Proteinaceous molecules and uses therefor

Abstract

This disclosure relates generally to proteinaceous molecules that bind to pathological transactive response DNA binding protein of 43 kDa (TDP-43), such as a TDP-43 aggregate, and their use for determining the presence of pathological TDP-43 in the brain of a subject. More particularly, this disclosure relates to proteinaceous molecules and conjugates comprising such proteinaceous molecules and a labelling moiety that bind to pathological TDP-43 and their use for determining if a subject has a neurodegenerative disease associated with TDP-43 pathology and, optionally, further treating said subject for the neurodegenerative disease.

Description

1006111282PROTEINACEOUS MOLECULES AND USES THEREFORRELATED APPLICATION

[0001] This application is related to Australian provisional patent application no. 2024903147 filed on 30 September 2024, the contents of which are incorporated herein by reference in their entirety.FIELD

[0002] This disclosure relates generally to proteinaceous molecules that bind to pathological transactive response DNA binding protein of 43 kDa (TDP-43), such as a TDP-43 aggregate, and their use for determining the presence of pathological TDP-43 in the brain of a subject. More particularly, this disclosure relates to proteinaceous molecules and conjugates comprising such proteinaceous molecules and a labelling moiety that bind to pathological TDP-43 and their use for determining if a subject has a neurodegenerative disease associated with TDP-43 pathology and, optionally, further treating said subject for the neurodegenerative disease.BACKGROUND

[0003] Amyotrophic lateral sclerosis (ALS) is a motor neuron disease affecting motor neurons in both the brain and the spinal cord. ALS is a fatal disease characterised by a loss of pyramidal cells in the cerebral motor cortex, anterior spinal motor neurons and brain stem motor neurons causing muscle weakness and atrophy. ALS typically shows rapid deterioration after onset, often leading to death within a few years.

[0004] Frontotemporal dementia (FTD) is characterised by progressive damage to the frontal and / or temporal lobes of the brain and is associated with progressive deterioration of decision-making abilities, control of behaviour and language. FTD is one of the most common forms of presenile dementia, with median life expectancy after diagnosis of less than 15 years.

[0005] ALS and FTD are both rapidly progressive and fatal neurodegenerative diseases with significant clinical, genetic and pathological overlap. ALS and FTD are typically classified as either familial (approximately 10% of cases, where one or more defined genetic mutations are implicated) or sporadic (approximately 90% of cases, in which etiology is typically not well understood), with the familial and sporadic forms1006111282 being clinically indistinguishable. ALS and FTD are neuropathologically characterised by deposition and aggregation of TDP-43 in neurons. Current research suggests that mis-localisation of nuclear TDP-43 to the cytoplasm triggers toxic events including aberrant phosphorylation and fragmentation of TDP-43 (Shenouda et al. (2018), Adv Neurobiol, 20:239-263). However, the molecular mechanisms that regulate physiological nucleo-cytoplasmic shuttling of TDP-43 during mRNA processing and drive cytoplasmic accumulation in disease remain largely unknown.

[0006] TDP-43 is a multifunctional RNA / DNA binding protein encoded by the TARDBP gene. It harbors two RNA recognition motifs and a large C-terminal glycine- rich domain that mediates protein-protein interactions. The glycine-rich domain contains the vast majority of pathological TARDBP mutations in familial ALS.

[0007] There is a lack of reliable diagnostic tools and assays to detect pathological TDP-43 and, consequently, identify diseases associated with TDP-43 pathology, such as FTD and ALS at an early stage. TDP-43 has been a target of interest for positron emission tomography (PET) tracer development in the past, but traditional approaches, such as the use of small molecules, have failed thus far. Importantly, the TDP-43 filament formed in diseases associated with TDP-43 pathology, such as ALS and FTD, rebuffs binding by traditional PET tracers (Arseni et al. (2022) Nature, 601 : 139-143). Alternative approaches, such as the use of anti-TDP-43 antibodies, peptides and new chemical structures have recently been considered, but have not yet resulted in a commercially available PET tracer.

[0008] There is a clear need for the development of new methods for detecting pathological TDP-43 and diagnosing associated diseases.SUMMARY

[0009] The present disclosure is predicated in part on the identification that particular peptides are able to bind to pathological TDP-43 and distinguish between subjects having pathological TDP-43 and those not having pathological TDP-43 in the brain. The inventors have found that such peptides are able to bind to pathological TDP-43 in the brain for a duration that is particularly amenable to identification via nuclear imaging, such as PET, after being systemically administered. Based on this activity, the inventors have conceived that these peptides and conjugates comprising these peptides and a labelling moiety will be useful for determining the presence of1006111282 pathological TDP-43 in the brain of a subject and determining if a subject has a neurodegenerative disease associated with TDP-43 pathology.

[0010] Accordingly, in one aspect, there is provided a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, including R, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;1006111282Xs is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof; wherein the proteinaceous molecule does not comprise the amino acid sequence of SEQ ID NO: 1 :RKQTIDNSQGA [SEQ ID NO: 1 ],

[0011] In some embodiments, Xi is selected from the group consisting of Q, A, N, T, S, K, H, D, E, G, I, L, V and modified forms thereof, such as Q; X2 is selected from the group consisting of T, A, S, C, M, N, Q, D, E, Y, G, I, L, V and modified forms thereof, such as T; X3 is selected from the group consisting of I, A, G, L, V, F, Y, S, T, C, M, N, Q and modified forms thereof, such as I; X4 is selected from the group consisting of D, A, E, N, Q, C, T, S, G, I, L, V and modified forms thereof, such as D or E; X5 is selected from the group consisting of G, A, V I, L, F, Y, S, T, C, M and modified forms thereof, such as G; and / or Xe is selected from the group consisting of A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N, Q and modified forms thereof, such as A or G.

[0012] In some embodiments, the proteinaceous molecule further comprises a labelling moiety. In particular embodiments, the labelling moiety is a fluorescent label or a radionuclide.

[0013] In some embodiments, the labelling moiety is a fluorescent label selected from the group consisting of fluorescein isothiocyanate (FITC), tetramethylrhodamine (TRITC), carboxytetramethylrhodamine (TAMRA), 2-(5-(1 -(6-(N-(2-maleimdylethyl)- amino)-6-oxohexyl)-1 ,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene)-1 ,3- pentadienyl)-1 -ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt (Cy5), 6-[1 ,1-dimethyl-2-[5- (1 ,1 ,3-trimethylbenzo[e]indol-3-ium-2-yl)penta-2,4-dienylidene]benzo[e]indol-3- yl]hexanoic acid;chloride (Cy5.5), 1 -(5-carboxypentyl)-2-[(1 E,3E,5E,7Z)-7-(1-ethyl-5- sulfo-3H-indol-2-ylidene)hepta-1 , 3, 5-trieny l]-3H-indol-1 -ium-5-sulfonate (Cy7), 9-(2(or 4)-(N-(2-maleimdylethyl)-sulfonam idyl)-4(or2)-sulfophenyl)-2,3,6,7, 12,13,16,17- octahydro-(1 -H,5H, 11 H, 15H-xantheno(2 , -3, 4-ij : 5, 6, 7-i 'j ')d iqu inol izin-18-ium salt(Texas Red), 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY), (N-(4,4-difluoro- 1 ,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene-2-yl)iodoacetamide (BODIPY1006111282507 / 545 IA), N-(4,4-difluoro-5,7-diphenyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl)- N'-iodoacetylethylene diamine (BODIPY 530 / 550 IA), Alexa Fluor 350, Alexa Fluor 405, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 514, Alexa Fluor 532, Alexa Fluor546, Alexa Fluor 555, Alexa Fluor 561 , Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor633, Alexa Fluor 635, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680, Alexa Fluor700, Alexa Fluor 750, Alexa Fluor 790, Atto 390, Atto 425, Atto 430LS, Atto 465,Atto 490LS, Atto 495, Atto 514, Atto 488, Atto 520, Atto 532, Atto 540Q, Atto 550, Atto 565, Atto 590, Atto 594, Atto 633, Atto 647, Atto 655, Atto Rho14, Atto Rho6G, Atto Rho3B, Atto Rho1 1 , Atto Rho12, Atto Rho13, Atto Thio12, Atto Rho101 , Atto 580Q, Atto 680, Atto 700, Atto 647N, Atto 610, Atto 612Q, Atto 620, Atto 0xa12, Atto 725, Atto 740, Atto MB2, DyLight 350, DyLight 405, DyLight 488, DyLight 550, DyLight 594, DyLight 633, DyLight 650, DyLight 680, DyLight 755 and DyLight 800.

[0014] In some embodiments, the labelling moiety is a radionuclide selected from the group consisting of11C,18F,13N,15O,124l,44Sc,64Cu,68Ga,82Rb,86Y,89Zr,133La,99mTc, m |n123|anc| 1311 |nparticular embodiments, the labelling moiety further comprises a radionuclide binding moiety, such as a chelator or a synthon.

[0015] In some embodiments, the labelling moiety is a pre-targeting moiety, such as one comprising an azidyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl or tetrazinyl group.

[0016] In some embodiments, the proteinaceous molecule comprises a cell penetrating moiety. In specific embodiments, the proteinaceous molecule comprises two cell penetrating moieties. In some embodiments, the cell penetrating moiety is a cell penetrating peptide, such as a peptide comprising at least two arginine residues.

[0017] In some embodiments, the cell penetrating peptide comprises two or three arginine residues.

[0018] In another aspect, there is provided a conjugate represented by Formula II:B-Z (II) wherein:B is a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:1006111282RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, including R, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof; andZ is a labelling moiety.1006111282

[0019] In some embodiments, Xi is selected from the group consisting of Q, A, N, T, S, K, H, D, E, G, I, L, V and modified forms thereof, such as Q; X2 is selected from the group consisting of T, A, S, C, M, N, Q, D, E, Y, G, I, L, V and modified forms thereof, such as T; X3 is selected from the group consisting of I, A, G, L, V, F, Y, S, T, C, M, N, Q and modified forms thereof, such as I; X4 is selected from the group consisting of D, A, E, N, Q, C, T, S, G, I, L, V and modified forms thereof, such as D or E; X5 is selected from the group consisting of G, A, V I, L, F, Y, S, T, C, M and modified forms thereof, such as G; and / or Xe is selected from the group consisting of A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N, Q and modified forms thereof, such as A or G.

[0020] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula III:Z1RKX1X2X3X4NSQX5X6Z2 (III) wherein:Xi to Xe are as defined for Formula II; andZ1 and Z2 are independently absent or a labelling moiety, wherein at least one of Z1 and Z2 is present.

[0021] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula IV:Y1Z1RKX1X2X3X4NSQX5X6Z2Y2 (IV) wherein:Xi to Xe are as defined for Formula II;Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;Z1 and Z2 are independently absent or a labelling moiety, wherein at least one of Z1 and Z2 is present.

[0022] In other embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula V:1006111282Z3Y1 Z4X7X8X9Z1 R KX1 X2X3X4N S QX5X6Z2X10X11 Xi 2X13X14X15X16X17X18Xi 9X20 X21X22X23X24X25Z5Y2Z6 (V) wherein:Xi to Xe are as defined for Formula II;X7 to X25 are independently absent or any amino acid residue or a modified form thereof;Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;Z1 to Ze are independently absent or a labelling moiety, wherein at least one of Z1 to Ze is present.

[0023] In some embodiments, X7, X13, X17 and X24 are independently selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; Xs, X9, X12, X15 and X20 are independently selected from the group consisting of acidic amino acid residues, including D, E and modified forms thereof; X10 and X14 are independently selected from the group consisting of aromatic amino acid residues, including Y, W, F and modified forms thereof; Xu and X22 are independently selected from the group consisting of am ide-containing amino acid residues, including Q, N and modified forms thereof; X and X21 are independently selected from the group consisting of hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; X18, X19 and X25 are independently selected from the group consisting of basic amino acid residues, including R, K, H and modified forms thereof; and X23 is selected from the group consisting of P and modified forms thereof.

[0024] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VI:Z3Y1Z4GDDZ1RKQTIDNSQGAZ2YQEAFDISKKEMQPTHZ5Y2Z6 (VI) wherein:Y1, Y2 and Z1 to Ze are as defined for Formula V.

[0025] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VII:1006111282Z3Y1GDDRKQTIDNSQGAYQEAFDISKKEMQPTHY2 (VII) wherein:Y1 and Y2 are independently a cell penetrating moiety; andZ3 is a labelling moiety.

[0026] In particular embodiments, the cell penetrating moiety is a cell penetrating peptide. In some embodiments, the cell penetrating peptide comprises at least two arginine residues, such as one comprising two or three arginine residues.

[0027] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VIII:Z3RRRGDDRKQTIDNSQGAYQEAFDISKKEMQPTHRR (VIII) wherein:Z3 is a labelling moiety.

[0028] In some embodiments, the labelling moiety is a fluorescent label or a radionuclide.

[0029] In some embodiments, the labelling moiety is a fluorescent label selected from the group consisting of FITC, TRITC, TAMRA, Cy5, Cy5.5, Cy7, Texas Red, BODIPY, BODIPY 507 / 545 IA, BODIPY 530 / 550 IA, Alexa Fluor 350, Alexa Fluor 405, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 514, Alexa Fluor 532, Alexa Fluor 546,Alexa Fluor 555, Alexa Fluor 561 , Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633,Alexa Fluor 635, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680, Alexa Fluor 700,Alexa Fluor 750, Alexa Fluor 790, Atto 390, Atto 425, Atto 430LS, Atto 465, Atto490LS, Atto 495, Atto 514, Atto 488, Atto 520, Atto 532, Atto 540Q, Atto 550, Atto 565, Atto 590, Atto 594, Atto 633, Atto 647, Atto 655, Atto Rho14, Atto Rho6G, Atto Rho3B, Atto Rho1 1 , Atto Rho12, Atto Rho13, Atto Thiol 2, Atto Rho101 , Atto 580Q, Atto 680, Atto 700, Atto 647N, Atto 610, Atto 612Q, Atto 620, Atto 0xa12, Atto 725, Atto 740, Atto MB2, DyLight 350, DyLight 405, DyLight 488, DyLight 550, DyLight 594, DyLight 633, DyLight 650, DyLight 680, DyLight 755 and DyLight 800.

[0030] In some embodiments, the labelling moiety is a radionuclide selected from the group consisting of11C,18F,13N,15O,124l,44Sc,64Cu,68Ga,82Rb,86Y,89Zr,133La,100611128299mTc, 111 |n123|anc| 1311 |nparticular embodiments, the labelling moiety further comprises a radionuclide binding moiety, such as a chelator or a synthon.

[0031] In some embodiments, the labelling moiety is a pre-targeting moiety, such as one comprising an azidyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl or tetrazinyl group.

[0032] In another aspect, there is provided a method of determining if a subject has a neurodegenerative disease associated with TDP-43 pathology comprising: a) administering to the subject the conjugate of Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, includingR, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, includingS, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;1006111282X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology.

[0033] In another aspect, there is provided a use of the conjugate of Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, includingR, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, includingS, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I,1006111282L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof, in the manufacture of an agent for determining if a subject has a neurodegenerative disease associated with TDP-43 pathology.

[0034] In another aspect, there is provided the conjugate of Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, including R, K, H and modified forms thereof; acidic amino acid residues, including D, E1006111282 and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof, for use in a method of determining if a subject has a neurodegenerative disease associated with TDP-43 pathology comprising: a) administering to the subject the conjugate of Formula II or the proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I;1006111282 b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology.

[0035] In some embodiments, the neurodegenerative disease associated with TDP- 43 pathology is amyotrophic lateral sclerosis (ALS) or frontotemporal dementia (FTD).

[0036] In some embodiments, step b) is carried out between about 30 minutes and about 6 hours after step a), such as between about 1 hour and about 3 hours after step a).

[0037] In a further aspect, there is provided a method for determining the presence of pathological TDP-43 in the brain of a subject, comprising: a) administering to the subject the conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that pathological TDP-43 is present in the brain.

[0038] In a further aspect, there is provided a use of the conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I in the manufacture of an agent for determining the presence of pathological TDP-43 in the brain of a subject.

[0039] In a further aspect, there is provided the conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I for use in a method for determining the presence of pathological TDP-43 in the brain of a subject, comprising: a) administering to the subject the conjugate according to Formula II or the proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I;1006111282 b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that pathological TDP-43 is present in the brain.

[0040] In some embodiments, the pathological TDP-43 is aggregated TDP-43.

[0041] In yet another aspect, there is provided a method of treating a neurodegenerative disease associated with TDP-43 pathology comprising: a) administering to the subject a conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology; and d) treating the subject for the neurodegenerative disease associated with TDP- 43 pathology.

[0042] In yet another aspect, there is provided a use of a conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I in the manufacture of a medicament for treating a neurodegenerative disease associated with TDP-43 pathology.

[0043] In yet another aspect, there is provided a conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I for use in a method of treating a neurodegenerative disease associated with TDP-43 pathology comprising: a) administering to the subject the conjugate according to Formula II or the proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I;1006111282 b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology; and d) treating the subject for the neurodegenerative disease associated with TDP- 43 pathology.

[0044] In some embodiments, the presence or absence of the conjugate or proteinaceous molecule is determined by nuclear imaging, such as positron emission tomography (PET), computed tomography (CT) or single-photon emission computed tomography (SPECT). In some embodiments, the nuclear imaging is PET.

[0045] Also provided, in another aspect, is a pharmaceutical composition comprising a proteinaceous molecule of Formula I or a conjugate of Formula II and a pharmaceutically acceptable carrier or diluent.BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 is a series of images showing mouse whole brain optical imaging 2 hours post tail vein injection of the test (3+2R0-Cy7) and control (3+2R -Cy7) conjugate into iTDP-43 and wild type (WT) mice (Figure 1A), and a graph showing the quantification of the mean fluorescence intensity in the brain (Figure 1 B).

[0047] Figure 2 is a series of brain slice images showing that 3+2R0-Cy7 (red) selectively targets iTDP-43 cells (DAPI; blue) in comparison to WT mice and control conjugates (3+2R -Cy7) (Figure 2A). Figure 2B shows that the 3+2R0-Cy7 signal is indicative of cytosolic inclusions in the CA1 region of the hippocampus, which is consistent with TDP-43 pathology.

[0048] Figure 3 is a graph showing the time course analysis of average brain signal after 3+2R0-Cy7 injection in WT mice (orange) and iTDP-43 mice (green) compared to uninjected mice (blue).

[0049] Figure 4 is a series of images showing the brain fluorescence 1 hour, 2 hours and 24 hours after 3+2R0-Cy7 injection in WT mice and iTDP-43 mice compared to uninjected mice.1006111282

[0050] Figure 5 is an image showing the binding of the core 11 amino acid sequence of 14-3-30 theta (green) to TDP-43 (grey).

[0051] Figure 6 is an image showing the binding of the core 11 amino acid sequence of 14-3-30 theta, with a D143E mutation (blue) to TDP-43 (grey).DETAILED DESCRIPTION1. Definitions

[0052] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present disclosure, preferred methods and materials are described. For the purposes of the present disclosure, the following terms are defined below.

[0053] The articles “a” and “an” are used herein to refer to one or to more than one (i.e. to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.

[0054] By “about” is meant a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by as much 15, 14, 13, 12, 11 , 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 % to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length.

[0055] The term “agent” includes a compound that induces a desired pharmacological and / or physiological effect. The term also encompasses pharmaceutically acceptable and pharmacologically active ingredients of those compounds specifically mentioned herein including but not limited to salts, esters, amides, prodrugs, active metabolites, analogues and the like. When the above term is used, then it is to be understood that this includes the active agent per se as well as pharmaceutically acceptable, pharmacologically active salts, esters, amides, prodrugs, metabolites, analogues, etc. The term “agent” is not to be construed narrowly but extends to small molecules, proteinaceous molecules such as peptides, polypeptides and proteins as well as compositions comprising them and genetic molecules such as RNA, DNA and mimetics and chemical analogues thereof as well as cellular agents.1006111282

[0056] The term "aggregate" and grammatical variants thereof as used in relation to TDP-43 refers to a body or accumulation of TDP-43 proteins, such as mutated or misfolded TDP-43. The aggregate may be insoluble and may be located in the cytoplasm of a cell, such as a neuron.

[0057] The term "alkenyl" as used herein refers to aliphatic hydrocarbon groups containing at least one carbon-carbon double bond (e.g. one carbon-carbon double bond) and which may be straight or branched. In some embodiments, the alkenyl group is unbranched. Suitable alkenyl groups have from 3 to 10 carbon atoms (i.e. C3- C10 alkenyl), including from 3 to 8 carbon atoms (i.e. C3-C8 alkenyl). Representative alkenyl groups include 1 -propenyl (-CH=CHCH3), 2-propenyl (-CH2CH=CH2), 1 - butenyl (-CH=CHCH2CH3), 2-butenyl (-CH2CH=CHCH3) and 3-butenyl (- CH2CH2CH=CH2). In particular embodiments, an alkenyl group may be optionally substituted by 1 to 3 substituents (e.g. 1 , 2 or 3 substituents), wherein the substituent is selected from the group consisting of halogen, C1-C3 alkyl, -OH, -NH2 or =0.

[0058] The term "alkyl" as used herein refers to a straight or branched aliphatic hydrocarbon group, including a C1-C3 alkyl and C1-2 alkyl unless otherwise noted. Examples of alkyl groups include methyl, ethyl, n-propyl or iso-propyl. In preferred embodiments, the alkyl group is unbranched (i.e. a straight aliphatic hydrocarbon group).

[0059] As used herein, the term "alkynyl" refers to aliphatic hydrocarbon groups containing at least one carbon-carbon triple bond (e.g. one carbon-carbon triple bond) and which may be straight or branched. In some embodiments, the alkynyl group is unbranched. Suitable alkynyl groups include C3-C10 alkynyl or C3-C8 alkynyl. Representative groups include 1 -propynyl, 2-propynyl, 1 -butynyl, 2-butynyl and 3- butynyl. In particular embodiments, an alkynyl group may be optionally substituted by 1 to 3 substituents (e.g. 1 , 2 or 3 substituents), wherein the substituent is selected from the group consisting of halogen, C1-3 alkyl, -OH, -NH2 or =0.

[0060] Amino acid residues are referred to herein interchangeably using their full name or the one or three letter codes standard in the art. Abbreviations used for unnatural or modified amino acid residues or derivatives thereof are defined herein where appropriate.1006111282

[0061] Amino acid residues are defined herein on the basis of the side chain classification in some instances. Families of amino acid residues having similar side chains have been defined in the art, which can be generally sub-classified as follows:TABLE 1AMINO ACID SUB-CLASSIFICATION1006111282

[0062] As used herein, the term “and / or” refers to and encompasses any and all possible combinations of one or more of the associated listed items, as well as the lack of combinations when interpreted in the alternative (or).

[0063] The term “any amino acid residue” is used herein to refer to any of the 20 naturally occurring amino acid residues and modified versions thereof, including residues with modified side chains, N-methyl amino acids, a-methyl amino acids, residues with acetylated N-termini, beta amino acids, D-amino acids, and the like.

[0064] The term "associated with" when used in relation to diseases associated with TDP-43 pathology, means that TDP-43 pathology (or pathological TDP-43) contributes, either directly or indirectly, to the pathogenesis or progression of the disease, including of one or more symptoms of the disease. The specified activity may, for example, directly lead to the pathogenesis (i.e. development) of the disease or the development of one or more symptoms of the disease (e.g. via the formation of TDP- 43 aggregates). Alternatively or in addition, TDP-43 pathology may result in the progression (i.e. worsening) of the disease or one or more symptoms of the disease.

[0065] Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, and variations such as “comprises” and “comprising”, will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps. Thus, the use of the term “comprising” and the like indicates that the listed integers are required or mandatory, but that other integers are optional and may or may not be present. By “consisting of” is meant including, and limited to, whatever follows the phrase “consisting of”. Thus, the phrase “consisting of” indicates that the listed elements are required or mandatory, and that no other elements may be present. By “consisting essentially of” is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified for the listed elements. Thus, the phrase “consisting essentially of” indicates that the listed elements are required or mandatory, but that other elements are optional and may or may not be present depending upon whether or not they affect the activity or action of the listed elements. In specific embodiments, the term "consisting essentially of', in the context of a specific amino acid sequence disclosed herein, includes within its scope about 1 to about 50 optional amino acids (and all integer optional amino acids in between) upstream of the specific amino acid sequence1006111282 and / or about 1 to about 50 optional amino acids (and all integer optional amino acids in between) downstream of the specific amino acid sequence.

[0066] The term "cycloalkenyl" refers to an unsaturated monocyclic or fused or spiro polycyclic carbocycle having from 6 to 16 carbon atoms and at least one carbon-carbon double bond. It includes monocyclic systems such as cyclohexenyl, cycloheptenyl, cyclooctenyl and the like, along with bicyclic systems. In particular embodiments, a cycloalkenyl group may be optionally substituted by 1 to 3 substituents, for example, 1 , 2 or 3 substituents, wherein the substituent is selected from Ci-Ce alkyl, halogen, hydroxy, -NH2, =0 or -C(O)-OH. In some embodiments, the cycloalkenyl group is unsubstituted. The term "heterocycloalkenyl" refers to a cycloalkenyl group having one or more ring atoms as heteroatoms, with the remainder of the ring atoms being carbon atoms. Suitable heteroatoms include nitrogen, oxygen and sulfur.

[0067] The term "cycloalkynyl" is used herein to refer to an unsaturated monocyclic or fused or spiro polycyclic carbocycle having from 6 to 16 carbon atoms and at least one carbon-carbon triple bond. It includes monocyclic systems such as cyclohexynyl, cyclooctynyl and the like, along with bicyclic and tricyclic systems, such as dibenzylcyclooctynyl. In particular embodiments, a cycloalkynyl group may be optionally substituted by 1 to 3 substituents, for example, 1 , 2 or 3 substituents, wherein the substituent is selected from C1-C6 alkyl, halogen, hydroxy, -NH2, =0 or -C(O)-OH. In some embodiments, the cycloalkynyl group is unsubstituted. The term "heterocycloalkynyl" refers to a cycloalkynyl group having one or more ring atoms as heteroatoms, with the remainder of the ring atoms being carbon atoms. Suitable heteroatoms include nitrogen, oxygen and sulfur.

[0068] By “derivative” is meant a molecule, such as a polypeptide or an amino acid residue, that has been derived from the basic molecule by modification, for example by conjugation or complexing with other chemical moieties or by post-translational modification techniques as would be understood in the art. The term “derivative” also includes within its scope alterations that have been made to a parent molecule including additions or deletions that provide for functionally equivalent molecules.

[0069] As used herein, the term “dosage unit form” refers to physically discrete units suited as unitary dosages for the subject to be treated, each unit containing a predetermined quantity of active material calculated to produce the desired effect in association with the required pharmaceutically acceptable vehicle.1006111282

[0070] By “effective amount”, in the context of diagnosing a disease, is meant the administration of an amount of a proteinaceous molecule or conjugate to an individual to be diagnosed that is effective for determining the presence or absence of a particular target, such as pathological TDP-43, thereby allowing the diagnosis of a condition. The effective amount will vary depending upon the health and physical condition of the individual to be diagnosed, the taxonomic group of individual to be treated, the formulation of the proteinaceous molecule or conjugate and other relevant factors. It is expected that the amount will fall in a relatively broad range that can be determined through routine trials.

[0071] The term "labelling moiety" is used herein to refer to any moiety that is capable of being detected when localised to a protein, cell or tissue (e.g. by imaging, fluorescence and the like). The term "labelling moiety" also includes a group that is capable of forming a covalent bond with a moiety that is capable of being detected when localised to a protein, cell or tissue in vivo, such as a pre-targeting moiety as described herein.

[0072] The term "pathological TDP-43" is used herein to refer to a form of TDP-43 that leads to or contributes to (i.e. is associated with) the development or progression of one or more diseases, disorders or conditions, such as a neurodegenerative disease as described herein. In particular embodiments, the pathological TDP-43 is a TDP-43 aggregate (also referred to herein as aggregated TDP-43). The pathological TDP-43 may comprise one or more mutations selected from the group consisting of A315T, Q331 K, Q343R, N345K, R361 S, N390D, N390S, M337V, G294A, G294V, G295S, A382T and G287S, and may be hyperphosphorylated (e.g. at serine 403 / 404 and 409 / 410) and / or ubiquitinated.

[0073] By “pharmaceutically acceptable carrier” is meant a pharmaceutical vehicle comprised of a material that is not biologically or otherwise undesirable, i.e., the material may be administered to a subject along with the selected active agent without causing any or a substantial adverse reaction. Carriers may include excipients and other additives such as diluents, fillers, detergents, wetting or emulsifying agents, pH buffering agents, preservatives and the like. Similarly, a “pharmaceutically acceptable” salt, ester, amide, prodrug or derivative of a compound as provided herein is a salt, ester, amide, prodrug or derivative that this not biologically or otherwise undesirable.1006111282

[0074] As used herein, the terms “polypeptide”, “proteinaceous molecule”, “peptide” and “protein” are used interchangeably to refer to a polymer of amino acid residues and to variants and synthetic analogues of the same. The term “proteinaceous” as used in for example "proteinaceous molecule" refers to the presence of at least a part of the molecule that resembles or is a protein, wherein "protein" is to be understood to include a chain of amino-acid residues at least two residues long, thus including a peptide, a polypeptide and a protein and an assembly of proteins or protein domains. These terms apply to amino acid polymers in which one or more amino acid residues is a synthetic non-naturally-occurring amino acid, such as a chemical analogue or modified form of a corresponding naturally-occurring amino acid, as well as to naturally-occurring amino acid polymers. These terms do not exclude modifications, for example, glycosylations, acetylations, phosphorylations, attachment of labelling or cell penetrating moieties and the like as described herein. Soluble forms of the subject proteinaceous molecules are particularly useful. Included within the definition are, for example, polypeptides containing one or more analogues or modified forms of an amino acid including, for example, unnatural amino acids.

[0075] The term "pre-targeting moiety" is used herein to refer to a moiety that is able to form a covalent bond with a label molecule (e.g. a radionuclide binding moiety or synthon comprising a radionuclide) in vivo, for example, via a biorthogonal reaction (e.g. Staudinger ligation, the strain promoted alkyne-azide cyclo-addition or the inverse electron demand Diels-Alder reaction). The pre-targeting approach involves the administration of a targeting molecule (e.g. a proteinaceous molecule of the disclosure) comprising a suitable functional group to a subject. After time has been allowed for blood circulation, accumulation at the target site (i.e. pathological TDP-43) and subsequent elimination of excess targeting molecule, a labelled molecule (e.g. a radionuclide containing molecule) comprising a complementary functionality to the functional group of the targeting molecule is administered to the subject. The labelled molecule and targeting molecule undergo a biorthogonal reaction to form a covalent bond there between.

[0076] As used herein, the terms “salts” and “prodrugs” include any pharmaceutically acceptable salt, ester, hydrate or any other compound which, upon administration to the recipient, is capable of providing (directly or indirectly) a proteinaceous molecule of the disclosure, or an active metabolite or residue thereof. The term “pharmaceutically acceptable salts” refers without limitation to derivatives of the1006111282 disclosed compounds wherein the parent compound is modified by converting an existing acid or base moiety to its salt form (e.g. by reacting the free base group with a suitable organic acid). Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. Representative acid addition salts include acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxy- ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, toluenesulfonate, undecanoate and valerate salts, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like, as well as nontoxic ammonium, quaternary ammonium, and amine cations, including, but not limited to ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like. The pharmaceutically acceptable salts of the present disclosure include the conventional non-toxic salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. The pharmaceutically acceptable salts can be synthesised from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in, for example, Remington: The Science and Practice of Pharmacy, Adeboye Adejare and Joseph Remington (Ed), Academic Press, London, 23rdEdition, 2021 ; Stahl and Wermuth (2002) Pharmaceutical Salts: Properties, Selection, and Use, Wiley-VCH; and Berge et al. (1977) Journal of Pharmaceutical Science, 66: 1 -19, each of which is incorporated herein by reference in its entirety.1006111282

[0077] The term “sequence identity” as used herein refers to the extent that sequences are identical on an amino acid-by-amino acid basis over a window of comparison. Thus, a “percentage of sequence identity” is calculated by comparing two optimally aligned sequences over the window of comparison, determining the number of positions at which the identical amino acid residue (e.g. Ala, Pro, Ser, Thr, Gly, Vai, Leu, lie, Phe, Tyr, Trp, Lys, Arg, His, Asp, Glu, Asn, Gin, Cys and Met) occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison (i.e. the window size), and multiplying the result by 100 to yield the percentage of sequence identity.

[0078] “Similarity” refers to the percentage number of amino acids that are identical or constitute conservative substitutions as defined in Tables 1 and 2 herein. Similarity may be determined using sequence comparison programs such as GAP (Deveraux et al. (1984), Nucleic Acids Research 12: 387-395). In this way, sequences of a similar or substantially different length to those cited herein might be compared by insertion of gaps into the alignment, such gaps being determined, for example, by the comparison algorithm used by GAP.

[0079] Terms used to describe sequence relationships between two or more polypeptides include “reference sequence,” “comparison window”, “sequence identity,” “percentage of sequence identity” and “substantial identity”. A “reference sequence” is at least 8 but frequently 10 to 30 amino acid residues in length. As two amino acid sequences may each comprise (1 ) a sequence (i.e. only a portion of the complete proteinaceous molecule) that is similar between the two proteinaceous molecules, and (2) a sequence that is divergent between the two proteinaceous molecules, sequence comparisons between two (or more) proteinaceous molecules are typically performed by comparing sequences of the two proteinaceous molecules over a “comparison window” to identify and compare local regions of sequence similarity. A “comparison window” refers to a conceptual segment of at least 6 contiguous positions in which a sequence is compared to a reference sequence of the same number of contiguous positions after the two sequences are optimally aligned. The comparison window may comprise additions or deletions (i.e. gaps) of about 20% or less as compared to the reference sequence (which does not comprise additions or deletions) for optimal alignment of the two sequences. Optimal alignment of sequences for aligning a comparison window may be conducted by computerized implementations of algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package1006111282Release 7.0, Genetics Computer Group, 575 Science Drive Madison, Wl, USA) or by inspection and the best alignment (i.e. resulting in the highest percentage homology over the comparison window) generated by any of the various methods selected. Reference also may be made to the BLAST family of programs as for example disclosed by Altschul et al. (1997) Nucl. Acids Res. 25: 3389. A detailed discussion of sequence analysis can be found in Unit 19.3 of Ausubel et al. (1998) Current Protocols in Molecular Biology, John Wiley & Sons Inc, Chapter 15; Lambert et al. (2003) Current Genomics, 4:131 -146; and Bawano et al. (2017) Bioinformatics, Volume 1: Data, Sequence Analysis and Evolution (Methods in Molecular Biology (1525)), Humana Press, pages 167-189.

[0080] The term “subject” as used herein refers to a vertebrate subject, particularly a mammalian or avian (bird) subject, for whom diagnosis is desired, for example, a subject suspected of having a disease as described herein. Suitable subjects include, but are not limited to, primates; avians (birds); livestock animals such as sheep, cows, horses, deer, donkeys and pigs; laboratory test animals such as rabbits, mice, rats, guinea pigs and hamsters; companion animals such as cats and dogs; and captive wild animals such as foxes, deer and dingoes. In particular embodiments, the subject is a primate, suitably a human.

[0081] As used herein, the terms “treatment”, “treating”, and the like, refer to obtaining a desired pharmacologic and / or physiologic effect. The effect may be therapeutic in terms of a partial or complete cure for a disease, disorder or condition and / or adverse effect or symptom attributable to the disease, disorder or condition. These terms also cover any treatment of a condition or disease in a subject, particularly in a human, and include: (a) inhibiting the disease or condition, i.e. arresting its development; or (b) relieving the disease or condition, i.e. causing regression of the disease or condition.

[0082] The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is not, and should not be taken as an acknowledgment or admission or any form of suggestion that the prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates.

[0083] Each embodiment described herein is to be applied mutatis mutandis to each and every embodiment unless specifically stated otherwise.10061112822. Abbreviations

[0084] The following abbreviations are used throughout the application: h = hour min = minute kDa = kilodaltonTDP-43 = transactive response DNA binding protein of 43 kDaPET = positron emission tomographyFITC = fluorescein isothiocyanateTRITC = tetramethylrhodamineTAMRA = carboxytetramethylrhodamineCy5 = cyanine 5Cy5.5 = cyanine 5.5Cy7 = cyanine 7BODIPY = 4,4-difluoro-4-bora-3a,4a-diaza-s-indaceneALS = amyotrophic lateral sclerosisFTD = frontotemporal dementiaCT = computed tomographySPECT = single-photon emission computed tomographyWT = wild typeDAPI = 4',6-diamidino-2-phenylindole1006111282PEG = poly(ethylene glycol)DCM = dichloromethaneSPPS = solid phase peptide synthesisHPLC = high performance liquid chromatographyFA = formic acid rt = retention timeLCMS = liquid chromatography-mass spectrometryFmoc = fluorenyl methoxycarbonylAA = amino acidDMF = N,N-dimethylformamideDIC = N,N’-diisopropylcarbodiimideDIPEA = N,N-diisopropylethylamine equiv. = equivalentsTFA = trifluoroacetic acidTIPS = triisopropylsilaneHOBt = hydroxybenzotriazoleSiFA = silicon-fluoride-acceptorMRI = magnetic resonance imaging3. Proteinaceous Molecules and Conjugates

[0085] The present disclosure provides proteinaceous molecules that bind to pathological TDP-43 (e.g. aggregated TDP-43) and can be used to determine if a1006111282 subject has a neurodegenerative disease associated with TDP-43 pathology, such as ALS or FTD.

[0086] As exemplified herein, the inventors have identified derivatives of a subsequence of the protein 14-3-30 that are able to bind to TDP-43 with greater affinity than the wild-type sequence. As also exemplified herein, the inventors have demonstrated that the use of a conjugate comprising a subsequence of 14-3-30, a cell penetrating moiety and a labelling moiety is able to bind to pathological TDP-43 (i.e. a TDP-43 aggregate) in the brain of a mouse expressing human A315T mutant TDP-43 and generate a pathological TDP-43 specific signal at about two hours after administration. This time frame is particularly suitable for imaging via a PET scan.

[0087] Accordingly, in one aspect, there is provided a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, including R, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M1006111282 and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof.

[0088] In some embodiments, the proteinaceous molecule does not comprise the amino acid sequence of SEQ ID NO: 1 .

[0089] In some embodiments, Xi is selected from the group consisting of S, T, A, G, Q, N, R, K, H, D, E, I, L, V and modified forms thereof. In some embodiments, Xi is selected from the group consisting of Q, A, N, T, S, K, H, D, E, G, I, L, V and modified forms thereof, such as Q, A, N, T, S, K, H, D, E, G, I, L or V. In specific embodiments, Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; and amide-containing amino acid residues, including Q, N and modified forms thereof; such as S, T, A, G, Q, N or a modified form thereof. In particular embodiments, Xi is Q or A, such as Q.

[0090] In some embodiments, X2 is selected from the group consisting of S, T, A, G, Q, N, D, E, I, L, V, M, Y, W, F, C and modified forms thereof. In some embodiments, X2 is selected from the group consisting of T, A, S, C, M, N, Q, D, E, Y, G, I, L, V and modified forms thereof, such as T, A, S, C, M, N, Q, D, E, Y, G, I, L or V. In particular embodiments, X2 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; such as S, T, A, G or a modified form thereof. In some embodiments, X2 is S, T, A or G; especially T, A or S. In some embodiments, X2 is T.1006111282

[0091] In some embodiments, X3 is selected from the group consisting of S, T, A, G, Q, N, I, L, V, M, Y, W, F, C and modified forms thereof. In exemplary embodiments, X3 is selected from the group consisting of I, A, G, L, V, F, Y, S, T, C, M, N, Q and modified forms thereof, such as I, A, G, L, V, F, Y, S, T, C, M, N or Q. In some embodiments, X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; and hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; such as S, T, A, G, I, L, V, M or a modified form thereof. In some embodiments, X3 is S, T, A, G, I, L, V or M. In some embodiments, X3 is I, L or A, such as I.

[0092] In some embodiments, X4 is selected from the group consisting of S, T, A, G, Q, N, D, E, I, L, V, M, C and modified forms thereof. In some embodiments, X4 is S, T, A, G, Q, N, D, E, I, L, V, M or C. In some embodiments, X4 is selected from the group consisting of D, A, E, N, Q, C, T, S, G, I, L, V and modified forms thereof, such as D, A, E, N, Q, C, T, S, G, I, L or V. In some embodiments, X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; and acidic amino acid residues, including D, E and modified forms thereof; such as S, T, A, G, D, E or a modified form thereof or S, T, A, G, D or E. In some embodiments, X4 is A, D or E. In particular embodiments, X4 is an acidic amino acid residue including D, E or a modified form thereof; such as D or E. In some embodiments, X4 is D. In alternative embodiments, X4 is E.

[0093] In some embodiments, X5 is selected from the group consisting of S, T, A, G, I, L, V, M, Y, W, F, C and modified forms thereof. In some embodiments, X5 is selected from the group consisting of G, A, V I, L, F, Y, S, T, C, M and modified forms thereof, such as G, A, V I, L, F, Y, S, T, C or M. In some embodiments, X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; and hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; such as S, T, A, G, I, L, V, M or a modified form thereof. In some embodiments, X5 is S, T, A, G, I, L, V or M, such as G, A or V. In some embodiments, X5 is G or A. In some embodiments, X5 is G. In other embodiments, X5 is A.

[0094] In some embodiments, Xe is selected from the group consisting of A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N, Q and modified forms thereof. In some embodiments, X6is A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N or Q. In some embodiments, Xe is a small amino acid residue, including S, T, A, G or a modified form1006111282 thereof; such as S, T, A, G or a modified form thereof. In some embodiments, Xe is S, T, A or G. In particular embodiments, Xe is A or G. In some embodiments, Xe is A. In other embodiments, Xe is G.

[0095] In some embodiments:Xi is Q, A, N, T, S, K, H, D, E, G, I, L or V;X2is T, A, S, C, M, N, Q, D, E, Y, G, I, L or V;X3is I, A, G, L, V, F, Y, S, T, C, M, N or Q;X4is D, A, E, N, Q, C, T, S, G, I, L or V;X5is G, A, V I, L, F, Y, S, T, C or M; and / orX6is A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N or Q.

[0096] In some embodiments:Xi is S, T, A, G, Q or N;X2 is S, T, A or G;X3is S, T, A, G, I, L, V or M;X4is S, T, A, G, D or E;X5is S, T, A, G, I, L, V or M; and / orXe is S, T, A or G.

[0097] In some embodiments:Xi is Q or A;X2 is T, A or S;X3 is I, L or A;X4is A, D or E;X5 is G, A or V; and / or1006111282Xe is A or G.

[0098] In particular embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino sequence of any one of SEQ ID NOs: 1 -11 :RKQTIDNSQGA [SEQ ID NO: 1];RKATIDNSQGA [SEQ ID NO: 2];RKQAIDNSQGA [SEQ ID NO: 3];RKQSIDNSQGA [SEQ ID NO: 4];RKQTADNSQGA [SEQ ID NO: 5];RKQTLDNSQGA [SEQ ID NO: 6];RKQTIANSQGA [SEQ ID NO: 7];RKQTIENSQGA [SEQ ID NO: 8];RKQTIDNSQAA [SEQ ID NO: 9];RKQTIDNSQVA [SEQ ID NO: 10]; orRKQTIDNSQGG [SEQ ID NO: 11],

[0099] In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino sequence of any one of SEQ ID NOs: 2-11 . In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino sequence of any one of SEQ ID NOs: 1 , 2, 8, 9 and 11. In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino acid sequence of SEQ ID NO: 1.

[0100] In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino acid sequence represented by Formula IX:X7X8X9RKX1X2X3X4NSQX5X6X10X11X12X13X14X15X16X17X18X19X20X21X22X23X24X25 (IX) wherein:1006111282Xi to Xe are as defined for Formula I above; andX7 to X25 are independently absent or any amino acid residue or a modified form thereof.

[0101] In some embodiments:X7, X13, X17 and X24 are independently selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof;Xs, X9, X12, X15 and X20 are independently selected from the group consisting of acidic amino acid residues, including D, E and modified forms thereof;X10 and X14 are independently selected from the group consisting of aromatic amino acid residues, including Y, W, F and modified forms thereof;X11 and X22 are independently selected from the group consisting of am ide-containing amino acid residues, including Q, N and modified forms thereof;X and X21 are independently selected from the group consisting of hydrophobic amino acid residues, including I, L, V, M and modified forms thereof;Xis, X19 and X25 are independently selected from the group consisting of basic amino acid residues, including R, K, H and modified forms thereof; and / orX23 is selected from the group consisting of P and modified forms thereof.

[0102] In some embodiments, X7 is S, T, A or G, especially G.

[0103] In some embodiments, Xs is D or E, especially D.

[0104] In some embodiments, X9 is D or E, especially D.

[0105] In some embodiments, X10 is Y, W or F, especially Y.

[0106] In some embodiments, Xu is Q or N, especially Q.

[0107] In some embodiments, X12 is D or E, especially E.

[0108] In some embodiments, X13 is S, T, A or G, especially A.

[0109] In some embodiments, X14 is Y, W or F, especially F.1006111282

[0110] In some embodiments, X15 is D or E, especially D.

[0111] In some embodiments, X16 is I, L, V or M, especially I.

[0112] In some embodiments, X17 is S, T, A or G, especially S.

[0113] In some embodiments, X18 is R, K or H, especially K.

[0114] In some embodiments, X19 is R, K or H, especially K.

[0115] In some embodiments, X20 is D or E, especially E.

[0116] In some embodiments, X21 is I, L, V or M, especially M.

[0117] In some embodiments, X22 is Q or N, especially Q.

[0118] In some embodiments, X23 is P.

[0119] In some embodiments, X24 is S, T, A or G, especially T.

[0120] In some embodiments, X25 is R, K or H, especially H.

[0121] In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino acid sequence represented by Formula X:GDDRKX1X2X3X4NSQX5X6YQEAFDISKKEMQPTH (X) wherein:Xi to Xe are as defined for Formula I above.

[0122] In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino acid sequence represented by Formula XI:X7X8X9R KQTI D N S Q G AX10X11 Xi 2X13X14X15X16Xi 7X18Xi 9X20X21 X22X23X24X25 (XI) wherein:X7 to X25 are as defined for Formula IX above.

[0123] In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino acid sequence of any one of SEQ ID NOs: 12-22:1006111282GDDRKQTIDNSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 12];GDDRKATIDNSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 13];GDDRKQAIDNSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 14];GDDRKQSIDNSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 15];GDDRKQTADNSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 16];GDDRKQTLDNSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 17];GDDRKQTIANSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 18];GDDRKQTIENSQGAYQEAFDISKKEMQPTH [SEQ ID NO: 19];GDDRKQTIDNSQAAYQEAFDISKKEMQPTH [SEQ ID NO: 20];GDDRKQTIDNSQVAYQEAFDISKKEMQPTH [SEQ ID NO: 21 ]; orGDDRKQTIDNSQGGYQEAFDISKKEMQPTH [SEQ ID NO: 22],

[0124] In some embodiments, the proteinaceous molecule comprises, consists or consists essentially of an amino acid sequence of SEQ ID NO: 12.

[0125] The proteinaceous molecule may, in some embodiments, be cyclised. In some embodiments, the proteinaceous molecule is cyclised through N-to-C cyclisation (head to tail cyclisation), preferably through an amide bond (e.g. an amide bond between the N- and C-termini of the linear peptide). In particular embodiments, the peptides have an amide-cyclised peptide backbone. In other embodiments, the peptides are cyclised using side chain to side chain cyclisation, preferably through a disulfide bond, a thioether bond such as a lanthionine bond, a triazole bond, a lactam bond or a dimethylene bond.

[0126] In some embodiments, the N- and C-termini are linked using a linking moiety. The linking moiety may be a peptide linker such that cyclisation produces an amide- cyclised peptide backbone. Variation within the peptide sequence of the linking moiety is possible, such that the linking moiety may be modified to alter the physicochemical properties of the proteinaceous molecules and potentially improve the therapeutic use of the proteinaceous molecules, for example, by improving stability. The linking moiety1006111282 will be of suitable length to span the distance between the N- and C-termini of the proteinaceous molecule without substantially altering the structural conformation of the proteinaceous molecule, for example, a peptidic linking moiety may be 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid residues in length. In some embodiments, longer or shorter peptidic linking moieties may be required. In alternative embodiments, the proteinaceous molecule is an acyclic molecule.

[0127] In some embodiments (where the proteinaceous molecules of the disclosure comprise an N- and C-terminus), the proteinaceous molecules of the disclosure have a primary, secondary or tertiary amide, a hydrazide, a hydroxamide or a free-carboxyl group at the C-terminus and / or a primary amine or acetamide at the N-terminus. In some embodiments, the proteinaceous molecules of the disclosure are cyclic peptides and, thus, may not comprise N- and / or C-terminal amino acid residues. In some embodiments, the proteinaceous molecules of the disclosure have a primary amide or a free carboxyl group (C-terminal acid) at the C-terminus and a primary amine at the N-terminus, especially a free carboxyl group at the C-terminus and a primary amine at the N-terminus.

[0128] The proteinaceous molecule of the disclosure may be between 10 amino acid residues and 50 amino acid residues in length (and all integer amino acid residues there between); especially between 10 amino acid residues and 40 amino acid residues in length, between 10 amino acid residues and 35 amino acid residues in length or between 11 amino acid residues and 30 amino acid residues in length. In some embodiments, the proteinaceous molecule of the disclosure is or is up to 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39 or 40 residues in length.

[0129] In some embodiments, the proteinaceous molecule of the disclosure selectively binds to pathological TDP-43, such as a mutated TDP-43 or a TDP-43 aggregate, over wild-type (i.e. physiologically functional) TDP-43.

[0130] In some embodiments, the proteinaceous molecule of Formula I, IX, X or XI as discussed supra has at least about 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence similarity to the amino acid sequence of any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12. In some embodiments, the proteinaceous molecule of Formula I, IX, X or XI as discussed supra has at least about 80%, 81 %, 82%, 83%, 84%, 85%,100611128286%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12. In such molecules, the variance occurs at one or more of Xi to X25 when present in the subject Formula.

[0131] The present disclosure also contemplates proteinaceous molecules that are variants of any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12. Such “variant” proteinaceous molecules include proteinaceous molecules derived from any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12, by deletion or addition of one or more amino acids (such as from 1 -50 amino acid residues and all integer amino acids there between) to the N-terminal and / or C-terminal end of the proteinaceous molecule, deletion or addition of one or more amino acids (such as from 1 -5 amino acid residues and all integer amino acids there between) at one or more sites in the proteinaceous molecule, or substitution of one or more amino acids at one or more sites in the proteinaceous molecule. For example, in some embodiments, the variant proteinaceous molecule comprises an addition of one amino acid residue or deletion of one amino acid residue.

[0132] The proteinaceous molecules of any one of SEQ ID NOs: 1 to 22 may be altered in various ways, including amino acid substitutions, deletions, truncations and insertions. Methods for such manipulations are generally known in the art. For example, amino acid sequence variants of any one of SEQ ID NOs: 1 to 22 may be prepared by mutagenesis of nucleic acids encoding the amino acid sequence of any one of SEQ ID NOs: 1 to 22. Methods for mutagenesis and nucleotide sequence alterations are well known in the art. Recursive ensemble mutagenesis can be used in combination with binding assays to identify active variants. Variant proteinaceous molecules may also be designed using medicinal chemistry approaches standard in the art.

[0133] Conservative substitutions, such as exchanging one amino acid with another having similar properties, may be particularly desirable. Variant proteinaceous molecules of the disclosure may contain conservative amino acid substitutions (e.g. 1 , 2 or 3 substitutions) at various locations along their sequence, as compared to a parent or reference amino acid sequence, such as any one of SEQ ID NOs: 1 to 22. A “conservative amino acid substitution” is one in which the amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid1006111282 residues having similar side chains have been defined in the art as discussed in detail below.

[0134] Acidic: The residue has a negative charge due to loss of a proton at physiological pH and the residue is attracted by aqueous solution so as to seek the surface positions in the conformation of a peptide in which it is contained when the peptide is in aqueous medium at physiological pH. Amino acids having an acidic side chain include glutamic acid and aspartic acid.

[0135] Basic: The residue has a positive charge due to association with protons at physiological pH or within one or two pH units thereof (e.g. histidine) and the residue is attracted by aqueous solution so as to seek the surface positions in the conformation of a peptide in which it is contained when the peptide is in aqueous medium at physiological pH. Amino acids having a basic side chain include arginine, lysine and histidine.

[0136] Charged: The residue is charged at physiological pH and, therefore, includes amino acids having acidic or basic side chains, such as glutamic acid, aspartic acid, arginine, lysine and histidine.

[0137] Hydrophobic: The residue is not charged at physiological pH and the residue is repelled by aqueous solution so as to seek the inner positions in the conformation of a peptide in which it is contained when the peptide is in aqueous medium at physiological pH. Amino acids having a hydrophobic side chain include tyrosine, valine, isoleucine, leucine, methionine, norleucine, phenylalanine and tryptophan. In particular embodiments, hydrophobic amino acids include valine, leucine and isoleucine.

[0138] Neutral / polar: The residues are not charged at physiological pH but the residue is not sufficiently repelled by aqueous solutions so that it would seek inner positions in the conformation of a peptide in which it is contained when the peptide is in aqueous medium at physiological pH. Amino acids having a neutral / polar side chain include asparagine, glutamine, cysteine, histidine, serine and threonine.

[0139] Amide-containing: The residues contain an amide in their side chain, such as glutamine and asparagine.

[0140] Aromatic: The residues contain an aromatic group in their side chain and include phenylalanine, tyrosine and tryptophan.1006111282

[0141] This description also characterises certain amino acids as “small” since their side chains are not sufficiently large, even if polar groups are lacking, to confer hydrophobicity. With the exception of proline, “small” amino acids are those with four carbons or less when at least one polar group is on the side chain and three carbons or less when not. Amino acids having a small side chain include glycine, serine, alanine and threonine. The gene-encoded secondary amino acid proline is a special case due to its known effects on the secondary conformation of peptide chains. For the purposes of the present disclosure, proline is not classified as a “small” amino acid unless otherwise specified.

[0142] The degree of attraction or repulsion required for classification as polar or non-polar is arbitrary and, therefore, amino acids specifically contemplated by the disclosure have been classified as one or the other. Most amino acids not specifically named can be classified on the basis of known behaviour.

[0143] Amino acid residues can be further sub-classified as cyclic or non-cyclic, and aromatic or non-aromatic, self-explanatory classifications with respect to the side-chain substituent groups of the residues, and as small or large. Dependent on their structural properties, amino acid residues may fall in two or more classes. For the naturally- occurring protein amino acids, sub-classification according to this scheme is presented in Table 1 in Section 1 supra.

[0144] Conservative amino acid substitution also includes groupings based on side chains. For example, a group of amino acids having aliphatic side chains is glycine, alanine, valine, leucine and isoleucine; a group of amino acids having aliphatic- hydroxyl side chains is serine and threonine; a group of amino acids having amide- containing side chains is asparagine and glutamine; a group of amino acids having aromatic side chains is phenylalanine, tyrosine, and tryptophan; a group of amino acids having basic side chains is lysine, arginine, and histidine; a group of amino acids having acidic side chains is glutamic acid and aspartic acid; and a group of amino acids having sulfur-containing side chains is cysteine and methionine. For example, it is reasonable to expect that replacement of an aspartic acid with a glutamic acid, a threonine with a serine, a lysine with an arginine, a tyrosine with a phenylalanine, an asparagine with a glutamine, or a similar replacement of an amino acid with a structurally related amino acid will not have a major effect on the properties of the resulting variant peptide of the disclosure. Whether an amino acid change results in a1006111282 proteinaceous molecule that binds to TDP-43 can readily be determined by assaying its activity. Conservative substitutions are shown in Table 2 under the headings of 'Exemplary Substitutions' and 'Preferred Substitutions'. Amino acid substitutions falling within the scope of the disclosure, are, in general, accomplished by selecting substitutions that do not differ significantly in their effect on maintaining (a) the structure of the peptide backbone in the area of the substitution, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. After the substitutions are introduced, the variants are screened for TDP-43 binding.TABLE 2EXEMPLARY AND PREFERRED AMINO ACID SUBSTITUTIONS1006111282

[0145] Alternatively, similar amino acids for making conservative substitutions can be grouped into three categories based on the identity of the side chains. The first group includes glutamic acid, aspartic acid, arginine, lysine and histidine, which all have charged side chains; the second group includes glycine, serine, threonine, cysteine, tyrosine, glutamine and asparagine; and the third group includes leucine, isoleucine, valine, alanine, proline, phenylalanine, tryptophan, methionine and norleucine, as described in Zubay, Biochemistry, third edition, Wm.C. Brown Publishers (1993).

[0146] Thus, a predicted non-essential amino acid residue in a proteinaceous molecule of the disclosure is typically replaced with another amino acid residue from the same side chain family. Alternatively, mutations can be introduced randomly along all or part of the coding sequence of a proteinaceous molecule of the disclosure, if it contains naturally occurring amino acid residues, such as by saturation mutagenesis, and the resultant mutants can be screened for an activity of the parent polypeptide, as described for example herein, to identify mutants which retain that activity. Following mutagenesis of the coding sequences, the encoded proteinaceous molecule can be expressed recombinantly and its activity determined. A “non-essential” amino acid residue is a residue that can be altered from the wild-type sequence of an embodiment proteinaceous molecule of the disclosure without abolishing or substantially altering one or more of its activities. Suitably, the alteration does not substantially alter one of1006111282 these activities; for example, the activity is at least 20%, 40%, 60%, 70% or 80% of that of the wild-type. By contrast, an “essential” amino acid residue is a residue that, when altered from the wild-type sequence of an embodiment proteinaceous molecule of the disclosure, results in abolition of an activity of the parent molecule such that less than 20% of the wild-type activity is present.

[0147] Accordingly, the present disclosure also contemplates variants of the proteinaceous molecules of any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12, wherein the variants are distinguished from the parent sequence by the addition, deletion, or substitution of one or more amino acid residues. In general, variants will display at least about 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence similarity to a parent or reference proteinaceous molecule sequence as, for example, set forth in any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12, as determined by sequence alignment programs described elsewhere herein using default parameters. Desirably, variants will have at least 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to a parent or reference proteinaceous molecule sequence as, for example, set forth in any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12, as determined by sequence alignment programs described herein using default parameters. Variants of any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12, which fall within the scope of a variant proteinaceous molecule of the disclosure, may differ from the parent molecule generally by at least 1 , but by less than 5, 4, 3, 2 or 1 amino acid residue(s). In some embodiments, a variant proteinaceous molecule of the disclosure differs from the corresponding sequence in any one of SEQ ID NOs: 1 to 22, especially SEQ ID NO: 1 or 12, by at least 1 , but by less than 5, 4, 3, 2 or 1 amino acid residue(s). In some embodiments, the amino acid sequence of the variant proteinaceous molecule of the disclosure comprises the proteinaceous molecule of Formula I, IX, X or XI.

[0148] If the sequence comparison requires alignment, the sequences are typically aligned for maximum similarity or identity. “Looped” out sequences from deletions or insertions, or mismatches, are generally considered differences. The differences are, suitably, differences or changes at a non-essential residue or a conservative substitution.1006111282

[0149] In some embodiments, calculations of sequence similarity or sequence identity between sequences are performed as follows:

[0150] To determine the percent identity of two amino acid sequences or of two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g. gaps can be introduced in one or both of a first and a second amino acid or nucleic acid sequence for optimal alignment and non-homologous sequences can be disregarded for comparison purposes). In some embodiments, the length of a reference sequence aligned for comparison purposes is at least 40%, more usually at least 50% or 60%, and even more usually at least 70%, 80%, 90% or 100% of the length of the reference sequence. The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide at the corresponding position in the second sequence, then the molecules are identical at that position. For amino acid sequence comparison, when a position in the first sequence is occupied by the same or similar amino acid residue (i.e. conservative substitution) at the corresponding position in the second sequence, then the molecules are similar at that position.

[0151] The percent identity between the two sequences is a function of the number of identical amino acid residues shared by the sequences at individual positions, taking into account the number of gaps and the length of each gap, which need to be introduced for optimal alignment of the two sequences. By contrast, the percent similarity between the two sequences is a function of the number of identical and similar amino acid residues shared by the sequences at individual positions, taking into account the number of gaps and the length of each gap, which need to be introduced for optimal alignment of the two sequences.

[0152] The comparison of sequences and determination of percent identity or percent similarity between sequences can be accomplished using a mathematical algorithm. In certain embodiments, the percent identity or similarity between amino acid sequences is determined using the Needleman and Wunsch algorithm (1970, J. Mol. Biol., 48: 444-453) which has been incorporated into the GAP program in the GCG software package (Devereaux, et al. (1984) Nucleic Acids Research, 12: 387-395), using either a Blosum 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1 , 2, 3, 4, 5, or 6. In some embodiments, the1006111282 percent identity or similarity between amino acid sequences can be determined using the algorithm of Meyers and Miller (1989, Cabios, 4: 11 -17) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. In some embodiments, the percent identity or similarity between amino acid sequences can be determined using the CLUSTAL W program (Thompson et al. (1994) Nucleic Acids Research, 22(11 ): 4673-4680).

[0153] The proteinaceous molecules of the disclosure may also encompass modified amino acid residues. Modified amino acid residues may include residues with modified side chains, N-methyl amino acids, a-methyl amino acids, residues with acetylated N-termini, beta amino acids, and the like. Modified amino acid residues also include D-isomers of the amino acids and the modified forms listed below.

[0154] Examples of side chain modifications include modifications of amino groups, such as by acetylation with acetic anhydride; acylation of amino groups with succinic anhydride and tetrahydrophthalic anhydride; amidination with methylacetimidate; carbamoylation of amino groups with cyanate; pyridoxylation of lysine with pyridoxal- 5-phosphate followed by reduction with sodium borohydride; reductive alkylation by reaction with an aldehyde followed by reduction with sodium borohydride; and trinitrobenzylation of amino groups with 2,4,6-trinitrobenzene sulfonic acid. The carboxyl group may be modified by carbodiimide activation through O-acylisourea formation followed by subsequent derivatisation, for example, to a corresponding amide. The guanidine group of arginine residues may be modified by formation of heterocyclic condensation products with reagents such as 2,3-butanedione, phenylglyoxal and glyoxal. Tryptophan residues may be modified, for example, by alkylation of the indole ring with 2-hydroxy-5-nitrobenzyl bromide or sulfonyl halides, or by oxidation with / V-bromosuccinimide. Tyrosine residues may be modified by nitration with tetranitromethane to form a 3-nitrotyrosine derivative.

[0155] Suitable modified arginine residues include, but are not limited to, Nw- carboxymethyl-L-arginine, Nw-carboxyethyl-L-arginine, Na-acetyl-L-arginine, di(phenylglyoxal)-L-arginine, N-methyl-arginine, alpha-methyl-arginine, [3-arginine, N’- nitro-L-arginine, N’,N”-dimethyl-L-arginine, N’,N”-diethyl-L-arginine, homoarginine, 2- amino-4-guanidino-butyric acid and citrulline.1006111282

[0156] Suitable modified lysine residues include, but are not limited to, Ns- carboxycarbonyl-L-lysine, Ns-succinim idyl-L-lysine, 2-am ino-6-(2- hydroxyacetamido)hexanoic acid, Ns-3-hydroxypropyl-L-lysine, ornithine, Ns- allyloxycarbonyl-L-lysine, N-methyllysine, a-methyllysine, [3-lysine, Na-acetyl-L-lysine, Ns-acetyl-L-lysine, Ns-methyl-L-lysine, Ns-dimethyl-L-lysine and Ns-formyl-L-lysine.

[0157] Representative modified histidine residues include, but are not limited to, N1 - methyl-L-histidine, N3-methyl-L-histidine, N-methyl-L-histidine, a-methylhistidine, homohistidine and [3-histidine.

[0158] Suitable modified alanine residues include, but are not limited to, N- methylalanine, a-methylalanine (2-aminoisobutyric acid), [3-alanine, Na-acetyl-L- alanine, a-aminobutyric acid (2-aminobutyric acid), homoalanine, cyclohexylalanine and [3-homoalanine.

[0159] Suitable modified leucine residues include, but are not limited to, a- methylleucine, N-methylleucine, [3-leucine, t-butylglycine, homoleucine, Na-acetyl-L- leucine, te / t-leucine and [3-homoleucine.

[0160] Suitable modified glutamine residues include, but are not limited to, a- methylglutamine, Na-methylglutamine, NY-methylglutamine, [3-glutamine, homoglutamine, Na-acetyl-L-glutamine and [3-homoglutamine.

[0161] Exemplary modified asparagine residues include Np-methyl-Np-methoxy- asparagine, a-methylasparagine, Na-methylasparagine, Np-methylasparagine, [3- asparagine, homoasparagine, Na-acetyl-L-asparagine and [3-homoasparagine.

[0162] Modified glycine residues include, but are not limited to, N-methylglycine, [3- homoglycine, sarcosine and Na-acetyl-L-glycine.

[0163] Modified serine residues may include N-methylserine, a-methylserine, [3- serine, Na-acetyl-L-serine, isoserine, O-methylserine, homoserine and (3-homoserine.

[0164] Exemplary modified threonine residues include N-methylthreonine, a- methylthreonine, [3-threonine, Na-acetyl-L-threonine, O-methylthreonine, homothreonine and [3-homothreonine.

[0165] Suitable modified methionine residues include, but are not limited to, norleucine, N-methylmethionine, a-methylmethionine, [3-methionine, Na-acetyl-L-1006111282 methionine, methionine sulfoxide, methionine sulfone, selenomethionine, homomethionine and [3-homomethionine.

[0166] Exemplary modified proline residues include a-methylproline, [3-proline, Na- acetyl-L-proline, 4-phenoxy-pyrrolidine-2-carboxylic acid, 5,5-dimethylpyrrolidine-2- carboxylic acid, 5-methylpyrrolidine-2-carboxylic acid, homoproline and [3- homoproline.

[0167] Suitable modified isoleucine residues include, but are not limited to, a- methylisoleucine, N-methylisoleucine, [3-isoleucine, homoisoleucine, Na-acetyl-L- isoleucine, [3-methylisoleucine and [3-homoisoleucine.

[0168] Modified valine residues may include, but are not limited to, norvaline, a- methylvaline, N-methylvaline, [3-valine, [3-homovaline and Na-acetyl-L-valine.

[0169] Suitable modified phenylalanine residues include, but are not limited to, a- methylphenylalanine, N-methyl-phenylalanine, [3-phenylalanine, [3- methylphenylalanine, [3,[3-dimethylphenylalanine, [3-hydroxyphenylalanine, homophenylalanine, Na-acetyl-L-phenylalanine, [3-homophenylalanine, 4-fluoro-L- phenylalanine, 4-methyl-L-phenylalanine, 3,3-diphenyl-L-alanine, 4,4'-biphenyl-L- alanine and 2-naphthyl-L-alanine.

[0170] Exemplary modified tyrosine residues include D-tyrosine, a-methyltyrosine, N-methyltyrosine, [3-tyrosine, [3-methyltyrosine, [3,[3-dimethyltyrosine, [3- hydroxytyrosine, homotyrosine, O-methylhomotyrosine, Na-acetyl-L-tyrosine, O- methyltyrosine, O-ethyltyrosine, m-tyrosine and [3-homotyrosine.

[0171] Suitable modified tryptophan residues include, but are not limited to, a- methyltryptophan, N-methyltryptophan, [3-tryptophan, [3-methyltryptophan, homotryptophan, N-formyl-tryptophan, 2-methyltryptophan, Na-acetyl-L-tryptophan and [3-homotryptophan.

[0172] Suitable modified glutamic acid residues include, but are not limited to, N- methylglutamicacid, a-methylglutamic acid, [3-glutamic acid, Na-acetyl-L-glutamicacid, glutamic acid y-methyl ester, y-carboxy glutamic acid, homoglutamic acid and [3- homoglutamic acid.1006111282

[0173] Suitable modified aspartic acid residues include, but are not limited to, N- methylaspartic acid, a-methylaspartic acid, [3-aspartic acid, Na-acetyl-L-aspartic acid, aspartic acid [3-methyl ester and [3-homoaspartic acid.

[0174] Suitable modified cysteine residues include, but are not limited to, N- methylcysteine, a-methylcysteine, N-acetylcysteine, [3-cysteine, [3-methylcysteine and homocysteine.

[0175] The proteinaceous molecules of the disclosure also encompass a proteinaceous molecule comprising unnatural amino acid residues and / or their derivatives during peptide synthesis and the use of cross-linkers and other methods which impose conformational constraints on the proteinaceous molecules.

[0176] Examples of incorporating unnatural amino acids and derivatives during peptide synthesis include, but are not limited to, use of any of the modified amino acid residues listed supra, 4-amino butyric acid, 6-aminohexanoic acid, 4-amino-3-hydroxy- 5-phenylpentanoic acid, 4-amino-3-hydroxy-6-methylheptanoic acid, phenylglycine, A / s-acetyl-L-ornithine, 2 -thienyl alanine, penicillamine and / or D-isomers of amino acids. A list of unnatural amino acids contemplated by the present disclosure is shown in Table 3, in addition to the modified resides discussed supra.TABLE 3EXEMPLARY UNNATURAL OR MODIFIED AMINO ACIDS1006111282100611128210061112821006111282

[0177] Additional amino acids or other substituents may be added to the N- or C- termini, if present, of the proteinaceous molecules of the disclosure. For example, the proteinaceous molecules of the disclosure may form part of a longer sequence with additional amino acids added to either or both of the N- and C-termini.

[0178] The proteinaceous molecule may comprise a labelling moiety. Suitable moieties include any moiety that is capable of being detected when localised to a protein, cell or tissue in the brain (e.g. by imaging, fluorescence and the like) or a group that is capable of forming a covalent bond with a moiety that is capable of being detected when localised to a protein, cell or tissue in the brain. For example, suitable labelling moieties include a fluorescent label (e.g. a fluorophore), a radionuclide, a pretargeting moiety or a contrast agent (e.g. a gadolinium-based agent such as gadoteric acid, gadobutrol, gadoteridol, gadopentetic acid, gadobenate dimeglumine, gadodiamide, gadoversetamide, and pharmaceutically acceptable salts and derivatives thereof). In particular embodiments, the labelling moiety is a fluorescent label, a radionuclide or a pre-targeting moiety.

[0179] In some embodiments, the proteinaceous molecule comprises one labelling moiety. In alternative embodiments, the proteinaceous molecule comprises more than one labelling moiety, such as two, three or four labelling moieties. When more than one labelling moieties is present, the labelling moieties may be the same (e.g. two radionuclides) or different (e.g. one fluorescent label and one radionuclide).

[0180] In some embodiments, the labelling moiety is a fluorescent label. Suitable fluorescent labels include, but are not limited to, FITC, TRITC, TAMRA, Cy5, Cy5.5, Cy7, Texas Red, a BODIPY dye, an Alexa Fluor dye, an Atto dye, a DyLight Fluor dye, fluorescein, rhodamine, rhodol, indocyanine green, pafolacianine, thiofluorescein, aminofluorescein, chlorofluorescein, methylfluorescein, sulfofluorescein, aminorhodol,1006111282 carboxyrhodol, chlororhodol, methylrhodol, sulforhodol, aminorhodamine, carboxyrhodamine, chlororhodamine, methylrhodamine, sulforhodamine, thiorhodamine, cyanine, indocarbocyanine, oxacarbocyanine, thiacarbocyanine, merocyanine, cyanine 2, cyanine 3.5, oxadiazole derivatives, pyridyloxazole, nitrobenzoxadiazole, benzoxadiazole, pyren derivatives, cascade blue, oxazine derivatives, Nile red, Nile blue, cresyl violet, oxazine 170, acridine derivatives, proflavin, acridine orange, acridine yellow, arylmethine derivatives, auramine, crystal violet, malachite green, tetrapyrrole derivatives, porphin, phtalocyanine, bilirubin 1 - dimethylaminonaphthyl-5-sulfonate, 1 -anilino-8-naphthalene sulfonate, 2-p-touidinyl- 6-naphthalene sulfonate, 3-phenyl-7-isocyanatocoumarin, N-(p-(2- benzoxazolyl)phenyl)maleimide, a stilbene, a pyrene, Fluorescein dT, 5-TAMRA- cadavarine, 2-aminoacridone, hexachlorofluorescein (HEX), MAX, tetrachlorofluorescein (TET), carboxy-X-rhodamine (ROX), TYE™ 563, TYE™ 665, TYE™ 7056-carboxyflourescein (FAM), a coumarin, 5-TMRIA (tetramethylrhodamine- 5-iodoacetamide), 2-(5-(1 -(6-(N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1 ,3-dihydro- 3,3-dimethyl-5-sulfo-2H-indol-2-ylidene)-1 ,3-propyldienyl)-1 -ethyl-3,3-dimethyl-5- sulfo-3H-indolium salt (Cy3), N,N'-dimethyl-N-(iodoacetyl)-N'-(7-nitrobenz-2-oxa-1 ,3- diazol-4-yl)ethylenediamine (IANBD amide), N-((2-(iodoacetoxy)ethyl)-N- methyl)amino-7-nitrobenz-2-oxa-1 ,3-diazole (IANBD ester), 6-acryloyl-2- dimethylaminonaphthalene (acrylodan), pyrene, 6-amino-2,3-dihydro-2-(2- ((iodoacetyl)amino)ethyl)-1 ,3-dioxo-1 H-benz(de)isoquinoline-5,8-disulfonic acid salt (lucifer yellow), 4-(5-(4-dimethylaminophenyl)oxazol-2-yl)phenyl-N-(2- bromoacetamidoethyl)sulfonamide (Dapoxyl®(2-bromoacetamidoethyl)sulfonamide)), 5-((((2-iodoacetyl)amino)ethyl)amino)naphthalene-1-sulfonic acid (1 ,5-IAEDANS), carboxy-X-rhodamine, 5 / 6-iodoacetamide (XRIA 5,6), cyan fluorescent protein, green fluorescent protein, yellow fluorescent protein, a squaraine dye, squarylium dye, phthalocyanine, IRDye800CW, H-benz[e]indolium, 2-[2-[3-[2-(1 ,3-dihydro-1 , 1 ,3- trimethyl-2H-benz[e]indol-2-ylidene)ethylidene]-1 -cyclohexen-1 -yl]ethenyl]-3-[6-[(2,5- dioxo-1 -pyrrolidinyl)oxy]-6-oxohexyl]-1 ,1 -dimethyl- (Cy7.5), CH1055, 5-aminolevulinic acid (ALA), protoporphyrin IX and derivatives thereof.

[0181] In some embodiments, the fluorescent label is selected from the group consisting of FITC, TRITC, TAMRA, Cy5, Cy5.5, Cy7, Texas Red, 4,4-difluoro-4-bora- 3a,4a-diaza-s-indacene (BODIPY), (N-(4,4-difluoro-1 ,3,5,7-tetramethyl-4-bora-3a,4a- diaza-s-indacene-2-yl)iodoacetamide (BODIPY 507 / 545 IA), N-(4,4-difluoro-5,7-1006111282 diphenyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl)-N'-iodoacetylethylene diamine (BODIPY 530 / 550 IA), BODIPY-FL-hydrazide, Alexa Fluor 350, Alexa Fluor 405, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 514, Alexa Fluor 532, Alexa Fluor 546, AlexaFluor 555, Alexa Fluor 561 , Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, AlexaFluor 635, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680, Alexa Fluor 700, AlexaFluor 750, Alexa Fluor 790, Atto 390, Atto 425, Atto 430LS, Atto 465, Atto 490LS,Atto 495, Atto 514, Atto 488, Atto 520, Atto 532, Atto 540Q, Atto 550, Atto 565, Atto 590, Atto 594, Atto 633, Atto 647, Atto 655, Atto Rho14, Atto Rho6G, Atto Rho3B, Atto Rho1 1 , Atto Rho12, Atto Rho13, Atto Thio12, Atto Rho101 , Atto 580Q, Atto 680, Atto 700, Atto 647N, Atto 610, Atto 612Q, Atto 620, Atto 0xa12, Atto 725, Atto 740, Atto MB2, DyLight 350, DyLight 405, DyLight 488, DyLight 550, DyLight 594, DyLight 633, DyLight 650, DyLight 680, DyLight 755 and DyLight 800. In particular embodiments, the fluorescent label is FITC or Cy7.

[0182] In some embodiments, the labelling moiety is a radionuclide. Suitable radionuclides include, but are not limited to,11C,18F,13N,15O,124l,44Sc,64Cu,67Ga,68Ga82Rb86Y89Zr133La99mTc111ln123l125l131l152Tb67Cu90Y149T161T177Lu186Re,188Re,212Bi,213Bi,211At,212Pb and225Ac. In some embodiments, the radionuclide is11C,18F,13N,15O,124l,44Sc,64Cu,68Ga,82Rb,86Y,89Zr,133La,99mTc,111ln,123l or131l.

[0183] In some embodiments, the radionuclide is bound to a radionuclide binding moiety. The radionuclide binding moiety is any moiety that covalently binds a radionuclide, such as a chelator or synthon.

[0184] Suitable chelators include, but are not limited to, DOTA (1 ,4,7,10- tetraazacyclododecanel ,4,7, 10-tetraacetic acid), DOTA-NHS-ester, p-SCN-Bn-DOTA (C-DOTA), DOTAGA, DOTAGA-anhydride, CB-DO2A (4,10-bis(carboxymethyl)- 1 ,4,7, 10-tetraazabicyclo[5.5.2]tetradecane), TCMC.

[0185] (1 ,4,7,10-tetrakis(carbamoylmethyl)-l,4,7,10-tetraazacyclododecane), p- SCN-Bn-TCMC, 3p-C-DEPA (2-[(carboxymethyl)]-[5-(4-nitrophenyl-1 -[4,7, 10-tris- (carboxymethyl)-l ,4,7, 10-tetraazacyclododecan-1 -yl]pentan-2-yl)-amino]acetic acid), 3p-C-DEPA-NCS, p-NH2-Bn-Oxo-DO3A, TETA (1 ,4,8,11 - tetraazacyclotetradecanel , 4, 8, 1 1 -tetraacetic acid), BAT, p-NH2-Bn-TE3A, C-TETA, CB-TE2A (4, 11 -bis-(carboxymethyl)-1 ,4,8, 11 - tetraazabicyclo[6.6.2]-hexadecane), CB-TE1A1 P, CB-TE2P, MM-TE2A, DM-TE2A, TE2A, Diamsar, SarAr (1 -A / -(4-1006111282Aminobenzyl)-3,6, 10,13,16,19-hexaazabicyclo[6.6.6]- eicosane-1 ,8-diamine), AmBaSar, BaBaSar, NOTA (1 ,4, 7-triazacyclononane-1 ,4,7- triacetic acid), p-SCN-Bn- NOTA, NODASA, NODAGA (1 ,4, 7-triazacyclononane,1 -glutaric acid-4, 7-acetic acid), NETA ({4-[2-(bis-carboxymethylamino)-ethyl]-7-carboxymethyl-[1 ,4,7]triazonan-1 -yl}- acetic acid), NETA-monoamide, C-NE3TA-NCS, C-NETA-NCS, 3p-C-NETA, TACN- TM (N,N', A / "-tris(2-mercaptoethyl)-1 ,4,7-triazacyclononane), DTPA(diethylenetriaminepentaacetic acid), p-SCN-Bn-1 B-DTPA, p-SCN-Bn-1 B4M-DTPA, CHX-A"-DTPA (2-(p-isothiocyanatobenzyl)-cyclohexyldiethylenetriaminepentaacetic acid), p-SCN-Bn-CHX-A"-DTPA, TRAP (1 ,4,7-triazacyclononane1 ,4,7-tris[methyl(2- carboxyethyl)phosphinic acid]), AAZTA (1 ,4-bis (hydroxycarbonyl methyl)-6- [bis(hydroxylcarbonyl methyl)] amino-6-methyl perhydro-1 ,4-diazepine), NOPO, H2dedpa (1 ,2-[[6-(carboxy)-pyridin-2-yl]-methylamino]ethane), H4octapa ( / V; / V'-bis(6- carboxy-2-pyridylmethyl)-ethylenediamine- / \ / , / \ / '-diacetic acid), H2azapa (A / ,A / '-[1 - benzyl-1 ,2,3-triazole-4-yl]methyl- / \ / , / \ / '-[6-(carboxy)pyridin-2-yl]-1 ,2-diaminoethane), Hsdecapa ( / V, / V"-[[6-(carboxy)pyridin-2-yl]methyl]-diethylenetriamine- / \ / , / \ / ', / \ / "-triacetic acid), p-SCN-Bn-H40ctapa, HBED ( / V, / V'-bis(2-hydroxybenzyl)-ethylenediamine- / \ / , / \ / '- diacetic acid), HBED-CC, (HBED-CC)TFP, SHBED (A / ; / V'-bis(2-hydroxy-5- sulfobenzyl)-ethylenediamine- / V; / \ / '-diacetic acid), BPCA, CP256 (4-acetylamino-4-[2- [(3-hydroxy1 ,6-dimethyl-4-oxo-1 ,4-dihydro-pyridin2-ylmethyl)-carbamoyl]-ethyl]- heptanedioic acid bis-[(3-hydroxy-1 ,6-dimethyl4-oxo-1 ,4-dihydro-pyridin-2-ylmethyl)- amide]), PCTA (3,6,9, 15-tetraazabicyclo[9.3.1 ]- pentadeca-1 (15),11 ,13-triene-3,6,9,- triacetic acid), p-SCN-Bn-PCTA, DFO (deferoxamine), p-SCN-Bn-DFO (p-SCN-Bn- deferoxamine; 1 -(4-isothiocyanatophenyl)-3-[6, 17 -di hydroxy-7, 10, 18,21 -tetraoxo-27- (N-acetylhydroxylamino)- 6,11 ,17, 22- tetraazaheptaeicosine] thiourea), Hephospha ( / V, / V methylenephosphonate)- / V, / V'-[6-(methoxycarbonyl)pyridin-2-yl]-methyl-1 ,2- diaminoethane), HEHA (1 ,4,7,10,13,16-hexaazacyclohexadecane-A / , A / ', A / ", A / '"A / ""A / '""-hexaacetic acid), p-SCN-Bn-HEHA, PEPA (1 ,4,7,10,13- pentaazacyclopentadecane-A / ,A / ',A / ",A / '" A / ""-pentaacetic acid), p-SCN-Bn-PEPA, DOTPI (1 ,4,7, 10-tetraazacyclododecane-1 ,4,7, 10-tetrakis[methylene(2- carboxyethylphosphinic acid)]), tropolone, malonate, 8-hydroxyquinoline and derivatives thereof. In particular embodiments, the chelator is selected from the group consisting of DOTA, DOTA-NHS-ester, p-SCN-Bn-DOTA, DOTAGA, DOTAGA- anhydride, CB-DO2A, TCMC, p-SCN-Bn-TCMC, 3p-C-DEPA, 3p-C-DEPA-NCS, p- NH2-Bn-Oxo-DO3A, TETA, BAT, p-NH2-Bn-TE3A, C-TETA, CB-TE2A, CB-TE1A1 P, CB-TE2P, MM-TE2A, DM-TE2A, TE2A, Diamsar, SarAr, AmBaSar, BaBaSar, NOTA,1006111282 p-SCN-Bn-NOTA, NODASA, NODAGA, NETA, NETA-monoamide, C-NE3TA-NCS, C-NETA-NCS, 3p-C-NETA, TACN-TM, DTPA, p-SCN-Bn-1 B-DTPA, p-SCN-Bn- 1 B4M-DTPA, CHX-A"-DTPA, p-SCN-Bn-CHX-A"-DTPA, TRAP, AAZTA, NOPO, H2dedpa, H4octapa, H2azapa, Hsdecapa, p-SCN-Bn-H40ctapa, HBED, HBED-CC, (HBED-CC)TFP, SHBED, BPCA, CP256, PCTA, p-SCN-Bn-PCTA, DFO, Hephospha, p-SCN-Bn-DFO, HEHA, p-SCN-Bn-HEHA, PEPA, p-SCN-Bn-PEPA, DOTPI, and derivatives thereof. In some embodiments, the chelator is DOTA or NOTA.

[0186] In alternative embodiments, the radionuclide binding moiety is a synthon. Suitable synthons include, but are not limited to, methyl iodide, N-succinimidyl benzoate, an amine and / or thiol reactive synthon, a boron or silicon-fluoride bond forming agent [such as a trifluoroborate synthon, silicon-fluoride-acceptor (SiFA) synthon, and the like] or a click chemistry synthon [such as a synthon comprising a functional group, such as an azidyl, alkenyl (e.g. C3-C10 alkenyl), cycloalkenyl (e.g. Ce- C cycloalkenyl), a heterocycloalkenyl (e.g. C5-C16 heterocycloalkenyl), alkynyl (e.g. C3-C10 alkynyl), cycloalkynyl (e.g. Ce-C cycloalkynyl), heterocycloalkynyl (e.g. a Cs- C heterocycloalkynyl) or tetrazinyl group],

[0187] In some embodiments, the labelling moiety is [11C]methyl iodide, N- succinimidyl-4-[18F]fluorobenzoate ([18F]SFB), 4-nitrophenyl-2-18F-fluoropropionate ([18F]NFP), 4-[18F]fluorobenzoic acid ([18F]FBA), N-[6-(4-[18F]fluoro- benzylidene)aminooxyhexyl]maleimide ([18F]FBAM), N-[2-(4-[18F]fluorobenz- amido)ethyl]maleimide ([18F]FBEM), [18F]FDG-maleimidehexyloxime ([18F]FDG- MHO), [18F]SiFA-p-maleimide, [18F]aryltrifluoroborate ([18F]ArBF3), [18F]fluoro-N-(prop- 2-ynyl)benzamide or 1 -(azidomethyl)-4-[18F]fluorobenzene.

[0188] The labelling moiety may, in some embodiments, be a pre-targeting moiety. Suitable pre-targeting moieties include any moiety that is able to form a covalent bond with a label molecule (e.g. a radionuclide binding moiety or synthon comprising a radionuclide) in vivo, for example, via a biorthogonal reaction (e.g. Staudinger ligation, the strain promoted alkyne-azide cyclo-addition and the inverse electron demand Diels-Alder reaction). Representative pre-targeting moieties may comprise a functional group, such as an azidyl, alkenyl (e.g. C3-C10 alkenyl), cycloalkenyl (e.g. Ce- C cycloalkenyl), a heterocycloalkenyl (e.g. C5-C16 heterocycloalkenyl), alkynyl (e.g. C3-C10 alkynyl), cycloalkynyl (e.g. Ce-C cycloalkynyl), heterocycloalkynyl (e.g. a Cs- C heterocycloalkynyl) or tetrazinyl group.1006111282

[0189] In some embodiments, the proteinaceous molecule comprises a cell penetrating moiety, such as a cell-penetrating peptide. Suitable cell penetrating peptides include, but are not limited to, basic poly(Arg) (e.g. a peptide comprising two or three Arg residues) and poly(Lys) peptides and basic poly(Arg) and poly(Lys) peptides containing non-natural analogues of Arg and Lys residues; a TAT peptide including GRKKRRQRRRPPQ (TAT; SEQ ID NO: 23); penetratin, RQIKIWFQNRRMKWKK (SEQ ID NO: 24); Pep-1 , KETWWETWWTEWSQPKKKRKV (SEQ ID NO: 25); pVEC, LLIILRRRIRKQAHAHSK (SEQ ID NO: 26); Transportan, GWTLNSAGYLLGKINLKALAALAKKIL (SEQ ID NO: 27); p28,LSTAADMQGWTDGMASGLDKDYLKPDD (SEQ ID NO: 28); W / R, RRWRRWWRRWWRRWRR (SEQ ID NO: 29); AlkCWKis, CWKis (SEQ ID NO: 30); Di-CWKis, K18WCCWK18 (SEQ ID NO: 31 ); DipaLytic, GLFEALEELWEAK (SEQ ID NO: 32); K RGD, KieGGCRGDMFGCAK RGD (SEQ ID NO: 33); P1 , KwGGCMFGCGG (SEQ ID NO: 34); P2, K ICRRARGDNPDDRCT (SEQ ID NO: 35); P3, KKWKMRRNQFWVKVQRbAK (B) bA (SEQ ID NO: 36); P3a, VAYISRGGVSTYYSDTVKGRFTRQKYNKRA (SEQ ID NO: 37); P9.3,IGRIDPANGKTKYAPKFQDKATRSNYYGNSPS (SEQ ID NO: 38); Plae, PLAEIDGIELTY (SEQ ID NO: 39); Kplae, K GGPLAEIDGIELGA (SEQ ID NO: 40); cKplae, K GGPLAEIDGIELCA (SEQ ID NO: 41 ); MGP,GALFLGFLGGAAGSTMGAWSQPKSKRKV (SEQ ID NO: 42); HA2, WEAK(LAKA)2- LAKH(LAKA)2LKAC (SEQ ID NO: 43); LARL46, (LARL)6NHCH3(SEQ ID NO: 44); Hel- 11 -7, KLLKLLLKLWLLKLLL (SEQ ID NO: 45); KK, (KKKK)2GGC (SEQ ID NO: 46); KWK, (KWKK)2GCC (SEQ ID NO: 47); RWR, (RWRR)2GGC (SEQ ID NO: 48); SV40 NLS7, PKKKRKV (SEQ ID NO: 49); NLS12, PEVKKKRKPEYP (SEQ ID NO: 50); NLS12a, TPPKKKRKVEDP (SEQ ID NO: 51 ); SV40 NLS13, GGGGPKKKRKVGG (SEQ ID NO: 52); AV NLS13, GGGFSTSLRARKA (SEQ ID NO: 53); AV RME NLS17, CKKKKKKSEDEYPYVPN (SEQ ID NO: 54); AV FP NLS28, CKKKKKKKSEDEYPYVPNFSTSLRARKA (SEQ ID NO: 55); SV40 N1 NLS24, LVRKKRKTEEESPLKDKDAKKSKQE (SEQ ID NO: 56); Loligomer, KgfoKtKsGGKs (SEQ ID NO: 57); the protein transduction domain of the HIV-1 Tat protein, GRKKRRQRRRPPQ (SEQ ID NO: 58); the Drosophila melanogaster Antennapedia domain Antp (amino acids 43-58), RQIKIWFQNRRMKWKK (SEQ ID NO: 59); Buforin II, TRSSRAGLQFPVGRVHRLLRK (SEQ ID NO: 60); hClock-(amino acids 35-47) (human Clock protein DNA-binding peptide), KRVSRNKSEKKRR (SEQ ID NO: 61 ); MAP (model amphipathic peptide), KLALKLALKALKAALKLA (SEQ ID NO: 62); K-1006111282FGF, AAVALLPAVLLALLAP (SEQ ID NO: 63); Ku70-derived peptide, such as VPMLKE (SEQ ID NO: 64), VPMLK (SEQ ID NO: 65), PMLKE (SEQ ID NO: 66) or PMLK (SEQ ID NO: 67); Prion, Mouse Prpe (amino acids 1 -28), MANLGYWLLALFVTMWTDVGLCKKRPKP (SEQ ID NO: 68); SynBI, RGGRLSYSRRRFSTSTGR (SEQ ID NO: 69); Transportan-10, AGYLLGKINLKALAALAKKIL (SEQ ID NO: 70); CADY, Ac- GLWRALWRLLRSLWRLLWRA-cysteamide (SEQ ID NO: 71 ); Pep-7, SDLWEMMMVSLACQY (SEQ ID NO: 72); HN-1 , TSPLNIHNGQKL (SEQ ID NO: 73); VT5, DPKGDPKGVTVTVTVTVTGKGDPKPD (SEQ ID NO: 74); plSL, RVIRVWFQNKRCKDKK (SEQ ID NO: 75); DPV3, RKKRRRESRKKRRRES (SEQ ID NO: 76); DPV6, GRPRESGKKRKRKRLKP (SEQ ID NO: 77); R9-TAT, GRRRRRRRRRPPQ (SEQ ID NO: 78); ARF 19-31 , RVRVFWHIPRLT (SEQ ID NO: 79); Bip4, VSALK (SEQ ID NO: 80); Melittin, GIGAVLKVLTTGLPALISWIKRKRQQ (SEQ ID NO: 81 ); or gH625, HGLASTLTRWAHYNALIRAF (SEQ ID NO: 82). In some embodiments, the cell penetrating moiety is a poly(Arg) peptide (e.g. a peptide comprising two or three Arg residues) TAT, penetratin, Pep-1 , pVEC, Transportan or p28. In particular embodiments, the cell penetrating moiety is a poly(Arg) peptide, such as RR, RRR, RRRR (SEQ ID NO: 83), RRRRR (SEQ ID NO: 84) or RRRRRR (SEQ ID NO: 85).

[0190] While the proteinaceous molecule may comprise a single cell penetrating moiety, in some embodiments, the proteinaceous molecule comprises two cell penetrating moieties.

[0191] The cell penetrating moiety and labelling moiety may be conjugated to the N- terminal or C-terminal amino acid residue of the proteinaceous molecule or through a side-chain of an amino acid residue, such as through the amino group in the side chain of an amine- or amide-containing amino acid residue, such as lysine, arginine, glutamine and asparagine [e.g. the amino group of a lysine side-chain (i.e. the s-amino group)], through the side chain of a modified amino acid residue (e.g. azido-lysine or propargyl glycine), or the carboxylic acid in the side chain of an acidic amino acid residue, such as glutamic acid or aspartic acid.

[0192] In particular embodiments, the cell penetrating moiety is attached to the N- terminal or C-terminal amino acid residue of the proteinaceous molecule via an amide bond. For example, when two cell penetrating moieties are present, one moiety may1006111282 be attached at the N-terminus and the other moiety may be attached at the C-terminus of the proteinaceous molecule.

[0193] In particular embodiments, the labelling moiety is attached to the N- or C- terminus of the proteinaceous molecule. For example, the labelling moiety may form an amide bond with the N-terminal amino group or the C-terminal carboxylic acid group. Alternatively, the labelling moiety may be conjugated through a side-chain of an amino acid residue. The labelling moiety may, alternatively, be conjugated through the cell penetrating moiety, for example, it may be attached to the N- or C-terminus of the cell penetrating moiety.

[0194] In some embodiments, the labelling moiety or cell penetrating moiety is attached to the side chain of an amino acid residue which is inserted into the backbone of the proteinaceous molecule. For example, the proteinaceous molecule may have an additional amino acid residue having a side chain comprising the labelling moiety or cell penetrating moiety inserted into the sequence of the proteinaceous molecule [e.g. K(labelling moiety or cell penetrating moiety)]. The insertion may occur at any point, such as between X9 and R and / or between Xe and X10 in Formula IX.

[0195] The cell penetrating moiety and labelling moiety may be directly attached to the proteinaceous molecule or may be attached using a suitable linker, such as a PEG moiety or one or more amino acid residues (e.g. a proteinaceous molecule comprising 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid residues). For example, the PEG moiety may include a linear or branched PEG having 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10 oxyethylene units.

[0196] Proteinaceous molecules with high levels of stability may be desired, for example, to increase the half-life of the proteinaceous molecule in a subject. Thus, in some embodiments, the proteinaceous molecules of the disclosure comprise a stabilising or protecting moiety. The stabilising or protecting moiety may be conjugated at any point on the proteinaceous molecule. The stabilising or protecting moiety may be any moiety which delays or prevents substantial degradation of the proteinaceous molecule. A skilled person will be well aware of suitable stabilising or protecting moieties which may be used. Exemplary stabilising or protecting moieties include, but are not limited to, a peptide or protein such as an albumin including human serum albumin or a fragment or variant thereof, a glycine-rich homo-amino-acid polymer, a PAS sequence comprising a combination of alanine, serine and proline residues, an albumin binding moiety, which comprises an albumin binding peptide, a bacterial1006111282 albumin binding domain, an albumin-binding antibody fragment, or any combinations thereof; a polymer such as a PEG, a polysialic acid or a derivative thereof, hydroxyethyl starch or a derivative thereof, ethylene glycol / propylene glycol copolymers, carboxymethylcellulose, dextran or polyvinyl alcohol; a glycan or polysaccharide; or a capping moiety, including an acetyl group, pyroglutamate or an amino group.

[0197] In some embodiments, the protecting or stabilising moiety is a PEG. The PEG can be of any molecular weight, and can be branched or unbranched. In some embodiments, the molecular weight is between about 1 kDa and about 100 kDa. Other sizes can be used, depending on the desired profile (e.g. the duration of sustained release desired, the effects, if any, on TDP-43 binding, the ease in handling and other known effects of the PEG to a peptide or protein). For example, the PEG can have an average molecular weight of about 1 , 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 500, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500 or 5000 kDa.

[0198] In some embodiments, the PEG has 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10 oxyethylene units.

[0199] When present, the acetyl group and / or pyroglutamate may be conjugated to the N-terminal amino acid residue of the proteinaceous molecule. In some embodiments, the N-terminus of the proteinaceous molecule is a pyroglutamide or acetamide. In some embodiments, the amino group is conjugated to the C-terminal amino acid residue of the proteinaceous molecule. In particular embodiments, the proteinaceous molecule of the disclosure has a primary amide at the C-terminus.

[0200] When present, the PEG may be, for example, conjugated to the N-terminal or C-terminal amino acid residue of the proteinaceous molecule or through a side-chain of an amino acid residue, including through the amine of a lysine side-chain. In particular embodiments, the PEG is attached through the C-terminal amino acid residue, such as through the a-amino group, or through the side chain of an amino acid such as a lysine side-chain (i.e. the s-amino group). The moiety may also be attached to the proteinaceous molecule through a side-chain of an alternative amino group in the side chain of an amine- or am ide-containing amino acid residue, such as arginine, glutamine and asparagine or the carboxylic acid of an acidic residue, such as aspartic acid or glutamic acid.1006111282

[0201] The present disclosure also contemplates conjugates comprising a proteinaceous molecule of the disclosure, such as a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I, IX, X or XI and a labelling moiety. The conjugates may further comprise a cell penetrating moiety.

[0202] Accordingly, in another aspect, there is provided a conjugate represented by Formula II:B-Z (II) wherein:B is a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I, IX, X or XI as described herein; andZ is a labelling moiety.

[0203] Suitable embodiments of the proteinaceous molecule and labelling moiety are as discussed supra.

[0204] The labelling moiety may be conjugated to the proteinaceous molecule at any point, such as via the N-terminal or C-terminal amino acid residue of the proteinaceous molecule (e.g. via an amide bond) or through a side-chain of an amino acid residue as discussed for the proteinaceous molecule above. For example, the labelling moiety may be conjugate through the amino group in the side chain of an amine- or amide- containing amino acid residue, such as lysine, arginine, glutamine and asparagine, through the side chain of a modified amino acid residue (e.g. azido-lysine or propargyl glycine), or the carboxylic acid in the side chain of an acidic amino acid residue, such as glutamic acid or aspartic acid.

[0205] In some embodiments, the labelling moiety or cell penetrating moiety is attached to the side chain of an amino acid residue which is inserted into the backbone of the proteinaceous molecule. For example, the proteinaceous molecule may have an additional amino acid residue having a side chain comprising the labelling moiety or cell penetrating moiety inserted into the sequence of the proteinaceous molecule [e.g. K(labelling moiety or cell penetrating moiety)]. The insertion may occur at any1006111282 point, such as between X9 and R and / or between Xe and X10 in Formula IX. The labelling moiety may be attached using a suitable linker as described elsewhere herein.

[0206] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula III:Z1RKX1X2X3X4NSQX5X6Z2 (III) wherein:Xi to Xe are as defined for Formula I, IX, X or XI as described herein (and Formula II above); andZ1 and Z2 are independently absent or a labelling moiety, wherein at least one of Z1 and Z2 is present.

[0207] In such embodiments, the labelling moiety is attached via the N- and / or C- terminal amino acids of the proteinaceous molecule (e.g. via an amide bond). The labelling moiety may be attached directly or using a suitable linker as described elsewhere herein. Suitable labelling moieties are as described herein.

[0208] In some embodiments, one of Z1 and Z2 is present and the other is absent. In some embodiments, Z1 is present and Z2 is absent, or Z2 is present and Z1 is absent.

[0209] The conjugate may further comprise a cell penetrating moiety. Accordingly, in some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula IV:Y1Z1RKX1X2X3X4NSQX5X6Z2Y2 (IV) wherein:Xi to Xe are as defined for Formula I, IX, X or XI as described herein (and Formula II above);Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;Z1 and Z2 are independently absent or a labelling moiety, wherein at least one of Z1 and Z2 is present.1006111282

[0210] Suitable labelling moieties and cell penetrating moieties are as described supra.

[0211] The labelling moiety is attached via the N- and / or C-terminal amino acids of the proteinaceous molecule (e.g. via an amide bond). The labelling moiety may be attached directly or using a suitable linker as described elsewhere herein. Suitable labelling moieties are as described herein.

[0212] Similarly, the cell penetrating moiety may be attached via the via the N- and / or C-terminal amino acids of the proteinaceous molecule (e.g. via an amide bond) or may be attached to the labelling moiety, for example, when the labelling moiety is incorporated a part of an additional amino acid residue as discussed herein, the cell penetrating moiety may be attached to the N-terminal amine of such residue via an amide bond. A skilled person will be well aware of other methods for attaching the cell penetrating moiety to a labelling moiety standard in the art.

[0213] In some embodiments, one of Zi and Z2 is present and the other is absent. In some embodiments, Z1 is present and Z2 is absent, or Z2 is present and Z1 is absent.

[0214] In particular embodiments, Y1 and Y2 are both present. In some embodiments, Y1 is a cell penetrating peptide comprising or consisting of the amino acid sequence RRR and Y2 is a cell penetrating peptide comprising or consisting of the amino acid sequence RR.

[0215] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula XII:Z7Y1Z1RKX1X2X3X4NSQX5X6Z2Y2Z8 (XII) [SEQ ID NO:107] wherein:Xi to Xe are as defined for Formula I, IX, X or XI as described herein (and Formula II above);Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;1006111282Zi, Z2, Z7 or Zs are independently absent or a labelling moiety, wherein at least one of Z1, Z2, Z7 and Zs is present.

[0216] Suitable labelling moieties and cell penetrating moieties are as described herein. The cell penetrating moiety and labelling moiety may be conjugated to the proteinaceous molecule as described above.

[0217] In some embodiments, one of Z1 , Z2, Z7 and Zs is present and the rest are absent. In some embodiments, Z1 and Z2 are absent. In particular embodiments, Z7 and / or Zs is present. In some embodiments, Z7 is present.

[0218] In particular embodiments, Y1 and Y2 are both present. In some embodiments, Y1 is a cell penetrating peptide comprising or consisting of the amino acid sequence RRR and Y2 is a cell penetrating peptide comprising or consisting of the amino acid sequence RR.

[0219] In other embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula V:Z3Y1 Z4X7X8X9Z1 R KX1 X2X3X4N S QX5X6Z2X10X11 Xi 2X13X14X15X16X17X18X19X20 X21X22X23X24X25Z5Y2Z6 (V) wherein:Xi to Xe are as defined for Formula I, IX, X or XI as described herein (and Formula II above);X7 to X25 are as defined for Formula IX or XI as described herein;Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;Z1 to Ze are independently absent or a labelling moiety, wherein at least one of Z1 to Ze is present.

[0220] Suitable labelling moieties and cell penetrating moieties are as described herein. The cell penetrating moiety and labelling moiety may be conjugated to the proteinaceous molecule as described above.1006111282

[0221] In some embodiments, one of Zi to Ze is present and the rest are absent. In some embodiments, Zi , Z2, Z4 and Z5 are absent. In particular embodiments, Z3 and / or Ze is present. In some embodiments, Z3 is present.

[0222] In particular embodiments, Y1 and Y2 are both present. In some embodiments, Y1 is a cell penetrating peptide comprising or consisting of the amino acid sequence RRR and Y2 is a cell penetrating peptide comprising or consisting of the amino acid sequence RR.

[0223] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VI:Z3Y1Z4GDDZ1RKQTIDNSQGAZ2YQEAFDISKKEMQPTHZ5Y2Z6 (VI) wherein:Y1, Y2 and Zi to Ze are as defined for Formula V.

[0224] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VII:Z3Y1GDDRKQTIDNSQGAYQEAFDISKKEMQPTHY2 (VII) wherein:Y1 and Y2 are independently a cell penetrating moiety; andZ3 is a labelling moiety.

[0225] Suitable labelling moieties and cell penetrating moieties are as described herein. The cell penetrating moiety and labelling moiety may be conjugated to the proteinaceous molecule as described above. In some embodiments, Y1 is a cell penetrating peptide comprising or consisting of the amino acid sequence RRR and Y2 is a cell penetrating peptide comprising or consisting of the amino acid sequence RR.

[0226] In some embodiments, the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VIII:Z3RRRGDDRKQTIDNSQGAYQEAFDISKKEMQPTHRR (VIII) wherein:1006111282Zs is a labelling moiety.

[0227] Suitable labelling moieties are as described herein. The labelling moiety may be conjugated to the proteinaceous molecule as described above.

[0228] The proteinaceous molecules or conjugates of the disclosure may be purified, e.g. at least 70, 75, 80, 85, 90, 91 , 92, 93, 94, 95, 96, 97, 98 or 99% pure. In a preferred embodiment, the preparation of proteinaceous molecule or conjugate has less than about 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 % (by dry weight), of molecules that are not the subject of this disclosure. When the proteinaceous molecule is recombinantly produced, it is also desirably substantially free of culture medium, i.e. , culture medium represents less than about 20, 15, 10, 5, 4, 3, 2 or 1 % of the volume of the preparation.

[0229] The proteinaceous molecules or conjugates of the disclosure may also be in the form of salts or prodrugs. The salts of the proteinaceous molecules or conjugates of the present disclosure are preferably pharmaceutically acceptable, but it will be appreciated that non-pharmaceutically acceptable salts also fall within the scope of the present disclosure.

[0230] The proteinaceous molecules or conjugates may be in crystalline form and / or in the form of solvates, for example, hydrates. Solvation may be performed using methods known in the art.

[0231] The present disclosure also contemplates nucleic acid molecules which encode a proteinaceous molecule of the disclosure. Thus, in a further aspect of the present disclosure, there is provided an isolated nucleic acid molecule comprising a polynucleotide sequence that encodes a proteinaceous molecule of the disclosure or is complementary to a polynucleotide sequence that encodes a proteinaceous molecule of the disclosure, such as the proteinaceous molecule comprising, consisting or consisting essentially of a sequence represented by Formula I, IX, X or XI or any one of SEQ ID NOs: 1 to 22 as described herein.

[0232] The isolated nucleic acid molecules of the present disclosure may be DNA or RNA. When the nucleic acid is in DNA form, it may be genomic DNA or cDNA. RNA forms of the nucleic acid molecules of the present disclosure are generally mRNA.1006111282

[0233] Although the nucleic acid molecules are typically isolated, in some embodiments the nucleic acid molecules may be integrated into, ligated to, or otherwise fused or associated with other genetic molecules, such as an expression vector. Generally an expression vector includes transcriptional and translational regulatory nucleic acid operably linked to the polynucleotide sequence. Accordingly, in another aspect of the disclosure, there is provided an expression vector comprising a polynucleotide sequence that encodes a proteinaceous molecule of the disclosure, such as a proteinaceous molecule comprising, consisting or consisting essentially of a sequence represented by Formula I, IX, X or XI or any one of SEQ ID NOs: 1 to 22 as described herein.

[0234] In some embodiments, the proteinaceous molecules of the disclosure may be produced inside a cell by introduction of one or more expression constructs, such as an expression vector, that comprise a polynucleotide sequence that encodes a proteinaceous molecule of the disclosure.

[0235] The disclosure contemplates recombinantly producing the proteinaceous molecules of the disclosure inside a host cell, such as a mammalian cell (e.g. Chinese hamster ovary (CHO) cell, mouse myeloma (NSO) cell, baby hamster kidney (BHK) cell or human embryonic kidney (HEK293) cell), yeast cell (e.g. Pichia pastoris cell, Saccharomyces cerevisiae cell, Schizosaccharomyces pom be cell, Hansenula polymorpha cell, Kluyveromyces lactis cell, Yarrowia Hpolytica cell or Arxula adeninivorans cell), insect cell (e.g. Spodoptera frugiperda cell, such as an Sf9 cell) or bacterial cell (e.g. Escherichia coli cell, Cory nebacteri urn glutamicum or Pseudomonas fluorescens cell).

[0236] The expression of natural or synthetic nucleic acids is typically achieved by operably linking a polynucleotide sequence encoding a proteinaceous molecule of the present disclosure to a regulatory element (e.g. a promoter, which may be either constitutive or inducible), suitably incorporating the construct into an expression vector and introducing the vector into a suitable host cell. Typical vectors contain transcription and translation terminators, transcription and translation initiation sequences and promoters useful for regulation of the expression of the nucleic acid. The vectors optionally comprise generic expression cassettes containing at least one independent terminator sequence, sequences permitting replication of the cassette in eukaryotes, prokaryotes or both (e.g. shuttle vectors), and selection markers for both prokaryotic1006111282 and eukaryotic systems. Vectors may be suitable for replication and integration in prokaryotes, eukaryotes, or both (refer to Ausubel et al. (1998) Current Protocols in Molecular Biology, eds., John Wiley & Sons, Inc.; Rosano and Ceccarelli (2014) Front Microbiol, 5:172; and Klint, et al. (2013) PLOS One, 8(5): e63865).

[0237] Expression vectors containing regulatory elements from eukaryotic viruses such as retroviruses are typically used for expression of nucleic acid sequences in eukaryotic cells. Exemplary vectors include SV40 vectors such as pSVT7 and pMT2, vectors derived from bovine papilloma virus such as pBV-1 MTHA, and vectors derived from Epstein Bar virus such as pHEBO and p2O5. Other exemplary vectors include pMSG, pAV009 / A+, pMTO10 / A+, pMAMneo-5, baculovirus pDSVE and any other vector allowing expression of proteins under the direction of the SV-40 early promoter, SV-40 later promoter, metallothionein promoter, murine mammary tumour virus promoter, Rous sarcoma virus promoter, polyhedrin promoter or other promoters shown effective for expression in eukaryotic cells.

[0238] While a variety of vectors may be used, it should be noted that viral expression vectors are useful for modifying eukaryotic cells because of the high efficiency with which the viral vectors transfect target cells and integrate into the target cell genome. Illustrative expression vectors of this type can be derived from viral DNA sequences including, but not limited to, adenovirus, adeno-associated viruses, herpessimplex viruses and retroviruses such as B, C, and D retroviruses as well as spumaviruses and modified lentiviruses.

[0239] The peptide-encoding portion of the expression vector may comprise a naturally-occurring sequence or a variant thereof, which has been engineered using recombinant techniques. In one example of a variant, the codon composition of a polynucleotide encoding a proteinaceous molecule of the present disclosure is modified to permit enhanced expression of the proteinaceous molecule in a mammalian host using methods that take advantage of codon usage bias, or codon translational efficiency in specific mammalian cell or tissue types. Briefly, these latter methods are based on the observation that translational efficiencies of different codons vary between different cells or tissues and that these differences can be exploited, together with codon composition of a gene, to regulate expression of a protein in a particular cell or tissue type. Thus, for the construction of codon-optimised polynucleotides, at least one existing codon of a parent polynucleotide is replaced with1006111282 a synonymous codon that has a higher translational efficiency in a target cell or tissue than the existing codon it replaces. Although it is preferable to replace all the existing codons of a parent nucleic acid molecule with synonymous codons which have that higher translational efficiency, this is not necessary because increased expression can be accomplished even with partial replacement. Suitably, the replacement step affects 5%, 10%, 15%, 20%, 25%, 30%, more preferably 35%, 40%, 50%, 60%, 70% or more of the existing codons of a parent polynucleotide.

[0240] The expression vector is compatible with the cell in which it is introduced such that the proteinaceous molecule of the present disclosure is expressible by the cell. The expression vector is introduced into the cell by any suitable means which will be dependent on the particular choice of expression vector and cell employed. Such means of introduction are well-known to those skilled in the art. For example, introduction can be effected by use of contacting (e.g. in the case of viral vectors), electroporation, transformation, transduction, conjugation or triparental mating, transfection, infection membrane fusion with cationic lipids, high-velocity bombardment with DNA-coated microprojectiles, incubation with calcium phosphate-DNA precipitate, direct microinjection into single cells, and the like. Other methods also are available and are known to those skilled in the art. Alternatively, the vectors are introduced by means of cationic lipids, e.g., liposomes. Such liposomes are commercially available (e.g. Lipofectin®, Lipofectamine™, and the like, supplied by Invitrogen, Waltham MA, USA).

[0241] The proteinaceous molecules and conjugates may be prepared using any suitable method, such as chemical synthesis or recombinant DNA techniques. In some embodiments, the proteinaceous molecules or conjugates are prepared using standard peptide synthesis methods, such as solution synthesis or solid-phase synthesis. The chemical synthesis may be performed manually or using an automated synthesiser. For example, the linear peptides may be synthesised using solid-phase peptide synthesis using either Boc or Fmoc chemistry, as described in Merrifield (1963) J Am Chem Soc, 85(14): 2149-2154; Schnolzer, et al. (1992) I nt J Pept Protein Res, 40: I SO- 193; Cardoso, et al. (2015) Mol Pharmacol, 88(2): 291 -303; and Kumar et al. (2020) ACS Omega, 5: 2345-2354, the entire contents of which are incorporated by reference. Following deprotection and cleavage from the solid support, the linear peptides are purified using suitable methods, such as preparative chromatography.1006111282

[0242] Cell penetrating moieties and labelling moieties may be incorporated using methods known in the art. For example, a cell penetrating peptide or amino acid residue containing a labelling moiety or a side chain to which a labelling moiety can be attached (optionally containing a protecting group that can be selectively removed, such as an allyl group) can be incorporated as part of the peptide backbone during standard solid-phase synthesis of the peptide. Where required, the amino acid to which the labelling moiety is to be attached may be selectively deprotected following purification of the linear peptide and the labelling moiety can be attached via formation of an amide bond between an amine and an acid using standard techniques or using click chemistry (e.g. when the amino acid comprises an azidyl or alkynyl group such as azido-lysine or propargyl glycine). In such embodiments, the labelling moiety may be modified to include an amine, acid, azidyl group or alkynyl group as appropriate to facilitate the reaction.

[0243] In some embodiments, the proteinaceous molecules of the disclosure may be cyclised. Cyclisation may be performed using several techniques, for example, as described in Davies (2003) J Pept Sci, 9: 471 -501 ; or Thongyoo et al. (2006) Chem Commun (Camb), 27: 2848-2850. For example, N-to-C cyclisation may be conducted in the solution phase, using a dilute solution of the linear peptide in the presence of a coupling agent such as BOP (1 -benzotriazole-tris-dimethyl aminophosphonium hexafluorophosphate), PyBOP (1 -benzotriazolyloxy-tris-pyrrolidino phosphonium hexafluorophosphate), PyAOP (7-azabenzotriazol-1 -yloxy tris pyrrolidino phosphonium hexafluorophosphate), AOP (7-azabenzotriazol-1 -yloxy-tris-dimethyl aminophosphonium hexafluorophosphate), HBTII (O-(benzotriazol-1 -yl)-1 ,1 ,3,3- tetramethyl uronium hexafluorophosphate), TBTII (O-(benzotriazol-1 -yl)-1 ,1 ,3,3- tetramethyl uronium tetrafluoroborate), HATLI (O-(7-azabenzotriazol-1 -yl)-1 ,1 ,3,3- tetramethyl uronium hexafluorophosphate), HAPyll (O-(7-azabenzotriazol-1 -yl)- 1 ,1 ,3,3-tetramethylene uronium hexafluorophosphate), HAPiplI (O-(7- azabenzotriazol-1 -yl)-1 ,1 ,3,3-pentamethylene uranium hexafluorophosphate), DCC ( / V, / V'-dicyclohexylcarbodiimide), DIC (A / , / V'-diisopropylcarbodiimide), and / or EDC [1 - ethyl-3-(3'-dimethylaminopropyl)carbodiimide hydrochloride]. The cyclised peptide may then be deprotected (i.e. the side chain protecting groups may then be removed) using standard techniques, followed by purification using suitable methods, such as preparative chromatography. Alternatively, N-to-C cyclisation may be achieved on resin using a suitable coupling agent, such as those described above, and a suitable1006111282 resin, such as a Kaiser oxime resin, and / or linker (e.g. a safety catch linker), or via native chemical ligation as described in Thongyoo et al. (2006) Chem Commun (Camb), 27: 2848-2850.

[0244] In some embodiments, the proteinaceous molecules of the disclosure are prepared using recombinant DNA techniques. For example, the proteinaceous molecules may be prepared by a procedure including the steps of: (a) preparing a construct comprising a polynucleotide sequence that encodes the proteinaceous molecule of the disclosure and that is operably linked to a regulatory element; (b) introducing the construct into a host cell; (c) culturing the host cell to express the polynucleotide sequence to thereby produce the encoded proteinaceous molecule of the disclosure; and (d) isolating the proteinaceous molecule of the disclosure from the host cell. The proteinaceous molecule of may be prepared recombinantly using standard protocols, for example, as described in Klint, et al. (2013) PLOS One, 8(5): e63865; Sambrook, etal. (1989) Molecular Cloning: A Laboratory Manual (Cold Spring Harbour Press), in particular Sections 16 and 17; Ausubel, et al. (1998) Current Protocols in Molecular Biology (John Wiley and Sons, Inc.), in particular Chapters 10 and 16; and Coligan, et al. (1997) Current Protocols in Protein Science (John Wiley and Sons, Inc.), in particular Chapters 1 , 5 and 6. Cell penetrating moieties and labelling moieties may then be incorporated using methods known in the art where required, e.g. for the conjugates of the disclosure.4. Compositions

[0245] In accordance with the present disclosure, the proteinaceous molecules and conjugates are useful in compositions and methods for determining the presence of pathological TDP-43 in the brain of a subject and determining if a subject has a neurodegenerative disease associated with TDP-43 pathology. This, in some embodiments, the proteinaceous molecules and conjugates may be in the form of a pharmaceutical composition, wherein the pharmaceutical composition comprises, consists or consists essentially of a proteinaceous molecule or conjugate of the present disclosure and a pharmaceutically acceptable carrier or diluent.

[0246] The proteinaceous molecule or conjugate may be formulated into the pharmaceutical composition as a neutral or salt form.1006111282

[0247] As will be appreciated by those skilled in the art, the choice of pharmaceutically acceptable carrier or diluent will be dependent on the route of administration and on the nature of the condition and subject to be treated. The particular carrier or delivery system and route of administration may be readily determined by a person skilled in the art. The carrier or delivery system and route of administration should be carefully selected to ensure that the activity of the proteinaceous molecule or conjugate is not depleted during preparation of the formulation and the proteinaceous molecule or conjugate is able to reach the site of action intact. The pharmaceutical compositions of the disclosure may be administered through a variety of routes including, but not limited to, oral, intranasal, transmucosal, intramuscular, subcutaneous, intrathecal, intraventricular, intracerebral, intravesical, epidural, intravenous or intraperitoneal administration; especially intravenous, intrathecal or intracerebral administration; more especially intravenous administration.

[0248] The pharmaceutical forms suitable for injectable use include sterile injectable solutions or dispersions and sterile powders for the preparation of sterile injectable solutions. Such forms should be stable under the conditions of manufacture and storage and may be preserved against reduction, oxidation and microbial contamination.

[0249] A person skilled in the art will readily be able to determine appropriate formulations for the proteinaceous molecules or conjugates using conventional approaches. Techniques for formulation and administration may be found in, for example, Remington: The Science and Practice of Pharmacy, Adeboye Adejare and Joseph Remington (Ed), Academic Press, London, 23rdEdition, 2021.

[0250] Identification of preferred pH ranges and suitable excipients, such as antioxidants, is routine in the art, for example, as described in Katdare and Chaubel (2006) Excipient Development for Pharmaceutical, Biotechnology and Drug Delivery Systems (CRC Press). Buffer systems are routinely used to provide pH values of a desired range and may include, but are not limited to, carboxylic acid buffers, such as acetate, citrate, lactate, tartrate and succinate; glycine; histidine; phosphate; tris(hydroxymethyl)aminomethane (Tris); arginine; sodium hydroxide; glutamate; and carbonate buffers. Suitable antioxidants may include, but are not limited to, phenolic compounds such as butylated hydroxytoluene and butylated hydroxyanisole; vitamin E; ascorbic acid; reducing agents such as methionine or sulfite; metal chelators such1006111282 as ethylene diamine tetraacetic acid (EDTA); cysteine hydrochloride; sodium bisulfite; sodium metabisulfite; sodium sulfite; ascorbyl palmitate; lecithin; propyl gallate; and alpha-tocopherol.

[0251] For injection, the proteinaceous molecule or conjugate may be formulated in an aqueous solution, suitably in physiologically compatible buffers such as Hanks’ solution, Ringer’s solution, dextrose solution or physiological saline buffer, such as phosphate buffered saline (PBS).

[0252] The compositions of the present disclosure may be formulated for administration in the form of liquids, containing acceptable diluents (such as saline and sterile water). Acceptable diluents and carriers are familiar to those skilled in the art and include, but are not restricted to, saline, sterile water, ethoxylated and nonethoxylated surfactants, fatty alcohols, fatty acids, hydrocarbon oils (such as palm oil, coconut oil, and mineral oil), silicon oils, pH balancers, cellulose derivatives, emulsifying agents such as non-ionic organic and inorganic bases, preserving agents, steroid alcohols, triglyceride esters, phospholipids such as lecithin and cephalin, polyhydric alcohol esters and / or fatty alcohol esters.

[0253] Alternatively, the proteinaceous molecule or conjugate can be formulated readily using pharmaceutically acceptable carriers well known in the art into dosages suitable for oral administration. Such carriers enable the proteinaceous molecules or conjugates of the disclosure to be formulated in dosage forms such as tablets, pills, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a subject. These carriers may be selected from sugars, chitosan, starches, cellulose and its derivatives, malt, gelatin, talc, calcium sulfate, vegetable oils, synthetic oils, polyols, alginic acid, phosphate buffered solutions, emulsifiers, isotonic saline and pyrogen-free water.

[0254] Suspensions of the proteinaceous molecule or conjugate may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol or dextran. Optionally, the suspension may also contain suitable stabilisers or agents that increase the solubility of the proteinaceous molecules or conjugates to allow for the preparation of highly concentrated solutions.1006111282

[0255] Sterile solutions may be prepared by combining the proteinaceous molecule or conjugate in the required amount in the appropriate solvent with other excipients as described above as required, followed by sterilisation, such as filtration. Generally, dispersions are prepared by incorporating the various sterilised active agents into a sterile vehicle which contains the basic dispersion medium and the required excipients as described above. Sterile dry powders may be prepared by vacuum - or freeze-drying a sterile solution comprising the active agents and other required excipients as described above.

[0256] The peptide or conjugate of the disclosure may be compounded for convenient and effective administration in effective amounts with a suitable pharmaceutically acceptable carrier in dosage unit form. In some embodiments, a dosage unit form may comprise the peptide or conjugate in an amount in the range of from about 0.25 pg to about 2000 mg. The peptide or conjugate may be present in an amount of, for example, from about 0.25 pg to about 2000 mg / mL of carrier. In embodiments where the pharmaceutical composition comprises one or more additional active ingredients, the dosages are determined by reference to the usual dose and manner of administration of the said ingredients.5. Methods of Use

[0257] The inventors have conceived that the proteinaceous molecules and conjugates of the disclosure will be useful for determining the presence of pathological TDP-43 in the brain of a subject and determining if a subject has a neurodegenerative disease associated with TDP-43 pathology.

[0258] Accordingly, in another aspect, there is provided a method of determining if a subject has a neurodegenerative disease associated with TDP-43 pathology comprising, consisting or consisting essentially of: a) administering to the subject a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X orXI) of the disclosure; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology.1006111282

[0259] The disclosure also provides a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X or XI) of the disclosure for use in determining if a subject has a neurodegenerative disease associated with TDP-43 pathology, the use of a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X or XI) of the disclosure for determining if a subject has a neurodegenerative disease associated with TDP-43 pathology, or a use of a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X or XI) of the disclosure in the manufacture of a medicament for determining if a subject has a neurodegenerative disease associated with TDP-43 pathology.

[0260] The neurodegenerative disease is any neurodegenerative disease that is characterised by, or otherwise associated with, TDP-43 pathology. Typically such diseases are characterised by or associated with cytoplasmic accumulation of nuclear TDP-43 and with aberrant phosphorylation and fragmentation of TDP-43. In some embodiments, the disease is associated with aggregated TDP-43.

[0261] Representative neurodegenerative diseases associated with TDP-43 pathology include, but are not limited to, ALS, FTD, Alzheimer's disease, Parkinson's disease, mild cognitive impairment, Lewy body disease, argyrophilic grain disease, corticobasal degeneration, chronic traumatic encephalopathy, hippocampal sclerosis, limbic predominant age-related TDP-43 encephalopathy and Huntington's disease. In particular embodiments, the disease is ALS or FTD. The ALS may be familial or sporadic ALS. The FTD may be familial or sporadic FTD.

[0262] Numerous pathological variants of TDP-43 are known to be associated with sporadic or familial ALS and FTD, including for example A315T, N345K, M337V, G294A, A382T and G287S mutations. However those skilled in the art will recognise that the scope of the present disclosure is not limited to determining the presence of pathological TDP-43 having one or more of these mutations or determining if a subject has a neurodegenerative disease associated with TDP-43 pathology which is harbouring one or more of these mutations.

[0263] In some embodiments, the subject is suspected of having a neurodegenerative disease associated with TDP-43 pathology. Symptoms of the neurodegenerative disease, such as ALS or FTD, typically include behavioural and physical deficits characteristic of or associated with the disease, such as disinhibition,1006111282 hyperactivity, motor deficits and reduced muscle strength. In some embodiments, the subject is displaying one or more of these symptoms.

[0264] Step b) may be carried out at a time point after step a) that allows the proteinaceous molecule or conjugate that has bound to pathological TDP-43 in the brain to be detected over unbound proteinaceous molecule or conjugate (i.e. unbound proteinaceous molecule or conjugate has been cleared from the brain). For example, in some embodiments, step b) is carried out between about 30 mins and about 24 hours (and all integer minutes there between) after step a), about 30 mins and about 18 hours after step a), about 30 mins and about 12 hours after step a), about 30 mins and about 8 hours after step a), about 30 mins and about 6 hours after step a), about 30 mins and about 5 hours after step a), about 30 mins and about 4 hours after step a), about 1 hour and about 5 hours after step a), about 1 hour and about 4 hours after step a) or about 1 hour and about 3 hours after step a). In some embodiments, step b) is carried out at about 30 mins or about 1 , 2, 3, 4, 5 or 6 hours after step a). In particular embodiments, step b) is carried out at about 2 hours after step a).

[0265] Embodiments of the proteinaceous molecule and conjugate are as discussed supra. In particular embodiments, the proteinaceous molecule comprises a labelling moiety as discussed herein. In some embodiments, the proteinaceous molecule comprises two labelling moieties as discussed herein (e.g. one fluorescent label and one radionuclide). In some embodiments, the proteinaceous molecule or conjugate comprise a cell penetrating moiety as discussed herein. Suitable labelling moieties and cell penetrating moieties are as discussed supra.

[0266] The method for determining the presence or absence of the conjugate or proteinaceous molecule will depend on the specific agent used (e.g. the identity of the labelling moiety). A skilled person will readily able to determine suitable methods. For example, the presence or absence of the conjugate or proteinaceous molecule may be determined using imaging.

[0267] In particular embodiments, the presence or absence of the conjugate or proteinaceous molecule is determined using imaging. Suitable methods include, but are not limited to, nuclear imaging, magnetic resonance imaging (MRI) (e.g. when the labelling moiety is a contrast agent) and optical imaging (e.g. for detection of fluorescence).1006111282

[0268] In some embodiments, the presence or absence of the conjugate or proteinaceous molecule is determined using nuclear imaging. In such embodiments, the labelling moiety comprises a radionuclide. After administration of the conjugate or labelled proteinaceous molecule, for example, by intravenous injection, the subject is placed on the scanner. As the injected radionuclide decays it emits a positron that annihilates with an electron, producing a pair of gamma rays or photons that travel in opposite directions. In general terms, the emitted photons are detected when they reach a scintillator material in a scanning device, creating a burst of light that is detected by photomultiplier tubes. Suitable nuclear imaging methods include PET, SPECT, CT, scintigraphy, SPECT / CT, PET / CT, PET / MRI or SPECT / MRI. In some embodiments, the nuclear imaging is PET, CT or SPECT. In particular embodiments, the nuclear imaging is PET. In some embodiments, the nuclear imaging is a combination of PET or SPECT with CT or MRI, such as SPECT / CT, PET / CT, PET / MRI or SPECT / MRI.

[0269] In some embodiments, the labelling moiety is a radionuclide selected from the group consisting of11C,18F,13N,15O,124l,44Sc,64Cu,68Ga,82Rb,86Y,89Zr and133La, and the presence or absence of the conjugate or proteinaceous molecule is determined using PET.

[0270] In some embodiments, the labelling moiety is a radionuclide selected from the group consisting of99mTc,111In,123l and131l, and the presence or absence of the conjugate or proteinaceous molecule is determined using SPECT.

[0271] In some embodiments, particularly when the labelling moiety is a fluorescent label, the presence or absence of the conjugate or proteinaceous molecule is determined using optical imaging, such as fluorescence. A skilled person will be well aware of suitable systems for optical imaging, such as the fluorescence systems described, for example, in Refaat et al. (2022) Journal of Nanobiotechnology, 20: 450, the entire content of which is incorporated herein by reference.

[0272] When the labelling moiety is a pre-targeting moiety, the method may further comprise step a1 ) [i.e. between steps a) and b)], wherein step a1 ) comprises administering to the subject a label molecule (e.g. a radionuclide binding moiety or synthon comprising a radionuclide as described herein) comprising a functional group that is capable of forming a covalent bond with the pre-targeting moiety in vivo. Suitable functional groups include, but are not limited to, an azidyl, alkenyl (e.g. C3-C101006111282 alkenyl), cycloalkenyl (e.g. Ce-C cycloalkenyl), a heterocycloalkenyl (e.g. C5-C16 heterocycloalkenyl), alkynyl (e.g. C3-C10 alkynyl), cycloalkynyl (e.g. Ce-C cycloalkynyl), heterocycloalkynyl (e.g. a C5-C16 heterocycloalkynyl) or tetrazinyl group. Exemplary functional group pairings (i.e. the functional groups present on the pretargeting moiety and the label molecule) include azidyl and alkynyl, azidyl and cycloalkynyl, azidyl and heterocycloalkynyl, alkenyl and tetrazinyl, cycloalkenyl and tetrazinyl, heterocycloalkenyl and tetrazinyl, and the like.

[0273] Step a1 ) may be carried out, for example, between about 15 mins and about 24 hours (and all integer minutes there between) after step a), about 30 mins and about 18 hours after step a), about 30 mins and about 12 hours after step a), about 30 mins and about 8 hours after step a), about 30 mins and about 6 hours after step a), about 30 mins and about 4 hours after step a), about 30 mins and about 2 hours after step a), about 1 hour and about 3 hours after step a) or about 1 hour and about 2 hours after step a). In some embodiments, step a1 ) is carried out at about 30 mins or about 1 , 2, 3 or 4 hours after step a). Step b) may then be carried out thereafter, such as between about 15 mins and about 4 hours (and all integer minutes there between) after step a1 ), about 30 mins and about 3 hours after step a1 ), about 30 mins and about 2 hours after step a1 ) or about 30 mins and about 1 hour after step a1 ). In some embodiments, step b) is carried out at about 15 or 30 mins, or about 1 , 2, 3 or 4 hours after step a1 ).

[0274] The methods may be used to monitor the progression of the neurodegenerative disease associated with TDP-43 pathology or the efficacy of a treatment (e.g. as discussed herein) over a period of time. Accordingly, the method may further comprise repeating steps a) and b) [and optionally step a1 ) when the labelling moiety is a pre-targeting moiety] at one or more subsequent time points, and comparing the conjugate or proteinaceous molecule detected in b) to evaluate whether there has been a change in the neurodegenerative disease (e.g. an increase or decrease in the amount of pathological TDP-43, such as a TDP-43 aggregate). Comparing the conjugate or proteinaceous molecule detected at the two or more time points may include comparing an amount of conjugate or proteinaceous molecule detected (e.g. emitted radiation, fluorescence intensity, etc.), or comparing the presence or absence of the conjugate or proteinaceous molecule. Accordingly, in some embodiments, step b) comprises determining an amount of the conjugate or proteinaceous molecule in the brain of the subject.1006111282

[0275] Suitable periods of time between the repetition of steps a) and b) [and optionally step a1 )] include, for example, 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11 months, or 1 , 1 .5, 2, 2.5 or 3 years. In some embodiments, steps a) and b) [and optionally step a1 )] are repeated more than once, such as at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 times. The repetition of these steps may be performed over several years, such as at least 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10 years.

[0276] In a further aspect, there is provided a method for determining the presence of pathological TDP-43 in the brain of a subject, comprising, consisting or consisting essentially of: a) administering to the subject a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X orXI) of the disclosure; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that pathological TDP-43 is present in the brain.

[0277] Also provided is a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X or XI) of the disclosure for use in determining the presence of pathological TDP-43 in the brain of a subject, a use of a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X or XI) of the disclosure for determining the presence of pathological TDP-43 in the brain of a subject, or a use of a conjugate (e.g. Formula II, III, IV, V, VI, VII, VIII or XII) or a proteinaceous molecule (e.g. Formula I, IX, X or XI) of the disclosure in the manufacture of a medicament for determining the presence of pathological TDP-43 in the brain of a subject.

[0278] In some embodiments, the pathological TDP-43 is aggregated TDP-43 or a TDP-43 aggregate. In some embodiments, the pathological TDP-43 comprises one or more mutations selected from the group consisting of A315T, Q331 K, Q343R, N345K, R361 S, N390D, N390S, M337V, G294A, G294V, G295S, A382T and G287S. In some embodiments, the pathological TDP-43 comprises one or more mutations selected from the group consisting of A315T, N345K, M337V, G294A, A382T and G287S.1006111282

[0279] Step b) may be carried out at a time point after step a) that allows the proteinaceous molecule or conjugate that has bound to pathological TDP-43 in the brain to be detected over unbound proteinaceous molecule or conjugate (i.e. unbound proteinaceous molecule or conjugate has been cleared from the brain). Suitable time points are as discussed supra.

[0280] Embodiments of the proteinaceous molecule, conjugate and methods for determining the presence or absence of the conjugate or proteinaceous molecule are as discussed supra.

[0281] When the labelling moiety is a pre-targeting moiety, the method may further comprise step a1 ) [i.e. between steps a) and b)], wherein step a1 ) comprises administering to the subject a label molecule (e.g. a radionuclide binding moiety or synthon comprising a radionuclide as described herein) comprising a functional group that is capable of forming a covalent bond with the pre-targeting moiety in vivo. Suitable functional groups and label molecules are as discussed supra.

[0282] Step a1 ) may be carried out, for example, between about 15 mins and about 24 hours (and all integer minutes there between) after step a), about 30 mins and about 18 hours after step a), about 30 mins and about 12 hours after step a), about 30 mins and about 8 hours after step a), about 30 mins and about 6 hours after step a), about 30 mins and about 4 hours after step a), about 30 mins and about 2 hours after step a), about 1 hour and about 3 hours after step a) or about 1 hour and about 2 hours after step a). In some embodiments, step a1 ) is carried out at about 30 mins or about 1 , 2, 3 or 4 hours after step a). Step b) may then be carried out thereafter, such as between about 15 mins and about 4 hours (and all integer minutes there between) after step a1 ), about 30 mins and about 3 hours after step a1 ), about 30 mins and about 2 hours after step a1 ) or about 30 mins and about 1 hour after step a1 ). In some embodiments, step b) is carried out at about 15 or 30 mins, or about 1 , 2, 3 or 4 hours after step a1 ).

[0283] The methods may further comprise treating the subject if they have been determined to have a neurodegenerative disease associated with TDP-43 pathology or pathological TDP-43 in the brain. Accordingly, in another aspect, there is provided a method of treating a neurodegenerative disease associated with TDP-43 pathology comprising, consisting or consisting essentially of:1006111282 a) administering to the subject a conjugate according to Formula II or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology; and d) treating the subject for the neurodegenerative disease associated with TDP- 43 pathology.

[0284] Suitable embodiments of steps a) to c) are as discussed supra, including the proteinaceous molecule, conjugate, neurodegenerative disease associated with TDP- 43 pathology, methods for determining the presence or absence of the conjugate or proteinaceous molecule, optional step a1 ) and time points between steps are as discussed supra.

[0285] Suitable treatments include, but are not limited to, riluzole, a neuroprotectant (e.g. edaravone, sodium phenylbutyrate / taurursodiol), a SOD1 inhibitor (e.g. tofersen), dextromethorphan / quinidine, an antidepressant (e.g. citalopram, escitalopram, paroxetine, trazodone or sertraline), an antipsychotic (e.g. olanzapine, quetiapine, risperidone, brexpiprazole or aripiprazole), an antiepileptic, a cholinesterase inhibitor (e.g. galantamine, rivastigmine or donepezil), an immunotherapy (e.g. lecanemab or donanemab), an N-methyl-D-aspartate antagonist (e.g. memantine) or a peptide described in WO 2021 / 026601 A1 .

[0286] The method may be used to monitor the progression of the neurodegenerative disease associated with TDP-43 pathology or the efficacy of the treatment over a period of time. Accordingly, the method may further comprise repeating steps a) and b) [and optionally step a1 ) when the labelling moiety is a pretargeting moiety] subsequent to treating the subject for the neurodegenerative disease, and comparing the conjugate or proteinaceous molecule detected in b) to evaluate whether there has been a change in the neurodegenerative disease (e.g. an increase or decrease in the amount of pathological TDP-43, such as a TDP-43 aggregate). Comparing the conjugate or proteinaceous molecule detected at the two or more time1006111282 points may include comparing an amount of conjugate or proteinaceous molecule detected (e.g. emitted radiation, fluorescence intensity, etc.), or comparing the presence or absence of the conjugate or proteinaceous molecule. Accordingly, in some embodiments, step b) comprises determining an amount of the conjugate or proteinaceous molecule in the brain of the subject.

[0287] Suitable periods of time between the repetition of steps a) and b) [and optionally step a1 )] include, for example, 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11 months, or 1 , 1 .5, 2, 2.5 or 3 years. In some embodiments, steps a) and b) [and optionally step a1 )] are repeated more than once, such as at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 times. The repetition of these steps may be performed over several years, such as at least 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10 years.

[0288] In some embodiments, treatment is continued until the conjugate or proteinaceous molecule is determined to be absent in step b).

[0289] The use of the proteinaceous molecule of the disclosure for targeting a labelling moiety to pathological TDP-43 in the brain is also contemplated herein.

[0290] The proteinaceous molecule or conjugate of the disclosure may also be used to identify or image a pathological TDP-43 expressing cell. In some embodiments, the cell is a cell of the central nervous system, especially a cell of the brain, such as a neuron or glial cell. In particular embodiments, the pathological TDP-43 is aggregated TDP-43.

[0291] Any one of the methods and uses described above may involve administration of an effective amount of the proteinaceous molecule or conjugate of the disclosure as described supra. The conjugate or proteinaceous molecule may be administered via any suitable route of administration, such as oral, intranasal, transmucosal, intramuscular, subcutaneous, intrathecal, intraventricular, intracerebral, intravesical, epidural, intravenous or intraperitoneal administration; especially intravenous, intrathecal or intracerebral administration. In some embodiments, the conjugate or proteinaceous molecule is administered via intravenous administration.

[0292] The dosage will depend on the subject and the route of administration. A skilled person will readily be able to determine suitable dosages. For example, the proteinaceous molecule or conjugate may be administered in an amount in the range of from about 0.25 pg to about 2000 mg. The proteinaceous molecule or conjugate1006111282 may be administered, for example, as an intravenous infusion over a suitable period of time, such as about 1 , 5, 10, 15, 20, 25, 30, 35, 40, 45, 50 or 55 mins, or about 1 or 2 hours.

[0293] In order that the disclosure may be readily understood and put into practical effect, particular preferred embodiments will now be described by way of the following non-limiting examples.EXAMPLESGENERAL METHODS

[0294] Solvents [e.g. N,N-dimethylformamide (DMF), dichloromethane (DCM) and acetonitrile (CH3CN)] were obtained from ChemSupply (ChemSupply, Australia). Amino acids, coupling reagents and resins for Fmoc-solid-phase peptide synthesis (SPPS) were obtained from either Chemlmpex (Chemlmpex International, US) or Sigma Aldrich unless otherwise stated. Automated solid phase peptide synthesis (SPPS) was performed on a CEM Liberty Blue™ Peptide Synthesiser (CEM, USA) according to the manufacturer’s operating instructions. Manual SPPS work up, as well fluorophore labelling, was performed in polypropylene syringes equipped with porous filter disc, purchased from Torviq.

[0295] Analytical and semi-preparative reversed-phase high performance liquid chromatography (HPLC) was performed on a Shimadzu LC20AD series HPLC system (Shimadzu, Japan) equipped with PDA detector (A = 200 - 800 nm), Autosampler and Fraction Collector. Peptides were analysed using an analytical Polaris 3 C18-A 150 x 4.6 mm column (Agilent Technologies, USA) at a flow rate of 1 .0 mL min’1, and purified using a semi-prep Polaris 3 C18-A 150 x 4.6 mm column (Agilent Technologies, USA) at a flow rate of 5.0 mL min’1. HPLC column was kept at 30°C and chromatograms were collected at 210 nm unless otherwise stated. Gradients for analytical HPLC were run over 17 minutes using a mobile phase composed of 0.1 % formic acid (FA) in H2O (Solvent A) and 0.1 % FA in CH3CN (Solvent B) in a linear gradient as indicated; gradients for semi-preparative HPLC were run over 17 minutes using a mobile phase composed of 0.1 % formic acid (FA) in H2O (Solvent A) and 0.1 % FA in CH3CN (Solvent B) in a linear gradient as indicated. Semi-preparative injections of 0.6 mL to 1.2 mL were carried out onto a 2.0 mL loop using autosampler module. Analysis of the chromatograms was conducted using LabSolutions proprietary software (Shimadzu,1006111282Japan) and retention times (rt, min) of pure peptides reported with the gradients specified.

[0296] Peptides were flash frozen in liquid nitrogen and lyophilized using ScanVac Cool Safe Touch (Labogene, Denmark). Mass spectra were obtained on a Shimadzu liquid chromatography-mass spectrometry (LCMS)-8050 LCMS system (Shimadzu, Japan) in positive electron spray [ESI+] mode, fitted with a Polaris 3 C18-A 50 x 4.6 mm column (Agilent Technologies, USA). Liquid chromatography system was operated using LCMS grade solvents and mobile phase of 0.1 % formic acid in water (Solvent A) and 0.1 % formic acid in acetonitrile (Solvent B) and a linear gradient of 5- 95% Solvent B over 5 min. Data analysis was performed using LabSolutions v5.91 proprietary software.EXAMPLE 1 - PEPTIDE AND CONJUGATE DESIGN AND PREPARATION

[0297] Conjugates were prepared based on the amino acid sequence of a-helix 6 of 14-3-39 (the theta isoform of the 14-3-3 protein), which has an amino acid sequence of SEQ ID NO: 1 . Conjugates based on the amino acid sequence of the corresponding region of a non-interacting protein, 14-3-3^ (the theta isoform of the 14-3-3 protein), were used as control conjugates (KKGIVDQSQQA, SEQ ID NO: 86). For conjugate design, extended sequences encompassing amino acid residues 135-164 of 14-3-36 and 14-3-3^ were used.

[0298] All conjugates were furnished with cell penetrating peptide sequence 3+2R where three arginines (R) are attached to the N-terminus and 2 arginines (R) to the C- terminus. Conjugates were labelled with Cyanine-7 (Cy7), a far-red fluorescent dye for in vivo imaging using LI-COR Pearl Imaging system, and FITC-Ahx, a green fluorescent dye for in vitro microscopy. The conjugate sequences are provided in Table 4.TABLE 4CONJUGATE SEQUENCES1006111282

[0299] Conjugates were synthesised as below and characterised using LCMS (refer to Table 5).TABLE 5CONJUGATE CHARACTERISATION1006111282Materials and Methods

[0300] Automated SPPS was carried out using standard fluorenyl methoxycarbonyl (Fmoc)-solid-phase peptide synthesis on low swell 100 - 200 mesh Wang resin (0.05 mmol reaction scale, 0.5 mmol g-1 loading) on a CEM Liberty Blue™ Peptide Synthesiser (CEM, USA). Initial amino acid loading: Wang resin (100-200 mesh; 0.65 mmol g-1 , 77 mg, 0.05 mmol) was weighed into a 10 mL polypropylene syringe equipped with a porous polypropylene frit, which was used as the reaction vessel. The resin was washed with DCM (3 x 5 mL) before being allowed to swell in DCM (5 mL) for at least 0.5 h prior to the loading of the first amino acid. A solution of Fmoc-AA-OH (4 equiv.) was dissolved in a mixture of dry DCM (2 mL), DMF (2 mL), hydroxybenzotriazole (HOBt) (4 equiv.) and N,N’-diisopropylcarbodiimide (DIC) (4 equiv.) and taken up into the syringe with resin and stirred overnight using an orbital shaker. The resin was then capped with acetic anhydride (0.1 mL) and N,N- diisopropylethylamine (DIPEA) (0.1 mL) in DCM (3 mL) for 30 min. The resin was then washed with DCM (3 x 4 mL) and DMF (3 x 4 mL). Automated peptide synthesiser: Resin was pre-swelled in 50 / 50 DMF and DCM for 1 hour. Amino acids were dissolved in DMF at a concentration of 0.2 M before being transferred to the synthesiser. Peptides were synthesised using sequential amid coupling from C- to N-terminus for 5 minutes at 75°C, using five equiv. of amino acid with 10 equiv. of activator (0.5 M DIC ( / V, / V'-diisopropylcarbodiimide) in DMF) and 5 equiv. of activator base [0.5 M Oxyma (ethyl cyanohydroxyiminoacetate), 0.05 M DIPEA in DMF], followed by Fmoc deprotection in 20% piperidine in DMF for 3 minutes at 75°C and 3x resin wash in DMF. Following final Fmoc deprotection, resin was removed from synthesiser, transferred to a syringe fitted with a propylene filter, and either labelled with fluorophore (see below) or washed with DMF (x3), DCM (x3) and methanol (x3). Double couplings were performed for arginine residues to ensure complete coupling. Cleavage: Peptide was cleaved from resin using cleavage cocktail of 92.5% TFA (trifluoroacetic acid), 2.5% TIPS (triisopropylsilane), 2.5% thioanisole, and 2.5% H2O for 3 hours at room temperature, precipitated in ice cold diethyl ether, dissolved in H2O, freeze dried, and purified using a Shimadzu LC-20AD HPLC (Shimadzu, Japan).

[0301] To a peptide bound resin in a syringe fitted with a propylene filter, fluorophore with carboxylic acid functionality, HOBt (3 equiv.), benzotriazol-1 -1006111282 yloxytripyrrolidinophosphonium hexafluorophosphate (PyBOP) (3 equiv.), and DIPEA (6 equiv.) in DMF (4 mL) was taken up to the syringe and agitated overnight in orbital shaker. Resin was then washed with DMF (x3), DCM (x3) and methanol (x3) and proceeded to cleavage.EXAMPLE 2 - IN VIVO IMAGING

[0302] The ability of the conjugates of Example 1 to detect TDP-43 aggregates was determined using iTDP-43 mice. Tail vein injection of 3+2R0-Cy7 (targeting) and 3+2R -Cy7 (non-targeting control) imaging agents into healthy, wild-type (WT) mice showed comparable signal in the brain after 2 hours while injection into iTDP-43 mice showed significantly higher signal for targeting peptide 3+2R0-Cy7 versus background signal for the non-targeting peptide (refer to Figures 1 A and 1 B). Importantly, 3+2R0- Cy7 signal in iTDP-43 mice is significantly higher than in healthy controls and microscopic imaging of brain slices show cytosolic 3+2R0-Cy7 signals only in iTDP-43 mice that bare TDP-43 pathology (refer to Figures 2A and 2B). This data demonstrates that the 3+2R0-Cy7 peptide yields a TDP-43 aggregate specific signal in the CNS over the time frame of a typical PET scan, making it an ideal candidate for the development of a PET ligand to neuroimage TDP-43.

[0303] iTDP-43 and WT mice were intravenously injected at 3.5 months old with 3+2R0-Cy7 (7.8 mg / kg, equivalent to 20 mM blood volume concentration). Mice were imaged and perfused at 1 h, 2 h, 24 h post injection and compared to non-injected WT mice (refer to Figures 3 and 4). While 3+2R0-Cy7 shows a typical accumulation and clearance in WT mice (refer to Figure 3, orange line, with a peak brain signal at 1 hour), a retention effect due to specific TDP-43 binding is observed in iTDP-43 mice (refer to Figure 3, green line, with a peak brain signal at 2 hours). Brain signal returns to background (which is equivalent to uninjected mice) after 24 hours. The TDP-43 specific PET imaging window is at about 2 hours post-injection in mice (refer to Figures 3 and 4).Materials and Methods

[0304] Transgenic mice with expression of neuronal transactivator [line mThy1.2- tTA(41 )] and human A315T mutant TDP-43 under control of the tetracyclineresponsive element promoter [line pTRE-TDP-43 A315T (6)] (iTDP-43 mice) have been described previously (Ke et al. (2015) Acta Neuropathol, 130: 661-678). iTDP-100611128243 and WT mice were maintained on a C57BI / 6 background and housed on a 12-hour light / dark cycle with access to standard chow and water ad libitum. Mice were aged to 3 months-of-age prior to testing. Mice were injected intravenously (tail vein) with 7.8 mg / kg 14-3-39-Cy7 (prepared in accordance with Example 1 ) or non-TDP43-binding control 14-3-3 -Cy7 (prepared in accordance with Example 1 ) dissolved in 0.9% NaCI. Mice were left for 1 h, 2 h or 24 h post-injection, then anaesthetised, shaved, and whole-body imaging was performed in Pearl LI-COR. Mice were then transcardially perfused with phosphate buffered saline, tissues removed (liver, kidney, heart, lung, spleen, brain) and imaged together and individually in the Pearl LI-COR for each animal. All images were normalised to 14-3-3 -Cy7-injected control, and signal specificity was confirmed using a non-injected control. All animal experiments have been approved by the Macquarie University Animal Ethics Committee.EXAMPLE 3 - IN SILICO BINDING OF VARIANT PEPTIDES

[0305] The core sequence of 14-3-30 (optimised using alpha fold) was docked onto an alpha fold generated model of TDP-43 to determine the binding affinity of the core sequence (refer to Table 6; 14-3-30). Amino acids in the 11 amino acid sequence were then systematically mutated to alanine, folding of resulting peptides determined using alpha-fold and resulting peptides docked to the same binding site to determine docking scores of mutants.

[0306] Binding of 14-3-30 core sequence (11 amino acids) to TDP-43 using Schrodinger Bioluminate suite gave a reference binding score of -9.68 (refer to Tables 6 and 7, and Figure 5). Q140A and G147A mutations were found to bind better than the original sequence, with docking scores of -9.89 and -9.87, respectively). All other mutations led to decreased binding affinity.TABLE 6IN SILICO DOCKING SCORES FOR BINDING OF ALANINE SCAN OF 14-3-3© CORE SEQUENCETO TDP-431006111282

[0307] Further mutations of the core sequence of 14-3-30 were investigated in silico. Q140, T141 , 1142, D143, G147 and A148 were mutated and docked onto an alpha fold generated model of TDP-43 to determine the binding affinity compared to the core sequence (refer to Table 7; 14-3-30). Folding of peptides was determined using alphafold and resulting peptides were docked to the same binding site to determine docking scores of mutants.TABLE 7IN SILICO DOCKING SCORES FOR BINDING OF 14-3-3© CORE SEQUENCE MUTANTS TO TDP-431006111282to bind better than the original sequence, with docking scores of -10.15 and -9.73, respectively.

[0309] The disclosure of every patent, patent application, and publication cited herein is hereby incorporated herein by reference in its entirety.

[0310] The citation of any reference herein should not be construed as an admission that such reference is available as “Prior Art” to the instant application.

[0311] Throughout the specification the aim has been to describe the preferred embodiments of the disclosure without limiting the disclosure to any one embodiment or specific collection of features. Those of skill in the art will therefore appreciate that, in light of the instant disclosure, various modifications and changes can be made in the particular embodiments exemplified without departing from the scope of the present disclosure. All such modifications and changes are intended to be included within the scope of the appended claims.

Claims

1. 1006111282CLAIMS1. A proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, includingR, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, includingS, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including10061112821. L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof:RKQTIDNSQGA [SEQ ID NO: 1 ],2. The proteinaceous molecule according to claim 1 , wherein Xi is selected from the group consisting of Q, A, N, T, S, K, H, D, E, G, I, L, V and modified forms thereof.

3. The proteinaceous molecule according to claim 2, wherein Xi is Q.

4. The proteinaceous molecule according to any one of claims 1 -3, wherein X2 is selected from the group consisting of T, A, S, C, M, N, Q, D, E, Y, G, I, L, V and modified forms thereof.

5. The proteinaceous molecule according to claim 4, wherein X2 is T.

6. The proteinaceous molecule according to any one of claims 1 -5, wherein X3 is selected from the group consisting of I, A, G, L, V, F, Y, S, T, C, M, N, Q and modified forms thereof.

7. The proteinaceous molecule according to claim 6, wherein X3 is I.

8. The proteinaceous molecule according to any one of claims 1 -7, wherein X4 is selected from the group consisting of D, A, E, N, Q, C, T, S, G, I, L, V and modified forms thereof.

9. The proteinaceous molecule according to claim 8, wherein X4 is D or E.

10. The proteinaceous molecule according to any one of claims 1 -9, wherein X5 is selected from the group consisting of G, A, V I, L, F, Y, S, T, C, M and modified forms thereof.11 . The proteinaceous molecule according to claim 10, wherein X5 is G.100611128212. The proteinaceous molecule according to any one of claims 1 -11 , wherein Xe is selected from the group consisting of A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N, Q and modified forms thereof.

13. The proteinaceous molecule according to claim 12, wherein Xe is A orG.

14. The proteinaceous molecule according to any one of claims 1 -13, further comprising a labelling moiety.

15. The proteinaceous molecule according to claim 14, wherein the labelling moiety is a fluorescent label or a radionuclide.

16. The proteinaceous molecule according to claim 15, wherein the labelling moiety is a fluorescent label selected from the group consisting of fluorescein isothiocyanate (FITC), tetramethylrhodamine (TRITC), carboxytetramethylrhodamine (TAMRA), 2-(5-(1 -(6-(N-(2-maleimdylethyl)- amino)-6-oxohexyl)-1 ,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene)-1 ,3- pentadienyl)-1 -ethyl-3,3-dimethyl-5-sulfo-3H-indolium salt (Cy5), 6-[1 , 1 -dimethyl- 2-[5-(1 ,1 ,3-trimethylbenzo[e]indol-3-ium-2-yl)penta-2,4- dienylidene]benzo[e]indol-3-yl]hexanoic acid;chloride (Cy5.5), 1 -(5- carboxypentyl)-2-[(1 E,3E,5E,7Z)-7-(1 -ethyl-5-sulfo-3H-indol-2-ylidene)hepta-1 ,3,5-trienyl]-3H-indol-1 -ium-5-sulfonate (Cy7), 9-(2(or 4)-(N-(2-maleimdylethyl)- sulfonamidyl)-4(or2)-sulfophenyl)-2,3,6,7, 12,13,16, 17-octahydro-(1 -H,5H,11 H,15H-xantheno(2,-3,4-ij:5,6,7-i'j')diquinolizin-18-ium salt (Texas Red), 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY), (N-(4,4-difluoro-1 ,3,5,7- tetramethyl-4-bora-3a,4a-diaza-s-indacene-2-yl)iodoacetamide (BODIPY 507 / 545 IA), N-(4,4-difluoro-5,7-diphenyl-4-bora-3a,4a-diaza-s-indacene-3- propionyl)-N'-iodoacetylethylene diamine (BODIPY 530 / 550 IA), Alexa Fluor 350, Alexa Fluor 405, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 514, Alexa Fluor 532, Alexa Fluor 546, Alexa Fluor 555, Alexa Fluor 561 , Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, Alexa Fluor 635, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680, Alexa Fluor 700, Alexa Fluor 750, Alexa Fluor 790, Atto 390, Atto 425, Atto 430LS, Atto 465, Atto 490LS, Atto 495, Atto 514, Atto 488, Atto 520, Atto 532, Atto 540Q, Atto 550, Atto 565, Atto 590, Atto 594, Atto 633, Atto 647, Atto 655, Atto Rho14, Atto Rho6G, Atto Rho3B, Atto Rho11 , Atto Rho12,1006111282Atto Rho13, Atto Thiol 2, Atto Rho101 , Atto 580Q, Atto 680, Atto 700, Atto 647N, Atto 610, Atto 612Q, Atto 620, Atto 0xa12, Atto 725, Atto 740, Atto MB2, DyLight 350, DyLight 405, DyLight 488, DyLight 550, DyLight 594, DyLight 633, DyLight 650, DyLight 680, DyLight 755 and DyLight 800.

17. The proteinaceous molecule according to claim 15, wherein the labelling moiety is a radionuclide selected from the group consisting of11C,18F,13N,15O,124l,44Sc,64Cu,68Ga,82Rb,86Y,89Zr,133La,99mTc,111In,123l and131l.

18. The proteinaceous molecule according to claim 17, wherein the labelling moiety further comprises a radionuclide binding moiety.

19. The proteinaceous molecule according to claim 18, wherein the radionuclide binding moiety is a chelator or a synthon.

20. The proteinaceous molecule according to claim 14, wherein the labelling moiety is a pre-targeting moiety.

21. The proteinaceous molecule according to claim 20, wherein the pretargeting moiety comprises an azidyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl or tetrazinyl group.

22. The proteinaceous molecule according to any one of claims 1 -21 , further comprising a cell penetrating moiety.

23. The proteinaceous molecule according to claim 20, comprising two cell penetrating moieties.

24. The proteinaceous molecule according to claim 20 or claim 21 , wherein the cell penetrating moiety is a cell penetrating peptide.

25. The proteinaceous molecule according to claim 22, wherein the cell penetrating peptide comprises at least two arginine residues.

26. The proteinaceous molecule according to claim 23, wherein the cell penetrating peptide comprises two or three arginine residues.

27. A conjugate represented by Formula II:B-Z (II)1006111282 wherein:B is a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, includingR, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, includingS, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including1006111282I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof; andZ is a labelling moiety.

28. The conjugate according to claim 27, wherein Xi is selected from the group consisting of Q, A, N, T, S, K, H, D, E, G, I, L, V and modified forms thereof.

29. The conjugate according to claim 28, wherein Xi is Q.

30. The conjugate according to any one of claims 27-29, wherein X2 is selected from the group consisting of T, A, S, C, M, N, Q, D, E, Y, G, I, L, V and modified forms thereof.31 . The conjugate according to claim 30, wherein X2 is T.

32. The conjugate according to any one of claims 27-31 , wherein X3 is selected from the group consisting of I, A, G, L, V, F, Y, S, T, C, M, N, Q and modified forms thereof.

33. The conjugate according to claim 32, wherein X3 is I.

34. The conjugate according to any one of claims 27-33, wherein X4 is selected from the group consisting of D, A, E, N, Q, C, T, S, G, I, L, V and modified forms thereof.

35. The conjugate according to claim 34, wherein X4 is D or E.

36. The conjugate according to any one of claims 27-35, wherein X5 is selected from the group consisting of G, A, V I, L, F, Y, S, T, C, M and modified forms thereof.

37. The conjugate according to claim 36, wherein X5 is G.

38. The conjugate according to any one of claims 27-37, wherein Xe is selected from the group consisting of A, G, I, L, P, V, F, W, Y, D, E, R, H, K, S, T, C, M, N, Q and modified forms thereof.

39. The conjugate according to claim 38, wherein Xe is A or G.100611128240. The conjugate according to claim 27, wherein the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula III:Z1 RKX1X2X3X4NSQX5X6Z2 (III) wherein:Xi to Xe are as defined in any one of claims 27-39; andZ1 and Z2 are independently absent or a labelling moiety, wherein at least one of Z1 and Z2 is present.

41. The conjugate according to claim 27, wherein the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula IV:Y1Z1 RKX1X2X3X4NSQX5X6Z2Y2 (IV) wherein:Xi to Xe are as defined in any one of claims 27-39;Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;Z1 and Z2 are independently absent or a labelling moiety, wherein at least one of Z1 and Z2 is present.

42. The conjugate according to claim 27, wherein the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula V:Z3Y1 Z4X7X8X9Z1 R KX1 X2X3X4N S QX5X6Z2X10X11 Xi 2X13X14X15X16X17X18Xi 9X20X21 X22X23X24X25Z5Y2Z6 (V) wherein:Xi to Xe are as defined in any one of claims 27-39;1006111282X? to X25 are independently absent or any amino acid residue or a modified form thereof;Y1 and Y2 are independently absent or a cell penetrating moiety, wherein at least one of Y1 and Y2 is present;Z1 to Ze are independently absent or a labelling moiety, wherein at least one of Z1 to Ze is present.

43. The conjugate according to claim 42, wherein:X7, X13, X17 and X24 are independently selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof;Xs, X9, X12, X15 and X20 are independently selected from the group consisting of acidic amino acid residues, including D, E and modified forms thereof;X10 and X14 are independently selected from the group consisting of aromatic amino acid residues, including Y, W, F and modified forms thereof;X11 and X22 are independently selected from the group consisting of am ide- containing amino acid residues, including Q, N and modified forms thereof;X and X21 are independently selected from the group consisting of hydrophobic amino acid residues, including I, L, V, M and modified forms thereof;X18, X19 and X25 are independently selected from the group consisting of basic amino acid residues, including R, K, H and modified forms thereof; andX23 is selected from the group consisting of P and modified forms thereof.

44. The conjugate according to claim 42, wherein the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VI:Z3Y1Z4GDDZ1RKQTIDNSQGAZ2YQEAFDISKKEMQPTHZ5Y2Z6 (VI) wherein:Y1, Y2 and Z1 to Ze are as defined in claim 42.100611128245. The conjugate according to claim 42, wherein the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VII:Z3Y1GDDRKQTIDNSQGAYQEAFDISKKEMQPTHY2 (VII) wherein:Y1 and Y2 are independently a cell penetrating moiety; andZ3 is a labelling moiety.

46. The conjugate according to any one of claims 41 -45, wherein the cell penetrating moiety is a cell penetrating peptide.

47. The conjugate according to claim 46, wherein the cell penetrating peptide comprises at least two arginine residues.

48. The conjugate according to claim 47, wherein the cell penetrating peptide comprises two or three arginine residues.

49. The conjugate according to claim 27, wherein the conjugate comprises, consists or consists essentially of an amino acid sequence represented by Formula VIII:Z3RRRGDDRKQTIDNSQGAYQEAFDISKKEMQPTHRR (VIII) wherein:Z3 is a labelling moiety.

50. The conjugate according to any one of claims 27-49, wherein the labelling moiety is a fluorescent label or a radionuclide.51 . The conjugate according to claim 50, wherein the labelling moiety is a fluorescent label selected from the group consisting of fluorescein isothiocyanate (FITC), tetramethylrhodamine (TRITC), carboxytetramethylrhodamine (TAMRA), 2-(5-(1-(6-(N-(2-maleimdylethyl)-amino)-6-oxohexyl)-1 ,3-dihydro-3,3-dimethyl-5- sulfo-2H-indol-2-ylidene)-1 ,3-pentadienyl)-1-ethyl-3,3-dimethyl-5-sulfo-3H- indolium salt (Cy5), 6-[1 ,1-dimethyl-2-[5-(1 ,1,3-trimethylbenzo[e]indol-3-ium-2-1006111282 yl)penta-2,4-dienylidene]benzo[e]indol-3-yl]hexanoic acid;chloride (Cy5.5), 1 -(5- carboxypentyl)-2-[(1 E,3E,5E,7Z)-7-(1 -ethyl-5-sulfo-3H-indol-2-ylidene)hepta- 1 ,3,5-trienyl]-3H-indol-1 -ium-5-sulfonate (Cy7), 9-(2(or 4)-(N-(2-maleimdylethyl)- sulfonamidyl)-4(or2)-sulfophenyl)-2,3,6,7, 12,13,16, 17-octahydro-(1 -H,5H,11 H,15H-xantheno(2,-3,4-ij:5,6,7-i'j')diquinolizin-18-ium salt (Texas Red), 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY), (N-(4,4-difluoro-1 ,3,5,7- tetramethyl-4-bora-3a,4a-diaza-s-indacene-2-yl)iodoacetamide (BODIPY 507 / 545 IA), N-(4,4-difluoro-5,7-diphenyl-4-bora-3a,4a-diaza-s-indacene-3- propionyl)-N'-iodoacetylethylene diamine (BODIPY 530 / 550 IA), Alexa Fluor 350, Alexa Fluor 405, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 514, Alexa Fluor 532, Alexa Fluor 546, Alexa Fluor 555, Alexa Fluor 561 , Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, Alexa Fluor 635, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680, Alexa Fluor 700, Alexa Fluor 750, Alexa Fluor 790, Atto 390, Atto 425, Atto 430LS, Atto 465, Atto 490LS, Atto 495, Atto 514, Atto 488, Atto 520, Atto 532, Atto 540Q, Atto 550, Atto 565, Atto 590, Atto 594, Atto 633, Atto 647, Atto 655, Atto Rho14, Atto Rho6G, Atto Rho3B, Atto Rho11 , Atto Rho12, Atto Rho13, Atto Thiol 2, Atto Rho101 , Atto 580Q, Atto 680, Atto 700, Atto 647N, Atto 610, Atto 612Q, Atto 620, Atto 0xa12, Atto 725, Atto 740, Atto MB2, DyLight 350, DyLight 405, DyLight 488, DyLight 550, DyLight 594, DyLight 633, DyLight 650, DyLight 680, DyLight 755 and DyLight 800.

52. The conjugate according to claim 50, wherein the labelling moiety is a radionuclide selected from the group consisting of11C,18F,13N,15O,124l,44Sc,64Cu,68Ga,82Rb,86Y,89Zr,133La,99mTc,111In,123l and131l.

53. The conjugate according to claim 52, wherein the labelling moiety further comprises a radionuclide binding moiety.

54. The conjugate according to claim 53, wherein the radionuclide binding moiety is a chelator or a synthon.

55. The conjugate according to any one of claims 27-49, wherein the labelling moiety is a pre-targeting moiety.

56. The conjugate according to claim 55, wherein the pre-targeting moiety comprises an azidyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl or tetrazinyl group.100611128257. A method of determining if a subject has a neurodegenerative disease associated with TDP-43 pathology comprising: a) administering to the subject the conjugate according to any one of claims 27-56 or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I:RKX1X2X3X4NSQX5X6 (I) wherein:Xi is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; basic amino acid residues, including R, K, H and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; and hydrophobic amino acid residues, including I, L, V and modified forms thereof;X2 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X3 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof;X4 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; amide-containing amino acid residues, including Q, N and modified forms thereof; acidic1006111282 amino acid residues, including D, E and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; and C and modified forms thereof;X5 is selected from the group consisting of small amino acid residues, including S, T, A, G and modified forms thereof; hydrophobic amino acid residues, including I, L, V, M and modified forms thereof; aromatic amino acid residues, including Y, W, F and modified forms thereof; and C and modified forms thereof; andXe is any amino acid residue or a modified form thereof; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology.

58. The method according to claim 57, wherein the neurodegenerative disease associated with TDP-43 pathology is amyotrophic lateral sclerosis (ALS) or frontotemporal dementia (FTD).

59. The method according to claim 57 or claim 58, wherein step b) is carried out between about 30 minutes and about 6 hours after step a).

60. The method according to claim 59, wherein step b) is carried out between about 1 hour and about 3 hours after step a).61 . A method for determining the presence of pathological TDP-43 in the brain of a subject, comprising: a) administering to the subject the conjugate according to any one of claims 27-56 or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I as defined in claim 57; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; and1006111282 c) if the conjugate or proteinaceous molecule is present, determining that pathological TDP-43 is present in the brain.

62. The method according to claim 61 , wherein the pathological TDP-43 is aggregated TDP-43.

63. A method of treating a neurodegenerative disease associated with TDP-43 pathology comprising: a) administering to the subject a conjugate according to any one of claims 27-56 or a proteinaceous molecule comprising, consisting or consisting essentially of an amino acid sequence represented by Formula I as defined in claim 57; b) determining the presence or absence of the conjugate or proteinaceous molecule in the brain of the subject; c) if the conjugate or proteinaceous molecule is present, determining that the subject has a neurodegenerative disease associated with TDP-43 pathology; and d) treating the subject for the neurodegenerative disease associated with TDP-43 pathology.

64. The method according to any one of claims 57-63, wherein the presence or absence of the conjugate or proteinaceous molecule is determined by nuclear imaging.

65. The method according to claim 64, wherein the nuclear imaging is positron emission tomography (PET), computed tomography (CT) or singlephoton emission computed tomography (SPECT).

66. The method according to claim 65, wherein the nuclear imaging is PET.

67. A pharmaceutical composition comprising a proteinaceous molecule according to any one of claims 1 -26 or a conjugate according to any one of claims 27-56 and a pharmaceutically acceptable carrier or diluent.

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