in terms of hydrophobic residue spacing and charged residue spacing; (iv) peptide CDLDVKKLPFFA (SEQ ID NO: 970), which aligns with ApoE residues 239 to 250 and / or Ap residues 21 to 24 in terms of hydrophobic residue spacing and charged residue spacing; (v) CRLDLDVKKLPFFA (SEQ ID NO: 1508), which aligns with ApoE residues 237 to 250 and / or Ap residues 21 to 24 in terms of hydrophobic residue spacing and charged residue spacing; and (vi) CKGLNGDVETWLKP (SEQ ID NO: 1377), which aligns with ApoE residues 254 to 267 in terms of hydrophobic residue spacing and charged residue spacing.

[0024] Figs. 2A-2C illustrate Surface Plasmon Resonance (SPR) testing of SPR Ap42 and ApoE binding in the absence (Fig. 2A) or presence of peptides (Figs. 2B, 2C). SPR testing was carried out on a Reichert SR700DC as previously described (Goni et al., “Anti-P-sheet Conformation Monoclonal Antibody Reduces Tau and Ap Oligomer Pathology in an Alzheimer's Disease Model,” Alzheimers Res Ther. 10(1): 10 (2018), which is hereby incorporated by reference in its entirety). As shown in Figs. 2B-2C, respectively, Peptide 127 (SEQ ID NO: 133) and Peptide 148 (SEQ ID NO: 518), having complementary matching of the ApoE structure of regions 127-137 and 148-162, respectively, completely block the binding of ApoE4 to Ap.

[0025] Figs. 3A-3C illustrate testing of Peptide 127 (SEQ ID NO: 133) for its ability to inhibit fibrillogenesis of Ap induced by pathological function of ApoE. Detection of in vitro thioflavin fluorescence was performed on the Bio Tek Cytation 7 (Agilent, Santa Clara, CA) machine using [Ap42]:[Pep] ratios of 1 : 1 (10 pM, Fig. 3A), 1 :2 (20 pM, Fig. 3B) and 1 :5 (50 pM, Fig. 3C) with or without ApoE4.

[0026] Figs. 4A-4B illustrate testing of Peptide 148 (SEQ ID NO: 518) for its ability to inhibit fibrillogenesis of Ap induced by pathological function of ApoE. Detection of in vitro thioflavin fluorescence was performed on the Bio Tek Cytation 7 (Agilent, Santa Clara, CA) machine using [Ap42]:[Pep] ratios of 1 : 1 (10 pM, Fig. 4A) and 1 :2 (20 pM, Fig. 4B) with or without ApoE4.

[0027] Figs. 5-7 illustrate the effects of the combination anti-fibrillogenic Peptides 127 (SEQ ID NO: 133) and 148 (SEQ ID NO: 518) on locomotor and behavioral testing following 4 month inoculations once every other week for each peptide in an Alzheimer’s Disease (AD) transgenic mouse model SwDI (APP / PS1 Swedish -Dutch-Iowa) with knock-in human apoE genes (either apoE3 or apoEE). Control animals aged and inoculated with vehicle alone (sterile saline).

[0028] Fig. 5 shows the results of Barnes Maze assessment, which confirmed significant cognitive rescue compared to control animals.

[0029] Fig. 6 shows the results of Radial Arm Maze assessment, which confirmed significant cognitive rescue compared to control animals.

[0030] Fig. 7 shows the results of locomotor testing (grip strength and rotarod), which confirmed no differences between test and control cohorts.

[0031] Fig. 8 illustrates, from left to right, a ribbon depiction of the backbone of the ApoE4 N-terminal domain, electrostatic surface of the ApoE4 N-terminal domain in the same orientation as the ribbon depiction, labeling of key amino acid positions from the peptide design scheme for each of Peptides 127 (SEQ ID NO: 133) and 148 (SEQ ID NO: 518), and the peptide design scheme for reference.

[0032] Fig. 9 illustrates, from left to right, a ribbon depiction of the backbone of the ApoE4 N-terminal domain, electrostatic surface of the ApoE4 N-terminal domain in the same orientation as the ribbon depiction, labeling of key amino acid positions from the peptide design scheme for each of Peptides 127 (SEQ ID NO: 133) and 148 (SEQ ID NO: 518) with key measured distances labeled.

[0033] Fig. 10A-10B illustrate the folded (top) and extended (bottom) peptide structure for Peptides 127 (SEQ ID NO: 133) and 148 (SEQ ID NO: 518), respectively, with dimensions in Angstroms labeled.

[0034] Fig. 11 illustrates docked locations of Peptides 127 (SEQ ID NO: 133) and 148 (SEQ ID NO: 518) on ApoE4 N-terminal domain. Peptides are shown in stick depiction in their docked location and labeled as indicated. The ApoE4 backbone is shown in ribbon depiction.DETAILED DESCRIPTION

[0035] One aspect of the disclosure relates to a peptide or peptoid, when feasible, including from 8 to 20 amino acids, and including a plurality of hydrophobic amino acids and a plurality of charged amino acids, wherein spacing of the plurality of hydrophobic amino acids and the plurality of charged amino acids aligns with the spacing of hydrophobic amino acids and oppositely charged amino acids in an ApoE protein or polypeptide and / or Ap peptide.

[0036] By “aligns with”, it is intended that the spacing of hydrophobic amino acids and charged amino acids in one of the ApoE protein or Ap peptide, and the peptide as disclosed herein is when (i) both the ApoE protein or Ap peptide, and the peptide are aligned in an N-to-C orientation, or alternatively (ii) the peptide and the ApoE protein or Ap peptide are aligned in reverse orientations (i.e., one is N-to-C and the other is C-to-N). The ApoE protein sequence used for comparison to the peptides as described herein can be any mammalian ApoE protein, but is preferably a human ApoE protein.

[0037] The human ApoE protein exists as multiple isoforms, including those known as ApoE2, ApoE3, and ApoE4.

[0038] The human ApoE3 amino acid sequence (SEQ ID NO: 1505) is shown below:KVEQAVETEP EPELRQQTEW QSGQRWELAL GRFWDYLRWV QTLSEQVQEE 50LLSSQVTQEL RALMDETMKE LKAYKSELEE QLTPVAEETR ARLSKELQAA 100QARLGADMED VCGRLVQYRG EVQAMLGQST EELRVRLASH LRKLRKRLLR 150DADDLQKRLA VYQAGAREGA ERGLSAIRER LGPLVEQGRV RAATVGSLAG 200QPLQERAQAW GERLRARMEE MGSRTRDRLD EVKEQVAEVR AKLEEQAQQI 250RLQAEAFQAR LKSWFEPLVE DMQRQWAGLV EKVQAAVGTS AAPVPSDNH 299See also Genbank Accession NP_000032, which is hereby incorporated by reference in its entirety. The Cysl 12 and Argl58 residues are shown in bold typeface in SEQ ID NO: 1505. The human ApoE2 variant possesses Cysl 12 and Cysl 58, whereas the human ApoE4 variant possesses Argl 12 and Argl58 (Phillips, “Apolipoprotein E Isoforms and Lipoprotein Metabolism,” IUBMB Life 66(9):616-623 (2014); see also Genbank Accessions ARQ79459 [Cloning vector pcDNA3-ApoE2] and ARQ79463 [Cloning vector pcDNA3-ApoE4A], each of which is hereby incorporated by reference in its entirety). Additional ApoE isoforms andvariants are known and identified by UniProt Accession P02649, which is hereby incorporated by reference in its entirety.

[0039] The Ap peptide is produced through the proteolytic processing of a transmembrane protein, amyloid precursor protein (APP), by P- and y-secretases. Multiple forms of the Ap peptide exist, including Ap40 peptide and Ap42 peptide. See Chen et al., “Amyloid Beta: Structure, Biology and Structure-based Therapeutic Development,” Acta Pharmacol Sin 38: 1205-1235 (2017), which is hereby incorporated by reference in its entirety.

[0040] The human Ap42 peptide has the amino acid sequence (SEQ ID NO: 1506) shown below:DAEFRHDSGY EVHHQKLVFF AEDVGSNKGA I IGLMVGGW IA 42See Genbank Accession 2BEG A; Liihrs el al.. “3D Structure of Alzheimer's Amyloid-beta(l- 42) Fibrils,” Proc Natl Acad Sci USA 102(48): 17342-17347 (2005), each of which is hereby incorporated by reference in its entirety. The human Ap42 peptide contains residues 1-40 of SEQ ID NO: 1506. See Qiu et al., “A / >42 and A / ?40: Similarities and Differences,” J. Peptide Sci. 21(7): 522-529 (2015), which is hereby incorporated by reference in its entirety.

[0041] As used herein, the peptides of the invention can contain exclusively naturally occurring amino acids, as well as one or more non-naturally occurring amino acids. Both L- amino acids, D-amino acids, and mixtures thereof are contemplated.

[0042] Naturally occurring amino acids include alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. Naturally occurring amino acids can be classified based on the nature of their side chains. For example, hydrophobic amino acid residues include, without limitation, glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), and tryptophan (Trp). Positively charged and peptoid residues include, without limitation, lysine (Lys), arginine (Arg), and histidine (His). Negatively charged residues include, without limitation, aspartic acid (Asp) and glutamic acid (Glu). Polar (uncharged) amino acid residues include, without limitation, serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

[0043] Non-naturally occurring amino acids include, without limitation, homo amino acids, N-methyl amino acids, alpha-methyl amino acids, beta (homo) amino acids, gamma amino acids, helix / turn stabilizing motifs, and peptoids. Exemplary non-natural amino acids include, without limitation, hydroxyproline (Hyp), hydroxylysine (Hyl), beta-alanine, citrulline (Cit), ornithine (Om), norleucine (Nle), norvaline (Nva), 3 -nitrotyrosine, nitroarginine, and pyroglutamic acid (Pyr). Any other non-naturally occurring amino acids can also be used.

[0044] Peptoids are N-substituted glycine oligomers, which recapitulate many desirable attributes of natural peptides and demonstrate a range of biological activities (Dohm et al., “Peptoids: Bio-Inspired Polymers as Potential Pharmaceuticals,” Curr. Pharm. Des. 17(25):2732-47 (2011); Mandity & Fulop, “An Overview of Peptide and Peptoid Foldamers in Medicinal Chemistry,” Expert Opin Drug Discov 10: 1163-1177 (2015), each of which is hereby incorporated by reference in its entirety). Advantages of peptoids over peptides include stability of their structure to excessive temperature (over 75 °C), salt, pH, and organic solvent. They are resistant to proteolytic degradation and can be excreted whole in urine, which is an important attribute of a pharmacologically useful peptide mimic. Furthermore, peptoids lack the hydrogen of secondary amides, and thus are unlikely to form the ?-sheet structure, which is associated with the toxicity of Ap and Ap-derived peptides.

[0045] Although the disclosed peptides can be 8 to 20 amino acids in length, preferred peptides are from 8 to 18 amino acids in length, or 8 to 16 amino acids in length, including from 8 to 15 amino acids in length. For example, for peptides that are 8 to 15 amino acids in length, it is contemplated that such peptides may contain 8 or 9 or 10 or 11 or 12 or 13 or 14 or 15 amino acids. The peptide length is sufficient to prevent tertiary structure and immunogenicity while assuring a proper secondary structure responsible for the activity of the peptides.

[0046] In certain embodiments, the peptides are identified to “consist of’ a recited sequence, in which case the peptide includes only the recited amino acid sequence(s) without any extraneous amino acids at the N- or C-terminal ends thereof. To the extent that a recited sequence is in the form of a consensus sequence where one or more of the denoted X or Xaa residues can be any of one or more amino acids, then multiple peptide sequences are embraced by a peptide consisting of such a recited sequence.

[0047] In certain other embodiments, peptides are identified to “consist essentially of’ a recited sequence, in which case the peptide includes the recited amino acid sequence(s) optionally with one or more extraneous amino acids at the N- and / or C-terminal ends thereof, which extraneous amino acids do not materially alter the ability of the peptide or peptoid to align with ApoE and / or Ap , and thereby interfere with ApoE and Ap binding. By ‘material alteration’, it is intended to mean that there is less than 20% variation, less than 15% variation, less than 10% variation, or less than 5% variation in the ability of the peptide or peptoid to align with ApoE and / or Ap , and thereby interfere with ApoE and Ap binding, when comparing a peptide possessing the one or more extraneous amino acids to an otherwise identical peptide lacking the one or more extraneous amino acids. In certain embodiments, the number of extraneous amino acids at the N- or C-terminal ends is up to 3 amino acids at one or both ends, up to 2 amino acids at one or both ends, or not more than 1 amino acid at one or both ends.

[0048] Wherever the word ‘about’ is employed herein, for example in the context of amounts, such as concentrations and / or doses of the peptides, molecular weights or pHs, it will be appreciated that such variables are approximate and as such may vary by ± 10%, for example ± 5% and preferably ± 2% (e.g. ± 1%) from the numbers specified herein. In this respect, the term ‘about 10%’ means e.g. ±10% about the number 10, i.e. between 9% and 11%.

[0049] According to one embodiment, the peptide or peptoid, when feasible, includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O (SEQ ID NO: 1), whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue or polar amino acid residue;X3 is a hydrophobic amino acid residue;X4is a hydrophobic amino acid residue;X5 is a positively charged amino acid residue;Xe is a positively charged amino acid residue;X7 is a hydrophobic amino acid residue;Xs is a negatively charged amino acid residue;X9 is a hydrophobic amino acid residue;X10 is a negatively charged amino acid residue; and wherein the peptide optionally comprises a C-terminal proline residue.

[0050] In the peptide according to SEQ ID NO: 1, it is contemplated that each hydrophobic amino acid residue at positions Xi, X3, X4, X7 and X9, and optionally at position X2, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), NMet, cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each positively charged amino acid residue at X5 and Xe is independently selected from lysine (Lys), arginine (Arg), and histidine (His); each negatively charged amino acid residue at Xs and Xio is independently selected from aspartic acid (Asp) and glutamic acid (Glu); and each polar amino acid residue, optionally at position X2, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

[0051] Exemplary peptides according to SEQ ID NO: 1 are shown in Table 1 below:Table 1

[0052] When feasible, also contemplated are peptoid variants of the peptides recited in Table 1.

[0053] These peptides contain 10 or 11 amino acid residues, and are designed to bind one or both of ApoE in the pocket, containing residues 127 to 137 (see Moon et al., “Human Apolipoprotein E Isoforms Are Differentially Sialylated and The Sialic Acid Moiety in ApoE2 Attenuates ApoE2-Ap Interaction and Ap Fibrillation,” Neurobiology of Disease 164: 105631 (2022), which is hereby incorporated by reference in its entirety), and Ap residues 21-24 (see Yang et al., “Cryo-EM Structures of Amyloid-P 42 Filaments from Human Brains,” Science 375: 167-172 (2022), which is hereby incorporated by reference in its entirety). The choice of Pro or Vai in position 7 depends on the need to avoid a P bend and have a regular hydrophobic amino acid or to protect the adjacent Lys / Arg residues, which are a prime target of tryptic enzymes. Specifically, the strategy of the Pro after the repeated positively charged amino acids, either Arg or Lys, is similar to its use in human IgGs to protect the hinge region (Berasain et al., “Specific Cleavage Sites on Human IgG Subclasses By Cruzipain, the Major Cysteine Proteinase from Trypanosoma cruzi,". Mol Biochem Parasitol. 130(1 ):23 -9 (2003); Berasain et al., “Fasciola hepatica. Parasite-secreted Proteinases Degrade All Human IgG Subclasses: Determination of the Specific Cleavage Sites and Identification of the Immunoglobulin Fragments Produced,” Exp Parasitol 94(2):99-l 10 (2000)(erratwm in. Exp Parasitol 100(3):208 (2002)), each of which is hereby incorporated by reference in its entirety). The same strategy is valid for the carboxyl-end, where a terminal Pro residue can prevent early catabolism due to carboxypeptidases.

[0054] According to one embodiment, the peptide or peptoid, when feasible, includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11-X12-X13--X14 (SEQ ID NO: 2) whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue;X3 is a negatively charged amino acid residue;X4 is a positively charged amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a positively charged amino acid residue;X7 is a positively charged amino acid residue;Xs is a hydrophobic amino acid residue;X9 is a hydrophobic amino acid residue or polar amino acid residue;X10 is a negatively charged amino acid residue;Xu is a negatively charged amino acid residue;X12 is a hydrophobic amino acid residue;X13 is a hydrophobic amino acid residue; and X14 is a hydrophobic amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

[0055] In the peptide according to SEQ ID NO: 2, it is contemplated that each hydrophobic amino acid residue at positions Xi, X2, X5, Xs, X12, X13, and X14, optionally at position X9, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr) and tryptophan (Trp); each positively charged amino acid residue at X4, Xe, and X? is independently selected from lysine (Lys), arginine (Arg), and histidine (His); each negatively charged amino acid residue at X3, X10, and Xu is independently selected from aspartic acid (Asp) and glutamic acid (Glu); and each polar amino acid residue, optionally at position X9, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

[0056] Exemplary peptides according to SEQ ID NO: 2 are shown in Table 2 below:Table 2

[0057] When feasible, also contemplated are peptoid variants of the peptides recited in Table 2.

[0058] These peptides contain 14 or 15 amino acid residues, and are designed to bind one or both of ApoE (at residues 148 to 162) and Ap (at residues 21-24). These peptides will tightly bind in a key -lock manner through the sequence around the Argl58, thereby disrupting the pocket and the pathogenic interaction to the Ap. The Pro or Vai in position 8 should be evaluated to balance either the structure or the protection of the Arg / Lys-Arg / Lys from tryptic proteases as described above for the peptides according to SEQ ID NO: 1 and Table 1. Positions 5-9 should also bind Ap around residues 21-24. The carboxyl-terminal Pro residue can prevent early catabolism due to carboxypeptidases.

[0059] According to one embodiment, the peptide or peptoid, when feasible, includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11 (SEQ ID NO: 3) whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue or polar amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a hydrophobic amino acid residue;X5 is a negatively charged amino acid residue;Xe is a negatively charged amino acid residue;X?is a hydrophobic amino acid residue;Xs is a negatively charged amino acid residue;X9 is a negatively charged amino acid residue;X10 is a negatively charged amino acid residue; and Xu is a hydrophobic amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

[0060] In the peptide according to SEQ ID NO: 3, it is contemplated that each hydrophobic amino acid residue at positions Xi, X3, X4, X7, and Xu, and optionally at position X2, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each negatively charged amino acid residue at X5, Xe, Xs, X9, and Xio is independently selected from aspartic acid (Asp), and glutamic acid (Glu); and each polar amino acid residue, optionally at position X2, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

[0061] Exemplary peptides according to SEQ ID NO: 3 are shown in Table 3 below:Table 3

[0062] When feasible, also contemplated are peptoid variants of the peptides recited in Table 3.

[0063] These peptides contain 11 or 12 amino acid residues, and are designed to mimic the structure of Ap 18-24. However, the most important feature is its potential for binding to the “signal tagged” repetition Leu-Arg-Lys (141-143 and 144-146 in ApoE) with the “evolutionary” backward “signal tagged” Leu-Arg-Lys (148-146). This promotes binding ApoE in a critical position, which includes residues 138 to 149. As with the previously described peptides, the carboxyl-terminal Pro residue can prevent early catabolism due to carboxypeptidases.

[0064] According to one embodiment, the peptide or peptoid, when feasible, includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11-X12-X13-X14 (SEQ ID NO: 4) whereinXi is optional and, when present, is a hydrophobic amino acid residue;X2 is optional and, when present, is a positively charged amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a negatively charged amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a negatively charged amino acid residue;X?is a hydrophobic amino acid residue;Xs is a positively charged amino acid residue;X9 is a positively charged amino acid residue;X10 is a hydrophobic amino acid residue;Xu is optional and, when present, is a hydrophobic amino acid residue;X12 is optional and, when present, is a hydrophobic amino acid residue;X13 is optional and, when present, is a hydrophobic amino acid residue; andX14 is optional and, when present, is a hydrophobic amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

[0065] In the peptide according to SEQ ID NO: 4, it is contemplated that each hydrophobic amino acid residue at positions X3, X5, X7, and X10, and optionally at positions Xi, Xu, X12, X13, and X14, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu),isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr) and tryptophan (Trp); each negatively charged amino acid residue at X4 and Xe is independently selected from aspartic acid (Asp), and glutamic acid (Glu); and each positively charged amino acid residue at Xs and X9, and optionally at position X2, is independently selected from lysine (Lys), arginine (Arg), and histidine (His).

[0066] Exemplary peptides according to SEQ ID NO: 4 are shown in Table 4 below:Table 4

[0067] When feasible, also contemplated are peptoid variants of the peptides recited in Table 4.

[0068] These peptides contain 8 to 15 amino acid residues, and are designed to bind one or both of ApoE and Ap. Residues 3-11 in SEQ ID NO: 4 are designed to reverse the orientation and spacing of charged and hydrophobic residues in SEQ ID NO: 1. By using the reverse template of SEQ ID NO: 1, an anti-parallel alpha-helix that pairs with the ApoE region containing residues 235-247 is obtained. These peptides should destabilize the pairing of both alpha-helixes in ApoE. As with the previously described peptides, the carboxyl-terminal Pro residue can prevent early catabolism due to carboxypeptidases. In addition, the subset of peptides that contain the optional PFFA residues near their carboxyl-terminal end are also believed to block the Ap binding to ApoE and prevent fibrillization; hence, these peptides may possess three modes of action for interfering with AP-ApoE pathogenic interaction. Additional sequences that can be modified to include PFFA residues near their carboxyl-terminal endinclude, without limitation, SEQ ID NOS: 955-960, 979-984, 1003-1008, 1027-1032, 1051- 1056, 1075-1080, 1099-1104, 1117-1140, 1147-1152, 1159-1164, 1171-1176, 1183-1188, 1195- 1200, 1207-1212, and 1219-1248. By way of example, SEQ ID NO: 1508 differs from SEQ ID NO: 960 by the addition of PFFA residues at the carboxyl-terminal end.

[0069] According to one embodiment, the peptide or peptoid, when feasible, includes the amino acid sequence of:X1-X2-X3-X4-X5-X6-X7-X8-X9-X1O-X11-X12-X13-X14 (SEQ ID NO: 1507) whereinXi is a hydrophobic amino acid residue;X2 is a positively charged amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a hydrophobic amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a hydrophobic amino acid residue;X7 is a negatively charged amino acid residue;Xs is a hydrophobic amino acid residue;X9 is a negatively charged amino acid residue;X10 is a hydrophobic amino acid residue or polar amino acid residue;Xu is a hydrophobic amino acid residue;X12 is a hydrophobic amino acid residue; andX13 is a positively charged amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

[0070] In the peptide according to SEQ ID NO: 1507, it is contemplated that each hydrophobic amino acid residue at positions Xi, X3, X4, X5, Xe, Xs, Xu, X12, and optionally at position X10, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr) and tryptophan (Trp); each negatively charged amino acid residue at X7 and X9 is independently selected from aspartic acid (Asp) and glutamic acid (Glu); each positively charged amino acid residue at X2 and X13 is independently selected from lysine (Lys), arginine (Arg), and histidine (His); and each polar amino acid residue, optionally at position X10, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

[0071] Exemplary peptides according to SEQ ID NO: 1507 are shown in Table 5 below:Table 5

[0072] When feasible, also contemplated are peptoid variants of the peptides recited in Table 5.

[0073] These peptides contain 13 or 14 amino acid residues, and are designed to bind ApoE region 254-266 / 267 and destabilize the pairing of both alpha-helixes in ApoE. As with the previously described peptides, the carboxyl-terminal Pro residue can prevent early catabolism due to carboxypeptidases.

[0074] The peptides of the disclosure can be synthesized using standard peptide synthesis operations. These include both FMOC (9-Fluorenylmethyloxy-carbonyl) and tBoc (tert-Butyl oxy carbonyl) synthesis protocols that can be carried out on automated solid phase peptide synthesis instruments including, without limitation, the Applied Biosystems 431 A, 433 A synthesizers and Peptide Technologies Symphony or large scale Sonata or CEM Liberty automated solid phase peptide synthesizers.

[0075] The peptoids of the disclosure can be synthesized using solid-phase synthesis protocols as well as solution phase synthesis protocols, a number of which are described in detail by Clapperton et al., “A Field Guide to Optimizing Peptoid Synthesis,” ACS Polym.Au 2(6):417-429 (2022), which is hereby incorporated by reference in its entirety.

[0076] Once the peptides or peptoids are synthesized, they are preferably purified (preferably at least about 80% or 85% pure, more preferably at least about 90% or 95% pure, most preferably at least about 99% pure) by any suitable techniques. Exemplary purification techniques include, without limitation, gel filtration, ion exchange chromatography, hydrophobic interaction chromatography, affinity chromatography, reverse phase chromatography, and combinations thereof.

[0077] Either before or after purification, modifications of the peptides or peptoids is also contemplated. Such modifications can be used to enhance the efficacy of the peptides or peptoids, improve systemic delivery thereof, and / or enhance serum half-life. Exemplary modifications include, without limitation, controlled glutaraldehyde polymerization (Goni etal., “Production of Monoclonal Antibodies to Pathologic P-sheet Oligomeric Conformers in Neurodegenerative Diseases,” Sci Rep 7:9881 (2017), which is hereby incorporated by reference in its entirety), as well as PEGylation, N-alkylation, mannosylation, and N-linked and O-linked glycosylation (Mehrotra et al., “Recent Progress in the Oral Delivery of Therapeutic Peptides andProteins: Overview of Pharmaceutical Strategies to Overcome Absorption Hurdles,” Adv Pharm Bull. 14(1): 11—33 (online Aug 2023), which is hereby incorporated by reference in its entirety).

[0078] A further aspect of the disclosure relates to pharmaceutical compositions that include a pharmaceutically acceptable carrier and one or more peptides or peptoids, which are each present in an effective amount, preferably in a purified form.

[0079] In certain embodiments, more than one peptide or peptoid can be included in the pharmaceutical compositions. The one or more peptides or peptoids can possess different amino acid sequences but target conformational disruption of the same apoE / Ap interaction sites. Alternatively, the one or more peptides or peptoids can possess different amino acid sequences and target disruption of two or more distinct apoE / Ap interaction sites.

[0080] In certain embodiments, two or more peptides are included in the pharmaceutical composition. By way of the accompanying examples, the use of two peptides that target disruption of distinct ApoE / Ap interaction sites — Peptide 127 (SEQ ID NO: 133) and Peptide 148 (SEQ ID NO: 518) — is shown to be effective in producing cognitive rescue in several behavioral tests. Other combinations of peptides include any peptide from Table 1 in combination with one or more peptides from Tables 2-5, any peptide from Table 2 in combination with one or more peptides from Table 3-5, any peptide from Table 3 in combination with one or more peptides from Table 4 or 5, and any peptide from Table 4 in combination with one or more peptides of Table 5.

[0081] In certain embodiments, two or more peptoids are included in the pharmaceutical composition. In other embodiments, at least one peptide and at least one peptoid are included in the pharmaceutical composition.

[0082] Pharmaceutically-acceptable formulations may thus comprise one or more peptides or peptoids as disclosed herein in admixture with a pharmaceutically-acceptable diluent or carrier, which may be selected with due regard to the intended route of administration and standard pharmaceutical practice. Such pharmaceutically-acceptable carriers may be chemically inert to the one or more peptides or peptoids and may have no detrimental side effects or toxicity under the conditions of use. Such pharmaceutically-acceptable carriers may also impart an immediate, or a modified, release of the one or more peptides or peptoids from the composition.

[0083] Suitable pharmaceutical formulations may be commercially available (z.e., previously known to be useful for peptide or peptoid delivery) or otherwise prepared according to techniques that are described in the literature, for example, Remington The Science and Practice of Pharmacy, 22ndedition, Pharmaceutical Press (2012) and Martindale - The Complete Drug Reference, 38thEdition, Pharmaceutical Press (2014), each of which is hereby incorporated by reference in its entirety. Otherwise, the preparation of suitable formulations including one ormore peptides or peptoids as disclosed herein may be achieved by the skilled person using routine techniques.

[0084] The pharmaceutical composition may be administered systemically, for example orally, intravenously or intraarterially (including by intravascular and other perivascular devices / dosage forms (e.g. stents)), intramuscularly, cutaneously, subcutaneously, transmucosally (e.g. sublingually or buccally), intramucosally, intradermally, transdermally, nasally, pulmonarily (e.g. tracheally or bronchially), intraperitoneally, lymphatically, directly to the central nervous system by injection or catheter, or by any other parenteral route, in the form of a pharmaceutical preparation comprising the peptides or peptoids in pharmaceutically acceptable dosage form(s).

[0085] Formulations for injection (whether for systemic, intradermal, intramucosal, subcutaneous and / or intramuscular administration, or otherwise) may thus be in the form of an aqueous formulation such as an a suspension and / or, more preferably a solution (e.g. an (optionally) buffered aqueous formulation, such as a physiological saline-containing formulation, a phosphate-containing formulation, an acetate-containing formulation, or a borate-containing formulation) or a freeze-dried powder that may be reconstituted with a vehicle, such as an aqueous vehicle prior to use (e.g. injection).

[0086] Formulations for injection may include other suitable excipients known to those skilled in the art, such as solvents (e.g. water), co-solvents, solubilizing agents (e.g. cyclodextrins), wetting agents, suspending agents, emulsifying agents, thickening agents, chelating agents, antioxidants, reducing agents, antimicrobial preservatives, bulking agents and / or protectants.

[0087] Formulations for injection are preferably buffered by standard techniques to physiologically-acceptable pH values (e.g. pHs of between about 4.5 and about 9.5, e.g. about 6 and about 9, such as between about 6.5 and about 8.5) using buffers and / or pH modifiers as described herein, and / or may further comprise tonicity-modifying agents (such as sodium chloride).

[0088] Administration by injection is also useful for administering the peptides or peptoids of the disclosure, in the form of a solution or suspension into, e.g., the dermis (such as intradermal or subcutaneous injection), the mucosa (i.e., intramucosal injection), a joint cavity, or intrathecally into the cerebrospinal fluid.

[0089] In certain embodiments, the pharmaceutical formulation may include nanoparticles (e.g., solid-lipid nanoparticles and polymer nanoparticles), carbon nanotubes, liposomes, micelles, or dendrimers that encapsulate, or are linked to, one or more of the disclosed peptides or peptoids. See Mittal et al., “Nanotechnology -based Drug Delivery for the Treatment of CNSDisorders,” Transl. Neurosci. 13(l):527-546 (2022), which is hereby incorporated by reference in its entirety. These pharmaceutical formulations are preferably injectable, inhalable, or suitable for intranasal or parenteral delivery.

[0090] To facilitate oral delivery, while also protecting the one or more peptides or peptoids from destruction in the gastrointestinal system, the pharmaceutical formulation may include one or more of (i) an enteric coating that surrounds or encapsulates a tablet or capsule, (ii) excipients such as absorption enhancers (e.g., medium-chain fatty acids, lectins), surfactants, permeation enhancers (e.g., chelating agents, surfactants or detergents, bile salts, salicylates, toxins, venom extract, fatty acids, various polymers), and / or (iii) carrier systems including nanoparticles, lipid- based carrier systems (e.g., liposomes, nanoemulsions, solid lipid nanocarriers), micelles, microspheres, self-nanoemulsifying drug delivery systems (or SNEDDS, which are a characteristic mixture of lipid phase, surfactant, and co-surfactant(s)), and Emisphere™ (which is a drug delivery system that use sodium N-(8-(2-hydroxybenzoyl) amino caprylate)). See Mehrotra et cd.. “Recent Progress in the Oral Delivery of Therapeutic Peptides and Proteins: Overview of Pharmaceutical Strategies to Overcome Absorption Hurdles,” Adv. Pharm Bull.14(1): 11—33 (2024), which is hereby incorporated by reference in its entirety.

[0091] The use of an implant to deliver one or more of the disclosed peptides or peptoids is also contemplated. These implants may use one or more biodegradable and biocompatible polymers including, without limitation, poly(lactic-co-glycolic acid) (PLGA), A-methyl-2- pyrrolidone (NMP), poly(glycolic acid) (PGA), poly(lactic acid) (PLA), polycaprolactone, poly di ox an one, polyesters, poly (ortho esterjs, polyanhydrides, and biodegradable polycarbonates, hydroxy acid, polyanhydride, polyamide, poly(ester amide), polyphosphoester, poly(alkyl cyanoacrylate), and natural sugars such as chitosan. See Rashid etal., “Recent Advances in Micro- and Nano-Drug Delivery Systems Based on Natural and Synthetic Biomaterials,” Polymers 15(23):4563 (2023), which is hereby incorporated by reference in its entirety. By way of example, PLGA-based implants may have various forms, such as implanted microspheres / nanoparticles, injectable in-situ hydrogels, and solid implants.

[0092] Administration of the disclosed peptides or peptoids, as well as pharmaceutical compositions containing the same, may be continuous or intermittent. The mode of administration may also be determined by the timing and frequency of administration, but is also dependent, in the case of therapeutic uses, on the severity of the condition.

[0093] Depending on the disorder, and the patient, to be treated, as well as the route of administration, peptides or peptoids of the disclosure may be administered at varying therapeutically effective doses to a patient in need thereof.

[0094] Similarly, the amount of the disclosed peptides or peptoids in a formulation will depend (in the case of therapeutic treatment) on the severity of the condition, and on the patient, to be treated, but may be determined by the skilled person.

[0095] In any event, the medical practitioner, or other skilled person, will be able to determine routinely the actual dosage, which will be most suitable for an individual patient, depending on the route of administration. The dosages mentioned herein are exemplary of the average case; there can, of course, be individual instances where higher or lower dosage ranges are merited, and such are within the scope of this invention.

[0096] Doses may be administered between up to several times daily, such as once daily, twice daily, three times daily, or four times daily. Alternatively, doses may be administered periodically (e.g., once weekly, twice weekly, once monthly, twice monthly, etc.).

[0097] Effective amounts of the peptide or peptoid will depend on, e.g., the mode of administration, the disease or condition being treated, and the stage of the disease. By way of example only, suitable peptide concentrations may range from about 0.1 pM to about 10 mM, preferably about 1 pM to about 5 mM, about 10 pM to about 2 mM, or about 50 pM to about 1 mM. The volume of the composition administered, and thus, dosage of the peptide administered can be adjusted by one of skill in the art to achieve optimized results.

[0098] In some embodiments, the effective amount used for the administering is between about 0.1 and 2000 mg / dose. For example, in some embodiments the effective amount used for the administering is between about 0.1 and 10 mg / dose, about 0.1 and 50 mg / dose, about 0.1 and 100 mg / dose, about 0.1 and 150 mg / dose, about 0.1 and 200 mg / dose, about 0.1 and 250 mg / dose, about 0.1 and 300 mg / dose, about 0.1 and 350 mg / dose, about 0.1 and 400 mg / dose, about 0.1 and 450 mg / dose, about 0.5 and 10 mg / dose, about 0.5 and 50 mg / dose, about 0.5 and 100 mg / dose, about 0.5 and 150 mg / dose, about 0.5 and 200 mg / dose, about 0.5 and 250 mg / dose, about 0.5 and 300 mg / dose, about 0.5 and 350 mg / dose, about 0.5 and 400 mg / dose, about 0.5 and 450 mg / dose, about 0.5 and 500 mg / dose, about 1 and 10 mg / dose, about 1 and 50 mg / dose, about 1 and 100 mg / dose, about 1 and 150 mg / dose, about 1 and 200 mg / dose, about 1 and 250 mg / dose, about 1 and 300 mg / dose, about 1 and 350 mg / dose, about 1 and 400 mg / dose, about 1 and 450 mg / dose, about 1 and 500 mg / dose, about 10 and 50 mg / dose, about 10 and 100 mg / dose, about 10 and 150 mg / dose, about 10 and 200 mg / dose, about 10 and 250 mg / dose, about 10 and 300 mg / dose, about 10 and 350 mg / dose, about 10 and 400 mg / dose, about 10 and 450 mg / dose, about 10 and 500 mg / dose, about 50 and 100 mg / dose, about 50 and 150 mg / dose, about 50 and 200 mg / dose, about 50 and 250 mg / dose, about 50 and 300 mg / dose, about 50 and 350 mg / dose, about 50 and 400 mg / dose, about 50 and 450 mg / dose, about 50 and 500 mg / dose, about 100 and 150 mg / dose, about 100 and 200 mg / dose, about 100 and 250 mg / dose, about 100and 300 mg / dose, about 100 and 350 mg / dose, about 100 and 400 mg / dose, about 100 and 450 mg / dose, about 100 and 500 mg / dose, about 150 and 200 mg / dose, about 150 and 250 mg / dose, about 150 and 300 mg / dose, about 150 and 350 mg / dose, about 150 and 400 mg / dose, about 150 and 450 mg / dose, about 150 and 500 mg / dose, about 200 and 250 mg / dose, about 200 and 300 mg / dose, about 200 and 350 mg / dose, about 200 and 400 mg / dose, about 200 and 450 mg / dose, about 200 and 500 mg / dose, about 250 and 300 mg / dose, about 250 and 350 mg / dose, about 250 and 400 mg / dose, about 250 and 450 mg / dose, about 250 and 500 mg / dose, about 300 and 350 mg / dose, about 300 and 400 mg / dose, about 300 and 450 mg / dose, about 300 and 500 mg / dose, about 350 and 400 mg / dose, about 350 and 450 mg / dose, about 350 and 500 mg / dose, about 400 and 450 mg / dose, about 400 and 500 mg / dose, about 450 and 500 mg / dose, or about 500 and 1000 mg / dose. The dose can be adjusted lower to identify the minimal effective dose, or tailored higher or lower depending on the needs of the patient.

[0099] In any event, the dose administered to a mammal, particularly a human, in the context of the present invention should be sufficient to affect a therapeutic response in the mammal over a reasonable timeframe (as described hereinbefore). One skilled in the art will recognize that the selection of the exact dose and composition and the most appropriate delivery regimen will also be influenced by inter alia the pharmacological properties of the formulation, and the physical condition and mental acuity of the recipient, as well as the age, condition, body weight, sex and response of the patient to be treated, as well as genetic differences between patients.

[0100] Also contemplated herein are therapeutic systems that include, as separate compositions, a first composition containing a peptide or peptoid of the present invention in a suitable carrier, and a second composition containing an effective amount of another therapeutic agent in a suitable carrier. These separate compositions can be coadministered according to a therapeutic protocol and dosing schedule. The other therapeutic agent can be any other known or later-developed agent for the treatment of Alzheimer’s Disease or other conditions involving formation of aberrant amyloid deposits, any other known or later-developed agent for the treatment of tauopathies. For examples, the peptides or peptoids disclosed herein can be used in combination with one or more immunotherapies that target Ap (e.g., an anti-Ap monoclonal antibody or active fragment thereof) or one or more immunotherapies that target pTau (e.g., an anti-tau monoclonal antibody or active fragment thereof).

[0101] Another aspect of the disclosure relates to a method of blocking Ap / ApoE interaction. The method includes the steps of contacting an ApoE protein with any peptide or peptoid described herein, whereby the peptide or peptoid binds to the ApoE protein and / or binds to Ap and thereby blocks interaction of ApoE with Ap.

[0102] Another aspect of the disclosure relates to a method of inhibiting A protein oligomerization. The method comprises contacting an ApoE protein and / or Ap protein with any peptide or peptoid described herein, whereby the peptide or peptoid binds to the ApoE and / or Ap protein and thereby blocks interaction of the ApoE protein with an Ap protein.

[0103] According to these aspects of the disclosure, in some embodiments the blocking of Ap / ApoE interaction and the inhibiting of Ap protein oligomerization can be carried out in vitro.

[0104] According to these aspect of the disclosure, in some embodiments, discussed infra, when a patient is treated for Alzheimer’s Disease or other conditions involving formation of aberrant amyloid deposits, the blocking of Ap / ApoE interaction and the inhibiting of Ap protein oligomerization is carried out in vivo.

[0105] In some embodiments, the ApoE protein is an ApoE4 protein.

[0106] In some embodiments, one of the disclosed peptides is used for the contacting. In some embodiments, one of the disclosed peptoids is used for the contacting. In other embodiments, two or more peptides or peptoids are used for the contacting, preferably where the two or more peptides or peptoids bind to different locations on the ApoE protein.

[0107] Another aspect of the disclosure relates to a method of inhibiting Ap plaque and / or tau aggregate formation. The method comprises contacting an ApoE protein and / or Ap protein with a peptide or peptoid as disclosed herein, whereby the peptide or peptoid binds to the ApoE protein and / or Ap protein and inhibits Ap plaque formation. By virtue of inhibiting Ap plaque formation, the contacting will also indirectly inhibit downstream tau aggregate formation.

[0108] As used herein, the term “tau” is synonymous with the tau protein and refers to any of the Tau protein isoforms (identified in, for example, UniProt as P10636, 1-9, which is hereby incorporated by reference in its entirety). Tau is a soluble microtubule-associated protein that is dynamically phosphorylated and dephosphorylated by a host of kinase enzymes during the cell cycle. Tau’s ability to stabilize microtubules is dependent on the extent of its phosphorylation. In its dephosphorylated form, the protein is able to interact with tubulin to stabilize microtubules and promote tubulin assembly into microtubules (which form the cytoskeleton of the cell and are the major constituents of the mitotic spindles that pull apart eukaryotic chromosomes in mitosis). In its phosphorylated form, tau is able to dissociate from microtubules, thereby permitting mitosis to occur. Thus, the phosphorylation of tau acts as a direct microtubule associationdissociation switch within the neuron (Pedersen et al., “Tau Immunotherapy for Alzheimer’s Disease,” Trends Mol. Med. 21(6): 394-402 (2015), which is hereby incorporated by reference herein in its entirety).

[0109] It is well established that a number of interactions between Ap and tau exist in the development of Alzheimer’s Disease. Ap has a driving effect on tau phosphorylation.Hyperphosphorylated tau dislodges from microtubules, and its affinity for other tau monomers induces binding between individual tau proteins to form tau oligomers. This process is mediated by Ap-activated CDK-5 and GSK-3p. These tau oligomers potentiate neuronal damage, which leads to neurodegeneration . Truncated tau lacking its C-terminal 20 amino acids assembles more rapidly into filaments than full-length tau. Ap also induces the generation of a 17-kDa tau fragment via the activation of calpain-1 in hippocampal neurons. The expression of this tau fragment leads to neurite degeneration and cell death in neurons and nonneuronal cell types. Cleaved tau protein self-aggregates and misfolds, and these self-aggregated and misfolded products are further assembled into soluble oligomers. Tau oligomers, which are an intermediate form of tau that occurs prior to neurofibrillary tangle formation, are toxic. See Zhang et al., “Interaction between Ap and Tau in the Pathogenesis of Alzheimer's Disease,” Ini. J. Biol. Sei. 17(9): 2181-2192 (2021), which is hereby incorporated by reference in its entirety.

[0110] Therefore, the methods of the present invention can further be used to treat a subject having a condition or disease that is associated with, or resulting from, Ap plaque formations or deposits. Such conditions include, but are not limited to, Alzheimer's disease, diffuse Lewy body disease, Down syndrome, cerebral amyloid angiopathy (CAA), British familial dementia, and Danish familial dementia, Creutzfeld-Jakob disease and related prion diseases, frontotemporal dementia (FTD).[oni] A patient having (or suspected of having) one of these conditions can be treated by administering to the patient an effective amount of a peptide or peptoid described herein, or a pharmaceutical composition described herein, wherein the administering is effective to inhibit amyloid plaque and / or associated tau aggregate formation, and thereby treat the patient. The subject or patient to which the present invention may be applicable can be any vertebrate species, preferably mammalian, including but not limited to, cows, horses, sheep, pigs, fowl (e.g., chickens), goats, cats, dogs, hamsters, mice, rats, rabbits, monkeys, chimpanzees, and humans. In a preferred embodiment, the patient is a human.

[0112] In carrying out the methods of the present disclosure, it is preferable to select a patient having or at risk of having one of the above-identified diseases or conditions. Subjects amenable to treatment include individuals at risk of disease but not showing symptoms, as well as patients presently showing symptoms. In the case of Alzheimer's disease, virtually anyone is at risk of suffering from Alzheimer's disease. Therefore, the present methods can be administered prophylactically to the general population without the need for any assessment of the risk of the subject patient. The present methods are especially useful for individuals who do have a known genetic risk of Alzheimer's disease. Such individuals include those having relatives who have experienced this disease, and those whose risk is determined by analysis of genetic orbiochemical markers. Genetic markers of risk toward Alzheimer's disease include mutations in the APP gene, particularly mutations at position 717 and positions 670 and 671 referred to as the Hardy and Swedish mutations, respectively. Other markers of risk include mutations in the presenilin genes, PSI and PS2, and ApoE4 gene, a family history of AD, and hypercholesterolemia or atherosclerosis. Individuals presently suffering from Alzheimer's disease can be recognized from characteristic dementia by the presence of risk factors described above. In addition, a number of diagnostic tests are available for identifying individuals who have AD. These include measurement of CSF tau and Ap42 levels. Elevated tau and decreased AP42 levels signify the presence of AD. Individuals suffering from Alzheimer's disease can also be diagnosed by Alzheimer's Disease and Related Disorders Association criteria.

[0113] The efficacy of the treatment disclosed herein can be determined by evaluating the Alzheimer's Disease (AD) symptoms of the patient and / or by measuring the Ap concentration in bodily fluids or organs (e.g., the patient’s blood or cerebrospinal fluid). Blood or cerebrospinal fluid Ap peptide measurements can be conducted using any method known in the art. For example, blood Ap peptide measurements can involve taking a blood or cerebrospinal fluid sample, optionally separating serum from red blood cells when a blood sample is used, and then using an appropriate assay to detect Ap peptide in the sample. Exemplary assay formats include, without limitation, radioimmunoassay, enzyme linked immunosorbent assay (ELISA) or chromatographic analysis. Alternatively, efficacy of treatment can be assessed via imaging techniques to assess amyloid plaque burden and spatial distribution of the plaques. A radiotracer called Pittsburgh Compound B is FDA approved for use in PET scan for the diagnosis of Alzheimer disease. This molecule preferentially targets and binds to amyloid beta plaques with high affinity and specificity and can estimate the plaque burden and distribution of Alzheimer disease.

[0114] In asymptomatic patients, treatment can begin at any age (e.g., 10, 20, 30 years of age). Usually, however, it is not necessary to begin treatment until a patient reaches 40, 50, 60, or 70 years of age. Treatment typically entails multiple dosages over a period of time. Treatment can be monitored by the assays or imaging techniques described above. In the case of potential Down's syndrome patients, treatment can begin antenatally by administering therapeutic agent to the mother or shortly after birth.

[0115] As used herein, “treat” or “treatment” generally refers to either (i) a reduction in the concentration or amount of Ap peptide or toxic pTau oligomers in a bodily fluid and / or organ, or (ii) a slowing of disease progression, or (iii) a slowing of the symptom progression (z.e., reduce the rate of cognitive decline), or (iv) improvement of disease symptoms (z.e., partially reversecognitive decline). “Treatment” also includes prophylactic treatment to those at risk for amyloid diseases, e.g., familial AD.EXAMPLES

[0116] The following Examples are presented to illustrate various aspects of the present application, but are not intended to limit the scope of the claimed application.Example 1 - Assessment of Peptide Disruption of ApoE4 / A|J42 Binding by Surface Plasmon Resonance

[0117] Measurements of Ap42 and ApoE4 binding in the absence or presence of one of peptides CSQNKKPDLDP (SEQ ID NO: 133) and IIDRARRVSEDVGLP (SEQ ID NO: 518) was determined by surface plasmon resonance (SPR) using previously described methods (Goni et al., “Anti-P-sheet Conformation Monoclonal Antibody Reduces Tau and Ap Oligomer Pathology in an Alzheimer's Disease Model,” Alzheimers Res Ther. 10(1): 10 (2018), which is hereby incorporated by reference in its entirety).

[0118] Experiments were performed using a Reichert SR7000DC refractometer system (Reichert Technologies, Depew, NY, USA) equipped with a CM5 sensor chip. Acetate buffer was used for immobilization of AP42 (ligand). Carboxymethyl dextran on a CM5 gold sensor chip was activated by injecting a solution of 10 mg / ml NHS and 40 mg / ml l-ethyl-3-(3- dimethylaminopropyl)carbodiimide over the sensor chip surface for 7 minutes at a flow rate of 20 pl / minute. Then the ligand, 50 pg / ml hexafluoroisopropanol -treated synthetic peptide AP42 (monomeric AP) was immobilized by injection over the surface for 20 minutes. The unreacted sites on the sensor chip surface were blocked by injection of 1 M ethanolamine, pH 8.5, for 10 minutes. Analytes containing ApoE4 alone, or ApoE4 pre-incubated for 30 minutes with CSQNKKPDLDP (Peptide 127, SEQ ID NO: 133) or IIDRARRVSEDVGLP (Peptide 148, SEQ ID NO: 518) (1 :3 ApoE:peptide molar ratio), in PBS with 0.05% Tween 20, pH 7.4, were separately injected over the surface, and the association reaction was monitored.

[0119] Fig. 2A confirms that ApoE4 binds strongly to Ap42 in the absence of any interference. However, Figs. 2B and 2C confirm that both of the tested peptides completely block the binding of ApoE4 to Ap42. The effectiveness of these peptides in blocking ApoE4 / Ap42 binding confirms that targeting of ApoE residues 127 to 137 or residues 148 to 162 as well as Ap residues 21 to 24 via peptides (having hydrophobically interspaced charged amino acids that align with the spacing of hydrophobic amino acids and oppositely charged amino acids in an ApoE protein or polypeptide) constitutes a viable approach for disrupting amyloid formation in vivo.Example 2 — Assessment of Peptide Disruption of ApoE4 / Ap42 Binding by in vitro Thioflavin Fluorescence

[0120] Thioflavin T is among the most widely used amyloid dye among the hundreds of amyloid reports published yearly, and it is widely used as an in vitro marker of amyloid formation. Detection of in vitro thioflavin fluorescence was performed on the Bio Tek Cytation 7 (Agilent, Santa Clara, CA) machine using [Ap42]:[Pep] ratios of 1 : 1 (10 pM, Fig. 3A), 1 :2 (20 pM, Fig. 3B) and 1 :5 (50 pM, Fig. 3C) with or without ApoE4. All the preparations were seeded on 96 well black plates for fluorescence, the system set for automatic readings every two minutes after brief shaking and recording and analysis in the proprietary program (Agilent). Fluorescence produced by fibrillogenesis of Ap is induced by pathological function of ApoE4, and disruption of fibrillogenesis induced by ApoE4 is revealed by significantly reduced fluorescence.

[0121] As shown in Figs. 3A-3C, Peptide 127 (SEQ ID NO: 133) displayed strong inhibition of AP42 fibrillation at 10 and 20 pM ([Ap42]:[Pep] ratios of 1 : 1 and 1 :2) with or without ApoE4. Peptide 127 blocks the conformation of the region 127-147 involved in the pathological pro-fibrillogenic function of ApoE4 on Ap. The concomitant interference with conformation on Ap 21-24 proved to be highly anti-fibrillogenic. Similarly, as shown in Figs. 4A-4B, Peptide 148 (SEQ ID NO: 518) displayed strong inhibition of Ap42 fibrillation at 10 and 20 pM ([Ap42]:[Pep] ratios of 1 : 1 and 1 :2) with or without ApoE4. Peptide 148 binding to region 148- 162 on ApoE prevents the conformational stabilization of the pathological function of ApoE4; thus, blocking the pro-fibrillogenic effect of ApoE4 on Ap. Because of the potential concomitant Ap 21-24 structure change by the middle section of Peptide 148; Peptide 148 also proved to be anti-AP-fibrillogenic.Example 3 - Assessment of Peptide Combination Therapy in Transgenic Mouse Alzheimer’s Disease Model

[0122] Mice for the Alzheimer’s Disease (AD) transgenic model SwDI (APP / PS1 Swedish - Dutch-Iowa) with knockin human apoE genes either apoE3 or apoE 4, were aged to 8 months old, genotyped to assure the Tg background; and started to be inoculated intraperitoneally (IP), 100 ug / animal / week; alternating one week with Peptide 127 (SEQ ID NO: 133) and the following week with Peptide 148 (SEQ ID NO: 158). The inoculum (200 uL / animal) was diluted in sterile saline. All stock peptides were initially diluted in sterile saline 5 mg / ml and filtered through 0.2 um disk filters. Control animals aged and inoculated with vehicle alone (sterile saline).

[0123] The inoculation regimen proceeded for 4 months, after which the animals were rested for a week and then underwent locomotor and behavioral tests that lasted until after 13 months of age. Afterwards, the animals were sedated, blood was collected by cardiac puncture and were thoroughly perfused with sterile PBS-heparin until no organ showed signs of blood. Organs; liver, kidney, heart and spleen; were collected and equal pieces were immediately frozen (for biochemistry) in a bed of dry ice covered with aluminum foil, or immerse in a formaldehyde- buffered (PLP) solution for eventual use in histochemistry. The animals were readily decapitated with scissors, the brains carefully taken and examined for the absence of red in vessels, separated in both hemispheres which one was readily frozen in dry ice (and kept with the other organs in a freezer at -80 °C) and the other immersed in PLP and kept at 4 °C for 20 hours when the solution was changed to a cryoprotectant (PBS-Glucose) for long term storage.

[0124] Compared to the control, the combination of Peptides 127 (SEQ ID NO: 133) and 148 (SEQ ID NO: 518) produced significant cognitive rescue in several behavioral tests (Barnes Maze and Radial Arm Maze, Figs. 5-6) with no differences in locomotor tests (grip strength and Rotarod, Fig. 7). Thus, the differences in test results were not due to any physical benefits, but solely due to cognitive benefits afforded by the peptide combination. Immunohistochemistry analysis of amyloid burden corroborates the behavioral results.Example 4 - Molecular Modeling Illustrates Peptide Efficacy

[0125] The Protein Data Bank (PDB) file for the ApoE N-terminal domain with PDB ID 1GS9 was opened in ICM-Pro software (Molsoft, LLC La Jolla CA). The electrostatic surface was calculated using the “View Electrostatic Potential” command in the software. The free peptides were rendered in the software using the “File:New:Peptide” command and subjected to a full conformational search and energy calculation to fold them using the “Molecular Mechanics: Sample Peptide” command. The Distance function in the software was used to measure the distance in Angstroms between the indicated amino acid residue landmarks on the structures. Biophysical complementarity between peptides 127 / 148 and the ApoE N-terminal domain 3D model was assessed using docking in the software using the “Docking” command. These modeling results are depicted in Figs. 8-11.

[0126] The residues noted in the claimed peptide designs were observed as to whether they were part of the surface or not. Distances in the folded and extended peptides were compared to the distances between target amino acid positions on the ApoE N-terminal domain to assess whether they matched geometrically. Docked peptides were observed as to their locations and whether they overlapped or not.

[0127] Although preferred embodiments have been depicted and described in detail herein, it will be apparent to those skilled in the relevant art that various modifications, additions, substitutions, and the like can be made without departing from the spirit of the invention and these are therefore considered to be within the scope of the invention as defined in the claims which follow.

Claims

WHAT IS CLAIMED:

1. A peptide or peptoid comprising from 8 to 20 amino acids and comprising a plurality of hydrophobic amino acids and a plurality of charged amino acids, wherein spacing of the plurality of hydrophobic amino acids and the plurality of charged amino acids aligns with the spacing of hydrophobic amino acids and oppositely charged amino acids in an ApoE protein or polypeptide.

2. The peptide or peptoid according to claim 1, wherein the peptide or peptoid aligns with ApoE when both are in an N-to-C orientation.

3. The peptide or peptoid according to claim 1, wherein the peptide or peptoid aligns with ApoE when in reverse orientations (one is N-to-C and the other is C-to-N).

4. The peptide or peptoid according to claim 1, wherein the ApoE protein or polypeptide is ApoE4.

5. The peptide or peptoid according to any one of claims 1 to 4, wherein the peptide comprises the amino acid sequence of:Xl-X2-X3-X4-X5-X6-X7-X8-X9-X10 whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue or polar amino acid residue;X3 is a hydrophobic amino acid residue;X4is a hydrophobic amino acid residue;X5 is a positively charged amino acid residue;Xe is a positively charged amino acid residue;X7 is a hydrophobic amino acid residue;Xs is a negatively charged amino acid residue;X9 is a hydrophobic amino acid residue;X10 is a negatively charged amino acid residue; and wherein the peptide optionally comprises a C-terminal proline residue.

6. The peptide or peptoid according to claim 5, wherein each hydrophobic amino acid residue at positions Xi, X3, X4, X7 and X9, and optionally at position X2, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each positively charged amino acid residue at X5 and Xe is independently selected from lysine (Lys), arginine (Arg), and histidine (His); each negatively charged amino acid residue at Xs and Xio is independently selected from aspartic acid (Asp) and glutamic acid (Glu); and each polar amino acid residue, optionally at position X2, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

7. The peptide or peptoid according to claim 5, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 5-132.

8. The peptide or peptoid according to claim 7 further comprising a proline (Pro) residue at the C-terminal end thereof.

9. The peptide or peptoid according to claim 8, which comprises the amino acid sequence of any one of SEQ ID NOS: 133-260.

10. The peptide or peptoid according to any one of claims 1 to 4, wherein the peptide comprises the amino acid sequence of:Xl-X2-X3-X4-X5-X6-X7-X8-X9-X10-Xl 1-X12-X13--X14 whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue;X3 is a negatively charged amino acid residue;X4 is a positively charged amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a positively charged amino acid residue;X7 is a positively charged amino acid residue;Xs is a hydrophobic amino acid residue;X9 is a hydrophobic amino acid residue or polar amino acid residue;X10 is a negatively charged amino acid residue;Xu is a negatively charged amino acid residue;X12 is a hydrophobic amino acid residue;X13 is a hydrophobic amino acid residue; andX14 is a hydrophobic amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

11. The peptide or peptoid according to claim 10, wherein each hydrophobic amino acid residue at positions Xi, X2, X5, Xs, X12, X13, and X14, optionally at position X9, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each positively charged amino acid residue at X4, Xe, and X?is independently selected from lysine (Lys), arginine (Arg), and histidine (His); each negatively charged amino acid residue at X3, X10, and Xu is independently selected from aspartic acid (Asp) and glutamic acid (Glu); and each polar amino acid residue, optionally at position X9, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

12. The peptide or peptoid according to claim 10, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 261-516.

13. The peptide or peptoid according to claim 12 further comprising a proline (Pro) residue at the C-terminal end thereof.

14. The peptide or peptoid according to claim 13, which comprises the amino acid sequence of any one of SEQ ID NOS: 517-772.

15. The peptide or peptoid according to any one of claims 1 to 4, wherein the peptide comprises the amino acid sequence of:Xl-X2-X3-X4-X5-X6-X7-X8-X9-X10-Xll whereinXi is a hydrophobic amino acid residue;X2 is a hydrophobic amino acid residue or polar amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a hydrophobic amino acid residue;Xs is a negatively charged amino acid residue;Xe is a negatively charged amino acid residue;X?is a hydrophobic amino acid residue;Xs is a negatively charged amino acid residue;X9 is a negatively charged amino acid residue;X10 is a negatively charged amino acid residue; andXu is a hydrophobic amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

16. The peptide or peptoid according to claim 15, wherein each hydrophobic amino acid residue at positions Xi, X3, X4, X7, and Xu, and optionally at position X2, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each negatively charged amino acid residue at X5, Xe, Xs, X9, and X10 is independently selected from aspartic acid (Asp), and glutamic acid (Glu); and each polar amino acid residue, optionally at position X2, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

17. The peptide or peptoid according to claim 15, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 773-860 thereof.

18. The peptide or peptoid according to claim 17 further comprising a proline (Pro) residue at the C-terminal end thereof.

19. The peptide or peptoid according to claim 18, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 861-948.

20. The peptide or peptoid according to any one of claims 1 to 4, wherein the peptide comprises the amino acid sequence of:Xl-X2-X3-X4-X5-X6-X7-X8-X9-X10-Xl 1-X12-X13-X14 whereinXi is optional and, when present, is a hydrophobic amino acid residue;X2 is optional and, when present, is a positively charged amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a negatively charged amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a negatively charged amino acid residue;X?is a hydrophobic amino acid residue;Xs is a positively charged amino acid residue;X9 is a positively charged amino acid residue;X10 is a hydrophobic amino acid residue;Xu is optional and, when present, is a hydrophobic amino acid residue;X12 is optional and, when present, is a hydrophobic amino acid residue;X13 is optional and, when present, is a hydrophobic amino acid residue; andX14 is optional and, when present, is a hydrophobic amino acid residue; wherein the peptide or peptoid optionally comprises a C-terminal proline residue.

21. The peptide or peptoid according to claim 20, wherein each hydrophobic amino acid residue at positions X3, X5, X7, and X10, and optionally at positions Xi, Xu, X12, X13, and X14, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each negatively charged amino acid residue at X4 and Xe is independently selected from aspartic acid (Asp) and glutamic acid (Glu); and each positively charged amino acid residue at Xs and X9, and optionally at position X2, is independently selected from lysine (Lys), arginine (Arg), and histidine (His).

22. The peptide or peptoid according to claim 20, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 949-1140.

23. The peptide or peptoid according to claim 22 further comprising a proline (Pro) residue at the C-terminal end thereof.

24. The peptide or peptoid according to claim 23, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 1141-1248.

25. The peptide or peptoid according to any one of claims 1 to 4, wherein the peptide comprises the amino acid sequence of:Xl-X2-X3-X4-X5-X6-X7-X8-X9-X10-Xl 1-X12-X13-X14whereinXi is a hydrophobic amino acid residue;X2 is a positively charged amino acid residue;X3 is a hydrophobic amino acid residue;X4 is a hydrophobic amino acid residue;X5 is a hydrophobic amino acid residue;Xe is a hydrophobic amino acid residue;X7 is a negatively charged amino acid residue;Xs is a hydrophobic amino acid residue;X9 is a negatively charged amino acid residue;X10 is a hydrophobic amino acid residue or polar amino acid residue;Xu is a hydrophobic amino acid residue;X12 is a hydrophobic amino acid residue; andX13 is a positively charged amino acid residue; wherein the peptide optionally comprises a C-terminal proline residue.

26. The peptide or peptoid according to claim 25, wherein each hydrophobic amino acid residue at positions Xi, X3, X4, X5, Xe, Xs, Xu, X12, and optionally at position X10, is independently selected from glycine (Gly), alanine (Ala), valine (Vai), leucine (Leu), isoleucine (He), proline (Pro), phenylalanine (Phe), methionine (Met), cysteine (Cys), tyrosine (Tyr), and tryptophan (Trp); each negatively charged amino acid residue at X7 and X<; is independently selected from aspartic acid (Asp) and glutamic acid (Glu); each positively charged amino acid residue at X2 and X13 is independently selected from lysine (Lys), arginine (Arg), and histidine (His); and each polar amino acid residue, optionally at position X10, is selected from serine (Ser), threonine (Thr), asparagine (Asn), and glutamine (Gin).

27. The peptide or peptoid according to claim 25, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 1249-1376.

28. The peptide or peptoid according to claim 27 further comprising a proline (Pro) residue at the C-terminal end thereof.

29. The peptide or peptoid according to claim 28, wherein the peptide comprises the amino acid sequence of any one of SEQ ID NOS: 1377-1504.

30. The peptide or peptoid according to any one of claims 1 to 29, wherein the peptide or peptoid comprises from 8 to 15 amino acids.

31. The peptide or peptoid according to any one of claims 1 to 29, wherein the peptide comprises D-amino acids.

32. The peptide or peptoid according to any one of claims 1 to 29, wherein the peptide comprises L-amino acids.

33. The peptide or peptoid according to any one of claims 1 to 29, wherein the peptide comprises a mixture of L-amino acids and D-amino acids.

34. The peptide or peptoid according to any one of claims 1 to 33, in the form of a peptide.

35. The peptide or peptoid according to any one of claims 1 to 33, in the form of a peptoid comprising N-substituted amino acids independently selected from NAla, NVal, NLeu, Nile, NPhe, NMet, NCys, NTrp, NLys, NArg, NHis, NAsp, NGlu, NSer, NThr, NAsn, and NGln.

36. A pharmaceutical composition comprising a peptide or peptoid according to any one of claims 1 to 35.

37. The pharmaceutical composition according to claim 36, which comprises two or more of the peptides or peptoids.

38. The pharmaceutical composition according to claim 36, which comprises two or more peptides.

39. The pharmaceutical composition according to claim 36, which comprises two or more peptoids.

40. The pharmaceutical composition according to claim 36, which comprises at least one peptide and at least one peptoid.

41. The pharmaceutical composition according to claim 36, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

42. The pharmaceutical composition according to claim 36, wherein the composition is in the form of a tablet, capsule, solution, suspension, gel, powder, or granules.

43. A method of blocking pathogenic Ap / ApoE interaction, the method comprising: contacting an ApoE protein and / or an Ap protein with a peptide or peptoid according to any one of claims 1 to 35, whereby the peptide or peptoid binds to the ApoE protein and / or the Ap protein disrupting its functional conformation thereby blocking pathogenic interaction of the ApoE protein with an Ap protein.

44. The method according to claim 43, wherein the ApoE protein is an ApoE4 protein.

45. The method according to claim 43, wherein the peptide is used for said contacting.

46. The method according to claim 43, wherein the peptoid is used for said contacting.

47. The method according to claim 43, wherein two or more peptides or peptoids are used for said contacting, the two or more peptides or peptoids binding to different locations on the ApoE protein.

48. The method according to claim 43, wherein said contacting is carried out in vitro.

49. The method according to claim 43, wherein said contacting is carried out in vivo.

50. A method of inhibiting Ap protein oligomerization, the method comprising: contacting an ApoE protein and / or an Ap protein with a peptide according to any one of claims 1 to 35, whereby the peptide binds to the ApoE protein and / or the Ap protein, and thereby blocks pathogenic interaction of the ApoE protein with the Ap protein or blocks the Ap region from where the P-sheet pathologic structure spreads.

51. The method according to claim 50, wherein the ApoE protein is an ApoE4 protein.

52. The method according to claim 50, wherein the peptide is used for said contacting.

53. The method according to claim 50, wherein the peptoid is used for said contacting.

54. The method according to claim 50, wherein two or more peptides or peptoids are used for said contacting, the two or more peptides or peptoids binding to different locations on the ApoE protein.

55. The method according to claim 50, wherein said contacting is carried out in vitro.

56. The method according to claim 50, wherein said contacting is carried out in vivo.

57. A method of inhibiting amyloid plaque and / or tau aggregate formation, the method comprising: contacting an ApoE protein and / or an Ap protein with a peptide according to any one of claims 1 to 35, whereby the peptide binds to the ApoE protein and / or an Ap protein, and inhibits amyloid plaque and / or subsequent tau aggregate formation.

58. The method according to claim 57, wherein the ApoE protein is an ApoE4 protein.

59. The method according to claim 57, wherein the peptide is used for said contacting.

60. The method according to claim 57, wherein the peptoid is used for said contacting.

61. The method according to claim 57, wherein two or more peptides or peptoids are used for said contacting, the two or more peptides or peptoids binding to different locations on the ApoE protein.

62. The method according to claim 57, wherein said contacting is carried out in vivo.

63. A method of treating Alzheimer’ s Disease, the method comprising: administering to a patient having Alzheimer’s Disease an effective amount of a peptide according to any one of claims 1 to 35, or a pharmaceutical composition according to any one ofclaims 36 to 42, wherein said administering is effective to inhibit amyloid plaque and / or subsequent tau aggregate formation and thereby treat the patient.

64. The method according to claim 63, wherein said administering is carried out daily.

65. The method according to claim 63, wherein said administering is carried out more than once daily.

66. The method according to claim 63, wherein the effective amount used for said administering is between about 0.1 and 500 mg / dose.

67. The method according to claim 63, wherein the peptide or a pharmaceutical composition containing the peptide is administered.

68. The method according to claim 63, wherein two or more peptides or a pharmaceutical composition containing the two or more peptides is administered.

69. The method according to claim 68, wherein the two or more peptides bind to different locations on the ApoE protein.

70. The method according to claim 63, wherein said administering is carried out orally, sublingually, intranasally, by inhalation, intravenously, intraperitoneally, intramuscularly, subcutaneously, intrathecally, intraperitoneally, or lymphatically.

71. The method according to claim 63, wherein said administering is carried out by insertion of an implant.

72. The method according to claim 63 further comprising: administering to the patient one or more of an anti-Ap monoclonal antibody or active fragment thereof and an anti-tau monoclonal antibody or active fragment thereof.

73. The method according to any one of claims 63 to 72, wherein said treating is effective to reduce a rate of cognitive decline or maintain cognitive function.

74. A method of treating a condition associated with Ap plaque formations or deposits, the method comprising: administering to a patient having such a condition an effective amount of peptide according to any one of claims 1 to 35, or a pharmaceutical composition according to any one of claims 36 to 42, wherein said administering is effective to inhibit amyloid plaque and / or tau aggregate formation and thereby treat the patient.

75. The method according to claim 74, wherein said administering is carried out daily.

76. The method according to claim 74, wherein said administering is carried out more than once daily.

77. The method according to claim 74, wherein the effective amount used for said administering is between about 0.1 and 500 mg / dose.

78. The method according to claim 74, wherein the peptide or a pharmaceutical composition containing the peptide is administered.

80. The method according to claim 75, wherein two or more peptides or a pharmaceutical composition containing the two or more peptides is administered.

81. The method according to claim 80, wherein the two or more peptides bind to different locations on the ApoE protein.

82. The method according to claim 74, wherein said administering is carried out orally, sublingually, intranasally, by inhalation, intravenously, intraperitoneally, intramuscularly, subcutaneously, intrathecally, intraperitoneally, or lymphatically.

83. The method according to claim 74, wherein said administering is carried out by insertion of an implant.

84. The method according to claim 74 further comprising: administering to the patient one or more of an anti-Ap monoclonal antibody or active fragment thereof and an anti-tau monoclonal antibody or active fragment thereof.

85. The method according to any one of claims 74 to 84, wherein said treating is effective to reduce a rate of cognitive decline or maintain cognitive function.

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