US 2015O151 001A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2015/0151001 A1 Squires (43) Pub. Date: Jun. 4, 2015

(54) COMPOSITIONS AND METHODS FOR Publication Classification DETECTING OR ELMINATING SENESCENT CELLS TODAGNOSE OR TREAT DISEASE (51) Int. Cl. A647/48 (2006.01) (71) Applicant: Cenexys, Inc., San Francisco, CA (US) A614.9/00 (2006.01) A615 L/08 (2006.01) (72) Inventor: Shayne Squires, Coalville, UT (US) (52) U.S. Cl. CPC ...... A61K47/48246 (2013.01); A61 K5I/08 (2013.01); A61K 49/0056 (2013.01) (21) Appl. No.: 14/557.316 (57) ABSTRACT Disclosed are agents (e.g., peptides, polypeptides, proteins, (22) Filed: Dec. 1, 2014 Small molecules, antibodies, and antibody fragments that tar get senescent cells) and methods of their use for imaging senescent cells in vivo and for treating or preventing cancer, Related U.S. Application Data age-related disease, tobacco-related disease, or other diseases and disorders related to or caused by cellular senescence in a (63) Continuation of application No. 12/809,952, filed on mammal. The methods include administering one or more of Feb. 21, 2012, filed as application No. PCT/US2008/ the agents of the invention to a mammal, e.g., a human. The O13913 on Dec. 19, 2008. agents, which specifically bind to senescent cells, can be (60) Provisional application No. 61/015,416, filed on Dec. labeled with a radioactive label or a therapeutic label, e.g., a 20, 2007. cytotoxic agent. Patent Application Publication Jun. 4, 2015 Sheet 1 of 5 US 201S/O151001 A1

12 -- - w 1 1 - a -Y - -1 Se 1 -

s - s 1. y - - r ...------p-r" --

x

peptide concentration micronolai)

.C. seesCertififiasts -->. to Seese foasts -&-prostate epithetitiin Patent Application Publication Jun. 4, 2015 Sheet 2 of 5 US 201S/O151001 A1

g

r Mt

8 2 25 peptide concentration (micromokai) -KY- Seiescent fiboiasts -C- onsertescent ffoliasis - V. prostate epitheiun Patent Application Publication Jun. 4, 2015 Sheet 3 of 5 US 201S/O151001 A1

/ -- - - ^ S$f ^ - ^ / // --O / u / u ---i? u-1 -...------y--- 5, 20 25 33 35 - 4.5 SO peptide-icin A Conjugate (microficial

-- SeesCet fibrobiasts -O-- noiseiescent fibrosiasts -v. prostate epitheti in Patent Application Publication Jun. 4, 2015 Sheet 4 of 5 US 201S/O151001 A1

FG. 4

Patent Application Publication Jun. 4, 2015 Sheet 5 of 5 US 201S/O151001 A1

S & Š ? % US 2015/O151 001 A1 Jun. 4, 2015

COMPOSITIONS AND METHODS FOR ere length and replicate indefinitely. The minority of cancers DETECTING OR ELMINATING SENESCENT that do not express telomerase have alternative lengthening of CELLS TO DIAGNOSE OR TREAT DISEASE telomere (ALT) mechanisms. 0006 Indirect evidence suggests some relationship STATEMENT REGARDING SEQUENCE between replicative senescence and aging. Cultured cells LISTING from old donors exhibit senescence after fewer population 0001. The Sequence Listing associated with this applica doublings than cells from young donors (Martin et al., Lab. tion is provided in text format in lieu of a paper copy, and is Invest. 23:86-92, 1970; Schneideret al., Proc. Nat. Acad. Sci. hereby incorporated by reference into the specification. The USA 73:3584-3588, 1976). Cells from short-lived species name of the text file containing the Sequence Listing is senesce after fewer population doublings than cells from 200201 401D1 SEQUENCE LISTING..txt. The text file is long-lived species (Rohme, D., Proc. Nat. Acad. Sci. USA 70.5 KB, was created on Nov. 29, 2014, and is being submit 78:5009-3320, 1981). Cultured cells from donors with ted electronically via EFS-Web. hereditary premature aging syndromes such as Werner's Syn drome show senescence after fewer replications than cells FIELD OF THE INVENTION from age-matched controls (Goldstein, Genetics of Aging, 171-224, 1978; Martin, Genetic Effects on Aging, 5-39, 0002 This invention relates to compositions and methods 1990). Whether replicative senescence actually contributes to for the detection and treatment of cancer, age-related dis aging or age-related symptoms in vivo is questionable on the eases, tobacco-related diseases, and other diseases and disor basis of theoretical estimates of the number of cell divisions ders related to or caused by cellular senescence. that occur in Vivo and the absence of strong empirical evi dence. BACKGROUND OF THE INVENTION 0007. There are, however, other pathways to senescence 0003. The definition of cellular senescence has undergone than replication. Collectively, these are often referred to as Some revision since the phenomenon was described by stress-induced premature senescence (SIPS). Oxidative Leonard Hayflick as cessation of replication of cultured stress can shorten telomeres (von Zglinicki, Trends Biochem. human cells after a finite number of population doublings Sci. 27:339-344, 2002) and hyperoxia has been shown to (Hayflicket al., Exp. Cell Res. 37:585-621, 1961). Senescent induce Senescence. Gamma irradiation of human fibroblasts cells remain metabolically active but do not divide and resist in early to mid G1 phase causes senescence in a p53-depen apoptosis for long periods of time (Goldstein, Science 249: dent manner (Di Leonardo et al., Genes Dev. 8:2540-2551, 1129-1133, 1990). Cellular senescence is characterized by 1994). Ultraviolet radiation also induces cellular senescence. growth cycle arrest in the G1 phase, absence of S phase, and Other agents that can induce cellular senescence include lifespan control by multiple dominant genes (Stanulis-Prae hydrogen peroxide (Krtolica et al., Proc. Nat. Acad. Sci. USA ger, Mech. Ageing Dev. 38:1-48, 1987). 98: 12072-12077, 2001), sodium butyrate, 5-azacytadine, and 0004 Cellular senescence differs from quiescence and ter transfection with the Ras oncogene (Tominaga, Mech. Ageing minal differentiation in several important aspects. Senescent Dev. 123(8):927-936, 2002). Chemotherapeutic agents cells have characteristic morphological changes such as including doxorubicin, cisplatin, and a host of others have enlargement, flattening, and increased granularity (Dimri et been shown to induce Senescence in cancer cells (Roninson, al., Proc. Nat. Acad. Sci. USA 92:9363-9367, 1995). Senes Cancer Res. 63:2705-2715, 2003). 5-bromodeoxyuridine cent cells do not divide even if stimulated by mitogens treatment results in cellular senescence in both normal and (Campisi, Trends Cell Biol. 11:S27-S31, 2001). Senescence malignant cells (Michishita et al., J. Biochem. 126:1052 involves activation of p53 and/or Rb and their regulators such 1059, 1999). Generally speaking, agents that damage DNA as p16'', p21, and ARF. Except when p53 or Rb is inac can cause cellular senescence. The existence of cellular tivated, senescence is irreversible. Senescent cells express senescence in vivo has been demonstrated. In a study by increased levels of plasminogen activator inhibitor (PAI) and Dimri et al., published in 1995, senescent fibroblasts were exhibit staining for B-galactosidase activity at pH 6 (Sharp shown to exhibit staining for B-galactosidase activity at pH 6. less et al., J. Clin. Invest. 113:160-168, 2004). Irreversible G1 These cells failed to incorporate tritiated thymidine and arrest is mediated by inactivation of cyclindependent kinase retained 3-galactosidase activity after replating but did not (CdK) complexes which phosphorylate Rb. P21 accumulates divide. Quiescent fibroblasts did not show staining Kerati in aging cells and inhibits Cdk4-CdK6. P16 inhibits CdK4 nocytes, umbilical vein endothelial cells, and mammary epi CdK6 and accumulates proportionally with B-galactosidase thelial cells all showed increased staining with increased activity and cell volume (Stein et al., Mol. Cell. Biol. 19:2109 population doublings. Immortalized cells and terminally dif 2117, 1999). P21 is expressed during initiation of senescence ferentiated keratinocytes did not show staining Staining was but need not persist; p 16 expression helps maintain senes performed on skin biopsies to test whether senescence is cence once initiated. observed in vivo. An age-dependent pattern in which an 0005 Replicative senescence, the type of senescence increased number of cells showed staining with increased originally observed by Hayflick, is related to the progressive donorage was observed in the dermis and epidermis (Dimriet shortening of telomeres with each cell division. Senescence is al., Proc. Nat. Acad. Sci. USA 92:9363-93.67, 1995). The induced when certain chromosomal telomeres reach a critical existence of an increase in the number of senescent fibroblasts length (Mathon and Lloyd, Nat. Rev. Cancer 3:203-213, has been shown in the lungs of Subjects with emphysema 2001; Martins, U.M. Exp Cell Res. 256:291-299, 2000). relative to subjects without emphysema (Müller et al., Resp. Senescence can be abrogated by the expression of telomerase Res. 7:32-41, 2006). which lengthens telomeres; human fibroblasts undergo repli 0008 Cellular senescence confers a number of functional cation indefinitely when transfected to express telomerase. changes on the cell that likely have clinical relevance. Senes Most cancers express telomerase in order to maintain telom cent endothelial cells secrete elevated levels of plasminogen US 2015/O151 001 A1 Jun. 4, 2015

activator inhibitor 1 (PAI-1; Kletsas et al., Ann. N.YAcad. Sci. SUMMARY OF THE INVENTION 908:11-25, 2000). Senescent fibroblasts over express colla 0012. This invention features compositions and methods genase and under express collagenase inhibitors (West et al., for the detection and imaging of senescent cells in vitro or in Exp Cell Res 184:138-147, 1989). Serial passages of human vivo in order to predict the risk of diseases or disorders related fibroblasts from a 25 year old donor showed increased to or caused by cellular senescence (e.g., cancer occurrence, elastase endopeptidase type activity (Homsy et al., Journal of cancer metastasis, cardiovascular disease, cerebrovascular Investigative Dermatology 91:472-477, 1988). Endothelial disease, Alzheimer's disease, and emphysema). The inven cells obtained from tissue overlying atherosclerotic plaques tion also features compositions and methods for treating, were observed to have a senescent morphology and express preventing, or inhibiting the development or progression of increased levels of PAI-1 and intracellular adhesion molecule diseases or disorders related to or caused by cellular senes 1 and decreased levels of nitric oxide (Davis et al., British cence (e.g., by administering an agent that results in the death Heart Journal 60:459-464, 1988, Arterioscler. Thromb. and removal of one or more, all, or Substantially all senescent 11:1678-1689, 1991: Finn et al., Circulation 105:1541-1544, cells or cells expressing one or more senescence markers 1976: Comi et al., Exp. Cell Res. 219:304-308, 1995: Chang from an organism). The compositions and methods can be et al., Proc. Nat. Acad. Sci. USA 92: 11190-11194, 1995). administered in order to prevent, ameliorate, inhibit the devel Indirect evidence that cellular senescence may play a role in opment of, or treat diseases or disorders related to or caused cardiovascular disease also is provided by the observation by cellular senescence (e.g., cancer, cardiovascular disease, that shorter leukocyte telomere length is associated with an cerebrovascular disease, Alzheimer's disease, emphysema, osteoarthritis, and other age-related diseases). The invention increased risk of premature myocardial infarction (Brouilette features peptides which bind with higher affinity to senescent et al., Arteriosclerosis, Thrombosis, and Vascular Biology cells than non-senescent cells. 23:842-846, 2003). 0013. In a first aspect, the invention features a peptide, 0009 Cancer cells are immortal, meaning that they can polypeptide, antibody, or antibody fragment agent having an replicate indefinitely without exhibiting senescence. A pre amino acid sequence set forth in any one of SEQID NOS:1-3 ponderance of opinion in the Scientific community says that and 5-8 or a peptide, polypeptide, antibody, antibody frag the teleological purpose of senescence is to prevent cancer by ment, or Small molecule agent that is capable of specifically limiting the number of cell divisions that can occur. This view binding to an antigen having an amino acid sequence having is Supported by experiments in mice showing that p53 knock at least 20 amino acids with at least 80% sequence identity to out results in increased cancerincidence and severity. Indirect any one of the amino acid sequences set forth in SEQ ID evidence that senescence Suppresses cancer occurrence NOS:11-23. In one embodiment, the antigen has an amino acid sequence having at least 20 amino acids of any one of the includes the observations that oncogenes immortalize or amino acid sequences set forth in SEQ ID NOS:11-23. In extend cellular lifespan and tumor suppressors Rb and p53, another embodiment, the antigen has any one of the amino which are critical for senescence, suffer a loss of function acid sequences set forth in SEQID NOS:11-23. mutation in cancer (Shay et al., Biochem. Biophys. Acta. 0014. In embodiments of the first aspect of the invention, 1071:1-7, 1991). the agent includes a detectable label, a therapeutic agent, a 0010 Senescent cells can also promote tumorigenesis. chelating agent, or a linker moiety. An agent of the first aspect Senescent stromal cells express tumor promoting as well as of the invention can be indirectly or directly attached to the tumor Suppressing factors that exert a paracrine effect on detectable label. The linker moiety can have the amino acid neighboring epithelial cells; Such effects include mitogenic sequence GGGC (SEQ ID NO:9), GGGS (SEQID NO:10), ity and antiapoptosis (Changet al., Proc. Nat. Acad. Sci. USA or GG. A detectable label can be a radioactive agent, fluores 97(8):4291-4296, 2000). Senescent fibroblasts have been cent agent, bioluminescent molecule, epitope tag, or heavy shown to stimulate premalignant and malignant epithelial metal. Radioactive agents include iodine, astatine, and bro mine labels that are attached to an amino acid of an agent of cells but not normal epithelial cells to form tumors in mice; the first aspect of the invention. A radioactive agent can also this occurred when as few as 10% of the fibroblasts were be technetium-99m. Fluorescent agents include fluorescein senescent (Krtolica et al., Proc. Nat. Acad. Sci. USA isothiocyanate (FITC), allophycocyanin (APC), phycoeryth 98:12072-12077, 2001). Tumor wafl/cipl/sdil promoting fac rin (PE), rhodamine, tetramethyl rhodamine isothiocyanate tors secreted by senescent cells are partly mediated by p21 (TRITC), fluorescent protein (GFP), enhanced GFP (eGFP), (Roninson, Cancer Res.63:2705-2715, 2003). A threshold of yellow fluorescent protein (YFP), cyan fluorescent protein senescent stromal cells appears to provide a milieu allowing (CFP), red fluorescent protein (RFP), and dsRed. A biolumi adjacent premalignant epithelial cells to Survive, migrate, and nescent molecule can be luciferase. Epitope tags include divide (Campisi, Nat. Rev. Cancer 3:339-349, 2003). c-myc, hemagglutinin, and histidine tags. Atherapeutic agent 0011. In summary, cellular senescence does occur in vivo can be a cytotoxic agent. Such as an alkylating agent, an and is a likely sequel to environmental insults. Its prevalence antibiotic, an antineoplastic agent, an antimetabolic agent, a increases with age at least in Some tissue compartments. ribosomalactivity inhibitor, an antiproliferative agent, a tubu Senescence confers functional changes on the cell which have lin inhibitor, a topoisomerase I or II inhibitor, a growth factor, been associated to some degree with various age-related dis an hormonal agonist or antagonists, an apoptotic agent, an eases (Chang et al., Proc. Nat. Acad. Sci. USA 97 (8):4291 immunomodulator, a radioactive agent, a phospholipase, or a 4296, 2000). Senescent cells also contribute to tumor forma cytotoxic peptide or lysin. A cytotoxic agent can also be ricin, tion. There exists a need for agents that are capable of doxorubicin, methotrexate, camptothecin, homocamptoth detecting senescent cells in vivo and for treating or preventing ecin, thiocolchicine, colchicine, combretastatin, combretas diseases and disorders related to or caused by cellular senes tin A-4, podophyllotoxin, rhizoxin, rhizoxin-d, dolistatin, CCCC. paclitaxel, CC1065, ansamitocin p3, maytansinoid, Streptol US 2015/O151 001 A1 Jun. 4, 2015

ysin O. Stoichactis toxin, phallolysin, staphylococcus alpha cular disease, Alzheimer's disease, osteoarthritis, cardiac toxin, holothurin A, digitonin, melittin, lysolecithin, car diastolic dysfunction, benign prostatic hypertrophy, aortic diotoxin, cerebratulus A toxin, or any derivative thereof. A aneurysm, and emphysema. chelating agent joined to an agent of the first aspect of the 0021. In a sixth aspect, the invention features a method for invention can be an ininocarboxylic reactive group, a identifying a peptide or polypeptide capable of detecting polyaminopolycarboxylic reactive group, diethylenetri senescent cells by (a) culturing cells to produce Senescent aminepentaacetic acid (DTPA), or 1,4,7,10-tetraazacy cells, (b) exposing the senescent cells to a phage peptide clododecane-1,4,7,10-tetraacetic acid (DOTA). In any library, (c) recovering the senescent cells and phage from said embodiment, the agent of the first aspect of the invention can phage peptide library bound to said senescent cells, (d) elut further include a pharmaceutically acceptable carrier. An ing and amplifying bound phage to produce amplified phage, agent of the first aspect of the invention can specifically bind (e) repeating steps (a)-(d) one or more times, and (f) recov to a senescent cell. Such as a senescent lung, breast, colon, ering a peptide or polypeptide expressed by the amplified prostate, gastric, hepatic, ovarian, esophageal, or bronchial phage that is capable of detecting senescent cells. epithelial or stromal cell, a senescent skin epithelial or stro 0022. In a seventh aspect, the invention features a method mal cell; a senescent glial cell; or a senescent vascular endot for identifying an antibody or antibody fragment capable of helial or stromal cell. specifically binding to a senescent cell-specific antigen by 0015. In a second aspect, the invention features a method contacting an antibody or antibody fragment with a polypep of imaging a senescent cell-containing region in a mammal, tide having at least 20 amino acids having at least 80% amino Such as a human, in Vivo, by administering to the mammal an acid sequence identity to any one of the amino acid sequences agent of the first aspect of the invention that is joined to a set forth in SEQID NOS:11-23, and identifying an antibody detectable label, allowing the agent to bind senescent cells or antibody fragment that binds the polypeptide with a disso and allowing unbound agent to be cleared from the body of ciation constant of less than 107 M. said mammal, and obtaining an image of the Senescent cell 0023. In an eighth aspect, the invention features a method containing region. Senescent cell-containing regions include for identifying an antibody or antibody fragment capable of the breast, prostate, gastrointestinal tract, liver, lungs, intrac specifically binding to a senescent cell-specific antigen by ranial space, nasopharynx, oropharynx, larynx, esophagus, administering a polypeptide having at least 20 amino acids mediastinum, abdomen and pelvis, any region of the body having at least 80% amino acid sequence identity to any one containing peripheral vasculature, and the entire body. An of the amino acid sequences set forth in SEQID NOS:11-23 image of the Senescent cell-containing region can be obtained to a mammal, allowing the mammal to generate a humoral by Scintigraphy. immune response to the polypeptide, and identifying an anti body or antibody fragment that binds the polypeptide with a 0016. In a third aspect, the invention features a method of dissociation constant of less than 107 M. Mammals suitable predicting cancer risk in a mammal. Such as a human, by for the identification of antibodies that specifically bind administering to the mammalanagent of the first aspect of the senescent cell-specific antigens include mice, hamsters, rats, invention that is joined to a detectable label and predicting an guinea pigs, chickens, goats, sheep, cows, horses, non-human elevated cancer risk in the mammal by detecting binding of primates, and humans. the agent to a senescent cell of the mammal. The method can 0024. In an ninth aspect, the invention features a method be used to predict the risk of prostate cancer, colon cancer, for identifying a small molecule capable of specifically bind lung cancer, squamous cell cancer of the head and neck, ing to a senescent cell-specific antigen by contacting a small esophageal cancer, hepatocellular carcinoma, gastric cancer, molecule with a polypeptide having at least 20 amino acids pancreatic cancer, ovarian cancer, and breast cancer. having at least 80% amino acid sequence identity to any one 0017. In a fourth aspect, the invention features a method of of the amino acid sequences set forth in SEQID NOS:11-23 treating or preventing disease in a mammal. Such as a human, and identifying a small molecule that binds the polypeptide by administering to the mammal an agent of the first aspect of with a dissociation constant of less than 107 M. the invention joined to a cytotoxic agent. 0025 Inantenth aspect, the invention features a method of 0018. In a fifth aspect, the invention features a method of making an antibody or antibody fragment by recombinantly treating or preventing disease in a mammal. Such as a human, expressing a nucleic acid sequence that encodes an amino by administering to the mammal a nucleic acid molecule acid sequence having one or more of SEQ ID NOs: 1-3 and encoding a cytotoxic agent and an agent of the first aspect of 5-8, wherein the antibody or antibody fragment specifically the invention. binds a senescent cell. 0026. In an eleventh aspect, the invention features a 0019. The nucleic acid molecule can be administered in a method of cellular therapy performed by administering to a vector, Such as an adenoviral vector. In one embodiment, the mammal. Such as a human, in need thereof an agent of the cytotoxic agent and agent of the first aspect of the invention invention prior to, concurrent with, or following administra are expressed as a single polypeptide chain. tion of a cellular therapeutic. 0020. In either the fourth or fifth aspects of the invention, 0027. In antwelfth aspect, the invention features a method diseases that can be treated or prevented include cancer, age of cellular therapy performed by contacting an agent of the related diseases, tobacco-related diseases, and skin wrinkles invention to a donor cell, tissue, or organ prior to, concurrent Cancers that can be treated or prevented include prostate with, or following administration of the cell, tissue, or organ cancer, colon cancer, lung cancer, squamous cell cancer of the to a mammal. Such as a human. A donor cell, tissue, or organ head and neck, esophageal cancer, hepatocellular carcinoma, can be an autologous, allogeneic, Syngeneic, or Xenogeneic gastric cancer, pancreatic cancer, ovarian cancer, and breast cell, tissue, or organ. A donor cell can be a stem cell. Such as cancer. Age-related or tobacco-related diseases include car a hematopoietic, umbilical cord blood, totipotent, multipo diovascular disease, cerebrovascular disease, peripheral vas tent, or pluripotent stem cell. US 2015/O151 001 A1 Jun. 4, 2015

0028. In a thirteenth aspect, the invention features a kit Somerase I and II inhibitors, hormonal agonists or antago containing an agent of the invention and one or more of a nists, immunomodulators, or agents that cause cell lysis detectable label, a therapeutic agent, a chelating agent, or a including naturally occurring or synthetic peptides. Cyto linker moiety. toxic agents may be cytotoxic when activated by light or 0029. The term “about” is used hereinto mean a value that infrared (Photofrin, IR dyes; Nat. Biotechnol. 19(4):327-331, is +10% of the recited value. 2001), may operate through other mechanistic pathways, or 0030. By “administration' or “administering is meant a be supplementary potentiating agents. method of providing a dosage of an agent of the invention to 0035. By “detectable label” is meant any type of label a mammal (e.g., a human), where the route is, e.g., topical, which, when attached to an agent of the invention, renders the oral, parenteral (e.g., intravenous, intraperitoneal, intrarte agent detectable. A detectable label may be toxic or non rial, intradermal, intramuscular, or Subcutaneous injection, toxic, and may have one or more of the following attributes, inhalation, optical drops, or implant), nasal, vaginal, rectal, or without restriction: fluorescence (Kieferet al., WO9740055), Sublingual application in admixture with a pharmaceutically color, toxicity (e.g., radioactivity, e.g., a Y-emitting radionu acceptable carrier adapted for such use. The preferred method clide, Auger-emitting radionuclide, B-emitting radionuclide, of administration can vary depending on various factors, e.g., an C.-emitting radionuclide, or a positron-emitting radionu the components of the pharmaceutical composition, site of clide), radiosensitivity, or photosensitivity. Although a the potential or actual disease (e.g., the location of lung, detectable label may be directly attached, for example, to an breast, colon, prostate, liver, brain, heart, etc.), and the sever amino acid residue of an agent of the invention, or indirectly ity of disease. attached, for example, by being complexed with a chelating 0031. By “analog is meant an agent that differs from, but group that is attached (e.g., linked via a covalent bond or is structurally, functionally, and/or chemically related to the indirectly linked) to an amino acid residue of an agent of the reference agent. The analog may retain the essential proper invention. A detectable label may also be indirectly attached ties, functions, or structures of the reference agent. Most to an agent of the invention by the ability of the label to be preferably, the analog retains at least one biological function specifically bound by a second molecule. One example of this of the reference agent. Generally, differences are limited so type of an indirectly attached label is a biotin label that can be that the structure or sequence of the reference agent and the specifically bound by a second molecule, streptavidin. The analog are similar overall. For example, a peptide analog and second molecule may also be linked to a moiety that allows its reference peptide may differ in amino acid sequence by neutron capture (e.g., a boron cage as described in, for one or more amino acid Substitutions, additions, and/or dele example, Kahl et al., Proc. Natl. Acad. Sci. USA 87:7265 tions, or the presence of one or more non-naturally occurring 7269, 1990). amino acid residues, in any combination. An analog of a 0036. A detectable label may also be a metal ion from peptide or polypeptide of the invention may be naturally heavy elements or rare earth ions, such as Gd", Fe", Mn", occurring, such as an allelic variant, or it may be a variant that or Cr" (see, e.g., Curter, Invest. Radiol. 33(10):752-761, is not known to occur naturally. Non-naturally occurring ana 1998). Preferred radioactive detectable labels are radioactive logs of peptides may be made by direct synthesis, by modi iodine labels (e.g., ‘I,’’I, ‘I,’’I, or ''I) that are capable fication, or by mutagenesis techniques. of being coupled to each D- or L-Tyr or D- or L-4-amino-Phe 0032. By “chelating agent' is meanta molecule that forms residues present in the agents of the invention. Preferred multiple chemical bonds with a single metal atom. Prior to non-radioactive detectable labels are the many known dyes forming the bonds, the chelating agent has more than one pair that are capable of being coupled to NH2-terminal amino acid ofunshared electrons. The bonds are formed by sharing pairs residues. of electrons with the metal atom. Chelating agents include, 0037 Preferred examples of detectable labels that may be for example, an iminodicarboxylic group or a polyaminopo toxic to cells include ricin, diphtheria toxin, and radioactive lycarboxylic group. Chelating agents may be attached to an detectable labels (e.g., 'I, 2I, *, *I, II, 77Lu, Cu, agent of the invention using the methods generally described 7Cu, 'Sim, 166Ho, 18.Re, 188Re, 21 At 212Bi, 225Ac, 7Ga, in Liu et al., Bioconjugate Chem. 12(4):653, 2001: Alter et al., Ga, 75 Br, 7°Br, 77Br, 117'Sn, 7Sc, 109Pd, 89Sr. 15°Gd, U.S. Pat. No. 5,753,627; and PCT Publication No. WO 149Pm, 142Pr '''Ag, 165 Dy 213Bi, 11 In, 114m In, 201 Ti, 195mpt, 91/01144; each of which is hereby incorporated by refer 'Pt, Y and 'Y). These compounds, and others described ence). An agent of the invention may be complexed, through herein may be directly or indirectly attached to an agent of the its attached chelating agent, to a detectable label, thereby invention or its analogs. A toxic detectable label may also be resulting in an agent that is indirectly labeled. Similarly, a chemotherapeutic agent (e.g., camptothecins, homocamp cytotoxic or therapeutic agents, may also be attached via a tothecins, 5-fluorouracil or adriamycin), or may be a radi chelating group to an agent of the invention. osensitizing agent (e.g., paclitaxel, gemcitabine, fluoropyri 0033. By “coupled' is meant the characteristic of a first midine, metronitoZil, or the deoxycytidine analog 2.2 molecule being joined to a second molecule by a covalent difluoro-2'-deoxycytidine (dFdCyd) to which is directly or bond or through noncovalent intermolecular attraction. indirectly attached an agent or analog thereof of the present 0034. By “cytotoxic agent' is meant any naturally occur invention. ring, modified, or synthetic compound that is toxic to cells. 0038 A detectable label, when coupled to an agent of the Such agents are useful in the treatment of neoplasms, and in invention emits a signal that can be detected by a signal the treatment of other symptoms or diseases characterized by transducing machine. In some cases, the detectable label can cell proliferation or a hyperactive cell population. Cytotoxic emit a signal spontaneously, such as when the detectable label agents can also be used to target undesirable cells or tissues is a radionuclide. In other cases the detectable label emits a other than neoplastic cells or tissues, e.g., senescent cells. signal as a result of being stimulated by an external field Such Cytotoxic agents include, but are not limited to, alkylating as when the detectable label is a relaxivity metal. Examples of agents, antibiotics, antimetabolites, tubulin inhibitors, topoi signals include, without limitation, gamma rays, X-rays, vis US 2015/O151 001 A1 Jun. 4, 2015 ible light, infrared energy, and radio waves. Examples of cine, norvaline, (2.3, or 4) 3-pyridyl-Ala, 1,4-diaminobutyric signal transducing machines include, without limitation, acid, and 1,3-diaminopropionic acid, respectively. In all gamma cameras including SPECT/CT devices, PET scan aspects of the invention, it is noted that when amino acids are ners, fluorimeters, and Magnetic Resonance Imaging (MRI) not designated as either D- or L-amino acids, the amino acid machines. is either an L-amino acid or could be either a D- or L-amino 0039. By “diagnostically effective amount” is meant a acid. Examples of peptides of the invention include those dose of detectably-labeled agent that, when administered peptides having the sequence of, or a sequence Substantially internally to a mammal, is quantitatively sufficient to be identical to, the sequences set forth in SEQID NOs: 1-3 and detected by a signal transducing machine external to the 5-8 and related sequences. These peptide sequences can also mammal (e.g., a gamma camera used in gamma Scintigraphy) be incorporated into a polypeptide, protein, antibody, oranti but typically is quantitatively insufficient to produce a phar body fragment. macological effect. 0045. By “reducing agent” is meant a chemical compound 0040. By “imaging agent' is meant a compound that, used to reduce another chemical compound by donating elec when administered to a living Subject, Such as a mammal trons, thereby becoming oxidized. (e.g., a human), allows the visualization of internal structures (e.g., cells, tissues, and organs) and, in Some cases can pro 0046 By “specifically binds” is meant that an agent of the vide information as to the function of a cell, tissue, or organ in invention recognizes and binds to a target (e.g., a senescent the subject. cell), but does not substantially recognize and bind to a non 0041. By “linker moiety' is meant a sequence of amino target (e.g., non-senescent cells), both in vivo and in a sample, acid residues, e.g., at least one, two, three, four, five, six, e.g., an in vitro biological sample, that includes, e.g., senes seven, eight, nine, ten, fifteen, twenty, thirty, forty, fifty, or cent cells. A desirable agent of the invention specifically more residues, that couples an agent of the invention (e.g., a binds to senescent cells. Preferably, the agents of the inven peptide, polypeptide, protein, Small molecule, antibody, or tion bind senescent cells with at least 2, 5, 10, 20, 100, or 1000 antibody fragment that target senescent cells) to, e.g., one or fold greater affinity than they bind to non-senescent cells. more of a detectable label, atherapeutic agent, and a chelating Alternatively, agents of the invention specifically bind to agent. senescent cells with a dissociation constant less than 10M, 0042. By “senescent cell' is meant a cell that is metaboli more preferably less than 107M, 10M, 10M, 10M, cally active but is permanently withdrawn from the cell cycle 10'M, or 10'’M, and most preferably less than 10M, (see, e.g., Campisi, Cell 120:513-522, 2005). Senescent cells 10M, or 10-5M. do not replicate and possess one or more of the following 0047. By “substantial sequence identity” or “substantially additional characteristics attributed to senescent cells: cell identical” is meant that a nucleic acid oramino acid sequence cycle arrest in the G1 phase; an enlarged, flattened morphol exhibits at least 50%, preferably 60%, 70%, 75%, or 80%, ogy, increased granularity; staining for B-galactosidase activ more preferably 85%, 90% or 95%, and most preferably 99% ity at pH 6: Senescence associated heterochromatic foci, and identity to a reference amino acid sequence (e.g., one or more characteristic gene expression that is in part regulated by p16 of the sequences set forth in SEQID NOs: 1-3 and 5-8). For and p21. Alternatively, a senescent cell is a cell that can be amino acid sequences, the length of comparison sequences induced to become senescent (e.g., by stress) or that expresses will generally be at least 5 amino acids, preferably at least 10 cell-surface markers characteristic of Senescent cells; these contiguous amino acids, more preferably at least 15, 20, 25. markers include senescent cell-specific antigens having the 30, 40, 50, 60, 80, 90, 100, 150, 200, 250, 300, or 350 con polypeptide sequences set forth in SEQID NOS:11-23. Sen tiguous amino acids, and most preferably the full-length esecent cell-specific antigens are those peptides, polypep amino acid sequence. Sequence identity is typically mea tides, or glycoproteins that are expressed on the cell Surface of sured using BLASTR (Basic Local Alignment Search Tool) senescent cells, but are absent or only weakly expressed on or BLASTR2 with the default parameters specified therein the cell Surface of non-senescent cells. (see, Altschul et al., J. Mol. Biol. 215:403-410, 1990); and 0043. By "peptide' is meant a polymer that includes two Tatiana et al., FEMS Microbiol. Lett. 174:247-250, 1999). or more amino acids joined to each other by a peptide bond or This software program matches similar sequences by assign a modified peptide bond. “Peptide' refers to both short chain ing degrees of homology to various Substitutions, deletions, polymers, commonly referred to as peptides, oligopeptides, and other modifications. Conservative substitutions typically or oligomers, having, e.g., about 10-50 linked amino acid include Substitutions within the following groups: glycine, residues, and to longer polymers having up to about 100 alanine, Valine, isoleucine, leucine; aspartic acid, glutamic amino acid residues in length. Peptides may contain amino acid, asparagine, glutamine; serine, threonine; lysine, argin acids other than the 20 gene-encoded amino acids, and link ine; and phenylalanine, tyrosine. ages other than peptide bonds and may include cyclic or 0048. By “therapeutic agent' is meant any compound that branched peptides. “Peptides' include amino acid sequences is used in the detection, diagnosis or treatment of disease. modified either by natural processes, or by chemical modifi Such compounds may be naturally-occurring, modified, or cation techniques that are well known in the art. Modifica synthetic. A therapeutic agent may be, for example, an agent tions may occur anywhere in a polypeptide, including the that causes apoptosis or necrosis of a cell (e.g., a senescent peptide backbone, the amino acid side-chains, and the amino cell) in an organism (e.g., a mammal. Such as a human). or carboxyl termini. thereby reducing the number of Such cells in the organism. 0044) The notations used herein for the peptide amino acid Therapeutic agents that reduce the number of senescent cells residues are those abbreviations commonly used in the art. in an organism may be, e.g., alkylating agents, antibiotics, The less common abbreviations Abu, Ava, B-Ala, hSer, Nile, antimetabolites, hormonal agonists or antagonists, anti- or Nva, Pal, Dab, and Dap stand for 2-amino-butyric acid, amino pro-apoptotic agents, immunomodulators, or Supplementary Valeric acid, beta-aminopropionic acid, homoserine, norleu potentiating agents. US 2015/O151 001 A1 Jun. 4, 2015

0049. By “treating, stabilizing, or preventing cancer is depends on mitochondrial dehydrogenase levels, and senes meant causing a reduction in the size of a tumor or in the cent cells have higher mitochondrial mass than their non number of cancer cells, slowing or preventing an increase in senescent counterparts (Martin-Ruiz et al., J. Biol. Chem. the size of a tumor or in cancer cell proliferation, increasing 279(17):17826-33, 2004). Consequently, baseline values for the disease-free survival time between the disappearance of a senescent cells are higher than for the other cell types. Treat tumor or other cancer and its reappearance, preventing an ment with SenL showed no significant effect on metabolic initial or Subsequent occurrence of a tumor or other cancer, or activity for any of the three cell types and therefore did not reducing an adverse symptom associated with a tumor or affect cell proliferation rates. other cancer. In a desired embodiment, the percent of tumor or 0055 FIG. 3 is a graph showing the cytotoxicity of SenC cancerous cells surviving the treatment is at least 20, 40, 60. (SEQ ID NO:5) conjugated to ricin A subunit as tested in 80, or 100% lower than the initial number of tumor or can senescent fibroblasts, non-senescent fibroblasts, and immor cerous cells, as measured using any standard assay, Such as talized prostate epithelial cells. Significantly more senescent those described herein. Desirably, the decrease in the number cells were killed than non-senescent cells, and no effect was of tumor or cancerous cells induced by administration of a observed on immortalized epithelium. compound of the invention is at least 2, 5, 10, 20, or 50-fold 0056 FIG. 4 (A and B) are fluorescent micrographs show greater than the decrease in the number of non-tumor or ing the specific binding of a peptide agent of the invention. non-cancerous cells. Desirably, the methods of the present Senescent cell binding peptide SenC (SEQ ID NO:5) was invention resultina decrease of 20, 40, 60, 80, or 100% in the conjugated to fluorescein and contacted with senescent fibro size of a tumor or number of cancerous cells as determined blasts (FIG. 4A) and non-senescent fibroblasts (FIG. 4B). using standard methods. Desirably, at least 20, 40, 60, 80.90, Both images were acquired using "/60 second exposure time. or 95% of the treated subjects have a complete remission in Senescent cells show perinuclear and cytoplasmic staining, which all evidence of the tumor or cancer disappears. Desir indicating significant internalization. Only faint Surface ably, the tumor or cancer does not reappear after more than 5, staining is visible on the non-senescent cells. 10, 15, or 20 years. 0057 FIG. 5 (A and B) are immunofluorescent micro 0050. By “treating, stabilizing, or preventing age-related graphs showing the cell Surface expression of cathepsin B on diseases” is meant causing a reduction in the number of senescent cells (FIG. 5A) and lack of expression on non symptoms, a decrease in severity of any, all, or Substantially senescent cells (FIG. 5B). all of the symptoms, a complete resolution of any, all, or Substantially all symptoms, or preventing the occurrence of DETAILED DESCRIPTION OF THE INVENTION any, all, or Substantially all symptoms associated with one or more of the age-related diseases including, but not limited to, 0058. The invention features agents of the invention (e.g., cardiovascular disease, cerebrovascular disease, peripheral peptides, polypeptides, proteins, Small molecules, antibodies, vascular disease, Alzheimer's disease, osteoarthritis, cardiac or antibody fragments that target senescent cells) capable of diastolic dysfunction, benign prostatic hypertrophy, and can specifically binding to senescent cells. Thus, agents of the cers that increase in incidence and prevalence with increasing invention can include, e.g., detectable labels, therapeutic patientage Such as cancer, e.g., breast cancer, prostate cancer, agents, chelating agents, and cytotoxic agents, and can be and colon cancer. used in the detection and treatment of diseases and conditions 0051. By “treating, stabilizing, or preventing tobacco-re associated with cellular senescence. lated diseases” is meant causing a reduction in the number of 0059. In an embodiment, agents of the invention (e.g., symptoms, a decrease in severity of any, all, or Substantially peptides, polypeptides, proteins, Small molecules, antibodies, all of the symptoms, a complete resolution of any, all, or orantibody fragments that target senescent cells) can be used Substantially all symptoms, or preventing the occurrence of to produce imaging agents by conjugation to a detectable any, all, or Substantially all symptoms associated with one or label and used for medical imaging. Images obtained using an more of the diseases associated with the use of Smoking imaging agent of the invention specifically show tissues, tobacco as a risk factor, including, but are not limited to, organs, and structures in the body (e.g., a human body) that cardiovascular disease, cerebrovascular disease, peripheral contain senescent cells by means of e.g., a signal emitted vascular disease, aortic aneurysms, emphysema, esophageal from the location of the cells, tissues, organs, and structures cancer, lung cancer, and squamous cell cancers of the head upon specific binding of the imaging agent to the senescent and neck. Tobacco-related diseases also include diseases that cells. The signal so obtained indicates the presence of senes are associated with the use of chewing tobacco, Such as Squa cent cells and allows semi-quantitative estimation of the rela mous cell cancers of the mouth. tive senescent cell content of tissues, organs, or structures. Structures, tissues, or organs that contain a threshold content 0052. Other features and advantages of the invention will of senescent cells are at an elevated risk for the Subsequent be apparent from the following description and from the development of diseases or disorders (e.g., cancer). Typically, claims. a subject deemed at risk for the subsequent development of BRIEF DESCRIPTION OF THE DRAWINGS diseases or disorders (e.g., cancer) exhibit an increase of at least 0.5%, more preferably 1, 1.5, 2, 2.5, 5, or 10%, and more 0053 FIG. 1 is a graph showing the cytotoxicity of SenL preferably at least 15% or more in the level of senescent cells (SEQ ID NO:4) when tested on senescent fibroblasts, non relative to a patient that does not have the disease or disorder. senescent fibroblasts, and immortalized prostate epithelial Use of one or more of the imaging agents of the invention cells. No cell death was observed in immortalized epithelium. allows for the determination of a patient’s risk for developing 0054 FIG. 2 is a graph showing the results of a WST assay a disease or disorder, Such as cancer or a neurodegenerative used to measure cell proliferation following treatment with disease. Patients determined to have an elevated risk of devel SenL on Senescent fibroblasts, non-senescent fibroblasts, and oping cancer or other diseases and disorders related to or immortalized prostate epithelial cells. The WST-1 assay caused by cellular senescence by the use of the imaging US 2015/O151 001 A1 Jun. 4, 2015 agents of the invention can be aggressively monitored, and thesis, A Practical Approach, Oxford University Press, New their cellular senescence related disease or disorder (e.g., a York, 2003); incorporated herein by reference in its entirety). cancer) canthereby be detected earlier, facilitating early treat The standard coupling techniques used to couple the amino ment. Surgical prophylactic treatment can also be adminis acids in order to form the agents of the invention are known in tered if necessary. the art (see, e.g., Chan and White, Supra). For example, a 0060. One or more of the imaging agents of the invention polyamide-Rink resin can be prepared by loading a polya can also be used to determine the Senescent cell content in the mide resin with Fmoc-Rink using chemical protocols well brain, which may include, e.g., senescent glial cells and known in the art (see, e.g., Chan and White, supra). The first senescent cerebrovascular cells, in order to predict the amino acid in the peptide sequence is coupled to the resin patient’s risk of developing, e.g., Alzheimer's disease or cere after removing Fmoc from the N-terminal amine of the resin brovascular disease. One or more of the imaging agents of the using piperidine. Once the coupling is complete, the resin is invention can also be used to predict the onset of vascular washed and Fmoc is removed from the coupled amino acid disease, including cardiovascular disease, by detecting the using piperidine. The resin is washed again, and the next senescent cell content in one or more vascular structures in a amino acid in the sequence is coupled to the previously patient. One or more of the imaging agents of the invention coupled amino acid. This process is repeated using the nec can also be used to predict the risk of developing emphysema essary amino acids until the desired peptide is formed. Fol by determining the senescent cell content in the lungs of a lowing the coupling of the final amino acid, the Fmoc group Smoker. is removed using piperidine. The terminal amine can be left as 0061. One or more of the agents of the invention (e.g., a free amine, or it can be acetylated. The peptide can be peptides, polypeptides, proteins, Small molecules, antibodies, cleaved from the resin using trifluoroacetic acid (TFA), tri or antibody fragments that target senescent cells) can also be isopropylsilane, and water according to techniques known in used to prepare compositions for therapeutic administration the art (see, e.g., Chan and White, Supra). The cleaved peptide (e.g., to treat, stabilize, inhibit the development or progres is then separated from the residue by filtration. The TFA is sion of, or prevent age-related diseases, tobacco-related dis typically evaporated to dryness followed by precipitation of eases, cancer, neurodegenerative diseases, and other diseases the peptide with diethyl ether. Typically, the final peptide and disorders related to or caused by cellular senescence) by product is purified using HPLC. Mass spectrometry is used to coupling of the agents of the invention to, e.g., therapeutic verify that the desired peptide is obtained. The agents of the and/or cytotoxic agents. Senescent cells have been observed invention (e.g., peptides, polypeptides, proteins, Small mol in skin to make up only a fraction of the stromal cell com ecules, antibodies, or antibody fragments that target senes partment even in old donors (Dimri et al., Proc. Nat. Acad. cent cells) can be readily prepared by automated Solid phase Sci. USA 92:9363-9367, 1995) and up to 16% of stromal peptide synthesis using any one of a number of well-known, fibroblasts in patients with emphysema (Bergner, Respiratory commercially available automated synthesizers, such as the Res. 7:32, 2006), but senescent cells can exert clinically sig Applied Biosystems ABI 433A peptide synthesizer. nificant paracrine effects even when they compose only 10% 0063. The agents of the invention (e.g., peptides, polypep of the content of stromal cells (Krtolica et al., Proc. Nat. Acad. tides, proteins, Small molecules, antibodies, or antibody frag Sci. USA 98: 12072-12077, 2001). Thus, eliminating senes ments that target senescent cells) can be labeled for fluores cent cells while leaving intact the majority of the cellular cence detection by labeling the agent with a fluorophore. Such compartment from which the senescent cells originate can as rhodamine or fluorescein, using techniques well known in ablate the harmful paracrine effects of senescent cells. The the art (see, e.g., Lohse et al., Bioconi. Chem. 8:503-509, secretion of elevated levels of collagenase and elastase and 1997). The senescent cell targeting agents of the invention depressed levels of collagenase inhibitors by senescent stro (e.g., peptides, polypeptides, proteins, Small molecules, anti mal cells implicates them in diseases or conditions that fea bodies, or antibody fragments that target senescent cells) can ture breakdown or decrease instructural integrity of the extra also be labeled with a radioactive metal or a relaxivity metal cellular matrix, including emphysema, aortic aneurysms, vas by coupling the agent to a metal chelating agent that chelates cular disease, osteoarthritis, and skin wrinkling Elimination a radioactive metal or relaxivity metal. Examples of chelating of Some, all, or substantially all senescent stromal cells in a agents include, but are not limited to, ininocarboxylic and patient (e.g., in a tissue or organ of a patient) can attenuate the polyaminopolycarboxylic reactive groups, diethylenetri progression, decrease the symptoms, or prevent the occur aminepentaacetic acid (DTPA), and 1,4,7,10-tetraazacy rence of these diseases and conditions. Senescent stromal clododecane-1,4,7,10-tetraacetic acid (DOTA). The chelat cells have the ability to stimulate tumorigenesis and, by weak ing agent can be coupled via its amino acid side chain directly ening the extracellular matrix, facilitate metastasis. Thus, the to the agents of the invention. Alternatively, an intervening elimination of some, all, or substantially all of the senescent amino acid sequence can be coupled using SPPS to both the stromal cells in a patient (e.g., in a tissue or organ of a patient) agents of the invention and the chelating agent. can decrease the risk of tumorigenesis and metastasis (e.g., by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, Senescent Cell-Specific Antigens 99% or more). 0064. The inventors have discovered that senescent cells 0062. The senescent cell targeting agents of the invention express senescent cell-specific antigens. These antigens can (e.g., peptides, polypeptides, proteins, Small molecules, anti be specifically targeted and bound by the diagnostic or thera bodies, or antibody fragments that target senescent cells) can peutic (e.g., cytotoxic) agents of the invention, as discussed be prepared by amino acid coupling using Solid phase peptide herein. The specific binding of the diagnostic or therapeutic synthesis (SPPS). As is known in the art, the amino acids to be agents of the invention only to senescent cells and not non used as Substrates to form the agents of the invention are senescent cells reduces the incidence of inaccurate diagnoses Fmoc-protected prior to incorporation into a peptide chain (e.g., using diagnostic agents) or bystander cell injury (e.g., (see, e.g., Chanand White, FMOC Solid Phase Peptide Syn resulting from cytotoxic agents). US 2015/O151 001 A1 Jun. 4, 2015

0065. Senescent cell-specific antigens that can be targeted Small Molecules of the Invention by agents of the invention include, e.g., mutant beta-actin (SEQ ID NO:11; GI: 28336) and beta-actin (ACTB) protein 0067. The invention also features small molecules that can (SEQ ID NO:12: GI: 15277503), which were identified as be used diagnostically or therapeutically based on their abil cell-surface, Senescent-specific antigens occurring in both ity to specifically bind senescent cells. For example, Small replicatively senescent and stress-induced prematurely molecules of the invention include those capable of specifi senescent cells, drug resistance-related protein LRP (SEQID cally binding to one or more of the Senescent cell-specific NO:13; GI: 1097308) and major vault protein (MVP: SEQID antigens described herein and those capable of mimicking the NO:14: GI: 19913410), which were identified as senescence physical, chemical, biological, and/or targeting characteris specific Surface proteins in the replicatively senescent cells tics of SEQ ID NOs: 1-3 and 5-8 and peptides that are sub only, and thyroid hormone binding protein precursor (SEQID stantially identical. Small molecules of the invention can be NO:15; GI: 339647), unnamed protein product (SEQ ID labeled or fused to diagnostic ortherapeutic linkers, markers, NO:20; GI: 35655), prolyl 4-hydroxylase, beta subunit pre cytotoxic agents, or other agents of the invention to aid in the cursor (P4HB; SEQ ID NO:16: GI: 20070125), chain A, diagnosis or treatment of diseases or disorders related to or human protein disulfide isomerase (PDI: SEQID NO:17: GI: caused by cellular senescence. Furthermore, Small molecules 159162689), electron-transfer-flavoprotein, beta polypeptide of the invention can specifically bind a polypeptide that has at (ETFB: SEQ ID NO:18; GI: 4503609), unnamed protein least 80% (80%, 90%, 95%, 99%, or 100%) amino acid product (SEQ ID NO:21; GI: 158257194), unnamed protein sequence identity to any one of the amino acid sequences set product (SEQID NO:22; GI: 158259937), ATP synthase, H" forth in SEQID NOS:11-23, or a fragment thereof. transporting, mitochondrial F1 complex, alpha Subunit pre Methods of Screening Small Molecules for Binding to cursor (SEQ ID NO:19; GI: 4757810), and cathepsin B Senescent Cells (CTSB; SEQ ID NO:23), which were identified on the cell Surface of stress induced prematurely senescent cells. 0068. The invention also features methods for the high 0066 Antibodies that specifically bind to the senescent throughput screening (HTS) of candidate Small molecule cell-specific antigens described herein are listed in Table 1. agents of the invention for their ability to bind senescent cells These antibodies, and any other polypeptide, antibody, anti or senescent cell-specific antigens, including, but not limited body fragment, or Small molecule that specifically binds a to, the polypeptides set forth in SEQID NOS:11-23, or frag senescent cell-specific antigen or fragment thereof, can be ments thereof. Candidate small molecules will also be incorporated in a diagnostic or therapeutic agent of the inven screened for their ability to specifically bind senescent cells. tion, as discussed herein. In addition, the antibodies listed in In general, candidate Small molecules preferably bind target Table 1 can be modified, e.g., by humanization, according to sequences with a dissociation constant less than 10 M for known methods, as is discussed below, for use in the diagno further consideration as an agent of the invention. sis and treatment of disease in a mammal, e.g., a human. 0069. Senescent cells of any origin can be used in HTS Alternatively, antibodies against the Senescent cell-specific binding assays and methods. In general, fluorescence and antigens described herein for use in the diagnosis and treat luminescence based assays (e.g., ELISA, colorimetric ment of disease in a mammal, e.g., a human, can be produced assays) are used to measure binding affinities of candidate according to methods known in the art, as is discussed below. Small molecules contacted against single or multiple target TABLE 1. Senescent Cell-Specific Antigen SEQID NO: Anti-Senescent Cell-Specific Antigen Antibodies Mutant beta-actin 1 See, e.g., Leavitt et al., Mol Cell Biol. 7: 2467-2476 (1987); Lin et al., PNAS USA 82: 6995-6999 (1985). Beta-actin (ACTB) protein 2 Clone AC-15 (mouse anti-human beta-actin antibody); Clone mAbcam 8226 (mouse anti-human beta-actin antibody). Drug resistance-related protein LRP 3 Clone 1032 (mouse anti-human LRP antibody); rabbit anti-human LRP antibody (see, e.g., Kitazono et al., J. Natl. Cancer Inst. 91: 1647-1653 (1999)). Major vault protein (MVP) 4 Clone 1014 (mouse anti-human major vault protein antibody); clone 2C431 (mouse anti-human major vault protein antibody). Thyroid hormone binding 5 See, e.g., Cheng et al., J. Biol. Chem. 262: 11221-11227 (1987). protein precursor Prolyl 4-hydroxylase, beta 6 Clone P4HB (Abcam Cat. No. ab70415: mouse anti-human prolyl subunit precursor (P4HB) 4-hydroxylase, beta Subunit precursor antibody). Chain A, human protein 7 Abcam Cat. No. ab48167 (rabbit anti-human PDI antibody); clone disulfide isomerase (PDI) 1D3 (Santa Cruz Cat. No. sc-59640; mouse anti-human PDI antibody). Electron-transfer-flavoprotein, beta 8 Abcam Cat. No. ab73986 (rabbit anti-human ETFP antibody. polypeptide (ETFP) ATP synthase, H transporting, 9 Clone 15H4 (Abcam Cat. No. ab14748; mouse anti-human ATP5A); see also, mitochondrial F1 complex, alpha subunit precursor e.g., Vantourout et al., Moi. Immunol. 45:485492 (2008). Unnamed protein product (GI: 35655) 2O See, e.g., Pihlajaniemi et al., EMBO.J. 6: 643-649 (1987). Unnamed protein product (GI: 15825.7194) 21 See, e.g., Altschul et al., Nucleic Acids Res. 25:3389-3402 (1997) Unnamed protein product (GI: 158259937) 22 See e.g., Heidebrecht et al., Mol Cancer Res. 1: 271-9 (2003) Cathepsin B (CTSB) 23 Clone ZZ12 (mouse anti-human cathepsin Bantibody); clone S 12 (goat anti-human cathepsin Bantibody). US 2015/O151 001 A1 Jun. 4, 2015

senescent cells. Upon the identification of a candidate Small conformation induced and Stabilized by binding to the target molecule from a first screening process, further scrutiny of molecule or epitope. Additional binding forces Such as ionic the binding affinity and ability of the candidate small mol bonds and Vander Waals interactions can also be considered ecule can be by means of a second, different HTS assay. This when synthesizing a small molecule of the invention. The could be accomplished, for example, by contacting the can likelihood of forming the desired conformation can be refined didate Small molecule with alternate senescent cell popula and/or optimized using molecular computational programs. tions to more precisely determine the binding affinity of the 0073 Organic compounds can be designed to be rigid, or molecule. A discussion of HTS methodologies is found in, to present hydrogen bonding groups on edge or plane, which e.g., Verkman, “Drug discovery in academia. Am. J. Physiol. can interact with complementary sites. Rebek, Science 235, Cell Physiol. 286, C465-C474 (2004) and Dove, “Screening 1478-1484 (1987) and Rebek et al., J. Am. Chem. Soc. 109, for content—the evolution of high throughput.” Nat. Biotech 2426-2431 (1987) have summarized these approaches and mol. 21:859-864 (2003). Examples of HTS screening methods the mechanisms involved in binding of compounds to regions for the discovery of useful Small molecule agents are found in, of proteins. e.g., U.S. Pat. Nos. 7.279,286 and 7.276,346, which are incor 0074 Synthetic methods can be used by one skilled in the porated by reference herein. art to make Small molecules that interact with functional 0070 Candidate small molecules that have undergone groups of proteins, glycoproteins, or epitopes present on the HTS screening may be further modified to empirically exterior or in the interior of senescent cells. improve senescent cell binding affinities according to the design considerations discussed below. Preparation of Antibody Compositions of the Invention 0075. The invention also provides antibody compositions Small Molecule Design that include one or more of the sequences set forth in SEQID 0071 Small molecules of the invention can also be gener NOS:1-3 and 5-8, and peptides substantially identical thereto, ated according to the principles of rational design. Computer which can be included in, e.g., one or more of the comple modeling technology allows visualization of the three-di mentarity determining regions (CDRS) of the antibody com mensional atomic structure of a selected molecule and the positions. The invention further provides antibody composi design of new compounds that will interact with senescent tions that specifically bind a senescent cell-specific antigen cells or senescent cell-specific antigens (e.g., the senescent having at least 80% (e.g., 80%, 90%, 95%, 99%, or 100%) cell-specific antigens discussed above (set forth in SEQ ID amino acid sequence identity to any one of the Senescent NOS:11-23), or fragments thereof). The three-dimensional cell-specific antigens set forth in SEQ ID NOS:11-23, or a construct typically depends on data from X-ray crystallo fragment thereof. graphic analyses or NMR imaging of the selected molecule or 0076. The antibodies of the invention may be recombinant epitope. A computer graphics system enables prediction of (e.g., chimeric or humanized), synthetic, or naturally occur how a candidate small molecule compound will bind to the ring. Antibodies orantibody fragments of the invention can be target senescent cell and allows experimental manipulation of recombinantly produced, or identified and isolated from a the structures of the Small molecule and target protein to mammal that has been induced to produce or that naturally perfect binding specificity. A prediction of what the mol produces antibodies that specifically bind to senescent cell ecule-protein interaction will be when Small changes are specific antigens. For example, a polypeptide having at least made in one or both can be determined by using molecular 80% (e.g., 85%, 90%, 95%, 99%, or 100%) amino acid mechanics Software and computationally intensive comput sequence identity to one of the Senescent cell-specific anti ers. An example of a molecular modeling system described gens set forth in SEQ ID NOS:11-23 can be administered generally above includes the CHARMm and QUANTA pro (e.g., by injection) to a mammal (e.g., a mouse, rat, hamster, grams (Polygen Corporation, Waltham, Mass.). CHARMm guinea pig, sheep, goat, cow, horse, non-human primate, or performs the energy minimization and molecular dynamics human) to provoke a humoral immune response against the functions, while QUANTA performs the construction, immunizing senescent cell-specific antigen. Blood, plasma, graphic modeling and analysis of molecular structure. or ascites from the immunized mammal can be screened to QUANTA allows interactive construction, modification, identify or harvestantibodies that specifically bind the immu visualization, and analysis of the behavior of molecules that nizing Senenescent cell-specific antigen. If desired, the amino intact with each other. Another molecular modeling program acid and/or nucleic acid sequences of the antibodies produced that can be used to identify small molecules for use in the can be determined. Furthermore, all or a portion of the nucleic methods of the invention is DOCK (Kuntz Laboratory, acid sequences of the antibodies can be incorporated into a UCSF). vector (e.g., an expression vector or a viral vector) for expres sion of the antibody in a cellofa Subject (e.g., a mammal. Such Small Molecule Synthesis as a human) following administration of the vector to the Subject, or any other uses consistent with the methods 0072 Small molecules of the invention can be organic or described herein. inorganic compounds, and even nucleic acids. Specific bind 0077. The invention features complete antibodies, diabod ing to senescent cells can be achieved by including chemical ies, bi-specific antibodies, antibody fragments, Fab frag groups having the correct spatial location and charge in the ments, F(ab')2 molecules, single chain Fv (scFv) molecules, Small molecule. In a preferred embodiment, compounds are tandem Sclv molecules, or antibody fusion proteins. Anti designed with hydrogen bond donor and acceptor sites bodies of the invention include, e.g., the IgG, IgA, IgM, Ig). arranged to be complementary to the targeted molecule or and IgE isotypes. Antibodies of the invention contain one or epitope. An agent is formed with chemical side groups more CDRS or binding peptides that bind to proteins, glyco ordered to yield the correct spatial arrangement of hydrogen proteins, or epitopes present on the exterior or in the interior bond acceptors and donors when the agent is in a specific of senescent cells. US 2015/O151 001 A1 Jun. 4, 2015

0078. Many of the antibodies, or fragments thereof, an injected antibody should be less than against a totally described herein can undergo non-critical amino-acid Substi foreign mouse antibody or a partially foreign chimeric anti tutions, additions or deletions in both the variable and con body; stant regions without loss of binding specificity or effector I0083. 2) Because the effector portion of the humanized functions, or intolerable reduction of binding affinity (i.e., antibody is human, it may interact better with other parts of below about 107 M). Usually, an antibody incorporating the human immune system; and Such alterations exhibits Substantial sequence identity to a I0084 3) Injected mouse antibodies have been reported to reference antibody from which it is derived. Occasionally, a have a much shorter half-life in the human circulation than the mutated antibody can be selected having the same specificity half-life exhibited by normal human antibodies (see, e.g., and increased affinity compared with a reference antibody Shaw et al., J. Immunol. 138:4534-4538 (1987)). Injected from which it was derived. Phage-display technology offers humanized antibodies have a half-life essentially equivalent powerful techniques for selecting Such antibodies. See, e.g., to naturally occurring human antibodies, allowing Smaller Dower et al. (WO 91/17271), McCafferty et al. (WO and less frequent doses. 92/01047), and Huse (WO92/06204), each of which is incor I0085. The substitution of one or more, e.g., mouse CDRs porated by reference herein. into a human variable domain framework is most likely to result in retention of their correct spatial orientation if the Antibody Fragments human variable domainframework adopts the same or similar conformation to the mouse variable framework from which 0079. In another embodiment of the invention, an agent of the CDRs originated. This is achieved by obtaining the human the invention is a fragment of an intact antibody described variable domains from human antibodies whose framework herein. Antibody fragments include separate variable heavy sequences exhibit a high degree of sequence identity with the chains, variable light chains, Fab, Fab'. F(ab'), Fabc, and Fv. murine variable framework domains from which the CDRs Fragments can be produced by enzymatic or chemical sepa were derived. The heavy and light chain variable framework ration of intact immunoglobulins. For example, a F(ab') frag regions can be derived from the same or different human ment can be obtained from an IgG molecule by proteolytic antibody sequences. The human antibody sequences can be digestion with pepsin at pH 3.0-3.5 using standard methods the sequences of naturally occurring human antibodies or can such as those described in Harlow and Lane, Antibodies: A be consensus sequences of several human antibodies. See, Laboratory Manual, Cold Spring Harbor Pubs., N.Y. (1988). e.g., Kettleborough et al., Protein Engineering 4:773 (1991); Fab fragments may be obtained from F(ab')2 fragments by Kolbinger et al., Protein Engineering 6:971 (1993). limited reduction, or from whole antibody by digestion with I0086) Suitable human antibody sequences are identified papain in the presence of reducing agents. Fragments can also by computer comparisons of the amino acid sequences of the be produced by recombinant DNA techniques. Segments of mouse variable regions with the sequences of known human nucleic acids encoding selected fragments are produced by antibodies. The comparison is performed separately for digestion of full-length coding sequences with restriction heavy and light chains but the principles are similar for each. enzymes, or by de novo synthesis. Often fragments are I0087 Methods of preparing chimeric and humanized anti expressed in the form of phage-coat fusion proteins. This bodies and antibody fragments are described in U.S. Pat. Nos. manner of expression is advantageous for affinity-sharpening 4,816,567, 5,530,101, 5,622,701, 5,800,815, 5,874,540, of antibodies. 5,914,110, 5,928,904, 6,210,670, 6,677,436, and 7,067,313 and U.S. Patent Application Nos. 2002/0031508, 2004/ 0265311, and 2005/0226876. Preparation of antibody or frag Humanized Antibodies ments thereof is further described in U.S. Pat. Nos. 6,331,415, 0080. The invention also includes humanized antibodies 6,818.216, and 7,067,313. Each of these patents is incorpo in which one or more of the CDRs are derived from a non rated herein by reference. human antibody sequence (e.g., those antibodies described in Diagnostic Agents Coupled to Agents of the Invention Table 1), and one or more, but preferably all, of the CDRs bind specifically to a protein, glycoprotein, or epitope present on I0088 Agents of the invention (e.g., peptides, polypep the exterior or in the interior of a senescent cell (e.g., the tides, proteins, Small molecules, antibodies, or antibody frag senescent cell-specific antigens described above (SEQ ID ments that target senescent cells) can be joined or coupled to NOS:11-23), or a fragment thereof). any molecule that can be used to label, detect, identify, screen, or diagnose senescent cells in a mammal (e.g., a human). 0081. A humanized antibody contains constant frame Diagnostic and therapeutic (e.g., cytotoxic) agents of the work regions derived Substantially from a human antibody invention that include a detectable label can be administered (termed an acceptor antibody), as well as, in some instances, in a pharmaceutical compound to a Subject (e.g., a cancer a majority of the variable region derived from a human anti patient) or these compositions can be encoded by a vector body. One or more of the CDRs (all or a portion thereof, as (e.g., a viral vector) and expressed in a cell of the Subject well as discreet amino acids Surrounding one or more of the following administration. CDRS) are provided from a non-human antibody, such as a I0089. An agent of the invention that incorporates a detect mouse antibody. The constant region(s) of the antibody, may able label can be used, for example, to type, grade, and track or may not be present. Humanized antibodies provide several the metastasis of a tumor in a Subject having cancer. The advantages over non-humanized antibodies for therapeutic or diagnostic agent can also include a therapeutic agent (e.g., diagnostic use in humans. These include: cytotoxic) of the invention. In this case, the diagnostic agent 0082) 1) The human immune system should not recognize facilitates the detection or measurement, or binding to senes the framework or constant region of the humanized antibody cent cells, of the therapeutic agent after administration to a as foreign, and therefore the antibody response against Such Subject. US 2015/O151 001 A1 Jun. 4, 2015

0090. A diagnostic or therapeutic agent of the invention 0095 Epitope tags are short amino acid sequences, e.g., that includes a detectable label can be produced recombi 5-20 amino acid residues in length, that can be incorporated nantly in vitro. In one embodiment of the invention, the into an agent of the invention as a detectable label to allow for detectable label is capable of detection based on an intrinsic detection once expressed in a cell, secreted from the cell, or property, e.g., fluorescence and bioluminescence, or based on bound to a target cell (e.g., a senescent cell). An agent of the its ability to bind with another molecule capable of being invention that incorporates an epitope tag as a diagnostic detected (e.g., an epitope tag. Such as c-myc, hemagglutinin, agent can be detected by virtue of its interaction with an and histamine tags). Peptides, polypeptides, or proteins with antibody, antibody fragment, or other binding molecule spe diagnostic properties can be altered (e.g., by making amino cific for the epitope tag. Nucleotide sequences encoding the acid substitutions, mutations, truncations, or additions) to epitope tag are produced either by cloning appropriate por facilitate incorporation into an agent of the invention. Desir tions of natural genes or by Synthesizing a polynucleotide that able alterations include, for example, changes to the amino encodes the epitope tag. An antibody, antibody fragment, or acid sequence that facilitate protein expression, longevity, other binding molecule that binds an epitope tag can directly cell secretion, and detectability. incorporate its own detectable label (e.g., a fluorochrome, 0091. The invention also features a nucleic acid molecule radiolabel, heavy metal, or enzyme Such as horseradish per encoding a peptidic detectable label as a fusion protein with oxidase) or serve as a target for a secondary antibody, anti an agent of the invention; the nucleic acid molecule can be body fragment, or other binding molecule that incorporates incorporated into a vector (e.g., an expression vector). Such Such a label. Exemplary epitope tags that can be used as a that, upon expression of the agent of the invention in a cell detectable label include c-myc (SEQID NO: 24), hemagglu transfected or transduced with the vector, expression of the tinin (HA; SEQID NO:25), and histidine tag (His: SEQID detectable label and agent of the invention are operably linked NO: 26). Furthermore, fluorescent (e.g., GFP) and biolumi (e.g., fused, contiguously-joined, or tethered together). nescent molecules, discussed above, can also serve as epitope tags, as antibodies, antibody fragments, and other binding 0092 Molecules that can be used as the detectable label as molecules are commercially available for the detection of discussed herein include, but are not limited to, radioactive these moieties. agents, fluorescent agents, bioluminescent molecules, 0096 Agents of the invention (e.g., peptides, polypep epitope tags, and heavy metals, each of which is discussed in tides, proteins, Small molecules, antibodies, or antibody frag detail below. ments that target senescent cells) can also be coupled to a 0093. Fluorescent agents include fluorochromes, such as chelating agent to form diagnostic agents of the invention. fluorescein isothiocyanate (FITC), allophycocyanin (APC), Diagnostic agents can be prepared by various methods phycoerythrin (PE), rhodamine, and tetramethylrhodamine depending upon the chelator chosen. A portion of an agent of isothiocyanate (TRITC). Other fluorescent molecules include the invention (e.g., a portion of a peptide, polypeptide, pro green fluorescent protein (GFP; SEQID NO: 27), enhanced tein, Small molecule, antibody, or antibody fragment) can be GFP (eGFP), yellow fluorescent protein (SEQ ID NO: 28: isolated from a natural Source, recombinantly produced, or YFP), cyan fluorescent protein (SEQ ID NO: 29; CFP), and synthesized. The portion of an agent of the invention is most red fluorescent protein (SEQ ID NO:30; RFP or DsRed). conveniently prepared by techniques generally established in Each of these fluorescent molecules can be used as detectable the art of peptide synthesis, Such as the Solid-phase approach. label in a diagnostic agent of the invention. Peptidic fluores Solid-phase synthesis involves the stepwise addition of cent molecules can be recombinantly expressed in a cell (e.g., amino acid residues to a growing peptide chain that is linked a blood cell. Such as a lymphocyte) following transfection or to an insoluble Support or matrix. Such as polystyrene. The transduction of the cell with an expression vector that encodes C-terminus residue of the peptide is first anchored to a com the nucleotide sequence of the fluorescent molecule. Upon mercially available Support with its amino group protected exposure of the fluorescent molecule to a stimulating fre with an N-protecting agent Such as a t-butyloxycarbonyl quency of light, the fluorescent molecule will emit light at a group (tBoc) or a fluorenylmethoxycarbonyl (FMOC) group. low, medium, or high intensity that can be observed by eye The amino-protecting group is removed with Suitable depro under a microscope or by an optical imaging device. Exem tecting agents such as TFA in the case of tBOC or piperidine plary fluorescent molecules suitable for use as the detectable for FMOC and the next amino acid residue (in N-protected label in a diagnostic agent of the invention are described in, form) is added with a coupling agent such as dicyclocarbo e.g., U.S. Pat. Nos. 7,417,131 and 7,413,874. diimide (DCC). Upon formation of a peptide bond, the 0094 Bioluminescent molecules can also be used as a reagents are washed from the support. After addition of the detectable label incorporated into a diagnostic agent of the final residue, the agent is cleaved from the Support with a invention. Bioluminescent molecules. Such as luciferase suitable reagent, such as trifluoroacetic acid (TFA) or hydro (e.g., firefly (SEQID NO:31), Renilla (SEQID NO:32), and gen fluoride (HF). Omphalotus luciferase) and aequorin, emit light as part of a 0097 Agents of the invention (e.g., peptides, polypep chemical reaction with a Substrate (e.g., luciferin and tides, proteins, Small molecules, antibodies, or antibody frag coelenterazine). In one embodiment of the invention, a vector ments that target senescent cells) and chelator components encoding a luciferase gene provides for the in Vivo, in vitro, or can be coupled to form a conjugate by reacting the free amino ex vivo detection of cells (e.g., blood cells, such as lympho group of the threonine residue of a peptide portion of an agent cytes) that have been transduced or transfected with an agent of the invention with an appropriate functional group of the of the invention. Exemplary bioluminescent molecules suit chelator, such as a carboxyl group or activated ester. For able for use as the detectable label in a diagnostic agent of the example, a conjugate may incorporate the chelator ethylene invention, and methods for their use are described in, e.g., diaminetetraacetic acid (EDTA), common in the art of coor U.S. Pat. Nos. 5,292,658, 5,670,356, 6,171,809, and 7,183, dination chemistry, when functionalized with a carboxyl sub O92. stituent on the ethylene chain. Synthesis of EDTA derivatives US 2015/O151 001 A1 Jun. 4, 2015 of this type are reported in Arya et al., Bioconjugate Chem and rhenium in their various forms (e.g., "TcO", istry, 2:323, 1991), wherein the four coordinating carboxyl mTcO", ReO", and ReO"). Incorporation of the metal groups are each blocked with a t-butyl group while the car within the conjugate can be achieved by various methods boxyl substituent on the ethylene chain is free to react with the common in the art of coordination chemistry. When the metal amino group of a peptide portion of the agent of the invention, is technetium-99m, the following general procedure may be thereby forming a conjugate. used to form a technetium complex. An agent-chelator con 0098. A conjugate may incorporate a metal chelator com jugate solution is formed initially by dissolving the conjugate ponent that is peptidic, i.e., compatible with Solid-phase pep in aqueous alcohol Such as ethanol. The Solution is then tide synthesis. In this case, the chelator may be coupled to the degassed to remove oxygen then thiol-protecting groups are agent of the invention (e.g., peptides, polypeptides, proteins, removed with a Suitable reagent, for example, with sodium Small molecules, antibodies, orantibody fragments that target hydroxide, and then neutralized with an organic acid, Such as senescent cells) in the same manner as EDTA described above acetic acid (pH 6.0–6.5). In the labeling step, a stoichiometric or more conveniently the chelator and agent of the invention excess of Sodium pertechnetate, obtained from a molybde are synthesized intoto starting from the C-terminal residue of num generator, is added to a solution of the conjugate with an the agent and ending with the N-terminal residue of the chela amount of a reducing agent Such as Stannous chloride suffi tOr cient to reduce technetium and heated. The labeled conjugate 0099 Conjugates may further incorporate a linking group may be separated from contaminants' "TcO, and colloidal component that serves to couple the agent of the invention "TcO. chromatographically, for example, with a C-18 Sep (e.g., peptides, polypeptides, proteins, Small molecules, anti Pak cartridge. bodies, or antibody fragments that target senescent cells) to 0103) In an alternative method, labeling of the agent of the the chelator while not adversely affecting either the targeting invention can be accomplished by a transchelation reaction. function of the agent or the metal binding function of the The technetium source is a solution oftechnetium complexed chelator. Suitable linking groups include amino acid chains with labile ligands facilitating ligand exchange with the and alkyl chains functionalized with reactive groups for cou selected chelator. Suitable ligands for transchelation include pling to both the agent and the chelator. An amino acid chain tartarate, citrate, and heptagluconate. In this instance the pre is the preferred linking group when the chelator is peptidic so ferred reducing reagent is sodium dithionite. It will be appre that the conjugate can be synthesized in toto by Solid-phase ciated that the conjugate may be labeled using the techniques techniques. described above, or alternatively the chelator itself may be 0100. An alkyl chain linking group may be incorporated in labeled and subsequently coupled to the agent of the inven the conjugate by reacting the amino group of the threonine tion to form the conjugate; a process referred to as the “prela residue of a peptide portion of an agent of the invention (e.g., beled ligand method. peptides, polypeptides, proteins, Small molecules, antibodies, 0104. Another approach for labeling agents of the inven or antibody fragments that target senescent cells) with a first tion (e.g., peptides, polypeptides, proteins, Small molecules, functional group on the alkyl chain, such as a carboxyl group antibodies, or antibody fragments that target senescent cells) or an activated ester. Subsequently the chelator is attached to involves immobilizing the agent-chelator conjugate on a the alkyl chain to complete the formation of the conjugate by Solid-phase Support through a linkage that is cleaved upon reacting a second functional group on the alkyl chain with an metal chelation. This is achieved when the chelator is coupled appropriate group on the chelator. The second functional to a functional group of the Support by one of the complexing group on the alkyl chain is selected from Substituents that are atoms. Preferably, a complexing Sulfur atom is coupled to the reactive with a functional group on the chelator while not Support which is functionalized with a Sulfur protecting group being reactive with the threonine residue of the agent. For Such as maleimide. example, when the chelator incorporates a functional group 0105. When labeled with a diagnostically useful metal, Such as a carboxyl group or an activated ester, the second agent-chelator conjugates of the invention can be used to functional group of the alkyl chain linking group can be an detect tissue at risk of developing cancer (e.g., lung cancer, amino group. It will be appreciated that formation of the breast cancer, colon cancer, and prostate cancer), age-related conjugate may require protection and deprotection of the diseases (e.g., cardiovascular disease, cerebrovascular dis functional groups present in order to avoid formation of ease, or Alzheimer's disease), tobacco-related diseases (e.g., undesired products. Protection and deprotection are accom emphysema, aortic aneurysms, esophageal cancer, or squa plished using protecting groups, reagents, and protocols com mous cell cancer of the head and neck), or other diseases and mon in the art of organic synthesis. Particularly, protection disorders relating to or caused by cellular senescence by and deprotection techniques employed in Solid phase peptide procedures established in the art of diagnostic imaging. An synthesis described above may be used. agent labeled with a radionuclide metal. Such as technetium 0101. An alternative chemical linking group to an alkyl 99 m, may be administered to a mammal (e.g., a human) by chain is polyethylene glycol (PEG), which is functionalized intravenous injection in a pharmaceutically acceptable solu in the same manner as the alkyl chain described above for tion Such as isotonic saline, or by other methods described incorporation in the conjugates. It will be appreciated that herein. The amount of a labeled agent of the invention (e.g., linking groups may alternatively be coupled first to the chela peptides, polypeptides, proteins, Small molecules, antibodies, tor and then to the agent of the invention (e.g., peptides, orantibody fragments that target senescent cells) appropriate polypeptides, proteins, Small molecules, antibodies, or anti for administration is dependent upon the distribution profile body fragments that target senescent cells). of the chosen conjugate in the sense that a rapidly cleared 0102. In accordance with one aspect of the invention, agent may be administered at higher doses than one that clears agent-chelator conjugates of the invention incorporate a diag less rapidly. Unit doses acceptable for imaging tissues that nostically useful metal capable of forming a complex. Suit contain senescent cells are in the range of, e.g., 5-40 mCi for able metals include, e.g., radionuclides, such as technetium a 70kg individual. In vivo distribution and localization can be US 2015/O151 001 A1 Jun. 4, 2015 tracked by standard techniques described herein at an appro Taxoid: Tecogalan Sodium; Tegafur, Teloxantrone Hydro priate time Subsequent to administration; typically between chloride; Temoporfin; Teniposide; Teroxirone; Testolactone: 30 minutes and 180 minutes depending upon the rate of Thiamiprine; Thioguanine: Thiotepa; Thymitaq, Tlazofurin; accumulation at the target site with respect to the rate of Tirapazamine; Tomudex; TOP53: Topotecan Hydrochloride: clearance at non-target tissue. Toremifene Citrate; Trestolone Acetate; Triciribine Phos phate: Trimetrexate; Trimetrexate Glucuronate; Triptorelin; Therapeutic or Cytotoxic Agents Coupled to Agents of the Tubulozole Hydrochloride; Uracil Mustard; Uredepa; Vap Invention reotide: Verteporfin; Vinblastine; Vinblastine Sulfate; Vinc ristine; Vincristine Sulfate; Vindesine; Vindesine Sulfate; 010.6 Any of the agents of the invention (e.g., peptides, Vinepidine Sulfate; Vinglycinate Sulfate; Vinleurosine Sul polypeptides, proteins, Small molecules, antibodies, or anti fate; Vinorelbine Tartrate; Vinrosidine Sulfate; Vinzolidine body fragments that target senescent cells) can be coupled to Sulfate; Vorozole; Zeniplatin: Zinostatin: Zorubicin Hydro any known cytotoxic or therapeutic moiety. Examples chloride: 2-Chlorodeoxyadenosine; 2 Deoxyformycin; include, e.g., antineoplastic agents such as: Acivicin; Aclaru 9-aminocamptothecin; raltitrexed: N-propargyl-5,8-didea bicin; Acodazole Hydrochloride: Acronine; Adozelesin; Zafolic acid; 2-chloro-2'-arabino-fluoro-2'-deoxyadenosine: Adriamycin; Aldesleukin; Altretamine; Ambomycin; A. met 2-chloro-2'-deoxyadenosine; anisomycin; trichostatin A; antrone Acetate; Aminoglutethimide; Amsacrine; Anastro hPRL-G129R; CEP-751; linomide; sulfur mustard; nitrogen Zole; Anthramycin; Asparaginase; Asperlin; AZacitidine; mustard (mechlorethamine); cyclophosphamide; melphalan; AZetepa, Azotomycin; Batimastat; BenZOdepa; Bicaluta chlorambucil; ifosfamide; busulfan; N-methyl-Nnitrosourea mide: Bisantrene Hydrochloride; Bisnafide Dimesylate: (MNU): N,N'-Bis(2-chloroethyl)-N-nitrosourea (BCNU): Bizelesin; Bleomycin Sulfate; Brequinar Sodium; Bropir N-(2-chloroethyl)-N' cyclohexyl-N-nitrosourea (CCNU); imine; BuSulfan; Cactinomycin; Calusterone; Camptothecin, N-(2-chloroethyl)-N'-(trans-4-methylcyclohexyl-N-ni Caracemide: Carbetimer; Carboplatin: Carmustine; Carubi cin Hydrochloride; Carzelesin; Cedefingol: Chlorambucil; trosourea (MeCCNU); N-(2-chloroethyl)-N'-(diethyl) eth Cirolemycin; Cisplatin: Cladribine; Combretestatin A-4: Cri ylphosphonate-N-nitrosourea (fotemustine); streptozotocin, snatol Mesylate; Cyclophosphamide: Cytarabine; Dacarba diacarbazine (DTIC); mitozolomide; temozolomide: zine; DACA (N-2-(Dimethyl-amino) ethylacridine-4-car thiotepa; mitomycin C: AZQ; adozelesin: Cisplatin: Carbo boxamide); Dactinomycin; Daunorubicin Hydrochloride; platin: Ormaplatin: Oxaliplatin; C1-973: DWA 2114R: Daunomycin; Decitabine; Dexormaplatin, DeZaguanine; JM216:JM335; Bis(platinum); tomudex; azacitidine: cytara Dezaguanine Mesylate; Diaziquone; Docetaxel; Dolasatins: bine; gemcitabine, 6-Mercaptopurine; 6-Thioguanine; Doxorubicin; Doxorubicin Hydrochloride; Droloxifene: Hypoxanthine; teniposide 9-amino camptothecin; Topote Droloxifene Citrate; Dromostanolone Propionate; Duazomy can; CPT-11; Doxorubicin; Daunomycin; Epirubicin; daru cin; Edatrexate. Eflornithine Hydrochloride; Ellipticine; bicin; mitoxantrone; losoxantrone; Dactinomycin (Actino Elsamitrucin; Enloplatin: Enpromate: Epipropidine: Epirubi mycin D); amsacrine; pyrazoloacridine; all-trans retinol; cin Hydrochloride; Erbulozole; Esorubicin Hydrochloride: 14-hydroxy-retro-retinol; all-trans retinoic acid; N-(4-Hy Estramustine; Estramustine Phosphate Sodium; Etanidazole; droxyphenyl) retinamide: 13-cis retinoic acid; 3-Methyl Ethiodized Oil I 131; Etoposide; Etoposide Phosphate; Eto TTNEB; 9-cis retinoic acid; fludarabine (2-F-ara-AMP); or prine: Fadrozole Hydrochloride; Fazarabine: Fenretinide; 2-chlorodeoxyadenosine (2-Cda). Floxuridine: Fludarabine Phosphate: Fluorouracil; 0107. Other therapeutic compounds include, but are not 5-FdUMP. Fluorocitabine; Fosquidone; Fostriecin Sodium; limited to, 20-pi-1.25 dihydroxyvitamin D3; 5-ethynyluracil; Gemcitabine; Gemcitabine Hydrochloride; Gold Au 198: abiraterone; aclarubicin; acylfulvene; adecypenol; adoZe Homocamptothecin: Hydroxyurea; Idarubicin Hydrochlo lesin; aldesleukin; ALL-TKantagonists; altretamine; amba ride; Ifosfamide; Ilmofosine; Interferon Alfa-2a: Interferon mustine; amidox; amifostine; aminolevulinic acid; amrubi Alfa-2b: Interferon Alfa-nl, Interferon Alfa-n3; Interferon cin; amsacrine; anagrelide; anastrozole; andrographolide; Beta-I a: Interferon Gamma-I b; I proplatin; Irinotecan angiogenesis inhibitors; antagonist D; antagonist G, antar Hydrochloride; Lanreotide Acetate; Letrozole; Leuprolide elix; anti-dorsalizing morphogenetic protein-1, antiandro Acetate; Liarozole Hydrochloride; Lometrexol Sodium; gen, prostatic carcinoma, antiestrogen; antineoplaston; anti Lomustine; Losoxantrone Hydrochloride; Masoprocol; May sense oligonucleotides; aphidicolinglycinate; apoptosis gene tansine; Mechlorethamine Hydrochloride; Megestrol modulators; apoptosis regulators; apurinic acid; ara-CDP Acetate; Melengestrol Acetate; Melphalan: Menogaril; Mer DL-PTBA, argininedeaminase; asulacrine; atamestane; atri captopurine; Methotrexate; Methotrexate Sodium; Meto mustine; axinastatin 1; axinastatin 2; axinastatin3; aZasetron; prine; Meturedepa; Mitindomide; Mitocarcin; Mitocromin: aZatoxin; azatyrosine; baccatin III derivatives; balanol; bati Mitogillin; Mitomalcin; Mitomycin; Mitosper; Mitotane: mastat; BCR/ABL antagonists; benzochlorins; benzoylstau Mitoxantrone Hydrochloride: Mycophenolic Acid; Nocoda rosporine; beta lactam derivatives; beta-alethine; betaclamy Zole; Nogalamycin; Ormaplatin: Oxisuran: Paclitaxel; Pegas cin B; betulinic acid; bFGF inhibitor; bicalutamide: pargase; Peliomycin; Pentamustine: PeploycinSulfate; Per bisantrene; bisaziridinylspermine; bisnafide; bistratene A: fosfamide; Pipobroman; Piposulfan; Piroxantrone bizelesin; breflate: bleomycin A2; bleomycin B2; bropir Hydrochloride; Plicamycin; Plomestane: Porfimer Sodium; imine; budotitane; buthionine Sulfoximine; calcipotriol: Porfiromycin; Prednimustine: Procarbazine Hydrochloride: calphostin C; camptothecin derivatives (e.g., 10-hydroxy Puromycin; Puromycin Hydrochloride; Pyrazofurin; camptothecin); canarypox IL-2; capecitabine; carboxamide Rhizoxin; Rhizoxin D; Riboprine; Rogletimide: Safingol: amino-triazole; carboxyamidotriazole; CaRest M3; CARN Safingol Hydrochloride: Semustine; SimtraZene; Sparfosate 700; cartilage derived inhibitor; carzelesin: casein kinase Sodium; Sparsomycin; Spirogermanium Hydrochloride; inhibitors (ICOS); castanospermine; cecropin B; cetrorelix: Spiromustine; Spiroplatin; Streptonigrin: Streptozocin, chlorins; chloroquinoxaline Sulfonamide; cicaprost; cis-por Strontium Chloride Sr 89: Sulofenur; Talisomycin; Taxane: phyrin, cladribine; clomifene analogues; clotrimazole; collis US 2015/O151 001 A1 Jun. 4, 2015 mycin A; collismycin B; combretastatin A4, combretastatin piritrexim; placetin A; placetin B; plasminogen activator analogue; conagenin, crambescidin 816; crisinatol; crypto inhibitor; platinum complex, platinum compounds; plati phycin 8; cryptophycin A derivatives; curacin A; cyclopen num-triamine complex; podophyllotoxin; porfimer Sodium; tanthraquinones; cycloplatam, cypemycin; cytarabine ocfos porfiromycin; propyl bis-acridone; prostaglandin J2, protea fate; cytolytic factor; cytostatin; dacliximab, decitabine; Some inhibitors; protein A-based immune modulator, protein dehydrodidemnin B: 2 deoxycoformycin (DCF); deslorelin; kinase C inhibitor, protein kinase C inhibitors, microalgal; dexifosfamide; dexraZoxane; dexVerapamil; diaziquone; protein tyrosine phosphatase inhibitors; purine nucleoside didemnin B; didox; diethylnorspermine; dihydro-5-azacyti phosphorylase inhibitors; purpurins; pyrazoloacridine; pyri dine; dihydrotaxol. 9-; dioxamycin; diphenyl spiromustine; doxylated hemoglobin polyoxyethylene conjugate; raf discodermolide; docosanol; dolasetron: doxifluridine; antagonists; raltitrexed; ramosetron: ras farnesyl protein droloxifene; dronabinol; duocarmycin SA; ebselen; ecomus transferase inhibitors; ras inhibitors; ras-GAP inhibitor; retel tine; edelfosine; edrecolomab; eflornithine; elemene; emite liptine demethylated; rhenium Re 186 etidronate; rhizoxin; fur; epirubicin; epothilones (A, R=H; B, R=Me); epith ribozymes: RII retinamide; rogletimide; rohitukine; ilones; epristeride; estramustine analogue; estrogen agonists; romurtide; roquinimex: rubiginone Bl; ruboxyl; Safingol; estrogen antagonists; etanidazole; etoposide: etoposide saintopin; SarCNU; sarcophytol A, sargramostim; Sdi 1 4'-phosphate (etopofos); exemestane; fadrozole; faZarabine; mimetics; semustine; senescence derived inhibitor 1; sense fenretinide; filgrastim; finasteride; flavopiridol; flezelastine: oligonucleotides; signal transduction inhibitors; signal trans fluasterone; fludarabine; fluorodaunorunicin hydrochloride: duction modulators; single chain antigen binding protein; forfenimex; formestane; fostriecin: fotemustine; gadolinium sizofuran; Sobuzoxane; sodium borocaptate; sodium pheny texaphyrin; gallium nitrate; galocitabine; ganirelix; gelati lacetate; Solverol; somatomedin binding protein; Sonermin; nase inhibitors; gemcitabine; glutathione inhibitors; hepsul sparfosic acid; spicamycin D; spiromustine; splenopentin; fam; heregulin; hexamethylene bisacetamide; homohar spongistatin 1; squalamine; stem cell inhibitor, stem-cell ringtonine (HHT); hypericin; ibandronic acid; idarubicin; division inhibitors; stipiamide; stromelysin inhibitors; sulfi idoxifene; idramantone, ilmofosine; illomastat; imidazoacri nosine; Superactive vasoactive intestinal peptide antagonist; dones; imiquimod; immunostimulant peptides; insulin-like Suradista; Suramin; Swainsonine; synthetic glycosaminogly growth factor-1 receptor inhibitor; interferonagonists; inter cans; tallimustine; tamoxifen methiodide; tauromustine; taZ ferons; interleukins; iobenguane; iododoxorubicin; arotene; tecogalan Sodium, tegafur, tellurapyrylium; telom ipomeanol, 4-, irinotecan; iroplact; irsogladine; isobenga erase inhibitors; temoporfin, temozolomide; teniposide; Zole: isohomohalicondrin B; itasetron; jasplakinolide: tetrachlorodecaoxide; tetrazomine; thaliblastine; thalido kahalalide F: lamellarin-N triacetate; lanreotide; leinamycin; mide; thiocoraline; thrombopoietin; thrombopoietin lenograstim; lentinan Sulfate; leptolstatin; letrozole; leuke mimetic; thymalfasin; thymopoietin receptor agonist; thy mia inhibiting factor, leukocyte alpha interferon; leuprolide-- motrinan; thyroid stimulating hormone; tin ethyl etiopurpu estrogen-progesterone; leuprorelin; levamisole; liarozole; rin; tirapazamine; titanocene dichloride; topotecan; topsen linear polyamine analogue; lipophilic disaccharide peptide; tin, toremifene; totipotent stem cell factor, translation lipophilic platinum compounds; lissoclinamide 7: lobaplatin: inhibitors; tretinoin; triacetyluridine; triciribine; trimetrex lombricine; lometrexol; lonidamine; losoxantrone; lovasta ate; triptorelin; tropisetron; turosteride; tyrosine kinase tin; loxoribine; lurtotecan; lutetium texaphyrin; lysofylline; inhibitors; tyrphostins; UBC inhibitors: ubenimex: urogeni lytic peptides; maytansine; mannostatin A; marimastat; tal sinus-derived growth inhibitory factor; urokinase receptor masoprocol; maspin; matrilysin inhibitors; matrix metallo antagonists; vapreotide; variolin B; vector system, erythro proteinase inhibitors; menogaril; merbarone; meterelin; cyte gene therapy; velaresol; Veramine; Verdins; verteporfin; methioninase; metoclopramide: MIF inhibitor; ifepristone; Vinorelbine; Vinxaltine; vitaxin; Vorozole; Zanoterone; Zeni miltefosine; mirimostim; mismatched double stranded RNA; platin: Zilascorb; and Zinostatin stimalamer. mithracin, mitoguaZone; mitolactol; mitomycin analogues; 0108. One or more of the agents of the invention (e.g., mitonafide; mitotoxin fibroblast growth factor-saporin; peptides, polypeptides, proteins, Small molecules, antibodies, mitoxantrone; mofarotene; molgramoStim; monoclonal anti or antibody fragments that target senescent cells) can also be body, human chorionic gonadotrophin; monophosphoryl coupled to a lytic peptide. Such lytic peptides induce cell lipid A+myobacterium cell wall sk; mopidamol; multiple death and include, but are not limited to, streptolysin O: drug resistance gene inhibitor, multiple tumor suppressor Stoichactis toxin; phallolysin; Staphylococcus alpha toxin; 1-based therapy; mustard anticancer agent; mycaperoxide B; holothurin A; digitonin; melittin; lysolecithin; cardiotoxin; mycobacterial cell wall extract; myriaporone; N-acetyldina and cerebratulus A toxin (Kemet al., J. Biol. Chem. 253(16): line; N-Substituted benzamides; nafarelin; nagrestip; nalox 5752-5757, 1978). Agents of the invention can also be one-pentazocine; napavin; naphterpin, nartograstim; nedapl coupled to a synthetic peptide that shares some sequence atin: nemorubicin; neridronic acid; neutral endopeptidase; homology or chemical characteristics with any of the natu nilutamide; nisamycin; nitric oxide modulators; nitroxide rally occurring peptide lysins. Such characteristics include, antioxidant; nitrullyn; 06-benzylguanine; octreotide; oki but are not limited to, linearity, positive charge, amphipath cenone; oligonucleotides; onapristone; ondansetron; icity, and formation of alpha-helical structures in a hydropho ondansetron; oracin; oral cytokine inducer, ormaplatin: bic environment (Leuschner et al., Biology of Reproduction oSaterone; Oxaliplatin, oxaunomycin; paclitaxel analogues; 73:860-865, 2005). Agents of the invention can also be paclitaxel derivatives; palauamine; palmitoylrhizoxin; pam coupled to an agent that induces complement-mediated cell idronic acid; panaxytriol; panomifene; parabactin; paZellip lysis such as, for example, the immunoglobulin F subunit. tine; pegaspargase; peldesine; pentosan polysulfate Sodium; Agents of the invention can also be coupled to any member of pentostatin: pentrozole; perflubron; perfosfamide: perillyl the phospholipase family of enzymes (including phospholi alcohol; phenazinomycin; phenylacetate; phosphatase pase A, phospholipase B, phospholipase C, or phospholipase inhibitors; picibanil; pilocarpine hydrochloride; pirarubicin; D) or to a catalytically-active subunit thereof. US 2015/O151 001 A1 Jun. 4, 2015

0109 Agents of the invention (e.g., peptides, polypep ine; Fludarabine-PO4 (25 mg/m/day, x5 days); mitox tides, proteins, Small molecules, or antibody fragments that antrone (12-14 mg/m); mitozolomide (>400 mg/m); Pen target senescent cells) can also be coupled to a radioactive to statin; or Tomudex. agent, including, but not limited to: Fibrinogen 'I; 0112 Any of the agents of the invention (e.g., peptides, Fludeoxyglucose 'F: Fluorodopa "F: Insulin ''I; Insulin polypeptides, proteins, Small molecules, antibodies, or anti ''I: Iobenguane 'I: Iodipamide Sodium '''I: Iodoantipy body fragments that target Senescent cells) can be coupled to rine ''I; Iodocholesterol ''I: Iodohippurate Sodium 'I; an antimetabolic agent. Antimetabolic agents include, but are Iodohippurate Sodium 'I: Iodohippurate Sodium 13 II s not limited to, the following compounds and their derivatives: Iodopyracet'I; Iodopyracet ''I: Iofetamine Hydrochloride azathioprine, cladribine, cytarabine, dacarbazine, fludarabine '*'I: Iomethin ''I: Iomethin ''I; Iothalamate Sodium 'I; phosphate, fluorouracil, gencitabine chlorhydrate, mercap Iothalamate Sodium '''I; tyrosine '''I: Liothyronine 'I: topurine, methotrexate, mitobronitol, mitotane, proguanil Liothyronine ''I; Merisoprol Acetate '7Hg; Merisoprol chlorohydrate, pyrimethamine, raltitrexed, trimetrexate glu curonate, urethane, vinblastine Sulfate, Vincristine Sulfate, Acetate 'Hg: Merisoprol 'Hg: Selenomethionine 7Se: etc. In other embodiments, the agents of the invention can be Technetium’"Tc Antimony Trisulfide Colloid; Technetium coupled to a folic acid-type antimetabolite, a class of agents "Tc Bicisate; Technetium "Tc Disofenin; Technetium that includes, for example, methotrexate, proguanil chlorhy "Tc Etidronate; Technetium '"Tc Exametazine; Techne drate, pyrimethanime, trimethoprime, or trimetrexate glucu tium’"Tc Furifosmin; Technetium’"Tc Gluceptate; Tech ronate, or derivatives of these compounds. netium '"Tc Lidofenin; Technetium "Tc Mebrofenin: 0113. In another embodiment, any of the agents of the Technetium’"Tc Medronate; Technetium’"Tc Medronate invention (e.g., peptides, polypeptides, proteins, Small mol Disodium; Technetium’"Tc Mertiatide; Technetium’"Tc ecules, antibodies, or antibody fragments that target senes Oxidronate; Technetium’"Tc Pentetate; Technetium’"Tc cent cells) can be coupled to a member of the anthracycline Pentetate Calcium Trisodium; Technetium’"Tc Sestamibi; family of neoplastic agents, including but not limited to acla Technetium "Tc Siboroxime; Technetium '"Tc; Succi rubicine chlorhydrate, daunorubicine chlorhydrate, doxoru mer; Technetium "Tc Sulfur Colloid; Technetium "Tc bicine chlorhydrate, epirubicine chlorhydrate, idarubicine Teboroxime; Technetium "Tc Tetrofosmin; Technetium chlorhydrate, pirarubicine, or Zorubicine chlorhydrate; a "Tc Tiatide; Thyroxine 'I: Thyroxine ''I: Tolpovidone camptothecin, or its derivatives or related compounds, such as ''I; Triolein I; or Triolein ''I. 10, 11 methylenedioxycamptothecin; or a member of the maytansinoid family of compounds, which includes a variety 0110. Therapeutic or cytotoxic agents may further of structurally-related compounds, e.g., ansamitocin P3, include, for example, anti-cancer Supplementary Potentiat maytansine, 2-N-demethylmaytanbutine, and maytanbicy ing Agents, including, but not limited to: Tricyclic anti-de clinol. pressant drugs (e.g., imipramine, desipramine, amitryptyline, 0114 Any of the agents of the invention (e.g., peptides, clomipramine, trimipramine, doxepin, nortriptyline, protrip polypeptides, proteins, Small molecules, antibodies, or anti tyline, amoxapine, and maprotiline); non-tricyclic anti-de body fragments that target senescent cells) can be modified or pressant drugs (e.g., Sertraline, traZodone, and citalopram); labeled to facilitate diagnostic ortherapeutic uses. Detectable Ca" antagonists (e.g., Verapamil, nifedipine, nitrendipine, labels such as a radioactive, fluorescent, heavy metal, or other and caroverine); Calmodulin inhibitors (e.g., prenylamine, agents may be bound to any of the agents of the invention. trifluoroperazine, and clomipramine); Amphotericin B; Tri Single, dual, or multiple labeling of an agent may be advan paranol analogs (e.g., tamoxifen); antiarrhythmic drugs (e.g., tageous. For example, dual labeling with radioactive iodina quinidine); antihypertensive drugs (e.g., reserpine); Thiol tion of one or more residues combined with the additional depleters (e.g., buthionine and sulfoximine) and Multiple coupling of for example, "Y via a chelating group to amine Drug Resistance reducing agents such as Cremaphor EL. containing side or reactive groups, would allow combination 0111. The agents of the invention (e.g., peptides, polypep labeling. This may be useful for specialized diagnostic needs tides, proteins, Small molecules, antibodies, or antibody frag Such as identification of widely dispersed Small neoplastic ments that target senescent cells) can also be administered cell masses. with cytokines, such as granulocyte colony stimulating fac 0115 Agents of the invention (e.g., peptides, polypep tor, or anticancer agents used in anti-cancer cocktails includ tides, proteins, Small molecules, antibodies, or antibody frag ing, e.g., those shown with their MTDs in parentheses: gem ments that target senescent cells), or analogs thereof, can also citabine (1000 mg/m); methotrexate (15gm/m i.v.+leuco. be modified, for example, by halogenation of the tyrosine <500 mg/m i.v. w/o leuco); 5-FU (500 mg/m/dayx5 days); residues of the peptide component. Halogens include fluo FUDR (100 mg/kgx5 in mice, 0.6 mg/kg/day in human i.a.); rine, chlorine, bromine, iodine, and astatine. Such haloge FdUMP. Hydroxyurea (35 mg/kg/d in man); Docetaxel (60 nated agents may be detectably labeled, e.g., if the halogen is 100 mg/m); discodermolide; epothilones; Vincristine (1.4 a radioisotope, such as, for example F, Br, 77Br ‘I, 'I, mg/m); vinblastine (escalating: 3.3-11.1 mg/m, or rarely to 'I, ‘I,’’I, I, or ''A. Halogenated agents of the inven 18.5 mg/m); vinorelbine (30 mg/m/wk); meta-pac: irinote tion contain a halogen covalently bound to at least one amino can (50-150 mg/m, 1x/wk depending on patient response); acid, and preferably to D-Tyrresidues in eachagent molecule. SN-38 (-100 times more potent than Irinotecan); 10-OH Other suitable detectable modifications include binding of campto; topotecan (1.5 mg/m/day in humans, 1xiv LDIOm other compounds (e.g., a fluorochrome such as fluorescein) to ice–75 mg/m): etoposide (100 mg/m in man); adriamycin; a lysine residue of the agent of the invention, or analog, flavopiridol; Cis-Pt (100 mg/m in man); carbo-Pt (360 particularly an agent or analog having a linker including mg/m in man); bleomycin (20 mg/m); mitomycin C (20 lysines. mg/m); mithramycin (30 Sug/kg); capecitabine (2.5 g/m 0116 Radioisotopes for radiolabeling any of the agents of orally); cytarabine (100 mg/m/day); 2-C1-2'deoxyadenos the invention (e.g., peptides, polypeptides, proteins, Small US 2015/O151 001 A1 Jun. 4, 2015 molecules, antibodies, or antibody fragments that target 0121 An agent of the invention (e.g., peptides, polypep senescent cells) include any radioisotope that can be tides, proteins, Small molecules, antibodies, or antibody frag covalently bound to a residue of the peptide component of the ments that target senescent cells) can also be coupled directly agent of the invention or analog thereof. The radioisotopes to a cytotoxic or therapeutic agent using known chemical can be selected from radioisotopes that emit either beta or methods, or the two moieties can be coupled via an indirect gamma radiation, or alternatively, any of the agents of the linkage. Such as through a chelating group. For example, the invention can be modified to containchelating groups that, for agent can be attached to a chelating group that is attached to example, can be covalently bonded to lysine residue(s) of the the cytotoxic or therapeutic agent. Chelating groups include, analog. The chelating groups can then be modified to contain but are not limited to, ininocarboxylic and polyaminopoly any of a variety of radioisotopes, such as gallium, indium, carboxylic reactive groups, diethylenetriaminepentaacetic technetium, ytterbium, rhenium, or thallium (e.g., ‘I,’Ga, acid (DTPA), and 1,4,7,10-tetraazacyclododecane-1,4,7,10 In, mTc, 69Yb. Re). tetraacetic acid (DOTA). For general methods, see, e.g., Liu et 0117. When one or more of the agents of the invention al., Bioconjugate Chem. 12(4):653, 2001; Cheng et al., WO (e.g., peptides, polypeptides, proteins, Small molecules, anti 89/12631; Kieffer et al., WO 93/12112: Albert et al., U.S. Pat. bodies, or antibody fragments that target Senescent cells) is No. 5,753,627; and WO91/01144 (each of which are hereby modified by attachment of a radioisotope, preferable radio incorporated by reference). isotopes are those having a radioactive half-life correspond I0122) When coupled to a therapeutic or cytotoxic agent, ing to, or longer than, the biological half-life of the agent specific targeting by the agents of the invention (e.g., pep used. More preferably, the radioisotope is a radioisotope of a tides, polypeptides, proteins, Small molecules, antibodies, or halogen atom (e.g. a radioisotope of fluorine, chlorine, bro antibody fragments that target senescent cells) allows selec mine, iodine, and astatine), even more preferably Br, 'Br, tive destruction of senescent cells. For example, the agents of 76 22 123 24 25 29 13 2. the invention can be used to target and destroy senescent cells Br, 144I, 14-I, 14“I, 14s I, 14 I, II, or 21 At of the lung, breast, prostate, and colon in order to prevent, 0118 Agents of the invention (e.g., peptides, polypep stabilize, inhibit the progression of, or treat cancers originat tides, proteins, Small molecules, antibodies, or antibody frag ing in these organs. Also, for example, the agents of the ments that target senescent cells) coupled to radioactive met invention can be used to target and destroy senescent cells of als are useful in radiographic imaging or radiotherapy. the vasculature, brain, liver, kidney, heart, lung, prostate, Preferred radioisotopes also include "Tc, Cr, Ga, Ga, colon, nasopharynx, oropharynx, larynx, bronchus, and skin 11 In 168Yb. 140La, 90Y, 88Y 153Sm, 156Ho, 165Dy, 6.Cu, in order to prevent, stabilize, inhibit the progression of, or 97.Ru, 10.Ru, 186Re, 188Re, 20Pb, 2 Bi, 22Bi, 213Bi, and treat age-related diseases or tobacco-related diseases or con '''Bi. The choice of metal can be determined based on the ditions relating to these organs. Any of the agents of the desired therapeutic or diagnostic application. invention (e.g., peptides, polypeptides, proteins, Small mol 0119 The agents of the invention (e.g., peptides, polypep ecules, antibodies, or antibody fragments that target senes tides, proteins, Small molecules, antibodies, or antibody frag cent cells) can be administered to a mammalian Subject. Such ments that target senescent cells), when coupled to a metal as a human, directly or in combination with any pharmaceu component, are useful as diagnostic and/or therapeutic tically acceptable carrier or salt known in the art. Pharmaceu agents. A detectable label may be a metal ion from heavy tically acceptable salts may include non-toxic acid addition elements or rare earthions, such as Gd", Fe", Mn", or Cr". salts or metal complexes that are commonly used in the phar Conjugates that include paramagnetic or Superparamagnetic maceutical industry. Examples of acid addition salts include metals are useful as diagnostic agents in MRI imaging appli organic acids such as acetic, lactic, pamoic, maleic, citric, cations. Paramagnetic metals that can be coupled to the malic, ascorbic, Succinic, benzoic, palmitic, Suberic, Sali agents of the invention include, but are not limited to, chro cylic, tartaric, methanesulfonic, toluenesulfonic, or trifluoro mium (III), manganese (II), iron (II), iron (III), cobalt (II), acetic acids or the like; polymeric acids such as tannic acid, nickel(II), copper(II), praseodymium(III), neodymium(III), carboxymethylcellulose, or the like; and inorganic acids Such Samarium (III), gadolinium (III), terbium (III), dysprosium as hydrochloric acid, hydrobromic acid, Sulfuric acid phos (III), holmium (III), erbium (III), and ytterbium (III). Prefer phoric acid, or the like. Metal complexes include Zinc, iron, ably, the polymer has a relaxtivity of at least 10, 12, 15, or 20 and the like. One exemplary pharmaceutically acceptable mM' sec'Z', wherein Z is the concentration of paramag carrier is physiological saline. Other physiologically accept netic metal. able carriers and their formulations are known to one skilled 0120 Chelating groups may be used to indirectly couple in the art and described, for example, in Remington's Phar detectable labels or other molecules to an agent of the inven maceutical Sciences, (18th edition), ed. A. Gennaro, 1990, tion (e.g., a peptide, polypeptide, protein, Small molecule, Mack Publishing Company, Easton, Pa. antibody, or antibody fragment). Chelating groups can link Agents of the Invention for Use in Cellular Therapy agents of the invention with radiolabels, such as a bifunc Applications tional stable chelator, or can be linked to one or more terminal or internal amino acid reactive groups. Conjugates can be 0123. One or more agents of the invention (e.g., peptides, linked via an isothiocyanate B-Ala or appropriate non polypeptides, proteins, Small molecules, antibodies, or anti C.-amino acid linker which prevents Edman degradation. body fragments that target senescent cells) can be used to Examples of chelators known in the art include, for example, improve the efficacy of a cellular therapy provided to a patient the ininocarboxylic and polyaminopolycarboxylic reactive (e.g., a mammal. Such as a human) in need thereof by deplet groups, ininocarboxylic and polyaminopolycarboxylic reac ing (i.e., killing) one or more, all, or Substantially all (e.g., tive groups, diethylenetriaminepentaacetic acid (DTPA), and 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid 40%, 50%, 60%, 70%, 80%, 90%. 95%, 99% or more) senes (DOTA). cent cells in the transferred donor cells, tissue, or organ prior US 2015/O151 001 A1 Jun. 4, 2015

to, concurrent with, or following administration of the cellu ing agents (e.g., magnesium Stearate) may also be used. Tab lar therapy. Alternatively, a patient (e.g., a mammal. Such as a lets and pills can additionally be prepared with enteric coat human) may be treated with one or more agents of the inven ings. tion prior to, concurrent with, or following administration of 0.126 Liquid dosage forms for oral administration include the cellular therapy. Cellular therapies include, without limi pharmaceutically acceptable emulsions, Solutions, Suspen tation, the autologous, allogeneic, Syngeneic, or Xenogeneic sions, syrups, and Soft gelatin capsules. These forms contain transplantation or engraftment of cells (e.g., blood, pancreatic inert diluents commonly used in the art, such as water oran oil islet cells, and stem cells (e.g., hematopoietic stem cells medium. Besides such inert diluents, compositions can also (HSC), umbilical cord blood stem cells (see, e.g., US Patent include adjuvants, such as wetting agents, emulsifying Application Publication 2002/0164794), pluripotent stem agents, and Suspending agents. cells (e.g., hoX11-expressing pluripotent stem cells described I0127. Formulations for parenteral administration include in US Patent Application Publication 2005/0158288), multi sterile aqueous or non-aqueous Solutions, Suspensions, or potent stem cells, and totipotent stem cells), tissues (e.g., skin emulsions. Examples of Suitable vehicles include propylene and adipose tissue), or organs (e.g., kidney, liver, heart, and glycol, polyethylene glycol, vegetable oils, gelatin, hydroge lung) to a patient (e.g., a mammal. Such as a human) in need nated naphthalenes, and injectable organic esters, such as thereof. Cellular therapies (e.g., cell (e.g., stem cells), tissue, ethyl oleate. Such formulations may also contain adjuvants, or organ transplantation or engraftment) are currently being Such as preserving, wetting, emulsifying, and dispersing developed for a wide range of therapeutic indications for agents. Biocompatible, biodegradable lactide polymer, lac degenerative and pathologic diseases, such as osteoarthritis tide/glycolide copolymer, or polyoxyethylene-polyoxypro (see, e.g., U.S. Patent Application Publication No. 2007/ pylene copolymers may be used to control the release of the 0264238), ischemia and cardiac tissue damage. Such as that compounds. Other potentially useful parenteral delivery sys caused by myocardial infarction (see, e.g., Dzau et al., U.S. tems for the agents of the invention include ethylene-vinyl Patent Application Publication No. 2007/0259425), type I acetate copolymer particles, osmotic pumps, implantable and type II diabetes (see, e.g., Uchida et al., U.S. Patent infusion systems, and liposomes. Application Publication No. 2007/0212732), renal dysfunc I0128 Liquid formulations can be sterilized by, for tion and multi-organ failure (see, e.g., Westenfelder, U.S. example, filtration through a bacteria-retaining filter, by Patent Application Publication No. 2007/0178071), and neu incorporating sterilizing agents into the compositions, or by rodegenerative diseases (see, e.g., Kim et al., U.S. Patent irradiating or heating the compositions. Alternatively, they Application Publication No. 2007/0054399). The preceding can also be manufactured in the form of sterile, Solid compo U.S. patent application Publications are hereby incorporated sitions which can be dissolved in sterile water or some other by reference in their entirety. sterile injectable medium immediately before use. I0129. Compositions for rectal or vaginal administration Administration and Dosage are preferably Suppositories which may contain, in addition to active Substances, excipients such as cocabutter or a Supposi 0.124 Pharmaceutical formulations of a therapeutically tory wax. Compositions for nasal or Sublingual administra effective amount of an agent of the invention (e.g., peptides, tion are also prepared with standard excipients known in the polypeptides, proteins, Small molecules, antibodies, or anti art. Formulations for inhalation may contain excipients, for body fragments that target senescent cells), or pharmaceuti example, lactose, or may be acqueous solutions containing, for cally acceptable salt-thereof, can be administered orally, example, polyoxyethylene-9-lauryl ether, glycocholate and parenterally (e.g., intramuscular, intraperitoneal, intrave deoxycholate, or may be oily solutions for administration in nous, or Subcutaneous injection, inhalation, intradermally, the form of nasal drops or spray, or as a gel. optical drops, or implant), nasally, vaginally, rectally, Sublin 0.130. The amount of active ingredient in the compositions gually, or topically. The pharmaceutical formulation can of the invention can be varied. One skilled in the art will include the agent of the invention in admixture with a phar appreciate that the exact individual dosages may be adjusted maceutically acceptable carrier adapted for the route of Somewhat depending upon a variety of factors, including the administration. agent being administered, the time of administration, the 0.125 Methods well known in the art for making pharma route of administration, the nature of the formulation, the rate ceutical formulations can be found, for example, in Reming of excretion, the nature of the subjects conditions, and the ton's Pharmaceutical Sciences (18th edition), ed. A. age, weight, health, and gender of the patient. In addition, the Gennaro, 1990, Mack Publishing Company, Easton, Pa. severity of the condition targeted by an agent of the invention Compositions intended for oral use may be prepared in Solid will also have an impact on the dosage level. Generally, or liquid forms according to any method known to the art for dosage levels of an agent of the invention of between 0.1 the manufacture of pharmaceutical compositions. The com ug/kg to 100 mg/kg of body weight are administered daily as positions may optionally contain Sweetening, flavoring, col a single dose or divided into multiple doses. Preferably, the oring, perfuming, and/or preserving agents in order to pro general dosage range is between 250 ug/kg to 5.0 mg/kg of vide a more palatable preparation. Solid dosage forms for oral body weight per day. Wide variations in the needed dosage are administration include capsules, tablets, pills, powders, and to be expected in view of the differing efficiencies of the granules. In such solid forms, the active compound is various routes of administration. For instance, oral adminis admixed with at least one inert pharmaceutically acceptable tration generally would be expected to require higher dosage carrier or excipient. These may include, for example, inert levels than administration by intravenous injection. Varia diluents, such as calcium carbonate, sodium carbonate, lac tions in these dosage levels can be adjusted using standard tose, Sucrose, starch, calcium phosphate, sodium phosphate, empirical routines for optimization, which are well known in or kaolin. Binding agents, buffering agents, and/or lubricat the art. In general, the precise therapeutically effective dosage US 2015/O151 001 A1 Jun. 4, 2015 can be determined by the attending physician in consideration mal skin fibroblasts. The phage were incubated with 2x10' of the above-identified factors. cells of normal skin fibroblast (CCD 1070 sk) at room tem 0131. An agent of the invention (e.g., a peptide, polypep perature for 60 minutes at room temperature with slow shak tide, protein, Small molecule, antibody, or antibody fragment ing in PBS with 0.5% BSA. At the end of incubation, the cells that targets senescent cells) can be administered in a Sustained were pelleted in a microcentrifuge at 1500 RPM for 2 minutes release composition, such as those described in, for example, and the Supernatant recovered. The Supernatant was used to U.S. Pat. No. 5,672,659 and U.S. Pat. No. 5,595,760. The use resuspend another 2x10 subtraction cells. This subtraction of immediate or Sustained release compositions depends on step was repeated 3 times for each round. the type of condition being treated. If the condition consists of 0.139. After the final subtraction step in each round, the an acute or over-acute disorder, a treatment with an immedi recovered phage Supernatant was used to suspend 5x10 ate release form will be preferred over a prolonged release selection cells. The bound phage were recovered and ampli composition. Alternatively, for preventative or long-term fied as described above. The process was repeated for a total treatments, a Sustained released composition will generally of 5 rounds of selection. After the fifth round of selection, be preferred. phage were titered and 20 well-separated plaques were 0132 An agent of the invention (e.g., a peptide, polypep picked, amplified, and sequenced. tide, protein, Small molecule, antibody, or antibody fragment that targets senescent cells) can be prepared in any suitable Results: manner. The agent may be isolated from naturally-occurring Sources, recombinantly produced, or produced synthetically, 0140. Three consensus sequences were obtained from the identified from a library of small molecules, or produced by a phage display selection procedure. These correspond to the combination of these methods. The synthesis of short pep SEQ ID NOS:1-3. SEQ ID NO:1 represented 9/20 clones, tides is well known in the art. See e.g., Stewart et al., Solid SEQ ID NO:2 represented 6/20 clones, and SEQ ID NO:3 Phase Peptide Synthesis (Pierce Chemical Co., 2d ed., 1984). represented 2/20 clones. A peptide portion of any of the agents of the invention can be synthesized according to standard peptide synthesis methods Example 2 known in the art. 0133. The present invention is illustrated by the following Human Study to Test the Prognostic Use of a examples, which are in no way intended to be limiting of the Senescent Cell Binding Agent Labeled with a invention. Radioisotope; Using a Senescent Cell Detecting Radiotracer to Predict Cancer Risk EXAMPLES 0.141. This study is designed to show that noninvasive, in Example 1 Vivo imaging of senescent cell content can be used to predict cancer risk in smokers. Six-hundred subjects will be enrolled. Discovery of Agents that Bind Senescent Cells Eighty percent of these subjects will be smokers; twenty percent will be nonsmokers. Subjects less than 18 years of Phage Selection Technique age, Subjects with a history of cancer, and pregnant Subjects will be excluded. The anticipated mean age of study Subjects 0134) Normal skin cell line CCD-1070Sk was obtained is 55 years. Each study Subject will undergo Scintigraphic from American Type Culture Collection (Bethesda, Md.). imaging using a radio-labeled peptide of the invention (SEQ The cells were grown in Eagle's Minimal Essential medium ID NO: 1) as the radiopharmaceutical. The radiopharmaceu with Earle's BSS, 2 mM L-glutamine, 1.0 mMSodium pyru tical will be prepared by reacting peptide-gly-gly-gly-ser Vate, 0.1 mM nonessential amino acids and 1.5 g/L Sodium DTPA with 111-In chloride. Each study subject will receive bicarbonate supplemented with 10% fetal bovine serum. an intravenously administered dose of 5 mCi of 111-In 0135 Cells were sub-cultured every 4-5 days till they labeled peptide. Anterior and posterior whole body planar became senescent. Cell senescence was confirmed by Senes images of the subjects will be obtained at 24 and 48 hours cence-associated beta-galactosidase staining following administration of the radio-labeled peptide. 0.136 An M13 phage peptide library, Ph.D.- 12, was SPECT imaging of the chest will also be obtained at 24 and 48 obtained from New England BioLabs (Beverly, Mass.), hours following administration of the radio-labeled peptide. which displays random 12-mer peptides. Scintigraphic images will be acquired on a SPECT/CT cam 0.137 For the selection step for round 1, an aliquot (10 uL) era using a medium energy collimator. Two radiologists will of the Ph.D.-12 complete phage library was incubated with read the SPECT/CT study of each subject. Regions of interest 5x10 cells of senescent fibroblasts in 1 mL PBS/0.5% BSA will be drawn around each lung and the mediastinum. The for -3.5 hours at room temperature with slow shaking on activity within each region will be determined for each sub Lab-Quake. At the end of the incubation, the cells were pel ject. Subjects will be followed for two years and monitored leted in a microcentrifuge at 1500 RPM for 2 minutes and the for the incidence of lung cancer. Kaplan-Meier curves will be supernatant removed. Cells were washed with PBS/1.0% drawn for the study population based upon cancer free Sur BSA/0.5% Tween (wash buffer) for a total of 4 washes using vival. An optimal threshold of activity within the regions of fresh tubes between washes. Phage that bound to the target interest will be determined such as to divide subjects that cells were eluted with 200 uL of 0.2 M glycine (pH 2.2) for 8 develop cancer from subjects who don't: a Kaplan-Meier minutes then neutralized with 30 u, of 1 M Tris-HCl (pH curve will be drawn for the set of subjects with activity levels 9.0). The number of phage bound was determined and the above the threshold and a Kaplan-Meier curve will be drawn remaining eluate was amplified. for subjects with activity levels below the threshold level. We 0.138. From the amplified eluate from Round 1, an aliquot expect to observe development of lung cancer in approxi (2x10' phage) was used for subtraction panning against nor mately twenty Subjects. Activity in pulmonary regions of US 2015/O151 001 A1 Jun. 4, 2015

interest will be significantly higher in Smokers than nonsmok cells in each of four random microscopic fields for each tissue ers. Smokers who develop lung cancer will have significantly sample. A two-tailed t-test will be used to evaluate the loss of higher activity in pulmonary regions of interest than Smokers senescent cells in the tissues of diabetic mice treated with who do not develop lung cancer. Cancer-free Survival curves senescent cell cytotoxic agent relative to the loss of senescent will be significantly different for subjects with a pulmonary cells in the tissues of diabetic control mice. region of interest activity level above the threshold value than for subjects with activities below the threshold value. Example 5 Example 3 Enhancement of Stem Cell Treatments by Pre-Treatment with Senescent Cell Binding Agents Using Senescent Cell Binding Agents Coupled to Coupled to Cytotoxic Agents: Effects on Subsequent Cytotoxic Agents to Eliminate Senescent Cells: StemCell Engraftment Effect on Subsequent Development of Cancer 0144. The following experiment can be used to show that 0142. The purpose of this study is to show that elimination exogenously administered stem cells engraft at higher rates of some senescent cells from an organism using the agents of into damaged tissue if the treated organism undergoes pre the invention (e.g., peptides, polypeptides, proteins, Small treatment with a senescent cell cytotoxic agent of the inven molecules, antibodies, or antibody fragments that target tion (e.g., a peptide agent) to reduce the content of senescent senescent cells) will reduce the risk of subsequently develop cells in the damaged tissue compartment. Removal of senes ing cancer. For example, a Senescent cell binding peptide. cent cells increases the engraftment of stem cells into dam (SEQ ID NO: 1) will be conjugated to a cytotoxic peptide aged tissue. having the sequence KFAKFAKKFAKFAKKFAKFAK 0145 Balb-C mice will undergo left anterior descending (SEQID NO:4; Leuschner and Hansel, Biology of Reproduc (LAD) artery ligation for 60 minutes to induce myocardial tion 73:860-865) via an amino acid linker sequence (e.g., a infarction. The mice will be pre-anesthetized in an isoflurane linker sequence selected from GGGC (SEQIDNO:9), GGGS inhalation chamber and receive an i.p. injection of Sodium (SEQID NO:10), and GG) at the C terminus of the senescent pentobarbital (25 mg/kg). The animals will be intubated and cell binding peptide. Study subjects will consist of 60 ventilated for the duration of the procedure. The LAD artery BALB/c mice, mean age 6 mo which includes 30 experimen will be identified following left lateral thoracotomy and peri tal animals and 30 controls. Experimental animals will cardectomy. Ligation will be performed on the proximal 2 receive an intravenously administered dose of 0.2 mg/Kg of mm portion of the LAD using a 9-0 ethilon Stitch. Mice will body weight of senescent cell binding peptide conjugate once be maintained at 23°C. with 12 hour light and 12 hour dark every three months for one year. Control animals will receive cycles for 6 days. Seven surviving mice will be used as experi an equal dose of a 37 amino acid control peptide. Kaplan mental animals and seven will be used as control animals. Meier curves representing tumor free survival will be con Experimental mice will receive tail vein injections of a senes structed for each group of mice. Approximately 30% of con cent cell cytotoxic agent conjugated to a lytic peptide trol mice will develop tumors by the end of the study period. sequence (SEQ ID NO:8; GVYHFAPLTPTPGGKFAK The study is expected to show significantly longer tumor free FAKKFAKFAK; 300 ug/dose) every second day for six days. Survival in the experimental group compared to the control 0146 Murine hematopoietic stem cells (HSC) will be group. obtained from StemCell Technologies Inc. Cells will be trans fected to express enhanced green fluorescent protein (EGFP). Example 4 Plasmid pEF-1 a-EGFP containing an EGFP gene under the control of human EF1, a promoter, and a neomycin-resistance Reduction of Senescent Cell Content in Diabetic cassette, will be constructed as follows: (1) the promoter Mice by Treatment with a Senescent Cell Cytotoxic region of pEGFP-N3 (Clontech, Palo Alto, Calif.) will be Agent removed by cutting out the Asel-NheI DNA fragment and 0143 Diabetes is induced in female CD-1 mice, 5-7 weeks joining the blunt-ended termini, and (2) human EF1, a pro old and 25-35 g in body weight, by intraperitoneal injection of moter from pEF-BOS (a fragment of HindIII and EcoRI 200 mg/Kg body weight of streptozotocin dissolved in DNA) will be inserted into the HindIII-EcoRI site of the sodium citrate saline buffer (pH 4.5). Tail vein blood glucose plasmid. Murine HSC will be transfected with pEF-1 a-EGFP will be measured 5 days after injection to ensure induction of by electroporation and selected in the presence of G418. A diabetes. Diabetic mice will be maintained at constant tem single clone that brightly expresses EGFP will be chosen and perature (23°C.) with 12 hour light and 12 hour dark cycles used for the experiments. The clone will be adapted to feeder for 16 weeks following confirmation of diabetes. Seven dia free conditions and maintained on gelatin-coated dishes in betic mice will receive a weekly tail vein injection of a senes Dulbecco's Modified Eagle's Media supplemented with 15% cent cell cytotoxic agent (SenL: SEQID NO:6; GVYHFA fetal calf serum, 2 mM Sodium pyruvate, 2 mM L-glutamine, PLTPTPGGGSKFAKFAKKFAKFAK; 300 g/dose), 1x nonessential amino acids, 1,000 units of 0.1 mM 2-mer comprising a senescent cell binding sequence linked to alytic captoethanol per mL, along with 100 units of streptomycin peptide sequence during the 16 weeks. Seven control animals and 100 ug of penicillin per mL. Cells will be collected after will receive an equivalent volume TV injection of normal trypsinization with EDTA and placed in aliquots of the saline. At the end of 16 weeks, all animals will be sacrificed. medium described above for mouse tail vein injection 1 hour Tissue cross sections will be prepared from aorta, lung, liver, later. and heart from Snap frozen tissue. Tissue samples will be 0147 HSC (10) will be injected via tail vein into each stained for SA-B-gal activity using the method of Campisi et experimental and control mouse. Ten days later, each animal al. Percentage of SA-B-gal positive cells will be determined will be sacrificed. Hearts will be excised and fixed in 2% for each tissue sample from each animal by counting 1000 paraformaldehyde in phosphate-buffered solution (PBS) for US 2015/O151 001 A1 Jun. 4, 2015 20

2 hours and cryoprotected in 30% sucrose overnight. Tissue (GGGC: SEQ ID NO:9) to produce SenR (SEQ ID NO:7). will be embedded in optimum cutting temperature medium Senescent fibroblasts, non-senescent fibroblasts, and prostate and sectioned at 5 um on a cryostat. Serial sections will be epithelial cells were then incubated with the peptide-ricin A stained with hematoxylin and eosin (H&E). Tissue will be conjugate (SenR). Increased cell death was observed in the examined with a fluorescent microscope. Percentage of GFP case of senescent cells treated with SenR than in the other cell positive cells will be determined for each cardiac tissue types (FIG.3). For example, an approximately 3 fold increase sample from each animal by counting 1000 cells in each of in cell death was measured in senescent fibroblast when com four random microscopic fields for each tissue sample. A pared to non-senescent fibroblast when treated with 50 uMof two-tailed t-test will be used to evaluate the hypothesis that SenR. exogenously administered HSC engraft at a higher rate into mice treated with senescent cell cytotoxic agent relative to Binding of Peptide to Senescent Cells Versus Non-Senescent untreated controls. Special attention will be paid to cardiac Cells tissue in the LAD territory (anterior wall) of each heart. 0151. In the absence of unlabeled peptide, radio-labeled Example 6 SenC bound to prostate epithelium, normal fibroblasts, and senescent cells at an average of 0.06%, 0.08%, and 0.32% of In Vitro Validation of the Use of Senescent Cell added dose, respectively, demonstrating that SenC binds to Binding Agents as Agents to Deliver Molecular senescent cells at a higher rate than to the other cell types Cargo to the Cytoplasm of Senescent Cells (P=0.001). In the presence of 20 uMunlabeled SenC, labeled SenC bound to prostate epithelium, normal fibroblasts, and 0148 Cytotoxicity of Senescent Cell Binding Peptide senescent cells at an average of 0.06%, 0.09%, and 0.25% of Conjugated to a Lytic Peptide Sequence Cytotoxic peptide added dose, respectively. There was no significant difference SenL (i.e., SEQID NO:6) was synthesized by conjugating a between binding rates in the presence or absence of unlabeled senescent cell binding agent (SEQID NO: 1) to alytic peptide SenC in the case of prostate epithelium and normal fibroblasts sequence (KFAKFAKKFAKFAK; SEQ ID NO:4) via a 4 (P=0.5 and 0.4 respectively), indicating that the binding to residue linker (GGGS: SEQID NO:10) and tested for differ these cell types is nonspecific. Labeled SenC did bind at ential cytotoxic activity in senescent fibroblasts, prostate epi significantly different rates to senescent cells in the absence thelial cells, and non-senescent fibroblasts. Senescent fibro blasts exhibited dose-dependent cell death at a significantly vs. presence of unlabeled SenC (P=0.04), indicative of spe higher rate than either non-senescent fibroblasts or prostate cific binding (see FIG. 4). epithelial cells (FIG. 1). The effect of SenL on cell prolifera Example 7 tion was also assessed using the Cell Proliferation Assay WST-1 (Roche, Mannheim, Germany). As shown in FIG. 2, no change in cell proliferation was seen in any of the three cell Isolation and Identification of Senescent types in response to treatment with SenL. The proliferation Cell-Specific Antigens assay uses WST-1 as a reagent; the reaction is catalyzed by 0152 Fibroblasts (CCD-1070Sk) were cultured as out mitochondrial dehydrogenases. Senescent cells have higher lined above and divided into three groups: replicatively senes mitochondrial mass than their non-senescent counterparts cent, stress induced prematurely senescent, and non-senes (Lee et al., J. Biomed. Sci. 9:517-26, 2002); consequently, cent. Cell Surface proteins of each group of cells were baseline WST assay values are higher for senescent cells. The conjugated to biotin using the Pierce Cell Surface Protein occurrence of cell death in response to treatment with SenL Isolation Kit (Thermo Scientific, product number 89881) fol and absence of change in proliferation rate indicate that SenL lowing the instructions of the manufacturer. Membrane pro causes cell death in non-proliferating cells, e.g. senescent teins were captured using neutravidin following membrane cells. dissolution and sent for 2D gel electrophoresis. Spots from 0149. It is worth noting that each cultured cell population each gel were analyzed to look for differences in protein contains a mixture of senescent and non-senescent cells at all expression among the three groups of fibroblasts. Protein population doublings, but that the relative proportion of spots occurring in the gels corresponding to senescent (rep senescent cells within the population increases stochastically licative or stress-induced) cells but not in gels corresponding with each population doubling (Martin-Ruiz et al., J. Biol. to non-senescent fibroblasts were identified as senescence Chem. 279(17):17826-33, 2004). Thus, the “senescent cells” specific antigens and identified using MALDI-TOF mass used in this cell killing experiment are predicted to contain a spectrometry. Subpopulation of non-senescent cells. Likewise, a fraction of 0153. Mutant beta-actin (SEQ ID NO:11; GI: 28336) and the “non-senescent cells' used in this experiment are pre ACTB protein (SEQID NO:12; GI: 15277503) were identi dicted to be senescent. Therefore, the observed difference in fied as cell-surface, senescence specific antigens occurring in cell killing between the two populations is reduced by the both replicatively senescent and stress-induced prematurely impure composition of each population with regard to senes senescent cells. Drug resistance-related protein LRP (SEQID cence. This explains why some cytotoxicity is observed in the NO:13; GI: 1097308) and major vault protein (SEQ ID “non-senescent” fibroblast population. It also explains why NO:14: GI: 19913410) were identified as senescence-specific the prostate epithelial cells (RWPE-1), which are not pre Surface proteins in the replicatively senescent cells only. dicted to have any senescent cells due to immortalization Senescence specific antigens that were identified in stress through HPV-18 transduction, show no cell death at all. induced prematurely senescent cells included thyroid hor mone binding protein precursor (SEQ ID NO:15; GI: In Vitro Cytotoxicity: Conjugation to Ricin-A 339647); unnamed protein product (SEQ ID NO:20; GI: 0150. A senescent cell binding agent (i.e., SEQID NO:1) 35655); prolyl 4-hydroxylase, beta subunit precursor (SEQ was conjugated to the ricin A Subunit via a 4 peptide linker ID NO:16: GI: 20070125); chain A, human protein disulfide US 2015/O151 001 A1 Jun. 4, 2015

isomerase (SEQID NO:17: GI: 159162689): electron-trans cold PBS. Secondary antibody (goat anti rabbit IgG conju fer-flavoprotein, beta polypeptide (SEQ ID NO:18; GI: gated to FITC) was diluted 1:100 in PBS with 0.2% BSA and 4503609); unnamed protein product (SEQ ID NO:21; GI: used to incubate cells for 30 minutes at room temperature. 158257194); unnamed protein product (SEQID NO:22; GI: Cells were washed three times with cold PBS. 158259937); ATP synthase, H transporting, mitochondrial F1 complex, alpha subunit precursor (SEQ ID NO:19; GI: Example 9 4757810), and cathepsin B (SEQID NO:23). Materials and Methods Methods Cell Culture Induction of Replicative Senescence 0159 All cells were obtained from AmericanType Culture 0154 CCD-1070Sk (fibroblasts) was cultured as Collection (Manassas, Va.). Each cell culture was grown at described herein. Cells were cultured until they underwent 68 37°C. in 5% CO.. CCD-1070Sk (fibroblasts) were grown in population doublings at which time they displayed typical minimum essential medium (Eagle) with 2 mM L-glutamine senescent morphology and underwent minimal further cell and Earle's BSS adjusted to contain 1.5 g/L sodium bicar growth in response to mitogens. bonate, 0.1 mM non-essential amino acids, and 1.0 mM sodium pyruvate, and supplemented with 10% fetal bovine Isolation of Cell Surface Proteins serum. RWPE-1 (non-cancerous prostate epithelial cells) 0155 Cell surface proteins were extracted from three were grown in keratinocyte-serum free medium Supple groups of cells (replicatively senescent fibroblasts, stress mented with 5 ng/mL human recombinant EGF and 0.05 induced prematurely senescent fibroblasts, and non-senes mg/mL bovine pituitary extract. cent fibroblasts). Each group of cells contained 10 cells. Cell surface proteins were isolated using the Pierce Cell Surface Chemical Induction of Cellular Senescence Protein Isolation Kit (Thermo Scientific, product number (0160 CCD-1070Sk (fibroblasts) was cultured as 8.9881), following the instructions of the manufacturer. Pro described above to 70% confluence. Cells were then treated tein isolates were analyzed using spectrophotometry which with 200 uM hydrogen peroxide (i.e., H.O.) for 2 hours. showed absorbances at 280 nm of 1.25, 1.375, and 1.347 for Media was then replaced with fresh media and cells were replicatively senescent cells, stress induced prematurely allowed to grow for 3 days. Cells were then harvested by senescent cells, and non-senescent cells respectively. Total trypsinization, split 1:3, and again grown to 70% confluence. volume of each protein isolate was 500 uL. Cells were then retreated with 200 uM hydrogen peroxide. Lower passage number fibroblasts frequently required two Identification of Cell Surface Proteins treatments with hydrogen peroxide, but higher passage fibro 0156 Cell surface protein isolates were sent for analysis blasts occasionally required only a single treatment. by the proteomics core at the University of Massachusetts Medical School (Worcester, Mass.). The analysis was carried Peptide Synthesis out by performing a buffer exchange for each protein isolate 0.161 Peptides were synthesized using standard FMOC sample followed by 2D gel electrophoresis. Each gel was protected chemistry. For in vitro cell cytotoxicity studies, compared to find protein spots that occurred in the gels cor peptide was synthesized with a C-terminal, cell lytic responding to the Senescent (replicative or stress-induced) sequence according to the following: GVYHFAPLTPT cells but not in the gels corresponding to the non-senescent PGGGS(KFAKFAK) (SEQ: ID NO:6; SenL). The peptide cells. Protein spots that occurred in the senescent cell Samples sequence GVYHFAPLTPTPGGGC (SEQ ID NO:5; SenC) but not in the non-senescent samples were digested and sent was synthesized for Subsequent conjugation to ricin-A Sub for mass spectrometry analysis for identification. unit and for radio-labeling with 99m-technetium. Example 8 Conjugation of FITC to Peptide Immunostaining for Cathepsin B Expression on the 0162 FITC was conjugated to the N terminus of SenC Surface of Senescent Cells according to the following: (1) A senescent cell binding pep tide (SenC: SEQID NO:5) was prepared at a concentration of O157 Senescent fibroblasts and non-senescent fibroblasts 5 mM in 125 uL of 0.5 M NaHCO, buffer, pH 9.5, and (2) were grown on cover slips and immunostained for cell Surface FITC was added to the peptide solution at a 1:5 molar ratio expression of cathepsin B. Images appear in FIGS. 5A and (peptide:FITC) and diluted to a final volume of 200 uL. The 5B, which show surface staining for cathepsin B on the senes solution was incubated in the dark for 2 hours. Peptide-FITC cent cells but not on their non-senescent counterparts. conjugate was purified on a P4 column using PBS, pH 7.2 as 0158 Fibroblasts (CCD1070Sk) were grown in culture as an eluent. detailed herein. Replicatively senescent cells were acquired by growing cells for 50 population doublings followed by plating on cover slips. Non-senescent cells were acquired by Cell Internalization plating mid passage fibroblasts on cover slips. Cells on cover (0163 Premature senescence offibroblasts (CCD-1070Sk) slips were allowed to attach overnight. Cells were fixed with was induced as outlined above. Non-senescent fibroblasts methanol and washed. Rabbit polyclonal antibody to cathe were grown in culture as detailed above. Cells were harvested psin B was diluted 1:100 in PBS with 0.2% BSA. Cells were and added to collagen-coated coverslips and grown in MEM incubated with primary antibody to cathepsin B for one hour plus 10% FBS overnight. Media was removed, and cells were at room temperature followed by washing three times with washed once with PBS. Minimal essential media without US 2015/O151 001 A1 Jun. 4, 2015 22

FBS was added to the cells. A senescent cell binding peptide form SenR. RicinA was obtained from manufacturer in solu (SenC: SEQID NO:5) conjugated to FITC was added to cells tion. A buffer exchange was performed with 0.1 MPBS/20% on coverslips at a concentration of 5 uM and incubated for 3 glycerol. Ricin A was conjugated with NHS-PEO-maleim hours. Cells were than washed five times with PBS followed ide cross linker (Pierce, Rockford, Ill.) at a 1:10 molar ratio by fixation with 1:1 methanol/acetone for 10 minutes at -20° for 30 minutes at room temperature. Derivatized ricin A was C. Coverslips were air dried and mounted in fluorescent purified on a P4 column using 0.1 MPBS/20% glycerolas an mounting media with DAPI and visualized with an Olympus elution buffer. Derivatized ricinA was combined with P12S at BX51 fluorescent microscope and DP70 digital camera with a 1:1 molar ratio and reacted for 2 hours at room temperature. excitation and emission wavelengths of 490 and 520 nm. (0167 CCD-1070Sk and RWPE-1 were grown in culture as detailed above. Senescence was chemically-induced as Cytotoxicity of Senescent Cell Binding Peptide Conjugated described above. Cells were trypsinized and suspended in to a Lytic Peptide Sequence culture media containing 10% FBS. Cells were centrifuged at (0164. CCD-1070Sk and RWPE-1 were grown in culture 1,000 rpm for 5 minutes. Supernatant was removed and cells as detailed above. Premature senescence of fibroblasts was were resuspended in 1 mL media without FBS. Cell suspen chemically induced as described above. Cells were trypsi sions were diluted to 20,000 cells/75 uL. Twenty-five micro nated and suspended in culture media containing 10% FBS. liters of appropriately-diluted peptide-ricin A conjugate was Cells were centrifuged at 1,000 rpm for 5 minutes. Superna added to each cell sample to give various concentrations. tant was removed and cells were resuspended in 1 mL media Each sample was prepared in triplicate. Cell Suspensions without FBS. Cell suspensions were diluted to 20,000 cells/ were transferred to a 96 well plate and incubated in the pres 75uL. Twenty-five microliters of appropriately-diluted agent ence of peptide-ricin A conjugate for 2 hours at 37° C. Per solution (SenL: SEQID NO:6) was added to cell samples to centage of cells killed (FIG. 3) was determined as outlined give various concentrations of 0, 0.1, 0.5, 1.0, 2.5, 5.0 or 10 above. uM. Each sample was prepared in triplicate. Cell Suspensions were transferred to a 96 well plate and incubated in the pres Radio-Labeling of Peptide Senescent Cell Binding Peptide ence of various concentrations of SenL for 2 hours at 37° C. Six samples of each cell type contained no agent. The assay 0168 A senescent cell binding agent (SEQID NO: 1) was plate was removed from the incubator, and 2 LL of lysis conjugated at its C terminus to the linker sequence GGGC solution (Tris 25 mM, pH 7.5, 0.5% triton X-100) was added (SEQ ID NO:9) by synthesizing both as a single construct to three samples of each cell type without agent to generate a (i.e., SenC: SEQID NO:5). The purpose of attaching GGGC positive control maximum LDH release. LDH release was was that it can be used to chelate reduced 99m-Tc for radio measured in each sample by adding 100LL of CytoTox-ONE labeling. A 2 Laliquot of conjugated senescent cell binding Reagent (Roche Applied Science) to each well and mixing on agent (3 M) was mixed with 40 uL of 0.25 Mammonium a plate shaker for 30 seconds. Samples were incubated at 22 acetate, 15 uL of tartrate buffer pH 8.7, 4 uL of stannous C. for 10 minutes. The reaction was terminated by adding 50 chloride in 100 mM of sodium tartrate, and 30 uL of 99m-Tc uL of Stop Solution (per 100 ul, of CytoTox-ONETM Reagent pertechnetate. The mixture was heated for 25 minutes at 95° added) to each well. Fluorescence was measured in each well C. Quality control was performed with Sep-Pak and was using an excitation wavelength of 530 nm and an emission always above 90% purity. A small aliquot was also injected on wavelength of 620 nm (Cytofluor 4000). The CytoTox-ONE a Waters 600 HPLC to check the radiological profile. Frac assay was shown to yield a quantity of fluorescent product tions were collected and read on a gamma counter (Perkin that is linearly proportional to the number of cells killed Elmer Wallac Wizard 1470). (correlation coefficient=0.99). The percentage of cells killed was calculated using the following formula: Cell Binding Assay (0169. Binding of radio-labeled SenC (SEQ ID NO:5) to 100%x (P-C) senescent and non-senescent cells was tested by competition % cytotoxicity= (M - C) with unlabeled SenC. Peptide solutions were prepared to contain 0 or 20 uM of unlabeled SenC, 15 nM radio-labeled SenC, and 0.2% BSA. Chemically-induced senescent fibro where P=LDH release in wells of peptide incubated cells; blasts, their non-senescent counterparts, and prostate epithe C=LDH release in wells of cells not incubated with peptide: lial cells were prepared as above, harvested, and centrifuged. and M-LDH release in wells incubated in lysis solution. The The cell pellets were resuspended in fresh media without formula is based upon the assumptions that, in a linear rela FBS, and cells were counted. Peptide solution containing 0 or tionship between CytoTox-ONE product development and 20 uM unlabeled SenC and constant concentrations of SenC number of cells killed, C is the y-intercept, and M is due to and 10 senescent cells were combined in a final volume of 100% cell killing. 200 uL of PBS in Eppendorf tubes and incubated for 4 hours. Cells were pelleted by centrifuging at 2500 rpm for 2 minutes Effect of SenL on Cell Proliferation and washed twice with PBS and 0.2% BSA. Pellets were 0.165 Cell proliferation was assayed using the Cell Prolif suspended in 5uL PBS and transferred to 12x75 mm tubes for eration Reagent WST-1 (Roche, Mannheim, Germany) by counting radioactivity using a gamma counter (Perkin-Elmer following the manufacturers instructions. Wallac Wizard 1470). In Vitro Cytotoxicity: Conjugation to Ricin-A Other Embodiments (0166 Peptide SenC (GVYHFAPLTPTPGGGC: SEQ ID 0170 All publications and patent applications mentioned NO:5) was conjugated to ricin A subunit (Sigma-Aldrich) to in this specification are herein incorporated by reference to US 2015/O151 001 A1 Jun. 4, 2015 23 the same extent as if each independent publication or patent is intended to cover any variations, uses, or adaptations of the application was specifically and individually indicated to be invention following, in general, the principles of the invention incorporated by reference. and including Such departures from the present disclosure 0171 While the invention has been described in connec that come within known or customary practice within the art tion with specific embodiments thereof, it will be understood to which the invention pertains and may be applied to the that it is capable of further modifications and this application essential features hereinbefore set forth.

SEQUENCE LISTING

<16 Os NUMBER OF SEO ID NOS: 35

<21 Oc SEO ID NO 1 <211 LENGTH: 12 <212> TYPE PRT <213> ORGANISM: Artificial Sequence <22 Os FEATURE; OTHER INFORMATION: Synthetic Construct <4 OOs SEQUENCE: 1 Gly Val Tyr His Phe Ala Pro Leu. Thr Pro Thr Pro 1. 5 1O

SEO ID NO 2 LENGTH: 12 TYPE PRT ORGANISM: Artificial Sequence FEATURE; OTHER INFORMATION: Synthetic Construct <4 OOs SEQUENCE: 2

Ser Phe Glin Ser His Lieu. Ile Glu Phe Ser Phe Glu 1. 5 1O

SEO ID NO 3 LENGTH: 12 TYPE PRT ORGANISM: Artificial Sequence FEATURE; OTHER INFORMATION: Synthetic Construct <4 OOs SEQUENCE: 3 Ala Pro Ile Lieu Lys Lieu Ala Pro Lieu. Ile His Pro 1. 5 1O

SEO ID NO 4 LENGTH: 14 TYPE PRT ORGANISM: Artificial Sequence FEATURE; OTHER INFORMATION: Syntheitc Construct <4 OOs SEQUENCE: 4 Llys Phe Ala Lys Phe Ala Lys Llys Phe Ala Lys Phe Ala Lys 1. 5 1O

SEO ID NO 5 LENGTH: 16 TYPE PRT ORGANISM: Artificial Sequence FEATURE; OTHER INFORMATION: Synthetic Construct <4 OOs SEQUENCE: 5 Gly Val Tyr His Phe Ala Pro Leu. Thr Pro Thr Pro Gly Gly Gly Cys 1. 5 1O 15 US 2015/O151 001 A1 Jun. 4, 2015 24

- Continued

<210s, SEQ ID NO 6 &211s LENGTH: 29 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic Construct <4 OOs, SEQUENCE: 6 Gly Val Tyr His Phe Ala Pro Leu. Thr Pro Thr Pro Gly Gly Ser Lys 1. 5 1O 15 Phe Ala Lys Phe Ala Lys Llys Phe Ala Lys Phe Ala Lys 2O 25

<210s, SEQ ID NO 7 &211s LENGTH: 16 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic Construct <4 OO > SEQUENCE: 7 Gly Val Tyr His Phe Ala Pro Leu. Thr Pro Thr Pro Gly Gly Gly Cys 1. 5 1O 15

<210s, SEQ ID NO 8 &211s LENGTH: 28 212. TYPE: PRT <213> ORGANISM: Artificial Sequence & 22 O FEATURE; <223> OTHER INFORMATION: Synthetic Construct <4 OOs, SEQUENCE: 8 Gly Val Tyr His Phe Ala Pro Leu. Thr Pro Thr Pro Gly Gly Llys Phe 1. 5 1O 15 Ala Lys Phe Ala Lys Llys Phe Ala Lys Phe Ala Lys 2O 25

<210s, SEQ ID NO 9 &211s LENGTH: 4 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic Construct <4 OOs, SEQUENCE: 9 Gly Gly Gly Cys 1.

<210s, SEQ ID NO 10 &211s LENGTH: 4 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic Construct

<4 OOs, SEQUENCE: 10 Gly Gly Gly Ser 1.

<210s, SEQ ID NO 11 &211s LENGTH: 375 212. TYPE: PRT <213> ORGANISM: Homo sapiens US 2015/O151 001 A1 Jun. 4, 2015 25

- Continued

<4 OOs, SEQUENCE: 11 Met Asp Asp Asp Ile Ala Ala Lieu Val Val Asp Asn Gly Ser Gly Met 1. 5 1O 15 Cys Lys Ala Gly Phe Ala Gly Asp Asp Ala Pro Arg Ala Val Phe Pro 2O 25 3O Ser Ile Val Gly Arg Pro Arg His Glin Gly Val Met Val Gly Met Gly 35 4 O 45 Glin Lys Asp Ser Tyr Val Gly Asp Glu Ala Glin Ser Lys Arg Gly Ile SO 55 6 O Lieu. Thir Lieu Lys Tyr Pro Ile Glu. His Gly Ile Val Thr Asn Trp Asp 65 70 7s 8O Asp Met Glu Lys Ile Trp His His Thr Phe Tyr Asn Glu Lieu. Arg Val 85 90 95

Ala Pro Glu Glu. His Pro Wall Leu Lleu. Thr Glu Ala Pro Lieu. ASn Pro 1OO 105 11 O Lys Ala Asn Arg Glu Lys Met Thr Glin Ile Met Phe Glu Thr Phe Asn 115 12 O 125 Thr Pro Ala Met Tyr Val Ala Ile Glin Ala Met Leu Ser Lieu. Tyr Ala 13 O 135 14 O Ser Gly Arg Thr Thr Gly Ile Val Met Asp Ser Gly Asp Gly Val Thr 145 150 155 160 His Thr Val Pro Ile Tyr Glu Gly Tyr Ala Leu Pro His Ala Ile Leu 1.65 170 175 Arg Lieu. Asp Lieu Ala Gly Arg Asp Lieu. Thir Asp Tyr Lieu Met Lys Ile 18O 185 19 O Lieu. Thr Glu Arg Gly Tyr Ser Phe Thr Thr Thr Ala Glu Arg Glu Ile 195 2OO 2O5 Val Arg Asp Ile Lys Glu Lys Lieu. Cys Tyr Val Ala Lieu. Asp Phe Glu 21 O 215 22O Gln Glu Met Ala Thr Ala Ala Ser Ser Ser Ser Lieu. Glu Lys Ser Tyr 225 23 O 235 24 O Glu Lieu Pro Asp Gly Glin Val Ile Thir Ile Gly Asn. Glu Arg Phe Arg 245 250 255 Cys Pro Glu Ala Lieu. Phe Gln Pro Ser Phe Leu Gly Met Glu Ser Cys 26 O 265 27 O Gly Ile His Glu Thir Thr Phe Asin Ser Ile Met Lys Cys Asp Val Asp 27s 28O 285 Ile Arg Lys Asp Lieu. Tyr Asp Asn. Thr Val Lieu. Ser Gly Gly. Thir Thr 29 O 295 3 OO Met Tyr Pro Gly Ile Ala Asp Arg Met Gln Lys Glu Ile Thr Ala Lieu. 3. OS 310 315 32O

Ala Pro Ser Thr Met Lys Ile Lys Ile Ile Ala Pro Pro Glu Arg Lys 3.25 330 335

Tyr Ser Val Trp Ile Gly Gly Ser Ile Leu Ala Ser Leu Ser Thr Phe 34 O 345 35. O

Gln Glin Met Trp Ile Ser Lys Glin Glu Tyr Asp Glu Ser Gly Pro Ser 355 360 365 Ile Val His Arg Lys Cys Phe 37 O 375

<210s, SEQ ID NO 12 US 2015/O151 001 A1 Jun. 4, 2015 26

- Continued

&211s LENGTH: 360 212. TYPE: PRT <213> ORGANISM: Homo Sapiens

<4 OOs, SEQUENCE: 12 Met Cys Lys Ala Gly Phe Ala Gly Asp Asp Ala Pro Arg Ala Val Phe 1. 5 1O 15 Pro Ser Ile Val Gly Arg Pro Arg His Glin Gly Val Met Val Gly Met 2O 25 3O Gly Glin Lys Asp Ser Tyr Val Gly Asp Glu Ala Glin Ser Lys Arg Gly 35 4 O 45 Ile Lieu. Thir Lieu Lys Tyr Pro Ile Glu. His Gly Ile Val Thr Asn Trp SO 55 6 O Asp Asp Met Glu Lys Ile Trp His His Thr Phe Tyr Asn. Glu Lieu. Arg 65 70 7s 8O

Wall Ala Pro Glu Glu. His Pro Wall Lieu. Lieu. Thr Glu Ala Pro Lieu. Asn 85 90 95 Pro Lys Ala Asn Lieu. Glu Lys Met Thr Glin Ile Met Phe Glu. Thir Phe 1OO 105 11 O Asn Thr Pro Ala Met Tyr Val Ala Ile Glin Ala Val Lieu Ser Lieu. Tyr 115 12 O 125 Ala Ser Gly Arg Thir Thr Gly Ile Val Met Asp Ser Gly Asp Gly Val 13 O 135 14 O Thr His Thr Val Pro Ile Tyr Glu Gly Tyr Ala Lieu Pro His Ala Ile 145 150 155 160 Lieu. Arg Lieu. Asp Lieu Ala Gly Arg Asp Lieu. Thir Asp Tyr Lieu Met Lys 1.65 17O 17s Ile Lieu. Thr Glu Arg Gly Tyr Ser Phe Thir Thr Thr Ala Glu Arg Glu 18O 185 19 O Ile Val Arg Asp Ile Lys Glu Lys Lieu. Cys Tyr Val Ala Lieu. Asp Phe 195 2OO 2O5 Glu Glin Glu Met Ala Thr Ala Ala Ser Ser Ser Ser Lieu. Glu Lys Ser 21 O 215 22O Tyr Glu Lieu Pro Asp Gly Glin Val Ile Thr Ile Gly Asn Glu Arg Phe 225 23 O 235 24 O Arg Cys Pro Glu Ala Lieu. Phe Glin Pro Ser Phe Leu Gly Met Glu Ser 245 250 255 Cys Gly Ile His Glu Thir Thr Phe Asin Ser Ile Met Lys Cys Asp Val 26 O 265 27 O Asp Ile Arg Lys Asp Lieu. Tyr Ala Asn Thr Val Lieu. Ser Gly Gly Thr 27s 28O 285 Thr Met Tyr Pro Gly Ile Ala Asp Arg Met Gln Lys Glu Ile Thr Ala 29 O 295 3 OO

Lieu Ala Pro Ser Thr Met Lys Ile Lys Ile Ile Ala Pro Pro Glu Arg 3. OS 310 315 32O

Lys Tyr Ser Val Trp Ile Gly Gly Ser Ile Leu Ala Ser Leu Ser Thr 3.25 330 335

Phe Glin Gln Met Trp Ile Ser Lys Glin Glu Tyr Asp Glu Ser Gly Pro 34 O 345 35. O Ser Ile Val His Arg Lys Cys Phe 355 360 US 2015/O151 001 A1 Jun. 4, 2015 27

- Continued

<210s, SEQ ID NO 13 &211s LENGTH: 896 212. TYPE: PRT <213> ORGANISM: Homo Sapiens

<4 OOs, SEQUENCE: 13 Met Ala Thr Glu Glu Phe Ile Ile Arg Ile Pro Pro Tyr His Tyr Ile 1. 5 1O 15 His Val Lieu. Asp Glin Asn. Ser Asn. Wal Ser Arg Val Glu Val Gly Pro 2O 25 3O Llys Thr Tyr Ile Arg Glin Asp Asn. Glu Arg Val Lieu. Phe Ala Pro Met 35 4 O 45 Arg Met Val Thr Val Pro Pro Arg His Tyr Cys Thr Val Ala Asn Pro SO 55 6 O Val Ser Arg Asp Ala Glin Gly Lieu Val Lieu. Phe Asp Val Thr Gly Glin 65 70 7s 8O Val Arg Lieu. Arg His Ala Asp Lieu. Glu Ile Arg Lieu Ala Glin Asp Pro 85 90 95 Phe Pro Leu Tyr Pro Gly Glu Val Lieu. Glu Lys Asp Ile Thr Pro Leu 1OO 105 11 O Glin Val Val Lieu Pro Asn. Thir Ala Lieu. His Lieu Lys Ala Lieu. Lieu. Asp 115 12 O 125 Phe Glu Asp Lys Asp Gly Asp Llys Val Val Ala Gly Asp Glu Trp Lieu. 13 O 35 14 O Phe Glu Gly Pro Gly Thr Tyr Ile Pro Arg Lys Glu Val Glu Val Val 145 150 155 160 Glu Ile Ile Glin Ala Thir Ile Ile Arg Glin Asn Glin Ala Lieu. Arg Lieu. 1.65 17O 17s Arg Ala Arg Lys Glu. Cys Trp Asp Arg Asp Gly Lys Glu Arg Val Thr 18O 185 19 O Gly Glu Glu Trp Leu Val Thir Thr Val Gly Ala Tyr Lieu Pro Ala Val 195 2OO 2O5

Phe G l Glu Val Lieu. Asp Lieu Val Asp Ala Val Ile Lieu. Thr Glu Lys 21 O 215 22O Thir Ala Lieu. His Lieu. Arg Ala Arg Arg Asn. Phe Arg Asp Phe Arg Gly 225 23 O 235 24 O Val Ser Arg Arg Thr Gly Glu Glu Trp Leu Val Thr Val Glin Asp Thr 245 250 255 Glu Ala His Val Pro Asp Wal His Glu Glu Val Lieu. Gly Val Val Pro 26 O 265 27 O Ile Thir Thr Lieu. Gly Pro His Asn Tyr Cys Val Ile Lieu. Asp Pro Val 27s 28O 285 Gly Pro Asp Gly Lys Asn Gln Lieu. Gly Glin Lys Arg Val Val Lys Gly 29 O 295 3 OO

Glu Lys Ser Phe Phe Leu Gln Pro Gly Glu Gln Lieu. Glu Glin Gly Ile 3. OS 310 315 32O

Glin Asp Val Tyr Val Lieu. Ser Glu Glin Glin Gly Lieu. Lieu. Lieu. Arg Ala 3.25 330 335 Lieu. Glin Pro Lieu. Glu Glu Gly Glu Asp Glu Glu, Llys Val Ser His Glin 34 O 345 35. O Ala Gly Asp His Trp Lieu. Ile Arg Gly Pro Lieu. Glu Tyr Val Pro Ser 355 360 365 US 2015/O151 001 A1 Jun. 4, 2015 28

- Continued

Ala Lys Val Glu Val Val Glu Glu Arg Glin Ala Ile Pro Lieu. Asp Glu 37 O 375 38O Asn Glu Gly Ile Tyr Val Glin Asp Wall Lys Thr Gly Llys Val Arg Ala 385 390 395 4 OO Val Ile Gly Ser Thr Tyr Met Lieu. Thr Glin Asp Glu Val Lieu. Trp Glu 4 OS 41O 415 Lys Glu Lieu Pro Pro Gly Val Glu Glu Lieu. Lieu. Asn Lys Gly Glin Asp 42O 425 43 O Pro Lieu Ala Asp Arg Gly Glu Lys Asp Thir Ala Lys Ser Lieu. Glin Pro 435 44 O 445 Lieu Ala Pro Arg Asn Llys Thr Arg Val Val Ser Tyr Arg Val Pro His 450 45.5 460 Asn Ala Ala Val Glin Val Tyr Asp Tyr Arg Glu Lys Arg Ala Arg Val 465 470 47s 48O Val Phe Gly Pro Glu Lieu Val Ser Leu Gly Pro Glu Glu Glin Phe Thr 485 490 495 Val Lieu. Ser Lieu. Ser Ala Gly Arg Pro Lys Arg Pro His Ala Arg Arg SOO 505 51O Ala Lieu. Cys Lieu Lleu Lieu. Gly Pro Asp Phe Phe Thr Asp Val Ile Thr 515 52O 525 Ile Glu Thir Ala Asp His Ala Arg Lieu. Glin Lieu Gln Lieu Ala Tyr Asn 53 O 535 54 O Trp His Phe Glu Val Asn Asp Arg Lys Asp Pro Gln Glu Thir Ala Lys 5.45 550 555 560 Lieu. Phe Ser Val Pro Asp Phe Val Gly Asp Ala Cys Lys Ala Ile Ala 565 st O sts Ser Arg Val Arg Gly Ala Val Ala Ser Val Thr Phe Asp Asp Phe His 58O 585 59 O Lys Asn. Ser Ala Arg Ile Ile Arg Thr Ala Val Phe Gly Phe Glu Thir 595 6OO 605 Ser Glu Ala Lys Gly Pro Asp Gly Met Ala Lieu Pro Arg Pro Arg Asp 610 615 62O Glin Ala Val Phe Pro Glin Asn Gly Lieu Val Val Ser Ser Val Asp Val 625 630 635 64 O Glin Ser Val Glu Pro Val Asp Glin Arg Thr Arg Asp Ala Lieu. Glin Arg 645 650 655

Ser Wall Glin Lieu. Ala Ile Glu Ile Thir Thr Asn. Ser Glin Glu Ala Ala 660 665 67 O Ala Lys His Glu Ala Glin Arg Lieu. Glu Glin Glu Ala Arg Gly Arg Lieu 675 68O 685 Glu Arg Gln Lys Ile Lieu. Asp Glin Ser Glu Ala Glu Lys Ala Arg Llys 69 O. 695 7 OO

Glu Lieu. Lieu. Glu Lieu. Glu Ala Lieu. Ser Met Ala Val Glu Ser Thr Gly 7 Os 71O 71s 72O

Thir Ala Lys Ala Glu Ala Glu Ser Arg Ala Glu Ala Ala Arg Ile Glu 72 73 O 73

Gly Glu Gly Ser Val Lieu. Glin Ala Lys Lieu Lys Ala Glin Ala Lieu Ala 740 74. 7 O

Ile Glu Thr Glu Ala Glu Lieu. Glin Arg Val Glin Llys Val Arg Glu Lieu. 7ss 760 765

Glu Lieu Val Tyr Ala Arg Ala Glin Lieu. Glu Lieu. Glu Val Ser Lys Ala US 2015/O151 001 A1 Jun. 4, 2015 29

- Continued

770 775

Glin Gln Lieu. Ala Glu Wall Glu Wall Phe Glin Met Thir Glu 78s 79 O 79.

Ala Ile Gly Pro Ser Thr Ile Arg Asp Luell Ala Wall Ala Gly Pro Glu 805 810 815

Met Glin Val Lys Lieu. Lieu. Glin Ser Luell Gly Luell Ser Thir Luell Ile 82O 825 83 O

Thir Asp Gly Ser Thr Pro Ile Asn Luell Phe ASn Thir Ala Phe Gly Luell 835 84 O 845

Lell Gly Met Gly Pro Glu Gly Glin Pro Luell Gly Arg Arg Wall Pro Wall 850 855 860

Ala Gln Pro Trp Gly Gly Asp Ile Pro Pro Wall Ser Gly Pro Ser 865 87O

Ser Ser Trp Arg Glin Pro Arg Gly Ala Cys Thir Ala Lell Thir Pro Asp 885 890 895

<210s, SEQ ID NO 14 &211s LENGTH: 893 212. TYPE: PRT <213> ORGANISM: Homo Sapiens <4 OOs, SEQUENCE: 14

Met Ala Thr Glu Glu Phe Ile Ile Arg Ile Pro Pro His Tyr Ile 1. 5 1O 15

His Val Lieu. Asp Glin Asn. Ser Asn Wall Ser Arg Wall Glu Wall Gly Pro 2O 25

Thir Tyr Ile Arg Glin Asp Asn Glu Arg Wall Lell Phe Ala Pro Met 35 4 O 45

Arg Met Val Thr Val Pro Pro Arg His Thir Wall Ala Asn Pro SO 55 6 O

Wall Ser Arg Asp Ala Glin Gly Lieu. Wall Luell Phe Asp Wall Thir Gly Glin 65 70

Wall Arg Lieu. Arg His Ala Asp Lieu Glu Ile Arg Lell Ala Glin Asp Pro 85 90 95

Phe Pro Leu Tyr Pro Gly Glu Val Luell Glu Lys Asp Ile Thir Pro Luell 1OO 105 11 O

Glin Wall Wall Leu Pro Asn. Thir Ala Luell His Luell Ala Luell Luell Asp 115 12 O 125

Phe Glu Asp Lys Asp Gly Asp Llys Wall Wall Ala Gly Asp Glu Trp Luell 13 O 35 14 O

Phe Glu Gly Pro Gly Thr Tyr Ile Pro Arg Lys Glu Wall Glu Wall Wall 145 150 155 160

Glu Ile Ile Glin Ala Thir Ile Ile Arg Glin ASn Glin Ala Luell Arg Luell 1.65 17O

Arg Ala Arg Lys Glu. Cys rp Asp Arg Asp Gly Glu Arg Wall Thir 18O 185 19 O

Gly Glu Glu Trp Leu Val Thr Thr Wall Gly Ala Lell Pro Ala Wall 195 2OO

Phe Glu Glu Val Lieu. Asp Lieu Val Asp Ala Wall Ile Lell Thir Glu 2 O 215 22O

Thir Ala Lieu. His Lieu. Arg Ala Arg Arg Asn Phe Arg Asp Phe Arg Gly 225 23 O 235 24 O US 2015/O151 001 A1 Jun. 4, 2015 30

- Continued Val Ser Arg Arg Thr Gly Glu Glu Trp Leu Val Thr Val Glin Asp Thr 245 250 255 Glu Ala His Val Pro Asp Wal His Glu Glu Val Lieu. Gly Val Val Pro 26 O 265 27 O Ile Thir Thr Lieu. Gly Pro His Asn Tyr Cys Val Ile Lieu. Asp Pro Val 27s 28O 285 Gly Pro Asp Gly Lys Asn Gln Lieu. Gly Glin Lys Arg Val Val Lys Gly 29 O 295 3 OO Glu Lys Ser Phe Phe Leu Gln Pro Gly Glu Gln Lieu. Glu Glin Gly Ile 3. OS 310 315 32O Glin Asp Val Tyr Val Lieu. Ser Glu Glin Glin Gly Lieu. Lieu. Lieu. Arg Ala 3.25 330 335 Lieu. Glin Pro Lieu. Glu Glu Gly Glu Asp Glu Glu, Llys Val Ser His Glin 34 O 345 35. O Ala Gly Asp His Trp Lieu. Ile Arg Gly Pro Lieu. Glu Tyr Val Pro Ser 355 360 365 Ala Lys Val Glu Val Val Glu Glu Arg Glin Ala Ile Pro Lieu. Asp Glu 37 O 375 38O Asn Glu Gly Ile Tyr Val Glin Asp Wall Lys Thr Gly Llys Val Arg Ala 385 390 395 4 OO Val Ile Gly Ser Thr Tyr Met Lieu. Thr Glin Asp Glu Val Lieu. Trp Glu 4 OS 41O 415 Lys Glu Lieu Pro Pro Gly Val Glu Glu Lieu Lieu. ASn Lys Gly Glin Asp 42O 425 43 O Pro Lieu Ala Asp Arg Gly Glu Lys Asp Thir Ala Lys Ser Lieu. Glin Pro 435 44 O 445 Lieu Ala Pro Arg Asn Llys Thr Arg Val Val Ser Tyr Arg Val Pro His 450 45.5 460 Asn Ala Ala Val Glin Val Tyr Asp Tyr Arg Glu Lys Arg Ala Arg Val 465 470 47s 48O Val Phe Gly Pro Glu Lieu Val Ser Leu Gly Pro Glu Glu Glin Phe Thr 485 490 495 Val Lieu. Ser Lieu. Ser Ala Gly Arg Pro Lys Arg Pro His Ala Arg Arg SOO 505 51O Ala Lieu. Cys Lieu Lleu Lieu. Gly Pro Asp Phe Phe Thr Asp Val Ile Thr 515 52O 525 Ile Glu Thir Ala Asp His Ala Arg Lieu. Glin Lieu Gln Lieu Ala Tyr Asn 53 O 535 54 O Trp His Phe Glu Val Asn Asp Arg Lys Asp Pro Glin Glu Thir Ala Lys 5.45 550 555 560 Lieu. Phe Ser Val Pro Asp Phe Val Gly Asp Ala Cys Lys Ala Ile Ala 565 st O sts Ser Arg Val Arg Gly Ala Val Ala Ser Val Thr Phe Asp Asp Phe His 58O 585 59 O

Lys Asn. Ser Ala Arg Ile Ile Arg Thr Ala Val Phe Gly Phe Glu Thir 595 6OO 605

Ser Glu Ala Lys Gly Pro Asp Gly Met Ala Lieu Pro Arg Pro Arg Asp 610 615 62O

Glin Ala Val Phe Pro Glin Asn Gly Lieu Val Val Ser Ser Val Asp Val 625 630 635 64 O

Glin Ser Val Glu Pro Val Asp Glin Arg Thr Arg Asp Ala Lieu. Glin Arg US 2015/O151 001 A1 Jun. 4, 2015 31

- Continued

645 650 655

Ser Wall Glin Lieu. Ala Ile Glu Ile Thir Thr Asn. Ser Glin Glu Ala Ala 660 665 67 O Ala Lys His Glu Ala Glin Arg Lieu. Glu Glin Glu Ala Arg Gly Arg Lieu 675 68O 685 Glu Arg Gln Lys Ile Lieu. Asp Glin Ser Glu Ala Glu Lys Ala Arg Llys 69 O. 695 7 OO Glu Lieu. Lieu. Glu Lieu. Glu Ala Lieu. Ser Met Ala Val Glu Ser Thr Gly 7 Os 71O 71s 72O Thir Ala Lys Ala Glu Ala Glu Ser Arg Ala Glu Ala Ala Arg Ile Glu 72 73 O 73 Gly Glu Gly Ser Val Lieu. Glin Ala Lys Lieu Lys Ala Glin Ala Lieu Ala 740 74. 7 O Ile Glu Thr Glu Ala Glu Lieu. Glin Arg Val Glin Llys Val Arg Glu Lieu. 7ss 760 765 Glu Lieu Val Tyr Ala Arg Ala Glin Lieu. Glu Lieu. Glu Val Ser Lys Ala 770 775 78O Glin Glin Lieu Ala Glu Val Glu Val Llys Llys Phe Lys Gln Met Thr Glu 78s 79 O 79. 8OO Ala Ile Gly Pro Ser Thir Ile Arg Asp Lieu Ala Val Ala Gly Pro Glu 805 810 815 Met Glin Val Lys Lieu. Lieu. Glin Ser Lieu. Gly Lieu Lys Ser Thr Lieu. Ile 82O 825 83 O Thr Asp Gly Ser Thr Pro Ile Asn Lieu Phe Asn Thr Ala Phe Gly Lieu. 835 84 O 845 Lieu. Gly Met Gly Pro Glu Gly Glin Pro Lieu. Gly Arg Arg Val Ala Ser 850 855 860 Gly Pro Ser Pro Gly Glu Gly Ile Ser Pro Glin Ser Ala Glin Ala Pro 865 87O 87s 88O Glin Ala Pro Gly Asp Asn His Val Val Pro Val Lieu. Arg 885 890

<210s, SEQ ID NO 15 &211s LENGTH: 508 212. TYPE: PRT <213> ORGANISM: Homo Sapiens <4 OOs, SEQUENCE: 15 Met Lieu. Arg Arg Ala Lieu. Lieu. Cys Lieu Ala Val Ala Ala Lieu Val Arg 1. 5 1O 15 Ala Asp Ala Pro Glu Glu Glu Asp His Val Lieu Val Lieu. Arg Llys Ser 2O 25 3O Asn Phe Ala Glu Ala Lieu Ala Ala His Llys Tyr Lieu. Lieu Val Glu Phe 35 4 O 45 Tyr Ala Pro Trp Cys Gly His Cys Lys Ala Lieu Ala Pro Glu Tyr Ala SO 55 6 O

Lys Ala Ala Gly Llys Lieu Lys Ala Glu Gly Ser Glu Ile Arg Lieu Ala 65 70 7s 8O

Llys Val Asp Ala Thr Glu Glu Ser Asp Lieu Ala Glin Glin Tyr Gly Val 85 90 95

Arg Gly Tyr Pro Thr Ile Llys Phe Phe Arg Asn Gly Asp Thr Ala Ser 1OO 105 11 O US 2015/O151 001 A1 Jun. 4, 2015 32

- Continued Pro Lys Glu Tyr Thr Ala Gly Arg Glu Ala Asp Asp Ile Val Asn Trp 115 12 O 125 Lieu Lys Lys Arg Thr Gly Pro Ala Ala Thir Thr Lieu. Arg Asp Gly Ala 13 O 135 14 O Ala Ala Glu Ser Lieu Val Glu Ser Ser Glu Val Ala Val Ile Gly Phe 145 150 155 160 Phe Lys Asp Val Glu Ser Asp Ser Ala Lys Glin Phe Lieu. Glin Ala Ala 1.65 17O 17s Glu Ala Ile Asp Asp Ile Pro Phe Gly Ile Thir Ser Asn. Ser Asp Wall 18O 185 19 O Phe Ser Lys Tyr Glin Lieu. Asp Lys Asp Gly Val Val Lieu. Phe Llys Llys 195 2OO 2O5 Phe Asp Glu Gly Arg Asn. Asn. Phe Glu Gly Glu Val Thir Lys Glu Asn 21 O 215 22O Lieu. Lieu. Asp Phe Ile Llys His Asn Glin Lieu Pro Lieu Val Ile Glu Phe 225 23 O 235 24 O Thr Glu Gln Thr Ala Pro Lys Ile Phe Gly Gly Glu Ile Llys Thr His 245 250 255 Ile Lieu. Lieu. Phe Lieu Pro Llys Ser Val Ser Asp Tyr Asp Gly Lys Lieu. 26 O 265 27 O Ser Asn. Phe Llys Thr Ala Ala Glu Ser Phe Lys Gly Lys Ile Lieu. Phe 27s 28O 285 Ile Phe Ile Asp Ser Asp His Thr Asp ASn Glin Arg Ile Lieu. Glu Phe 29 O 295 3 OO Phe Gly Lieu Lys Lys Glu Glu. Cys Pro Ala Val Arg Lieu. Ile Thr Lieu. 3. OS 310 315 32O Glu Glu Glu Met Thr Lys Tyr Llys Pro Glu Ser Glu Glu Lieu. Thir Ala 3.25 330 335 Glu Arg Ile Thr Glu Phe Cys His Arg Phe Lieu. Glu Gly Lys Ile Llys 34 O 345 35. O Pro His Lieu Met Ser Glin Glu Arg Ala Gly Asp Trp Asp Llys Glin Pro 355 360 365 Val Llys Val Pro Val Gly Lys Asn. Phe Glu Asp Wall Ala Phe Asp Glu 37 O 375 38O Lys Lys Asn Val Phe Val Glu Phe Tyr Ala Pro Trp Cys Gly His Cys 385 390 395 4 OO Lys Glin Lieu Ala Pro Ile Trp Asp Llys Lieu. Gly Glu Thir Tyr Lys Asp 4 OS 41O 415 His Glu Asn. Ile Val Ile Ala Lys Met Asp Ser Thr Ala Asn. Glu Val 42O 425 43 O Glu Ala Val Llys Val His Ser Phe Pro Thr Lieu Lys Phe Phe Pro Ala 435 44 O 445

Ser Ala Asp Arg Thr Val Ile Asp Tyr Asn Gly Glu Arg Thir Lieu. Asp 450 45.5 460

Gly Phe Llys Llys Phe Lieu. Glu Ser Gly Gly Glin Asp Gly Ala Gly Asp 465 470 47s 48O

Asp Asp Asp Lieu. Glu Asp Lieu. Glu Glu Ala Glu Glu Pro Asp Met Glu 485 490 495

Glu Asp Asp Asp Gln Lys Ala Val Lys Asp Glu Lieu. SOO 505 US 2015/O151 001 A1 Jun. 4, 2015 33

- Continued

<210s, SEQ ID NO 16 &211s LENGTH: 508 212. TYPE: PRT <213> ORGANISM: Homo sapiens

<4 OOs, SEQUENCE: 16 Met Lieu. Arg Arg Ala Lieu. Lieu. Cys Lieu Ala Val Ala Ala Lieu Val Arg 1. 5 1O 15 Ala Asp Ala Pro Glu Glu Glu Asp His Val Lieu Val Lieu. Arg Llys Ser 2O 25 3O Asn Phe Ala Glu Ala Lieu Ala Ala His Llys Tyr Lieu. Lieu Val Glu Phe 35 4 O 45 Tyr Ala Pro Trp Cys Gly His Cys Lys Ala Lieu Ala Pro Glu Tyr Ala SO 55 6 O Lys Ala Ala Gly Llys Lieu Lys Ala Glu Gly Ser Glu Ile Arg Lieu Ala 65 70 7s 8O Llys Val Asp Ala Thr Glu Glu Ser Asp Lieu Ala Glin Glin Tyr Gly Val 85 90 95 Arg Gly Tyr Pro Thr Ile Llys Phe Phe Arg Asn Gly Asp Thr Ala Ser 1OO 105 11 O Pro Lys Glu Tyr Thr Ala Gly Arg Glu Ala Asp Asp Ile Val Asn Trp 115 12 O 125 Lieu Lys Lys Arg Thr Gly Pro Ala Ala Thir Thr Lieu Pro Asp Gly Ala 13 O 135 14 O Ala Ala Glu Ser Lieu Val Glu Ser Ser Glu Val Ala Val Ile Gly Phe 145 150 155 160 Phe Lys Asp Val Glu Ser Asp Ser Ala Lys Glin Phe Lieu. Glin Ala Ala 1.65 17O 17s Glu Ala Ile Asp Asp Ile Pro Phe Gly Ile Thir Ser Asn. Ser Asp Wall 18O 185 19 O Phe Ser Lys Tyr Glin Lieu. Asp Lys Asp Gly Val Val Lieu. Phe Llys Llys 195 2OO 2O5 Phe Asp Glu Gly Arg Asn. Asn. Phe Glu Gly Glu Val Thir Lys Glu Asn 21 O 215 22O Lieu. Lieu. Asp Phe Ile Llys His Asn Glin Lieu Pro Lieu Val Ile Glu Phe 225 23 O 235 24 O Thr Glu Gln Thr Ala Pro Lys Ile Phe Gly Gly Glu Ile Llys Thr His 245 250 255 Ile Lieu. Lieu. Phe Lieu Pro Llys Ser Val Ser Asp Tyr Asp Gly Lys Lieu. 26 O 265 27 O Ser Asn. Phe Llys Thr Ala Ala Glu Ser Phe Lys Gly Lys Ile Lieu. Phe 27s 28O 285 Ile Phe Ile Asp Ser Asp His Thir Asp Asn Glin Arg Ile Lieu. Glu Phe 29 O 295 3 OO

Phe Gly Lieu Lys Lys Glu Glu. Cys Pro Ala Val Arg Lieu. Ile Thr Lieu. 3. OS 310 315 32O

Glu Glu Glu Met Thr Lys Tyr Llys Pro Glu Ser Glu Glu Lieu. Thir Ala 3.25 330 335 Glu Arg Ile Thr Glu Phe Cys His Arg Phe Lieu. Glu Gly Lys Ile Llys 34 O 345 35. O Pro His Lieu Met Ser Glin Glu Lieu Pro Glu Asp Trp Asp Llys Glin Pro 355 360 365 US 2015/O151 001 A1 Jun. 4, 2015 34

- Continued Val Llys Val Lieu Val Gly Lys Asn. Phe Glu Asp Wall Ala Phe Asp Glu 37 O 375 38O Lys Lys Asn Val Phe Val Glu Phe Tyr Ala Pro Trp Cys Gly His Cys 385 390 395 4 OO Lys Glin Lieu Ala Pro Ile Trp Asp Llys Lieu. Gly Glu Thir Tyr Lys Asp 4 OS 41O 415 His Glu Asn. Ile Val Ile Ala Lys Met Asp Ser Thr Ala Asn. Glu Val 42O 425 43 O Glu Ala Val Llys Val His Ser Phe Pro Thr Lieu Lys Phe Phe Pro Ala 435 44 O 445 Ser Ala Asp Arg Thr Val Ile Asp Tyr Asn Gly Glu Arg Thir Lieu. Asp 450 45.5 460 Gly Phe Llys Llys Phe Lieu. Glu Ser Gly Gly Glin Asp Gly Ala Gly Asp 465 470 47s 48O Asp Asp Asp Lieu. Glu Asp Lieu. Glu Glu Ala Glu Glu Pro Asp Met Glu 485 490 495 Glu Asp Asp Asp Gln Lys Ala Val Lys Asp Glu Lieu. SOO 505

<210s, SEQ ID NO 17 &211s LENGTH: 120 212. TYPE: PRT <213> ORGANISM: Homo Sapiens < 4 OO SEQUENCE: 17 Asp Ala Pro Glu Glu Glu Asp His Val Lieu Val Lieu. Arg Llys Ser Asn 1. 5 1O 15 Phe Ala Glu Ala Lieu Ala Ala His Llys Tyr Lieu. Lieu Val Glu Phe Tyr 2O 25 3O Ala Pro Trp Cys Gly. His Cys Lys Ala Lieu Ala Pro Glu Tyr Ala Lys 35 4 O 45 Ala Ala Gly Lys Lieu Lys Ala Glu Gly Ser Glu Ile Arg Lieu Ala Lys SO 55 6 O Val Asp Ala Thr Glu Glu Ser Asp Lieu Ala Glin Glin Tyr Gly Val Arg 65 70 7s 8O Gly Tyr Pro Thr Ile Llys Phe Phe Arg Asn Gly Asp Thr Ala Ser Pro 85 90 95 Lys Glu Tyr Thir Ala Gly Arg Glu Ala Asp Asp Ile Val Asn Trp Lieu. 1OO 105 11 O Llys Lys Arg Thr Gly Pro Ala Ala 115 12 O

<210s, SEQ ID NO 18 &211s LENGTH: 255 212. TYPE: PRT <213> ORGANISM: Homo Sapiens

<4 OOs, SEQUENCE: 18 Met Ala Glu Lieu. Arg Val Lieu Val Ala Wall Lys Arg Val Ile Asp Tyr 1. 5 1O 15

Ala Wall Lys Ile Arg Val Llys Pro Asp Arg Thr Gly Val Val Thir Asp 2O 25 3O Gly Val Llys His Ser Met Asn Pro Phe Cys Glu Ile Ala Val Glu Glu 35 4 O 45 US 2015/O151 001 A1 Jun. 4, 2015 35

- Continued Ala Val Arg Lieu Lys Glu Lys Llys Lieu Val Lys Glu Val Ile Ala Val SO 55 6 O Ser Cys Gly Pro Ala Glin Cys Glin Glu Thir Ile Arg Thr Ala Lieu Ala 65 70 7s 8O Met Gly Ala Asp Arg Gly Ile His Val Glu Val Pro Pro Ala Glu Ala 85 90 95 Glu Arg Lieu. Gly Pro Lieu. Glin Val Ala Arg Val Lieu Ala Lys Lieu Ala 1OO 105 11 O Glu Lys Glu Lys Val Asp Lieu Val Lieu. Lieu. Gly Lys Glin Ala Ile Asp 115 12 O 125 Asp Asp Cys Asn Glin Thr Gly Glin Met Thr Ala Gly Phe Lieu. Asp Trip 13 O 135 14 O Pro Glin Gly Thr Phe Ala Ser Glin Val Thr Lieu. Glu Gly Asp Llys Lieu. 145 150 155 160 Llys Val Glu Arg Glu Ile Asp Gly Gly Lieu. Glu Thir Lieu. Arg Lieu Lys 1.65 17O 17s Lieu Pro Ala Val Val Thr Ala Asp Lieu. Arg Lieu. Asn. Glu Pro Arg Tyr 18O 185 19 O Ala Thr Lieu Pro Asn. Ile Met Lys Ala Lys Llys Llys Lys Ile Glu Val 195 2OO 2O5 Ile Llys Pro Gly Asp Lieu. Gly Val Asp Lieu. Thir Ser Llys Lieu. Ser Val 21 O 215 22O Ile Ser Val Glu Asp Pro Pro Glin Arg Thr Ala Gly Val Llys Val Glu 225 23 O 235 24 O Thir Thr Glu Asp Lieu Val Ala Lys Lieu Lys Glu Ile Gly Arg Ile 245 250 255

<210s, SEQ ID NO 19 &211s LENGTH: 553 212. TYPE: PRT <213> ORGANISM: Homo sapiens <4 OOs, SEQUENCE: 19 Met Lieu. Ser Val Arg Val Ala Ala Ala Val Val Arg Ala Lieu Pro Arg 1. 5 1O 15 Arg Ala Gly Lieu Val Ser Arg Asn Ala Lieu. Gly Ser Ser Phe Ile Ala 2O 25 3O Ala Arg Asn Phe His Ala Ser Asn Thr His Leu Gln Lys Thr Gly Thr 35 4 O 45 Ala Glu Met Ser Ser Ile Lieu. Glu Glu Arg Ile Lieu. Gly Ala Asp Thr SO 55 6 O Ser Val Asp Lieu. Glu Glu Thr Gly Arg Val Lieu. Ser Ile Gly Asp Gly 65 70 7s 8O Ile Ala Arg Val His Gly Lieu. Arg Asin Val Glin Ala Glu Glu Met Val 85 90 95

Glu Phe Ser Ser Gly Lieu Lys Gly Met Ser Lieu. Asn Lieu. Glu Pro Asp 1OO 105 11 O

Asn Val Gly Val Val Val Phe Gly Asn Asp Llys Lieu. Ile Lys Glu Gly 115 12 O 125

Asp Ile Val Lys Arg Thr Gly Ala Ile Val Asp Val Pro Val Gly Glu 13 O 135 14 O

Glu Lieu. Lieu. Gly Arg Val Val Asp Ala Lieu. Gly Asn Ala Ile Asp Gly 145 150 155 160 US 2015/O151 001 A1 Jun. 4, 2015 36

- Continued

Lys Gly Pro Ile Gly Ser Llys Thr Arg Arg Arg Val Gly Lieu Lys Ala 1.65 17O 17s Pro Gly Ile Ile Pro Arg Ile Ser Val Arg Glu Pro Met Glin Thr Gly 18O 185 19 O Ile Lys Ala Val Asp Ser Lieu Val Pro Ile Gly Arg Gly Glin Arg Glu 195 2OO 2O5 Lieu. Ile Ile Gly Asp Arg Glin Thr Gly Lys Thir Ser Ile Ala Ile Asp 21 O 215 22O Thir Ile Ile Asin Glin Lys Arg Phe Asn Asp Gly Ser Asp Glu Lys Llys 225 23 O 235 24 O Llys Lieu. Tyr Cys Ile Tyr Val Ala Ile Gly Glin Lys Arg Ser Thr Val 245 250 255 Ala Glin Lieu Val Lys Arg Lieu. Thir Asp Ala Asp Ala Met Lys Tyr Thr 26 O 265 27 O Ile Val Val Ser Ala Thr Ala Ser Asp Ala Ala Pro Lieu. Glin Tyr Lieu. 27s 28O 285 Ala Pro Tyr Ser Gly Cys Ser Met Gly Glu Tyr Phe Arg Asp Asn Gly 29 O 295 3 OO Llys His Ala Lieu. Ile Ile Tyr Asp Asp Lieu. Ser Lys Glin Ala Val Ala 3. OS 310 315 32O Tyr Arg Gln Met Ser Lieu. Lieu. Lieu. Arg Arg Pro Pro Gly Arg Glu Ala 3.25 330 335 Tyr Pro Gly Asp Val Phe Tyr Lieu. His Ser Arg Lieu. Lieu. Glu Arg Ala 34 O 345 35. O Ala Lys Met Asn Asp Ala Phe Gly Gly Gly Ser Lieu. Thir Ala Lieu Pro 355 360 365 Val Ile Glu Thr Glin Ala Gly Asp Val Ser Ala Tyr Ile Pro Thr Asn 37 O 375 38O Val Ile Ser Ile Thr Asp Gly Glin Ile Phe Leu Glu Thr Glu Lieu. Phe 385 390 395 4 OO Tyr Lys Gly Ile Arg Pro Ala Ile Asin Val Gly Lieu. Ser Val Ser Arg 4 OS 41O 415 Val Gly Ser Ala Ala Glin Thr Arg Ala Met Lys Glin Val Ala Gly Thr 42O 425 43 O Met Lys Lieu. Glu Lieu Ala Glin Tyr Arg Glu Val Ala Ala Phe Ala Glin 435 44 O 445 Phe Gly Ser Asp Lieu. Asp Ala Ala Thr Glin Gln Lieu Lleu Ser Arg Gly 450 45.5 460 Val Arg Lieu. Thr Glu Lieu Lleu Lys Glin Gly Glin Tyr Ser Pro Met Ala 465 470 47s 48O Ile Glu Glu Glin Val Ala Val Ile Tyr Ala Gly Val Arg Gly Tyr Lieu. 485 490 495

Asp Llys Lieu. Glu Pro Ser Lys Ile Thr Llys Phe Glu Asn Ala Phe Lieu SOO 505 51O Ser His Val Val Ser Glin His Glin Ala Lieu. Lieu. Gly. Thir Ile Arg Ala 515 52O 525

Asp Gly Lys Ile Ser Glu Glin Ser Asp Ala Lys Lieu Lys Glu Ile Val 53 O 535 54 O

Thir Asn. Phe Lieu Ala Gly Phe Glu Ala 5.45 550 US 2015/O151 001 A1 Jun. 4, 2015 37

- Continued

<210s, SEQ ID NO 2 O &211s LENGTH: 508 212. TYPE: PRT <213> ORGANISM: Homo sapiens 22 Os. FEATURE: <221 > NAMEAKEY: misc feature <222s. LOCATION: (12) ... (12) <223> OTHER INFORMATION: Xaa can be any naturally occurring amino acid <4 OOs, SEQUENCE: 2O Met Lieu. Arg Arg Ala Lieu. Lieu. Cys Lieu Pro Trp Xaa Ala Lieu Val Arg 1. 5 1O 15 Ala Asp Ala Pro Glu Glu Glu Asp His Val Lieu Val Lieu. Arg Llys Ser 2O 25 3O Asn Phe Ala Glu Ala Lieu Ala Ala His Llys Tyr Pro Pro Val Glu Phe 35 4 O 45 His Ala Pro Trp Cys Gly. His Cys Lys Ala Lieu Ala Pro Glu Tyr Ala SO 55 6 O Lys Ala Ala Gly Llys Lieu Lys Ala Glu Gly Ser Glu Ile Arg Lieu Ala 65 70 7s 8O Llys Val Asp Ala Thr Glu Glu Ser Asp Lieu Ala Glin Glin Tyr Gly Val 85 90 95 Arg Gly Tyr Pro Thr Ile Llys Phe Phe Arg Asn Gly Asp Thr Ala Ser 1OO 105 11 O Pro Llys Glu Tyr Thr Ala Gly Arg Glu Ala Asp Asp Ile Val ASn Trp 115 12 O 125 Lieu Lys Lys Arg Thr Gly Pro Ala Ala Thir Thr Lieu Pro Asp Gly Ala 13 O 135 14 O Ala Ala Glu Ser Lieu Val Glu Ser Ser Glu Val Ala Val Ile Gly Phe 145 150 155 160 Phe Lys Asp Val Glu Ser Asp Ser Ala Lys Glin Phe Lieu. Glin Ala Ala 1.65 17O 17s Glu Ala Ile Asp Asp Ile Pro Phe Gly Ile Thir Ser Asn. Ser Asp Wall 18O 185 19 O Phe Ser Lys Tyr Glin Lieu. Asp Lys Asp Gly Val Val Lieu. Phe Llys Llys 195 2OO 2O5 Phe Asp Glu Gly Arg Asn. Asn. Phe Glu Gly Glu Val Thir Lys Glu Asn 21 O 215 22O Lieu. Lieu. Asp Phe Ile Llys His Asn Glin Lieu Pro Lieu Val Ile Glu Phe 225 23 O 235 24 O Thr Glu Gln Thr Ala Pro Lys Ile Phe Gly Gly Glu Ile Llys Thr His 245 250 255 Ile Lieu. Lieu. Phe Lieu Pro Llys Ser Val Ser Asp Tyr Asp Gly Lys Lieu. 26 O 265 27 O

Ser Asn. Phe Llys Thr Ala Ala Glu Ser Phe Lys Gly Lys Ile Lieu. Phe 27s 28O 285

Ile Phe Ile Asp Ser Asp His Thir Asp Asn Glin Arg Ile Lieu. Glu Phe 29 O 295 3 OO

Phe Gly Lieu Lys Lys Glu Glu. Cys Pro Ala Val Arg Lieu. Ile Thr Lieu. 3. OS 310 315 32O

Glu Glu Glu Met Thr Lys Tyr Llys Pro Glu Ser Glu Glu Lieu. Thir Ala 3.25 330 335 Glu Arg Ile Thr Glu Phe Cys His Arg Phe Lieu. Glu Gly Lys Ile Llys US 2015/O151 001 A1 Jun. 4, 2015 38

- Continued

34 O 345 35. O Pro His Lieu Met Ser Glin Glu Lieu Pro Glu Asp Trp Asp Llys Glin Pro 355 360 365 Val Llys Val Lieu Val Gly Lys Asn. Phe Glu Asp Wall Ala Phe Asp Glu 37 O 375 38O Lys Lys Asn Val Phe Val Glu Phe Tyr Ala Pro Trp Cys Gly His Cys 385 390 395 4 OO Lys Glin Lieu Ala Pro Ile Trp Asp Llys Lieu. Gly Glu Thir Tyr Lys Asp 4 OS 41O 415 His Glu Asn. Ile Val Ile Ala Lys Met Asp Ser Thr Ala Asn. Glu Val 42O 425 43 O Glu Ala Val Llys Val His Gly Phe Pro Thr Lieu. Gly Phe Phe Pro Ala 435 44 O 445 Ser Ala Asp Arg Thr Val Ile Asp Tyr Asn Gly Glu Arg Thir Lieu. Asp 450 45.5 460 Gly Phe Llys Llys Phe Lieu. Glu Ser Gly Gly Glin Asp Gly Ala Gly Asp 465 470 47s 48O Val Asp Asp Lieu. Glu Asp Lieu. Glu Glu Ala Glu Glu Pro Asp Met Glu 485 490 495 Glu Asp Asp Asp Gln Lys Ala Val Lys Asp Glu Lieu. SOO 505

<210 SEQ ID NO 21 &211s LENGTH: 255 212. TYPE: PRT <213> ORGANISM: Homo sapiens <4 OOs, SEQUENCE: 21 Met Ala Glu Lieu. Arg Val Lieu Val Ala Wall Lys Arg Val Ile Asp Tyr 1. 5 1O 15 Ala Wall Lys Ile Arg Val Llys Pro Asp Arg Thr Gly Val Val Thir Asp 2O 25 3O Gly Val Llys His Ser Met Asn Pro Phe Cys Glu Ile Ala Val Glu Glu 35 4 O 45 Ala Val Arg Lieu Lys Glu Lys Llys Lieu Val Lys Glu Val Ile Ala Val SO 55 6 O Ser Cys Gly Pro Ala Glin Cys Glin Glu Thir Ile Arg Thr Ala Lieu Ala 65 70 7s 8O Met Gly Ala Asp Arg Gly Ile His Val Glu Val Pro Pro Ala Glu Ala 85 90 95 Glu Arg Lieu. Gly Pro Lieu. Glin Val Ala Arg Val Lieu Ala Lys Lieu Ala 1OO 105 11 O Glu Lys Glu Lys Val Asp Lieu Val Lieu. Lieu. Gly Lys Glin Ala Ile Tyr 115 12 O 125

Asp Asp Cys Asn Glin Thr Gly Glin Met Thr Ala Gly Phe Lieu. Asp Trip 13 O 135 14 O

Pro Glin Gly Thr Phe Ala Ser Glin Val Met Leu Glu Gly Asp Llys Lieu. 145 150 155 160

Llys Val Glu Arg Glu Ile Asp Gly Gly Lieu. Glu Thir Lieu. Arg Lieu Lys 1.65 17O 17s

Lieu Pro Ala Val Val Thr Ala Asp Lieu. Arg Lieu. Asn. Glu Pro Arg Tyr 18O 185 19 O US 2015/O151 001 A1 Jun. 4, 2015 39

- Continued Ala Thr Lieu Pro Asn. Ile Met Lys Ala Lys Llys Llys Lys Ile Glu Val 195 2OO 2O5 Ile Llys Pro Gly Asp Lieu. Gly Val Asp Lieu. Thir Ser Llys Lieu. Ser Val 21 O 215 22O Ile Ser Val Glu Asp Pro Pro Glin Arg Thr Ala Gly Val Llys Val Glu 225 23 O 235 24 O Thir Thr Glu Asp Lieu Val Ala Lys Lieu Lys Glu Ile Gly Arg Ile 245 250 255

<210s, SEQ ID NO 22 &211s LENGTH: 503 212. TYPE: PRT <213> ORGANISM: Homo sapiens <4 OOs, SEQUENCE: 22 Met Ser Ser Ile Lieu. Glu Glu Arg Ile Lieu. Gly Ala Asp Thir Ser Val 1. 5 1O 15 Asp Lieu. Glu Glu Thr Gly Arg Val Lieu. Ser Ile Gly Asp Gly Ile Ala 2O 25 3O Arg Val His Gly Lieu. Arg Asn Val Glin Ala Glu Glu Met Val Glu Phe 35 4 O 45 Ser Ser Gly Lieu Lys Gly Met Ser Lieu. Asn Lieu. Glu Pro Asp Asn. Wall SO 55 6 O Gly Val Val Val Phe Gly Asn Asp Llys Lieu. Ile Lys Glu Gly Asp Ile 65 70 75 8O Val Lys Arg Thr Gly Ala Ile Val Asp Val Pro Val Gly Glu Glu Lieu 85 90 95 Lieu. Gly Arg Val Val Asp Ala Lieu. Gly Asn Ala Ile Asp Gly Lys Gly 1OO 105 11 O Pro Ile Gly Ser Llys Thr Arg Arg Arg Val Gly Lieu Lys Ala Pro Gly 115 12 O 125 Ile Ile Pro Arg Ile Ser Val Arg Glu Pro Met Gln Thr Gly Ile Llys 13 O 135 14 O Ala Val Asp Ser Lieu Val Pro Ile Gly Arg Gly Glin Arg Glu Lieu. Ile 145 150 155 160 Ile Gly Asp Arg Glin Thr Gly Lys Thir Ser Ile Ala Ile Asp Thir Ile 1.65 17O 17s Ile Asin Glin Lys Arg Phe Asn Asp Gly Ser Asp Glu Lys Llys Llys Lieu. 18O 185 19 O Tyr Cys Ile Tyr Val Ala Ile Gly Gln Lys Arg Ser Thr Val Ala Glin 195 2OO 2O5 Lieu Val Lys Arg Lieu. Thir Asp Ala Asp Ala Met Lys Tyr Thir Ile Val 21 O 215 22O

Val Ser Ala Thr Ala Ser Asp Ala Ala Pro Lieu. Glin Tyr Lieu Ala Pro 225 23 O 235 24 O Tyr Ser Gly Cys Ser Met Gly Glu Tyr Phe Arg Asp Asn Gly Lys His 245 250 255

Ala Lieu. Ile Ile Tyr Asp Asp Lieu. Ser Lys Glin Ala Wall Ala Tyr Arg 26 O 265 27 O

Glin Met Ser Lieu Lleu Lieu. Arg Arg Pro Pro Gly Arg Glu Ala Tyr Pro 27s 28O 285 Gly Asp Val Phe Tyr Lieu. His Ser Arg Lieu. Lieu. Glu Arg Ala Ala Lys 29 O 295 3 OO US 2015/O151 001 A1 Jun. 4, 2015 40

- Continued

Met Asn Asp Ala Phe Gly Gly Gly Ser Lieu. Thir Ala Lieu Pro Val Ile 3. OS 310 315 32O Glu Thr Glin Ala Gly Asp Val Ser Ala Tyr Ile Pro Thr Asn Val Ile 3.25 330 335 Ser Ile Thr Asp Gly Glin Ile Phe Leu Glu Thr Glu Lieu Phe Tyr Lys 34 O 345 35. O Gly Ile Arg Pro Ala Ile Asin Val Gly Lieu. Ser Val Ser Arg Val Gly 355 360 365 Ser Ala Ala Glin Thr Arg Ala Met Lys Glin Val Ala Gly. Thir Met Lys 37 O 375 38O Lieu. Glu Lieu Ala Glin Tyr Arg Glu Val Ala Ala Phe Ala Glin Phe Gly 385 390 395 4 OO Ser Asp Lieu. Asp Ala Ala Thr Glin Glin Lieu. Lieu. Ser Arg Gly Val Arg 4 OS 41O 415 Lieu. Thr Glu Lieu Lleu Lys Glin Gly Glin Tyr Ser Pro Met Ala Ile Glu 42O 425 43 O Glu Glin Val Ala Val Ile Tyr Ala Gly Val Arg Gly Tyr Lieu. Asp Llys 435 44 O 445 Lieu. Glu Pro Ser Lys Ile Thr Llys Phe Glu Asn Ala Phe Leu Ser His 450 45.5 460 Val Val Ser Glin His Glin Ala Lieu. Lieu. Gly. Thir Ile Arg Ala Asp Gly 465 470 47s 48O Lys Ile Ser Glu Glin Ser Asp Ala Lys Lieu Lys Glu Ile Val Thr Asn 485 490 495 Phe Lieu Ala Gly Phe Glu Ala SOO

<210s, SEQ ID NO 23 &211s LENGTH: 339 212. TYPE: PRT <213> ORGANISM: Homo sapiens <4 OOs, SEQUENCE: 23 Met Trp Gln Lieu. Trp Ala Ser Lieu. Cys Cys Lieu. Lieu Val Lieu Ala Asn 1. 5 1O 15 Ala Arg Ser Arg Pro Ser Phe His Pro Lieu. Ser Asp Glu Lieu Val Asn 2O 25 3O Tyr Val Asn Lys Arg Asn Thr Thr Trp Glin Ala Gly His Asn Phe Tyr 35 4 O 45 Asn Val Asp Met Ser Tyr Lieu Lys Arg Lieu. Cys Gly. Thir Phe Lieu. Gly SO 55 6 O Gly Pro Llys Pro Pro Glin Arg Val Met Phe Thr Glu Asp Leu Lys Lieu. 65 70 7s 8O

Pro Ala Ser Phe Asp Ala Arg Glu Gln Trp Pro Gln Cys Pro Thir Ile 85 90 95

Lys Glu Ile Arg Asp Glin Gly Ser Cys Gly Ser Cys Trp Ala Phe Gly 1OO 105 11 O

Ala Val Glu Ala Ile Ser Asp Arg Ile Cys Ile His Thr Asn Ala His 115 12 O 125

Val Ser Val Glu Val Ser Ala Glu Asp Lieu. Lieu. Thir Cys Cys Gly Ser 13 O 135 14 O

Met Cys Gly Asp Gly Cys Asn Gly Gly Tyr Pro Ala Glu Ala Trp Asn US 2015/O151 001 A1 Jun. 4, 2015 41

- Continued

145 150 155 160 Phe Trp Thr Arg Lys Gly Lieu Val Ser Gly Gly Lieu. Tyr Glu Ser His 1.65 17O 17s Val Gly Cys Arg Pro Tyr Ser Ile Pro Pro Cys Glu. His His Val Asn 18O 185 19 O Gly Ser Arg Pro Pro Cys Thr Gly Glu Gly Asp Thr Pro Llys Cys Ser 195 2OO 2O5 Lys Ile Cys Glu Pro Gly Tyr Ser Pro Thr Tyr Lys Glin Asp Llys His 21 O 215 22O Tyr Gly Tyr Asn Ser Tyr Ser Val Ser Asn Ser Glu Lys Asp Ile Met 225 23 O 235 24 O Ala Glu Ile Tyr Lys Asn Gly Pro Val Glu Gly Ala Phe Ser Val Tyr 245 250 255 Ser Asp Phe Leu Lleu Tyr Lys Ser Gly Val Tyr Gln His Val Thr Gly 26 O 265 27 O Glu Met Met Gly Gly His Ala Ile Arg Ile Lieu. Gly Trp Gly Val Glu 27s 28O 285 Asn Gly Thr Pro Tyr Trp Leu Val Ala Asn Ser Trp Asn Thr Asp Trp 29 O 295 3 OO Gly Asp Asn Gly Phe Phe Lys Ile Lieu. Arg Gly Glin Asp His Cys Gly 3. OS 310 315 32O Ile Glu Ser Glu Val Val Ala Gly Ile Pro Arg Thr Asp Glin Tyr Trp 325 330 335 Glu Lys Ile

<210s, SEQ ID NO 24 &211s LENGTH: 10 212. TYPE: PRT <213> ORGANISM: Artificial sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic construct <4 OOs, SEQUENCE: 24 Glu Gln Lys Lieu. Ile Ser Glu Glu Asp Lieu. 1. 5 1O

<210s, SEQ ID NO 25 &211s LENGTH: 9 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic construct <4 OOs, SEQUENCE: 25 Tyr Pro Tyr Asp Val Pro Asp Tyr Ala 1. 5

<210s, SEQ ID NO 26 &211s LENGTH: 6 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic construct

<4 OOs, SEQUENCE: 26

His His His His His His 1. 5 US 2015/O151 001 A1 Jun. 4, 2015 42

- Continued

<210s, SEQ ID NO 27 &211s LENGTH: 238 212. TYPE: PRT <213> ORGANISM: Aequorea victoria <4 OOs, SEQUENCE: 27 Met Ser Lys Gly Glu Glu Lieu Phe Thr Gly Val Val Pro Ile Leu Val 1. 5 1O 15 Glu Lieu. Asp Gly Asp Wall Asn Gly His Llys Phe Ser Val Ser Gly Glu 2O 25 3O Gly Glu Gly Asp Ala Thr Tyr Gly Llys Lieu. Thir Lieu Lys Phe Ile Cys 35 4 O 45 Thir Thr Gly Lys Lieu Pro Val Pro Trp Pro Thr Lieu Val Thir Thr Phe SO 55 6 O Ser Tyr Gly Val Glin Cys Phe Ser Arg Tyr Pro Asp His Met Lys Glin 65 70 7s 8O His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val Glin Glu Arg 85 90 95 Thir Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg Ala Glu Val 1OO 105 11 O Llys Phe Glu Gly Asp Thir Lieu Val Asn Arg Ile Glu Lieu Lys Gly Ile 115 12 O 125 Asp Phe Lys Glu Asp Gly Asn. Ile Lieu. Gly His Llys Lieu. Glu Tyr Asn 13 O 135 14 O Tyr Asn. Ser His Asn Val Tyr Ile Met Ala Asp Llys Gln Lys Asn Gly 145 150 155 160 Ile Llys Val Asn. Phe Lys Ile Arg His Asn. Ile Glu Asp Gly Ser Val 1.65 17O 17s Glin Lieu Ala Asp His Tyr Glin Glin Asn Thr Pro Ile Gly Asp Gly Pro 18O 185 19 O Val Lieu. Lieu Pro Asp Asn His Tyr Lieu. Ser Thr Glin Ser Ala Lieu. Ser 195 2OO 2O5 Lys Asp Pro Asn. Glu Lys Arg Asp His Met Val Lieu. Lieu. Glu Phe Val 21 O 215 22O Thir Ala Ala Gly Ile Thr His Gly Met Asp Glu Lieu. Tyr Lys 225 23 O 235

<210s, SEQ ID NO 28 &211s LENGTH: 194 212. TYPE: PRT <213> ORGANISM: Aliiyibrio fischeri

<4 OOs, SEQUENCE: 28 Met Phe Lys Gly Ile Val Glu Gly Ile Gly Ile Ile Glu Lys Ile Asp 1. 5 1O 15

Ile Tyr Thr Asp Lieu. Asp Llys Tyr Ala Ile Arg Phe Pro Glu Asn Met 2O 25 3O

Lieu. Asn Gly Ile Llys Lys Glu Ser Ser Ile Met Phe ASn Gly Cys Phe 35 4 O 45

Lieu. Thr Val Thr Ser Val Asn Ser Asn Ile Val Trp Phe Asp Ile Phe SO 55 6 O

Glu Lys Glu Ala Arg Llys Lieu. Asp Thr Phe Arg Glu Tyr Llys Val Gly 65 70 7s 8O US 2015/O151 001 A1 Jun. 4, 2015 43

- Continued Asp Arg Val Asn Lieu. Gly Thr Phe Pro Llys Phe Gly Ala Ala Ser Gly 85 90 95 Gly His Ile Lieu. Ser Ala Arg Ile Ser Cys Val Ala Ser Ile Ile Glu 1OO 105 11 O Ile Ile Glu Asn Glu Asp Tyr Glin Glin Met Trp Ile Glin Ile Pro Glu 115 12 O 125 Asn Phe Thr Glu Phe Lieu. Ile Asp Lys Asp Tyr Ile Ala Val Asp Gly 13 O 135 14 O Ile Ser Lieu. Thir Ile Asp Thr Ile Lys Asn Asn Glin Phe Phe Ile Ser 145 150 155 160 Lieu Pro Lieu Lys Ile Ala Glin Asn. Thir Asn Met Lys Trp Arg Llys Llys 1.65 17O 17s Gly Asp Llys Val Asn Val Glu Lieu. Ser Asn Lys Ile Asn Ala Asn Glin 18O 185 19 O Cys Trp

<210s, SEQ ID NO 29 &211s LENGTH: 225 212. TYPE: PRT &213s ORGANISM: Montastraea cavernosa

<4 OOs, SEQUENCE: 29 Met Ser Val Ile Llys Ser Wal Met Lys Ile Llys Lieu. Arg Met Asp Gly 1. 5 1O 15 Ile Val Asn Gly His Llys Phe Met Ile Thr Gly Glu Gly Glu Gly Lys 2O 25 3O Pro Phe Glu Gly Thr His Thr Ile Ile Leu Lys Val Lys Glu Gly Gly 35 4 O 45 Pro Leu Pro Phe Ala Tyr Asp Ile Lieu. Thir Thr Ala Phe Glin Tyr Gly SO 55 6 O Asn Arg Val Phe Thr Lys Tyr Pro Lys Asp Ile Pro Asp Tyr Phe Lys 65 70 7s 8O Gln Ser Phe Pro Glu Gly Tyr Ser Trp Glu Arg Ser Met Thr Phe Glu 85 90 95 Asp Glin Gly Val Cys Thr Val Thir Ser Asp Ile Llys Lieu. Glu Gly Asp 1OO 105 11 O Cys Phe Phe Tyr Glu Ile Arg Phe Tyr Gly Val Asn Phe Pro Ser Ser 115 12 O 125 Gly Pro Val Met Gln Lys Llys Thr Lieu Lys Trp Glu Pro Ser Thr Glu 13 O 135 14 O Asn Met Tyr Val Arg Asp Gly Val Lieu. Lieu. Gly Asp Wal Ser Arg Thr 145 150 155 160 Lieu. Lieu. Lieu. Glu Gly Asp Llys His His Arg Cys Asn. Phe Arg Ser Thr 1.65 17O 17s Tyr Gly Ala Lys Lys Gly Val Val Lieu Pro Glu Tyr His Phe Val Asp 18O 185 19 O His Arg Ile Glu Ile Lieu. Ser His Asp Lys Asp Tyr Asn Thr Val Glu 195 2OO 2O5

Val Tyr Glu Asn Ala Val Ala Arg Pro Ser Met Lieu Pro Val Lys Ala 21 O 215 22O

Lys 225 US 2015/O151 001 A1 Jun. 4, 2015 44

- Continued

<210s, SEQ ID NO 3 O &211s LENGTH: 23 O 212. TYPE: PRT <213> ORGANISM: Discosoma sp. <4 OOs, SEQUENCE: 30 Met Ser Cys Ser Lys Asn Val Ile Lys Glu Phe Met Arg Phe Llys Val 1. 5 1O 15 Arg Met Glu Gly Thr Val Asn Gly His Glu Phe Glu Ile Lys Gly Glu 2O 25 3O Gly Glu Gly Arg Pro Tyr Glu Gly His Cys Ser Val Lys Lieu Met Val 35 4 O 45 Thr Lys Gly Gly Pro Leu Pro Phe Ala Phe Asp Ile Leu Ser Pro Glin SO 55 6 O Phe Glin Tyr Gly Ser Lys Val Tyr Val Llys His Pro Ala Asp Ile Pro 65 70 7s 8O Asp Tyr Lys Llys Lieu. Ser Phe Pro Glu Gly Phe Llys Trp Glu Arg Val 85 90 95 Met Asin Phe Glu Asp Gly Gly Val Val Thr Val Ser Glin Asp Ser Ser 1OO 105 11 O Lieu Lys Asp Gly Cys Phe Ile Tyr Glu Val Llys Phe Ile Gly Val Asn 115 12 O 125 Phe Pro Ser Asp Gly Pro Val Met Glin Arg Arg Thr Arg Gly Trp Glu 13 O 135 14 O Ala Ser Ser Glu Arg Lieu. Tyr Pro Arg Asp Gly Val Lieu Lys Gly Asp 145 150 155 160 Ile His Met Ala Lieu. Arg Lieu. Glu Gly Gly Gly His Tyr Lieu Val Glu 1.65 17O 17s Phe Llys Ser Ile Tyr Met Val Lys Llys Pro Ser Val Glin Leu Pro Gly 18O 185 19 O Tyr Tyr Tyr Val Asp Ser Llys Lieu. Asp Met Thir Ser His Asn. Glu Asp 195 2OO 2O5 Tyr Thr Val Val Glu Gln Tyr Glu Lys Thr Glin Gly Arg His His Pro 21 O 215 22O Phe Ile Llys Pro Leu Gln 225 23 O

<210s, SEQ ID NO 31 &211s LENGTH: 550 212. TYPE: PRT <213> ORGANISM: Photinus pyralis <4 OOs, SEQUENCE: 31 Met Glu Asp Ala Lys Asn. Ile Llys Lys Gly Pro Ala Pro Phe Tyr Pro 1. 5 1O 15 Lieu. Glu Asp Gly Thr Ala Gly Glu Glin Lieu. His Lys Ala Met Lys Arg 2O 25 3O Tyr Ala Leu Val Pro Gly. Thir Ile Ala Phe Thr Asp Ala His Ile Glu 35 4 O 45

Val Asn Ile Thr Tyr Ala Glu Tyr Phe Glu Met Ser Val Arg Lieu Ala SO 55 6 O Glu Ala Met Lys Arg Tyr Gly Lieu. Asn. Thir Asn His Arg Ile Val Val 65 70 7s 8O US 2015/O151 001 A1 Jun. 4, 2015 45

- Continued

Cys Ser Glu Asn Ser Lieu. Glin Phe Phe Met Pro Val Lieu. Gly Ala Leu 85 90 95 Phe Ile Gly Val Ala Val Ala Pro Ala Asn Asp Ile Tyr Asn. Glu Arg 1OO 105 11 O

Glu Lieu Lleu. Asn. Ser Met Asn. Ile Ser Glin Pro Thir Wal Wall Phe Wall 115 12 O 125 Ser Lys Lys Gly Lieu. Glin Lys Ile Lieu. Asn Val Glin Llys Llys Lieu Pro 13 O 135 14 O Ile Ile Glin Lys Ile Ile Ile Met Asp Ser Lys Thr Asp Tyr Glin Gly 145 150 155 160 Phe Glin Ser Met Tyr Thr Phe Val Thr Ser His Leu Pro Pro Gly Phe 1.65 17O 17s Asn Glu Tyr Asp Phe Val Pro Glu Ser Phe Asp Arg Asp Llys Thir Ile 18O 185 19 O Ala Lieu. Ile Met Asn Ser Ser Gly Ser Thr Gly Ser Pro Lys Gly Val 195 2OO 2O5 Ala Lieu Pro His Arg Thr Ala Cys Val Arg Phe Ser His Ala Arg Asp 21 O 215 22O Pro Ile Phe Gly Asn Glin Ile Ile Pro Asp Thr Ala Ile Leu Ser Val 225 23 O 235 24 O Val Pro Phe His His Gly Phe Gly Met Phe Thir Thr Lieu. Gly Tyr Lieu. 245 250 255 Ile Cys Gly Phe Arg Val Val Lieu Met Tyr Arg Phe Glu Glu Glu Lieu 26 O 265 27 O Phe Lieu. Arg Ser Lieu. Glin Asp Tyr Lys Ile Glin Ser Ala Lieu. Lieu Val 27s 28O 285 Pro Thr Lieu Phe Ser Phe Phe Ala Lys Ser Thr Lieu. Ile Asp Llys Tyr 29 O 295 3 OO Asp Lieu. Ser Asn Lieu. His Glu Ile Ala Ser Gly Gly Ala Pro Lieu. Ser 3. OS 310 315 32O Lys Glu Val Gly Glu Ala Val Ala Lys Arg Phe His Lieu Pro Gly Ile 3.25 330 335 Arg Glin Gly Tyr Gly Lieu. Thr Glu Thir Thr Ser Ala Ile Lieu. Ile Thr 34 O 345 35. O Pro Glu Gly Asp Asp Llys Pro Gly Ala Val Gly Llys Val Val Pro Phe 355 360 365 Phe Glu Ala Lys Val Val Asp Lieu. Asp Thr Gly Llys Thr Lieu. Gly Val 37 O 375 38O Asn Glin Arg Gly Glu Lieu. CyS Val Arg Gly Pro Met Ile Met Ser Gly 385 390 395 4 OO Tyr Val Asn Asp Pro Glu Ala Thr Asn Ala Lieu. Ile Asp Lys Asp Gly 4 OS 41O 415 Trp Lieu. His Ser Gly Asp Ile Ala Tyr Trp Asp Glu Asp Glu. His Phe 42O 425 43 O

Phe Ile Val Asp Arg Lieu Lys Ser Lieu. Ile Llys Tyr Lys Gly Cys Glin 435 44 O 445

Wall Ala Pro Ala Glu Lieu. Glu Ser Ile Lieu. Lieu. Glin His Pro Asn. Ile 450 45.5 460

Phe Asp Ala Gly Val Ala Gly Lieu Pro Gly Asp Asp Ala Gly Glu Lieu. 465 470 47s 48O

Pro Ala Ala Val Val Val Lieu. Glu. His Gly Lys Thr Met Thr Glu Lys US 2015/O151 001 A1 Jun. 4, 2015 46

- Continued

485 490 495 Glu Ile Val Asp Tyr Val Ala Ser Glin Val Thir Thr Ala Lys Llys Lieu. SOO 505 51O Arg Gly Gly Val Val Phe Val Asp Glu Val Pro Llys Gly Lieu. Thr Gly 515 52O 525 Llys Lieu. Asp Ala Arg Lys Ile Arg Glu Ile Lieu. Ile Lys Ala Lys Llys 53 O 535 54 O Gly Gly Lys Ser Llys Lieu. 5.45 550

<210s, SEQ ID NO 32 &211s LENGTH: 311 212. TYPE: PRT <213> ORGANISM: Renilla reniformis

<4 OOs, SEQUENCE: 32 Met Thr Ser Llys Val Tyr Asp Pro Glu Glin Arg Lys Arg Met Ile Thr 1. 5 1O 15 Gly Pro Gln Trp Trp Ala Arg Cys Lys Gln Met Asn Val Lieu. Asp Ser 2O 25 3O Phe Ile Asn Tyr Tyr Asp Ser Glu Lys His Ala Glu Asn Ala Val Ile 35 4 O 45 Phe Lieu. His Gly Asn Ala Ala Ser Ser Tyr Lieu. Trp Arg His Val Val SO 55 6 O Pro His Ile Glu Pro Val Ala Arg Cys Ile Ile Pro Asp Lieu. Ile Gly 65 70 7s 8O Met Gly Lys Ser Gly Llys Ser Gly Asn Gly Ser Tyr Arg Lieu. Lieu. Asp 85 90 95 His Tyr Lys Tyr Lieu. Thir Ala Trp Phe Glu Lieu. Lieu. Asn Lieu Pro Llys 1OO 105 11 O Lys Ile Ile Phe Val Gly His Asp Trp Gly Ala Cys Lieu Ala Phe His 115 12 O 125 Tyr Ser Tyr Glu. His Glin Asp Llys Ile Lys Ala Ile Val His Ala Glu 13 O 135 14 O Ser Val Val Asp Val Ile Glu Ser Trp Asp Glu Trp Pro Asp Ile Glu 145 150 155 160 Glu Asp Ile Ala Lieu. Ile Llys Ser Glu Glu Gly Glu Lys Met Val Lieu. 1.65 17O 17s Glu Asn Asn Phe Phe Val Glu Thr Met Leu Pro Ser Lys Ile Met Arg 18O 185 19 O Llys Lieu. Glu Pro Glu Glu Phe Ala Ala Tyr Lieu. Glu Pro Phe Lys Glu 195 2OO 2O5 Lys Gly Glu Val Arg Arg Pro Thir Lieu. Ser Trp Pro Arg Glu Ile Pro 21 O 215 22O

Lieu Val Lys Gly Gly Llys Pro Asp Val Val Glin Ile Val Arg Asn Tyr 225 23 O 235 24 O

Asn Ala Tyr Lieu. Arg Ala Ser Asp Asp Lieu Pro Llys Met Phe Ile Glu 245 250 255

Ser Asp Pro Gly Phe Phe Ser Asn Ala Ile Val Glu Gly Ala Lys Llys 26 O 265 27 O Phe Pro Asn Thr Glu Phe Val Llys Val Lys Gly Lieu. His Phe Ser Glin 27s 28O 285 US 2015/O151 001 A1 Jun. 4, 2015 47

- Continued Glu Asp Ala Pro Asp Glu Met Gly Llys Tyr Ile Llys Ser Phe Val Glu 29 O 295 3 OO Arg Val Lieu Lys Asn. Glu Glin 3. OS 310

<210s, SEQ ID NO 33 &211s LENGTH: 21 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic construct <4 OOs, SEQUENCE: 33 Llys Phe Ala Lys Phe Ala Lys Llys Phe Ala Lys Phe Ala Lys Llys Phe 1. 5 1O 15 Ala Lys Phe Ala Lys 2O

<210s, SEQ ID NO 34 &211s LENGTH: 30 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic construct <4 OOs, SEQUENCE: 34 Gly Val Tyr His Phe Ala Pro Leu. Thr Pro Thr Pro Gly Gly Gly Ser 1. 5 1O 15 Llys Phe Ala Lys Phe Ala Lys Llys Phe Ala Lys Phe Ala Lys 2O 25

<210s, SEQ ID NO 35 &211s LENGTH: 5 212. TYPE: PRT <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic construct 22 Os. FEATURE: <221s NAME/KEY: MOD RES &222s. LOCATION: 5 223 OTHER INFORMATION: Xaa = DTPA

<4 OOs, SEQUENCE: 35 Gly Gly Gly Ser Xaa 1. 5

1.-86. (canceled) 92. The method of claim 87, wherein the therapeutic treat 87. A method for treating a senescence associated condi ment is a cytotoxic agent. tion in a Subject comprising: (a) detecting senescent cell 93. The method of claim 92, wherein the cytotoxic agent is content in the Subject; and (b) administering to the Subject a a small molecule. therapeutic treatment that selectively kills senescent cells. 88. The method of claim 87, wherein detecting comprises 94. The method of claim 92, wherein the cytotoxic agent is administering to the Subject an agent capable of binding to the coupled to a second agent capable of targeting a senescent senescent cell content wherein the agent is labeled with a cell. detectable label. 95. The method of claim 87, wherein the senescence asso 89. The method of claim 88, wherein the detectable label is ciated condition is cancer. fluorescent, bioluminescent, radioactive, an epitope tag, or a 96. The method of claim 87, wherein the senescence asso heavy metal. ciated condition is osteoarthritis. 90. The method of claim 87, wherein the therapeutic treat ment is an apoptotic agent. 97. The method of claim 87, wherein the senescence asso 91. The method of claim 90, wherein the apoptotic agent is ciated condition is atherosclerosis. a small molecule. k k k k k