JP2016538286A - Methods and pharmaceutical compositions for treating pancreatic cancer - Google Patents

Abstract

The present invention relates to methods and pharmaceutical compositions for treating pancreatic cancer. In particular, the invention relates to OX1R agonists for use in the treatment of pancreatic cancer in a subject in need thereof.

Description

  The present invention relates to methods and pharmaceutical compositions for treating pancreatic cancer.

BACKGROUND OF THE INVENTION Pancreatic cancer is an aggressive disease with a very poor prognosis. This is one of the most malignant cancers and is characterized by insidious onset, usually with a delayed diagnosis and a low survival rate after diagnosis. For example, pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death in the United States. Despite recent treatment advances, long-term survival in PDAC is often limited to patients who have undergone surgery at an early stage of the disease. The biological aggressiveness of PDAC is due in part to the resistance of this tumor to chemotherapy. Currently, the standard of treatment is limited to systemic chemotherapy with gemcitabine, which is intended to mitigate and has an unwilling minimum life-prolonging effect. Most recently, the administration of 5-fluorouracil, leucovorin, irinotecan and oxaliplatin (forfilinox) has demonstrated clinically and statistically meaningful life-prolonging effects on single-agent gemcitabine (Conroy et al. , N. Engl. J. Med., 364: 1817-1825 (2011)), and albumin-bound paclitaxel, which appears to target the desmoplastic stroma characteristic of pancreatic ductal adenocarcinoma (PDAC) Introduction of nanoparticles (nad-paclitaxel) (Garber, K., J. Natl. Cancer Inst., 102: 448-450 (2010)), an innovative combination of classical cytotoxics and improved delivery However, it raised the expectation that it could have a substantial impact on the efficacy of chemotherapy in advanced PDAC. Thus, despite minor advances in pancreatic cancer treatment, there remains a need for improved therapies and more creative approaches that develop and provide effective treatments for pancreatic cancer.

  Orexin (hypocretin) includes two neuropeptides produced in the hypothalamus, orexin A (OX-A) (33 amino acid peptide) and orexin B (OX-B) (28 amino acid peptide) (Sakurai T. et al., Cell, 1998, 92, 573-585). Orexin has been found to promote food consumption in mice, suggesting a physiological role for these peptides as mediators in central feedback mechanisms that control dietary behavior. Orexin controls sleep and wakefulness, which is an potentially potentially novel therapeutic approach for patients with narcolepsy or insomnia. Orexin has also been shown to play a role in arousal, reward, learning and memory. Two orexin receptors have been cloned and characterized in mammals. They belong to the superfamily of G protein-coupled receptors (7 transmembrane receptors) (Sakurai T. et al., Cell, 1998, 92, 573-585): Orexin-1 receptor (OX1R or HCTR1) is OX-A The orexin-2 receptor (OX2R or HCTR2) can bind to OX-A and OX-B. Recent studies have shown that activation of OX1R by orexin is robust in vitro and in vivo, even in colon cancer cells, even though they are resistant to the most commonly used drugs in colon cancer chemotherapy. It was shown that it can promote apoptosis (Voisin T, El Firar A, Fasseu M, Rouyer-Fessard C, Descatoire V, Walker F, Paradis V, Bedossa P, Henin D, Lehy T, Laburthe M. Aberrant expression of OX1 receptors for orexins in colon cancers and liver metastases: an openable gate to apoptosis. Cancer Res. 2011 May 1; 71 (9): 3341-51). Of note, all primary colorectal tumors expressed OX1R regardless of their location or Duke stage, but were negative for adjacent normal colon cells and control normal tissues. Thus, this study supports that OX1R is an Achilles's heel of colon cancer (even if it is chemoresistant) and that OX1R agonists can be new candidates for colon cancer treatment It is a suggestion.

The present invention relates to methods and pharmaceutical compositions for treating pancreatic cancer. In particular, the present invention relates to an OX1R agonist for use in the treatment of pancreatic cancer in a subject in need thereof.

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an OX1R agonist for use in the treatment of pancreatic cancer in a subject in need thereof.

  As used herein, the term (tem) “OX1R” has its general meaning in the art and refers to the orexin seven-transmembrane receptor OX1R. According to the present invention, OX1R promotes apoptosis in human pancreatic cancer cell lines through mechanisms that are not associated with Gq-mediated phospholipase C activation or cellular calcium transients. Orexin actually induces tyrosine phosphorylation of two tyrosine-based motifs in OX1R, ITIM and ITSM, which leads to recruitment of the phosphotyrosine phosphatase SHP-2, although its activity Is involved in mitochondrial apoptosis (Voisin T, El Firar A, Rouyer-Fessard C, Gratio V, Laburthe M. A hallmark of immunoreceptor, the tyrosine-based inhibitory motif ITIM, is present in the G protein- coupled receptor OX1R for orexins and drives apoptosis: a novel mechanism.FASEB J. 2008 Jun; 22 (6): 1993-2002 .; El Firar A, Voisin T, Rouyer-Fessard C, Ostuni MA, Couvineau A, Laburthe M. Discovery of a functional immunoreceptor tyrosine-based switch motif in a 7-transmembrane-spanning receptor: role in the orexin receptor OX1R-driven apoptosis. FASEB J. 2009 Dec; 23 (12): 4069-80.doi: 10.1096 / fj. 09-131367. Epub 2009 Aug 6.). An exemplary amino acid sequence of OX1R is shown as SEQ ID NO: 1.

Orexin receptor-1 OX1R (SEQ ID NO: 1)
1 mepsatpgaq mgvppgsrep spvppdyede flrylwrdyl ypkqyewvli aayvavfvva
61 lvgntlvcla vwrnhhmrtv tnyfivnlsl advlvtaicl pasllvdite swlfghalck
121 vipylqavsv svavltlsfi aldrwyaich pllfkstarr argsilgiwa vslaimvpqa
181 avmecssvlp elanrtrlfs vcderwaddl ypkiyhscff ivtylaplgl mamayfqifr
241 klwgrqipgt tsalvrnwkr psdqlgdleq glsgepqprg raflaevkqm rarrktakml
301 mvvllvfalc ylpisvlnvl krvfgmfrqa sdreavyacf tfshwlvyan saanpiiynf
361 lsgkfreqfk aafscclpgl gpcgslkaps prssashksl slqsrcsisk isehvvltsv
421 ttvlp

  Thus, as used herein, the term “OX1R agonist”, whether natural or not, can bind to OX1R and is an activity in a signaling pathway that is independent of transient calcium release. Refers to any compound that promotes OX1R activity consisting of oxidization (involved in the recruitment of SHP-2 and the induction of cellular apoptosis).

  In some embodiments, the OX1R agonist is a small organic molecule. The term “small organic molecule” refers to a molecule of a size comparable to organic molecules commonly used in pharmaceuticals. This term excludes biopolymers (eg, proteins, nucleic acids, etc.). Preferred small organic molecules range in size up to about 5000 Da, more specifically up to 2000 Da, most specifically up to about 1000 Da.

  In certain embodiments, the OX1R agonist is an antibody or portion thereof.

  As used herein, “antibody” includes both natural and non-natural antibodies. Specifically, “antibodies” include polyclonal and monoclonal antibodies, and monovalent and divalent fragments thereof. Furthermore, “antibody” includes chimeric antibodies, total synthetic antibodies, single chain antibodies and fragments thereof. The antibody can be a human antibody or a non-human antibody. Non-human antibodies may be humanized by recombinant methods in order to reduce immunogenicity in humans.

  In one embodiment of the antibodies described herein or portions thereof, the antibodies are monoclonal antibodies. In one embodiment of the antibodies described herein or portions thereof, the antibodies are polyclonal antibodies. In one embodiment of the antibodies described herein or portions thereof, the antibodies are humanized antibodies. In one embodiment of the antibodies described herein or portions thereof, the antibodies are chimeric antibodies. In one embodiment of the antibodies or portions thereof described herein, the portion of the antibody comprises the light chain of the antibody. In one embodiment of the antibodies described herein or portions thereof, the portion of the antibody comprises the heavy chain of the antibody. In one embodiment of the antibodies or portions thereof described herein, the portion of the antibody comprises the Fab portion of the antibody. In one embodiment of the antibodies described herein or portions thereof, the portion of the antibody comprises the F (ab ') 2 portion of the antibody. In one embodiment of the antibodies described herein or portions thereof, the portion of the antibody comprises the Fc portion of the antibody. In one embodiment of the antibodies or portions thereof described herein, the portion of the antibody comprises the Fv portion of the antibody. In one embodiment of the antibodies described herein or portions thereof, the portion of the antibody comprises the variable domain of the antibody. In one embodiment of the antibodies described herein or portions thereof, the portion of the antibody comprises a CDR domain of the antibody.

  Antibodies are prepared according to conventional methodologies. Monoclonal antibodies can be made using the method of Kohler and Milstein (Nature, 256: 495, 1975). In order to prepare monoclonal antibodies useful in the present invention, mice or other suitable host animals are treated with the antigen form of OX1R at appropriate intervals (eg, every 2 weeks, every week, every 2 months or 1 Immunize every month). Within one week of sacrifice, the animals may receive a final antigen “boost”. It is often preferable to use an immunological adjuvant during immunization. Suitable immunological adjuvants include Freund's complete adjuvant, Freund's incomplete adjuvant, alum, Ribi adjuvant, Hunter's Titermax, saponin adjuvants such as QS21 or QuilA, or CpG containing immunostimulatory oligonucleotides. Other suitable adjuvants are well known in the art. Animals may be immunized by subcutaneous, intraperitoneal, intramuscular, intravenous, intranasal or other routes. A given animal may be immunized by multiple forms with multiple forms of antigen. Briefly, recombinant OX1R may be provided using a recombinant cell line. Specifically, OX1R may be provided in the form of human cells in which OX1R is expressed on the surface. Following the immunization regimen, lymphocytes are isolated from the spleen, lymph nodes and other tissues of the animal and fused with an appropriate myeloma cell line using reagents such as polyethylene glycol to form hybridomas. Following the fusion, standard methods are used as described in the literature (Coding, Monoclonal Antibodies: Principles and Practice: Production and Application of Monoclonal Antibodies in Cell Biology, Biochemistry and Immunology, 3rd edition, Academic Press, New York, 1996). In use, cells are placed in a medium that allows growth for hybridomas but not for fusion partners. Following culture of the hybridoma, the cell supernatant is analyzed for the presence of antibodies having the desired specificity, ie, selectively binding to the antibodies. Suitable analytical techniques include ELISA, flow cytometry, immunoprecipitation and western blotting. Other screening techniques are well known in the art. A preferred technique is to confirm the binding of the antibody to a normally conformed and folded antigen, such as non-denaturing ELISA, flow cytometry and immunoprecipitation.

  As is well known in the art, only a small part of an antibody molecule, the paratope, is significantly involved in binding the antibody to its epitope (generally, Clark, WR (1986) The (See Experimental Foundations of Modern Immunology Wiley & Sons, Inc., New York; Roitt, I. (1991) Essential Immunology, 7th Ed., Blackwell Scientific Publications, Oxford). For example, the Fc 'and Fc regions are complement cascade effectors but are not involved in antigen binding. An antibody that is enzymatically cleaved in the pFc 'region, called an F (ab') 2 fragment, or an antibody produced without the pFc 'region retains the antibody binding site of both normal antibodies. Similarly, an antibody with an Fc region enzymatically cleaved, called a Fab fragment, or an antibody produced without an Fc region retains one antibody binding site of a normal antibody molecule. Proceeding further, the Fab fragment consists of an antibody light chain and a portion of an antibody heavy chain (denoted Fd) covalently linked. Fd fragments are a major determinant of antibody specificity (a single Fd fragment may be accompanied by up to 10 different light chains, although antibody specificity has not changed) In addition, even when the Fd fragment is isolated, it retains the epitope binding ability.

  As is well known in the art, the antibody binding region of an antibody includes a complementarity determining region (CDR) that directly interacts with an epitope of an antigen and a framework region that maintains the tertiary structure of the paratope ( FR) exists (see generally Clark, 1986; Roitt, 1991). Both the heavy chain Fd fragment and the light chain of an IgG immunoglobulin have four framework regions (FR1 to FR4), each separated by three complementarity determining regions (CDR1 to CDRS). CDRs, specifically the CDRS region, more specifically the heavy chain CDRS, are largely involved in antibody specificity.

  It is now well established in the art that non-CDR regions of mammalian antibodies can be replaced with similar regions in homologous or heterologous antibodies while maintaining the epitope specificity of the original antibody. This is most clearly shown in the development and use of “humanized” antibodies in which non-human CDRs are covalently linked to human FRs and / or Fc / pFc ′ regions to produce functional antibodies.

  In certain embodiments, the present invention provides compositions and methods comprising humanized forms of antibodies. As used herein, “humanized” describes an antibody in which some, most or all of the amino acids outside the CDR regions are replaced with corresponding amino acids from a human immunoglobulin molecule. Methods for humanization include US Pat. No. 4,816,567, US Pat. No. 5,225,539, US Pat. No. 5,585,089, US Pat. No. 5,693,761, US Pat. No. 5,693,762 and US Pat. No. 5,859,205, which are incorporated herein by reference. Are included in, but are not limited to. Said US Pat. No. 5,585,089 and US Pat. No. 5,693,761 and WO 90/07861 also propose four possible criteria that can be used in the design of humanized antibodies. The initial proposal is that the acceptor uses a framework from a specific human immunoglobulin that is significantly homologous to the immunoglobulin of the donor to be humanized, or a consensus framework from multiple human antibodies, It was that. The second proposal is to select the donor amino acid over the acceptor if an amino acid in the human immunoglobulin framework is unusual and the donor amino acid at that position is typical in the human sequence It was possible. A third proposal was that at the position immediately adjacent to the three CDRs in the humanized immunoglobulin chain, the donor amino acid could be selected over the acceptor amino acid. The fourth proposal is that in a three-dimensional model of the antibody, the amino acid is predicted to have a side chain atom within 3 cm of the CDR and is in a framework position that is predicted to be able to interact with that CDR. Of amino acids. The above methods are merely illustrative of several methods that one skilled in the art may employ to make humanized antibodies. Those skilled in the art will be familiar with other methods for humanizing antibodies.

  In one embodiment of the humanized form of the antibody, some, most or all of the amino acids outside the CDR regions are replaced with corresponding amino acids from a human immunoglobulin molecule, but within one or more CDR regions. Some, most or all of the amino acids are unchanged. Minor amino acid additions, deletions, insertions or modifications are permissible as long as the antibody does not interfere with the ability to bind to a given antibody. Suitable human immunoglobulin molecules will include IgG1, IgG2, IgG3, IgG4, IgA and IgM molecules. “Humanized” antibodies retain similar antigenic specificity as the original antibody. However, using a humanization method, the “directed evolution” method described by Wu et al. (Mol. Biol. 294: 151, 1999, the contents of which are incorporated herein by reference) is used. Can be used to increase the affinity and / or specificity of antibody binding.

  Fully human monoclonal antibodies can also be prepared by immunizing mice that are transgenic for most of the human immunoglobulin heavy and light chain loci. US Pat. No. 5,591,669, US Pat. No. 5,598,369, US Pat. No. 5,545,806, US Pat. No. 5,545,807 and US Pat. No. 6,150,584, and references cited therein, the contents of which are hereby incorporated by reference. Please refer to. These animals have been genetically modified so that there is a functional defect in the production of endogenous (eg murine) antibodies. These animals are further modified to contain all or part of a human germline immunoglobulin locus so that immunization of these animals results in the production of full-length human antibodies against the antigen of interest. Following immunization of these mice (eg, XenoMouse (Abgenix), HuMAb mice (Medarex / GenPharm)), monoclonal antibodies can be prepared according to standard hybridoma technology. These monoclonal antibodies have human immunoglobulin amino acid sequences and therefore will not elicit a human anti-mouse antibody (KAMA) response when administered to humans.

  There are also in vitro methods for the production of human antibodies. These include phage display technology (US Pat. No. 5,565,332 and US Pat. No. 5,573,905) and in vitro stimulation of human B cells (US Pat. No. 5,229,275 and US Pat. No. 5,567,610). The contents of these patents are incorporated herein by reference.

  Thus, as will be apparent to those skilled in the art, the present invention also includes F (ab ′) 2, Fab, Fv and Fd fragments; Fc and / or FR and / or CDR1 and / or CDR2 and / or light chain CDR3. Chimeric antibody in which the region is replaced with a homologous human or non-human sequence; the FR and / or CDR1 and / or CDR2 and / or light chain CDR3 region is replaced with a homologous human or non-human sequence A chimeric F (ab ′) 2 fragment antibody; a chimeric Fab fragment antibody in which the FR and / or CDR1 and / or CDR2 and / or light chain CDR3 regions are replaced with homologous human or non-human sequences; and FR and / or Or a chimeric Fd fragment in which the CDR1 and / or CDR2 regions are replaced with homologous human or non-human sequences It provides antibodies. The present invention also includes so-called single chain antibodies.

  The various antibody molecules and fragments may be derived from any of the commonly known immunoglobulin classes, including but not limited to IgA, secretory IgA, IgE, IgG and IgM. IgG subclasses are well known to those of skill in the art and include, but are not limited to, human IgG1, IgG2, IgG3 and IgG4.

  In another embodiment, the antibody according to the invention is a single domain antibody. The term “single domain antibody” (sdAb) or “VHH” refers to a single heavy chain variable domain of an antibody of the type that may be found in camelid mammals, originally lacking a light chain. Such VHH is also called “nanobody (registered trademark)”. According to the invention, the sdAb may in particular be a llama sdAb.

  In one embodiment of the reagents described herein, the reagent is a polypeptide. In certain embodiments, the polypeptide is a functional equivalent of orexin-A or orexin-B.

  The term “orexin-A” as used herein has its general meaning in the art and refers to the amino acid sequence shown in SEQ ID NO: 2.

Orexin-A (SEQ ID NO: 2): _p eplpdccrqk tcscrlyell hgagnhaagi ltlx

  The term “orexin-B” as used herein has its general meaning in the art and refers to the amino acid sequence shown in SEQ ID NO: 3.

  Orexin-B (SEQ ID NO: 3): 1 fsgppglqgr lqrllqasgn haagiltm

  As used herein, a “functional equivalent of orexin” is a polypeptide that can bind OX1R and thereby promote OX1R activity according to the present invention. The term “functional equivalent” includes orexin-A and orexin-B fragments, variants and muteins. Thus, the term “functional equivalent” includes an amino acid sequence such that a protein analog retains the ability to bind to OX1R and promote OX1R activity according to the present invention (eg, apoptosis of cancer cells). Or any equivalent of orexin (ie, orexin-A or orexin-B) obtained by, for example, deletion, substitution or addition of one or more amino acids. Amino acid substitutions may be made, for example, by point mutation of DNA encoding the amino acid sequence.

  In some embodiments, the functional equivalent is 80% or more homologous to the corresponding protein. In a preferred embodiment, the functional equivalent is 90% or more homologous when assessed by any conventional analysis algorithm, such as Pileup sequence analysis software (Program Manual for the Wisconsin Package, 1996). The term “functionally equivalent fragment” as used herein may also mean any fragment or collection of fragments of orexin that binds to OX1R and promotes OX1R activity according to the present invention. Accordingly, the present invention provides a polypeptide comprising contiguous amino acids having a sequence corresponding to at least a portion of orexin-A or orexin-B that binds to OX1R and promotes OX1R activity according to the present invention. .

  Functionally equivalent fragments can belong to the same protein family as the human orexins identified herein. “Protein family” means a group of proteins that share a common function and exhibit a common sequence homology. Homologous proteins may be derived from species other than humans. Specifically, the homology rate between functionally equivalent protein sequences over the entire amino acid sequence of the complete protein is 25% or more. More specifically, the homology rate over the entire amino acid sequence of the protein or protein fragment is 50% or more, and more specifically, the homology rate is 75% or more. More specifically, the homology rate over the entire sequence is greater than 80%. More specifically, the homology rate over the entire sequence is greater than 90%. More specifically, the homology rate over the entire sequence is greater than 95%.

  As will be apparent to those skilled in the art, the polypeptides of the present invention may be produced by any suitable means. In order to produce a sufficient amount of a polypeptide or functional equivalent thereof for use in accordance with the present invention, conveniently a recombinant host cell containing the polypeptide of the invention is cultured under suitable conditions. Thus, expression can be achieved. Specifically, the polypeptide is produced by expression from the encoding nucleic acid molecule by recombinant means. Systems for cloning and expression of polypeptides in different host cells are well known. When expressed recombinantly, the polypeptide is specifically produced by expression from the encoding nucleic acid in a host cell. Any host cell may be used depending on the individual requirements of the particular system. Suitable host cells include bacteria, mammalian cells, plant cells, yeast and baculovirus systems. Mammalian cell lines available in the art for expression of heterologous polypeptides include Chinese hamster ovary cells, Hela cells, baby hamster kidney cells and many others. Bacteria are also preferred hosts for recombinant protein production because of the ease with which bacteria can be manipulated and grown. In general, the preferred bacterial host is E. coli.

  In some embodiments, the polypeptide of the present invention is an immunoadhesin.

  The term “immunoadhesin” as used herein refers to an antibody-like molecule that combines the binding specificity of a heterologous protein (“adhesin” capable of binding to OX1R) and the effector function of an immunoglobulin constant domain. . Structurally, an immunoadhesin comprises a fusion of an amino acid sequence with the desired binding specificity for OX1R (ie, “heterologous”) and an immunoglobulin constant domain sequence. The adhesin portion of an immunoadhesin molecule is typically a contiguous amino acid sequence that includes at least a binding site for OX1R. In one embodiment, the adhesin comprises a polypeptide characterized by SEQ ID NO: 2 or SEQ ID NO: 3. The immunoglobulin constant domain sequences in the immunoadhesins are IgG-1, IgG-2, IgG-3 or IgG-4 subtype, IgA (including IgA-1 and IgA-2), IgE, IgD or IgM, It may be obtained from any immunoglobulin.

  The immunoglobulin sequence is typically (but not necessarily) an immunoglobulin constant domain (Fc region). Immunoadhesins can have many of the valuable chemical and biological properties of human antibodies. An immunoadhesin can consist of a sequence of a human protein with the desired specificity, linked to the appropriate human immunoglobulin hinge and constant domain (Fc) sequences, so that the binding specificity of interest can be achieved using all human components. Can be achieved. Such immunoadhesins are minimally immunogenic for the patient and are safe for chronic or repeated use.

  In one embodiment, the Fc region is a native sequence Fc region. In one embodiment, the Fc region is a variant Fc region. In yet another embodiment, the Fc region is a functional Fc region. The term “Fc region” as used herein is used to define the C-terminal region of an immunoglobulin heavy chain and includes native Fc regions and variant Fc regions. Although the boundaries of the Fc region of an immunoglobulin heavy chain can vary, the Fc region of a human IgG heavy chain is usually defined as extending from the amino acid residue at position Cys226 or from Pro230 to its carboxyl terminus. The immunoadhesin adhesion portion and immunoglobulin sequence portion may be linked by a minimal linker. The immunoglobulin sequence is typically (but not necessarily) an immunoglobulin constant domain. The immunoglobulin part in the chimera of the present invention may be derived from IgG1, IgG2, IgG3 or IgG4 subtypes, IgA, IgE, IgD or IgM, but is IgG1 or IgG3.

  Polypeptides of the invention, fragments thereof and fusion proteins according to the invention (eg immunoadhesins) are glycosylated (eg N-linked or O-linked glycosylation), myristylated, palmitylated, acetylated and phosphorylated (eg It may indicate translational modifications including but not limited to serine / threonine or tyrosine.

  In certain embodiments, it is contemplated that the polypeptides used in the therapeutic methods of the invention may be modified to improve their therapeutic effectiveness. Such modifications of the therapeutic compound may be used to reduce toxicity, increase circulation time, or alter biodistribution. For example, in combination with various drug carrier vehicles that alter biodistribution, the toxicity of potential important therapeutic compounds can be significantly reduced. For example, adding a dipeptide can improve the penetration of circulating drugs through the blood-retinal barrier into the eye using endogenous transporters.

  A strategy for improving drug survival is the use of water-soluble polymers. Various water-soluble polymers have been shown to alter biodistribution, improve the mode of cellular uptake, alter permeability through the physiological barrier, and alter clearance rates from the organism. In order to achieve targeting and sustained release effects, water-soluble polymers have been synthesized that contain drug moieties as terminal groups on the polymer chain, as part of the main chain, or as pendant groups.

  Polyethylene glycol (PEG) has been widely used as a drug carrier due to its high biocompatibility and ease of modification. Attachment to various drugs, proteins and liposomes has been shown to improve residence time and reduce toxicity. PEG can be attached to the active agent through the hydroxyl group at the chain end and through other chemical methods, but PEG itself is limited to a maximum of two active agents per molecule. In another approach, copolymers of PEG and amino acids retain the biocompatible properties of PEG but have the added advantage of multiple attachment points per molecule (resulting in greater drug loading). In addition, it has been studied as a new biocompatible material that can be adapted to various applications by synthetic design.

  Those skilled in the art are aware of PEGylation techniques for effective modification of drugs. For example, drug delivery polymers consisting of alternating polymers of PEG and trifunctional monomers such as lysine have been used by VectraMed (Plainsboro, N.J.). PEG chains (typically 2000 Daltons or less) are attached to the α- and e-amino groups of lysine via stable urethane linkages. Such copolymers retain the desirable properties of PEG while providing reactive pendant groups (lysine carboxylic acid groups) at tightly controlled predetermined intervals along the polymer chain. Reactive pendant groups can be used for derivatization, crosslinking or conjugation with other molecules. These polymers are useful in the production of stable, long-circulating prodrugs by altering the molecular weight of the polymer, the molecular weight of the PEG segment, and the cleavable linkage between the drug and the polymer. The molecular weight of the PEG segments affects the spacing of the drug / binding group complex and the amount of drug per conjugate molecular weight (smaller PEG segments result in greater drug loading). In general, increasing the molecular weight of the block copolymer conjugate will increase the circulation half-life of the conjugate. Nevertheless, the conjugate must be easily degraded or have a molecular weight (eg, less than 60 kDa) below threshold-limiting glomular filtration.

  In addition, the prodrug is a prodrug until it is released from the polymer backbone by a specific trigger (typically enzymatic activity) in the target tissue versus the polymer backbone, which is important for maintaining circulation half-life and biodistribution A linker for maintaining the form may be used. For example, this mode of tissue activated drug delivery is particularly useful when delivery to a specific site in the biodistribution is required, where the therapeutic agent is released at or near the site of the lesion. Libraries of linking groups for use in activated drug delivery are known to those skilled in the art and are based on enzyme kinetics, prevalence of the active enzyme and the cleavage specificity of the enzyme specific for the selected disease. It is possible. Such linkers may be used to modify the proteins or protein fragments described herein for therapeutic delivery.

  In one embodiment, the OX1R agonist is an aptamer. Aptamers are a class of molecules that, from a molecular recognition point of view, represent alternatives to antibodies. Aptamers are oligonucleotide or oligopeptide sequences that have the ability to recognize virtually any class of target molecule with a high degree of affinity and specificity. Such ligands can be isolated through a random sequence library Systematic Evolution of Ligands by EXponential enrichment (SELEX). This random sequence library can be obtained by combinatorial chemical synthesis of DNA. In this library, each member is a linear oligomer (finally chemically modified) with a unique sequence. Peptide aptamers consist of a conformationally constrained antibody variable region displayed by a platform protein (such as E. coli thioredoxin A) and are selected from a two-hybrid combinatorial library.

  The term “pancreatic cancer” or “pancreatic cancer” as used herein relates to pancreatic cell-derived cancer. Specifically, pancreatic cancer includes pancreatic adenocarcinoma (eg, pancreatic ductal adenocarcinoma), other cancers of the pancreatic exocrine region (eg, serous cystadenoma), acinar cell carcinoma, intraductal papillary mucinous tumor (IPMN) And pancreatic neuroendocrine tumors (eg insulinomas).

  In some embodiments, a therapeutically effective amount of an OX1R agonist of the invention is administered to the subject.

  “Therapeutically effective amount” means an amount of OX1R sufficient to treat pancreatic cancer at a reasonable risk-to-effect ratio applicable to any medical treatment. It will be appreciated that the total daily usage of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The particular therapeutically effective dosage level for any particular subject is the disorder being treated and the severity of the disorder; the activity of the particular compound used; the particular composition used, the age of the subject, In body weight, general health, sex and diet; administration time, route of administration and excretion rate of specific compounds used; duration of treatment; drugs used in combination or simultaneously with specific polypeptides used; and in the medical field It will vary depending on a variety of factors, including well-known similar factors. For example, it is well known in the art to start the compound dosage at a level lower than that required to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved. Is within the scope of the technology. However, the daily dosage of the product can vary over a wide range, ranging from 0.01 to 1,000 mg (per day per adult). Specifically, for adjustment of symptoms to the subject to be treated, the composition is 0.01, 0.05, 0.1, 0.5, 1.0, 2.5, 5 0.0, 10.0, 15.0, 25.0, 50.0, 100, 250 and 500 mg of active ingredient. The medicament typically contains about 0.01 mg to about 500 mg, specifically 1 mg to about 100 mg of the active ingredient. An effective amount of the drug is usually supplied at a dosage level of from 0.0002 mg / kg to about 20 mg / kg of body weight per day, in particular from about 0.001 mg / kg to 7 mg / kg of body weight per day.

  The OX1R agonists of the present invention may be combined with their pharmaceutically acceptable excipients and optionally a sustained release matrix such as a biodegradable polymer to form a therapeutic composition.

  “Pharmaceutically” or “pharmaceutically acceptable” refers to molecular entities and compositions that do not produce side effects, allergic reactions or other harmful reactions when administered to mammals, particularly humans, as appropriate. Point to. A pharmaceutically acceptable carrier or excipient refers to any form of filler, diluent, encapsulating material or formulation aid that is non-toxic solid, semi-solid or liquid.

  In the pharmaceutical composition for oral, sublingual, intramuscular, intravenous, transdermal, topical or rectal administration of the present invention, the active ingredient alone or in combination with other active ingredients, in unit dosage form, It can be administered to animals and humans as a mixture with conventional pharmaceutical supports. Suitable unit dosage forms include tablets, gel capsules, powders, granules and oral route dosage forms such as oral suspensions or solutions, sublingual and buccal dosage forms, aerosols, implants, subcutaneous, transdermal, topical , Intraperitoneal, intramuscular, intravenous, subcutaneous, transdermal, intrathecal and nasal dosage forms as well as rectal dosage forms.

  Specifically, the pharmaceutical composition contains a pharmaceutically acceptable vehicle for injectable formulations. These are specifically isotonic sterile salt solutions (monosodium phosphate or disodium phosphate, sodium chloride, potassium, calcium or magnesium, etc., or mixtures of such salts), or, optionally, sterile water or saline May be a dry (especially freeze-dried) composition that allows for the construction of an injectable solution.

  Dosage forms suitable for injectable use include sterile aqueous solutions or dispersions; formulations containing sesame oil, peanut oil or aqueous propylene glycol; and sterile powders for the immediate preparation of sterile injectable solutions or dispersions . In any case, the dosage form must be sterile and must be fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi.

  Solutions containing the compounds of the present invention as the free base or pharmacologically acceptable salt can be prepared in water suitably mixed with a surfactant such as hydroxypropylcellulose. Dispersions can also be prepared in glycerin, liquid polyethylene glycols and mixtures thereof, and oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

  The OX1R agonists of the present invention can be formulated into the composition in neutral or salt form. Pharmaceutically acceptable salts include, for example, acid addition salts formed with inorganic acids such as hydrochloric acid or phosphoric acid, or organic acids such as acetic acid, oxalic acid, tartaric acid, mandelic acid, etc. Included). Salts formed with free carboxyl groups also include inorganic bases such as sodium hydroxide, potassium hydroxide, ammonium hydroxide, calcium hydroxide or iron (III), and isopropylamine, trimethylamine, histidine, procaine, etc. It can be derived from an organic base.

  The carrier can also be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetable oils. For example, proper fluidity can be maintained by the use of a coating such as lecithin, by maintenance of the required particle size (in the case of dispersions) and by the use of surfactants. Various antibacterial and antifuCASK agents, such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, etc., can be used to prevent the action of microorganisms. In many cases, it will be preferable to include isotonic agents, for example, sugars or sodium chloride. Prolonged absorption in injectable compositions can be brought about by the use of agents delaying absorption in the composition, such as aluminum monostearate and gelatin.

  Sterile injectable solutions are prepared by adding the required amount of active polypeptide, optionally together with various other ingredients listed above, to a suitable solvent followed by filtered sterilization. Generally, dispersions are prepared by adding the various sterilized active ingredients to a sterilized medium that contains the basic dispersion medium and the other required ingredients from those listed above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred method of preparation is vacuum drying and lyophilization techniques, which combine the active ingredient powder with the additional desired ingredients (if any). Are produced from their previously sterile filtered solutions.

  Once formulated, the solution will be administered in a manner compatible with the dosage formulation and in an amount that is therapeutically effective. The formulation can be easily administered in various dosage forms, such as the injectable solution format, but drug release capsules and the like can also be used.

  For parenteral administration in aqueous solution, for example, the solution should be buffered if necessary, and the liquid diluent should first be made isotonic with sufficient saline or glucose. These particular aqueous solutions are particularly suitable for intravenous, intramuscular, subcutaneous and intraperitoneal administration. In this regard, sterile aqueous media that may be used will be known to those of skill in the art in light of the present disclosure. For example, a single dose could be dissolved in 1 mL of isotonic NaCl solution, added to 1000 mL of subcutaneous infusion fluid, or injected into the proposed infusion site. Some variation in dosage will necessarily occur depending on the condition of the subject being treated. In any event, the person responsible for administration will determine the appropriate dose for the individual subject.

  The OX1R agonist of the present invention is about 0.0001-1.0 milligrams, or about 0.001-0.1 milligrams, or about 0.1-1.0 milligrams (even about 10 milligrams) per dose. Can be formulated into a therapeutic mixture. Multiple doses are also possible.

  In addition to the compounds of the present invention formulated for parenteral administration, such as intravenous or intramuscular injection, other pharmaceutically acceptable forms include, for example, tablets or other solids for oral administration; liposomes Formulations; sustained release capsules; and other currently used forms.

  In some embodiments, the OX1R agonists of the invention are used in combination with chemotherapeutic agents. Chemotherapeutic agents include alkylating agents such as thiotepa and cyclophosphamide; alkyl sulfonates such as busulfan, improsulfan and piperosulfan; such as benzodopa, carbocon, meturedopa and uredopa Aziridines; ethyleneimines and methylamelamines, including altretamine, triethylenemelamine, trietylenephosphoramide, triethiylenethiophosphoramide and trimethylolomelamine; acetogenins (Especially bratacin and bratasinone); camptothecin (including the synthetic analog topotecan); bryostatin; calistatin; CC-1065 (its adzelesin, calzeresin and Cryptophysin (especially cryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin (including the synthetic analogues KW-2189 and CB1-TM1); eluterobin; pancratistatin Sarcodictin; spongistatin; nitrogen such as chlorambucil, chlornafazine, chlorophosphamide, estamustine, ifosfamide, mechloretamine, mechloretamine oxide hydrochloride, melphalan, nobenvicin, phenesterine, prednisotin, trophosphamide, uracil mustard Mustards; nitrosoureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine and ranimustine; antibiotics such as enediyne antibiotics (eg Calicheamicins, especially calicheamicin gamma II and calicheamicin omega II; dynemicins including dynemicin A; bisphosphonates such as clodronate; esperamicins; and neocardinostatin chromophore and related chromoprotein enediyne antibiotics Chromophore), aclacinomycins, actinomycin, anthromycin (authrarnycin), azaserine, bleomycins, kactinomycin, carabicin, caminomycin, cardinophilin, chromomycin, dactinomycin, daunorubicin, Detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin (morpholinodoxorubicin, cyanomorpholinodoxorubicin, 2-pyrrolinodoxorubicin And deoxyxorubicin), epirubicin, esorubicin, idarubicin, marcelomycin, mitomycins such as mitomycin C, mycophenolic acid, nogaramycin, olivomycins, pepromycin, potfiromycin, puromycin, quelamycin , Rhodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, dinostatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); such as denopterin, methotrexate, pteropterin, trimethrexate Folic acid analogues; purine analogues such as fludarabine, 6-mercaptopurine, thiaminpurine, thioguanine; ancitabine, azacitidine Pyrimidine analogues such as 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxyfluridine, enocitabine, furoxyuridine; androgens such as carsterone, drmostanolone propionate, epithiostanol, mepithiostan, test lactone; Anti-adrenal drugs such as trirostan; folic acid supplements such as frolinic acid; acegraton; aldophosphamide glycoside; aminolevulinic acid; eniluracil; amsacrine; vesturacil; bisantren; edatraxate Defofamine; demecorsin; diazicon; efflorithine; ellipticine acetate; gallium nitrate; hydroxyurea; Lonnan; lonidainine; maytansinoids such as maytansine and ansamitocin; mitoguazone; mitoxantrone; mopidanmol; nitraerine; pentostatin; phenamet; 2-ethylhydrazide; procarbazine; PSK polysaccharide complex; razoxan; lysoxin; schizophyllan; spirogermanium; tenuazonic acid; triazicon; 2,2 ', 2 "-trichlorotriethylamine; trichothecenes (especially T-2 toxin, veraccurin A A), Loridin A and Anguidine); Urethane; Vindesine; Dacarbazine; Mannomustine; Mitobronitol; Mitractol; Pipobrommann; Gasitocin (gac) ytosine); arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids such as paclitaxel and doxetaxel; chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; Coordination complex; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; vinorelbine; novantron; teniposide; edatrexate; daunomycin; aminopterin; Topoisomerase inhibitor RFS2000; retinoids such as difluoromethylomithine (DMFO), retinoic acid Capecitabine; and pharmaceutically acceptable salts of said of everything, including but acids or derivatives, but is not limited thereto.

  A further object of the present invention is a method of treating pancreatic cancer in a subject in need thereof, i) determining the expression level of OX1R in a tumor tissue sample obtained from said subject. Ii) comparing the expression level measured in step i) with a reference value; and iii) if the level measured in step i) is higher than the reference value, It relates to a method comprising the step of administering a therapeutically effective amount of an OX1R agonist.

  The expression level of OX1R may be measured by any method known in the art. For example, methods for measuring the amount of mRNA are well known in the art. Typically, nucleic acids contained in a sample (eg, a cell or tissue prepared from a patient) are first extracted according to standard methods, eg, using a lytic enzyme or chemical solution, or manufacturer's instructions To extract with a nucleic acid binding resin. Next, the extracted mRNA is detected by hybridization (eg, Northern blot analysis) and / or amplification (eg, RT-PCR). Preferably, quantitative or semi-quantitative RT-PCR is preferred. Real-time quantitative or semi-quantitative RT-PCR is particularly advantageous. Alternatively, immunohistochemistry (IHC) methods may be used. IHC specifically provides a method for detecting a target in a sample or tissue specimen in situ. In IHC, the overall cellular integrity of the sample is maintained, thus allowing detection of both the presence and location of the target of interest (ie OX1R). Typically, the sample is fixed in formalin, embedded in paraffin, cut and sectioned for staining and subsequent observation with a light microscope. Current IHC methods use either direct labeling or secondary antibody-based or hapten-based labeling. Examples of known IHC systems include, for example, EnVision ™ (DakoCytomation), Powervision ™ (Immunovision, Springdale, AZ), NBA ™ kit (Zymed Laboratories Inc., South San Francisco, CA), HistoFine® (Nichirei Corp, Tokyo, Japan) is included. In certain embodiments, the tumor tissue section may be incubated with an antibody against OX1R and then mounted on a slide or other support. Microscopic observations may then be performed on the sample mounted on a suitable solid support. For the preparation of micrographs, the section containing the sample may be mounted on a glass slide or other flat support and the presence of the protein of interest may be emphasized by selective staining.

  The “reference value” may be a “threshold value” or a “cutoff value”. Typically, the “threshold” or “cutoff value” can be determined experimentally, empirically or theoretically. As will be appreciated by those skilled in the art, the threshold can also be arbitrarily selected based on experimental and / or clinical status. The threshold must be determined to obtain optimized sensitivity and specificity in light of the test function and the balance of risk versus effect (the clinical significance of false positives and false negatives). Typically, optimized sensitivity and specificity (as well as threshold) can be determined using receiver operating characteristic (ROC) curves based on experimental data. Typically, the threshold is an OX1R expression level (or ratio or score) measured in a tumor tissue sample from one or more subjects having an amount of OX1R sufficient to receive treatment with an OX1R agonist. Is derived from In addition, retrospective measurement of OX1R expression levels (or ratios or scores) in appropriately stored historical subject samples may be used to establish these thresholds.

  A further object of the present invention is a method for screening a drug for treating pancreatic cancer, comprising i) providing a plurality of test substances; ii) determining whether the test substance is an OX1R agonist. And iii) a method comprising the step of positively selecting the test substance which is an OX1R agonist.

  Typically, the screening method of the present invention comprises the step of providing appropriate cells expressing orexin-1 receptor on the surface. Such cells include cells from mammals, yeast, Drosophila or E. coli. Specifically, cells are transfected with a polynucleotide encoding orexin-1 receptor to express the receptor. The expressed receptor is then contacted with a test substance and optionally an orexin-1 receptor ligand (eg orexins) to activate functional responses such as SHP-2 mobilization and induction of cellular cellular apoptosis. Observe the change. Functional assays include El Firar A, Voisin T, Rouyer-Fessard C, Ostuni MA, Couvineau A, Laburthe M. Discovery of a functional immunoreceptor tyrosine-based switch motif in a 7-transmembrane-spanning receptor: role in the orexin receptor OX1R-driven apoptosis. FASEB J. 2009 Dec; 23 (12): 4069-80. Doi: 10.1096 / fj.09-131367. Epub 2009 Aug 6. Specifically, the comparing step may comprise comparing the activity elicited by the test substance with the activity elicited by a well-known OX1R agonist such as orexin. Specifically, a substance that can have an activity similar to or better than that of a known OX1R agonist is positively selected.

  Typically, the screening methods of the invention can also include screening for test substances that can bind to orexin-1 receptor present on the cell surface. Typically, the test substance is labeled (eg with a radioactive label) and the binding is compared to a well-known OX1R agonist such as orexin. The preparation is incubated with labeled OX1R, the test substance complex bound to NGAL is isolated and characterized according to routine methods known in the art. Alternatively, OX1R may be bound to a solid support and binding molecules solubilized from the cells bound to the column and then eluted and characterized according to routine methods. In another embodiment, an intracellular compartment may be prepared from a cell expressing a molecule that binds NGAL, such as a signal transduction or regulatory pathway molecule regulated by NGAL. The preparation is incubated with labeled NGAL in the presence or absence of the candidate compound. The ability of the candidate compound to bind to the binding molecule is reflected in a decrease in binding to the labeled ligand.

  Typically, candidate compounds are selected from the group consisting of small organic molecules, peptides, polypeptides or oligonucleotides.

  A positively selected test substance may be subjected to a further selection step in view of further assaying its properties for the treatment of pancreatic cancer. For example, positively selected candidate compounds may be subjected to further selection steps in view of further assaying their properties in animal models of pancreatic cancer.

  In order to develop drugs that may be useful in the treatment of pancreatic cancer, the assay may be performed using high-throughput screening techniques to identify test substances. High throughput screening techniques may be performed using multi-well plates (eg, 96, 389 or 1536 well plates) to perform multiple assays using automated robotic systems. Thus, a large library of test substances can be assayed in a highly efficient manner. More specifically, stably transfected cells growing in wells of microtiter plates (96 or 384 wells) can be adapted for high throughput screening of compound libraries. The compounds in the library will be applied one at a time in an automated fashion to the wells of the microtiter dish containing the transgenic cells. Once test substances that activate apoptotic signals are identified, they can be positively selected for further characterization. These assays offer several advantages. Exposing the test substance to whole cells allows its activity to be evaluated in the natural situation where the test substance can act. Because this assay can be easily performed in a microtiter plate format, the described assay can be performed by an automated robotic system, which can test a large number of samples within a reasonably short period of time. Enable. The assay of the present invention can be used in a screen to assess the activity of a compound or extract that has not been previously tested, in which case a single concentration is tested and compared to a control. The assay of the present invention can also be used to assess the relative potency of compounds by testing a range, eg, 100 μM-1 μM, and calculating the concentration at which apoptosis is maximal.

  The invention is further illustrated by the following figures and examples. However, these examples and drawings should not be construed as limiting the scope of the present invention in any manner.

FIG. 1 shows tumor shrinkage induced by orexin-A injection in nude mice xenografted with AsPC-1 cells.

Examples Orexin is a hypothalamic peptide involved in sleep / wake control. We have shown that orexin promotes active apoptosis in colorectal cancer cells. Cell death is mediated by the orexin 1 receptor (OX1R) through the original mechanism involving the presence of two ITIMs (immunoreceptor tyrosine inhibitory motif) in the OX1R sequence and the recruitment and activation of the tyrosine phosphatase SHP-2 Is done. OX1R, a class IGPCR, is abnormally expressed in primary colorectal tumors and liver metastases. Pancreatic ductal adenocarcinoma (PAC) is a very malignant tumor with poor prognosis. Chemotherapy treatments show poor response rates. We have shown OX1R expression in most pancreatic adenocarcinomas by immunohistochemistry, suggesting ectopic expression of OX1R in 98% of the tested PACs.

  The purpose of this study was to investigate the presence of OX1R in 1 / human PAC cell lines and analyze the effects of orexin-A associated with apoptosis; 2 / OX1R to investigate tumor growth in response to orexin-A It was to develop an in vivo ectopic xenograft model from a cell line expressing. In three PAC cell lines (AsPC-1, HPAF-II and SW1990), the expression of OX1R was examined at the level of mRNA (RT-PCR), protein (immunocytochemistry) and function. The development of an animal model (ectopic xenograft) from a cell line expressing OX1R has made it possible to investigate the effects of orexin-A on tumor growth. Resected tumors were analyzed by immunohistochemistry. Only AsPC-1 cells express OX1R.

  Treatment with orexin-A promoted cell growth inhibition by 32% by promoting mitochondrial apoptosis. We used the SHP inhibitor NSC-87877 to demonstrate the ability of this inhibitor to reverse orexin-induced apoptosis in AsPC-1 cells. Orexin-A injection into nude mice xenografted with AsPC-1 cells reduced tumor progression by 49% when treated. All tumors corresponded to poorly differentiated adenocarcinoma expressing cytokeratin 7, CA9 (a marker of hypoxia) and OX1R. Induction of apoptosis was observed in tumors treated with orexin-A (caspase-3 activated).

  In conclusion, this study showed the antitumor and pro-apoptotic effects of orexin in PAC. In this connection, the orexin receptor represents a new promising target in antitumor treatment and / or preclinical diagnosis of the pancreas.

References Throughout this application, various references describe the state of the field to which the present invention relates. The disclosures of these references are incorporated into this disclosure by reference.

Claims (13)

  A method for the treatment of pancreatic cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of an OX1R agonist.   The method of claim 1, wherein the OX1R agonist is a small organic molecule.   The method of claim 1, wherein the OX1R agonist is an antibody.   2. The method of claim 1, wherein the OX1R agonist is selected from the group consisting of a chimeric antibody, a humanized antibody or an all human monoclonal antibody.   The method of claim 1, wherein the OX1R agonist is a polypeptide.   The method of claim 1, wherein the OX1R agonist is a functional equivalent of orexin-A or orexin-B.   The method according to claim 1, wherein the OX1R agonist is a polypeptide having 80% or more identity with SEQ ID NO: 2 or 3.   The method of claim 1, wherein the OX1R agonist is an immunoadhesin.   The method of claim 1, wherein the OX1R agonist is an aptamer.   The pancreatic cancer is a pancreatic adenocarcinoma such as pancreatic duct cancer, serous cystadenoma, acinar cell carcinoma, other tumors of the pancreatic exocrine pancreas such as intraductal papillary mucinous tumor (IPMN), and pancreatic neuroendocrine tumors such as insulinoma The method of claim 1, wherein the method is selected from the group consisting of:   The method of claim 1, wherein the subject is also administered a chemotherapeutic agent.   A method of treating pancreatic cancer in a subject in need thereof, comprising: i) measuring the expression level of OX1R in a tumor tissue sample obtained from said subject; ii) measured in step i) Comparing the expression level with a reference value; and iii) administering to the subject a therapeutically effective amount of an OX1R agonist if the level measured in step i) is higher than the reference value. Method.   A method of screening for a drug for treating pancreatic cancer, comprising i) providing a plurality of test substances; ii) determining whether the test substance is an OX1R agonist; and iii) being an OX1R agonist Selecting the test substance positively.