Sphingosine1-phosphate lyase polypeptides, polynucleotides and modulating agents and methods of use therefor

Abstract

Compositions, methods and kits for diagnosing and treating cancer are provided Therapeutic compositions may comprise agents that modulate the expression or activity of a sphingosine-1-phosphate lyase (SPL). Such compositions may be administered to a mammal afflicted with cancer. Diagnostic methods and kits may employ an agent suitable for detecting alterations in endogenous SPL. Such methods and kits may be used to detect the presence of a cancer or to evaluate the prognosis of a known disease. SPL polypeptides, polynucleotides and antibodies are also provided.

1 . An isolated polynucleotide comprising a sequence selected from the group consisting of: 
 (a) a sequence recited in SEQ ID NO:1;    (b) a sequence recited in SEQ ID NO:3;    (c) nucleotide sequences that hybridize to a polynucleotide complementary to either of the foregoing sequences under moderately stringent conditions, wherein the nucleotide sequences encode polypeptides having sphingosine-1-phosphate lyase activity; and    (d) nucleotide sequences that encode a polypeptide encoded by any of the foregoing sequences.    
     
     
         2 . An isolated polynucleotide encoding a polypeptide recited in SEQ ID NO:2, or a variant of such a polypeptide that has sphingosine-1-phosphate lyase activity.  
     
     
         3 . An isolated polynucleotide encoding a polypeptide comprising a sequence recited in SEQ ID NO:4, or a variant of such a polypeptide that has sphingosine-1-phosphate lyase activity.  
     
     
         4 . A recombinant expression vector comprising a polynucleotide according to any one of claims  1 - 3 .  
     
     
         5 . A host cell transformed or transfected with an expression vector according to  claim 4 .  
     
     
         6 . An isolated polynucleotide comprising at least 100 nucleotides complementary to a sequence recited in SEQ ID NO:1 or 3.  
     
     
         7 . A method for preparing a sphingosine-1-phosphate lyase, the method comprising culturing a host cell transformed or transfected with a polynucleotide according to any one of claims  1 - 3  under conditions promoting expression of the polynucleotide and recovering a sphingosine-1-phosphate lyase.  
     
     
         8 . A polypeptide comprising an amino acid sequence encoded by a polynucleotide according to  claim 1 .  
     
     
         9 . A polypeptide comprising an amino acid sequence recited in SEQ ID NO:2 or 4, or a variant thereof, wherein the polypeptide has sphingosine-1-phosphate lyase activity.  
     
     
         10 . A method for identifying an agent that modulates sphingosine-1-phosphate lyase activity, comprising: 
 (a) contacting a candidate agent with cells that express sphingosine-1-phosphate lyase; and    (b) subsequently measuring the level of sphingosine-1-phosphate lyase or mRNA encoding sphingosine-1-phosphate lyase in the cells, relative to a predetermined level in the absence of candidate agent, and therefrom identifying an agent that modulates sphingosine-1-phosphate lyase activity.    
     
     
         11 . A method for identifying an agent that modulates sphingosine-1-phosphate lyase activity, comprising: 
 (a) contacting a candidate agent with a polypeptide comprising a sequence recited in any one of SEQ ID NOs: 2, 4, 6 or 8, or a variant of such a sequence having sphingosine-1-phosphate lyase activity, wherein the step of contacting is carried out under conditions and for a time sufficient to allow the candidate modulator to interact with the polypeptide; and    (b) subsequently measuring the ability of the polypeptide to degrade sphingosine-1-phosphate or a derivative thereof, relative to an ability in the absence of candidate agent, and therefrom identifying an agent that modulates sphingosine-1-phosphate lyase activity.    
     
     
         12 . A method according to  claim 11 , wherein the step of contacting is performed by incubating a cell expressing the polypeptide with the candidate modulator, and wherein the step of measuring the ability to degrade sphingosine-1-phosphate is performed using an in vitro assay and a cellular extract.  
     
     
         13 . A pharmaceutical composition comprising an agent that modulates sphingosine-1-phosphate lyase activity in combination with a pharmaceutically acceptable carrier.  
     
     
         14 . A composition according to  claim 13 , wherein the sphingosine-1-phosphate lyase comprises a sequence recited in any one of SEQ ID NOs:2, 4 or 6.  
     
     
         15 . A composition according to  claim 13 , wherein the agent inhibits expression of an endogenous sphingosine-1-phosphate lyase gene.  
     
     
         16 . A composition according to  claim 15 , wherein the agent comprises a polynucleotide.  
     
     
         17 . A composition according to  claim 13 , wherein the agent inhibits the ability of an endogenous sphingosine-1-phosphate lyase to degrade sphingosine-1-phosphate.  
     
     
         18 . A composition according to  claim 17 , wherein the agent comprises an antibody or an antigen-binding fragment thereof.  
     
     
         19 . A method for modulating sphingosine-1-phosphate lyase activity, comprising contacting a sphingosine-1-phosphate lyase with an effective amount of an agent that modulates sphingosine-1-phosphate lyase activity, wherein the step of contacting is performed under conditions and for a time sufficient to allow the agent and the sphingosine-1-phosphate lyase to interact.  
     
     
         20 . A method according to  claim 19 , wherein the step of contacting is performed by incubating a cell expressing the polypeptide with the agent.  
     
     
         21 . A method according to  claim 19 , wherein the agent inhibits expression of an endogenous sphingosine-1-phosphate lyase gene.  
     
     
         22 . A method according to  claim 21 , wherein the agent comprises a polynucleotide.  
     
     
         23 . A method according to  claim 19 , wherein the agent is capable of inhibiting the ability of a polypeptide comprising a sequence recited in any one of SEQ ID NOs: 2, 4 or 6 to degrade sphingosine-1-phosphate.  
     
     
         24 . A method for inhibiting the growth of a cancer cell, comprising contacting a cancer cell with an agent that inhibits sphingosine-1-phosphate lyase activity.  
     
     
         25 . A method according to  claim 24 , wherein the agent inhibits expression of an endogenous sphingosine-1-phosphate lyase gene.  
     
     
         26 . A method according to  claim 25 , wherein the agent comprises a polynucleotide according to  claim 6 .  
     
     
         27 . A method according to  claim 24 , wherein the agent is capable of inhibiting the ability of a polypeptide comprising a sequence recited in any one of SEQ ID NOs: 2, 4 or 6 to degrade sphingosine-1-phosphate.  
     
     
         28 . A method according to  claim 24 , wherein the cancer cell is a breast cancer cell.  
     
     
         29 . A method for inhibiting the development and/or metastasis of a cancer in a mammal, comprising administering to a mammal an agent that inhibits sphingosine-1-phosphate lyase activity.  
     
     
         30 . A method according to  claim 29 , wherein the agent inhibits expression of an endogenous sphingosine-1-phosphate lyase gene.  
     
     
         31 . A method according to  claim 30 , wherein the agent comprises a polynucleotide according to  claim 6 .  
     
     
         32 . A method according to  claim 29 , wherein the agent is capable of inhibiting the ability of a polypeptide comprising a sequence recited in any one of SEQ ID NOs: 2, 4 or 6 to degrade sphingosine-1-phosphate.  
     
     
         33 . A method according to  claim 29 , wherein the agent is linked to a targeting component.  
     
     
         34 . A method according to  claim 33 , wherein the targeting component is an anti-tumor antibody.  
     
     
         35 . A method according to  claim 33 , wherein the targeting component binds to an estrogen receptor.  
     
     
         36 . A method according to  claim 29 , wherein the mammal is afflicted with breast cancer.  
     
     
         37 . A method for diagnosing a cancer in a mammal, comprising detecting an alteration in an endogenous sphingosine-1-phosphate lyase gene in a sample obtained from a mammal, and therefrom diagnosing a cancer in the mammal.  
     
     
         38 . A method according to  claim 37 , wherein the alteration is a deletion.  
     
     
         39 . A method according to  claim 37 , wherein the cancer is breast cancer and the sample is a breast tumor biopsy.  
     
     
         40 . A method for evaluating a cancer prognosis, comprising determining the presence or absence of an alteration in an endogenous sphingosine-1-phosphate lyase gene in a sample obtained from a mammal afflicted with cancer, and therefrom determining a prognosis.  
     
     
         41 . A method according to  claim 40 , wherein the alteration is a deletion.  
     
     
         42 . A method according to  claim 41 , wherein the deletion comprises amino acid residues 354-433 of SEQ ID NO:4.  
     
     
         43 . A method according to  claim 40 , wherein the cancer is breast cancer and the sample is a breast tumor biopsy.  
     
     
         44 . An isolated antibody that binds to a polypeptide having a sequence recited in any one of SEQ ID NOs: 2, 4 or 6.  
     
     
         45 . A monoclonal antibody that binds to a polypeptide having a sequence recited in any one of SEQ ID NOs: 2, 4 or 6.  
     
     
         46 . An antibody according to  claim 44  or  claim 45 , wherein the antibody inhibits the ability of a polypeptide having a sequence recited in any one of SEQ ID NOs: 2, 4 or 6 to degrade sphingosine-1-phosphate.  
     
     
         47 . A method for detecting sphingosine-1-phosphate lyase in a sample, comprising: 
 (a) contacting a sample with an antibody according to  claim 44  or  claim 45  under conditions and for a time sufficient to allow the antibody to bind to sphingosine-1-phosphate lyase; and    (b) detecting in the sample the presence of sphingosine-1-phosphate lyase bound to the antibody.    
     
     
         48 . A kit for detecting sphingosine-1-phosphate lyase in a sample, comprising an antibody according to  claim 44  or  claim 45  and a buffer or detection reagent.  
     
     
         49 . A kit for detecting an alteration in a sphingosine-1-phosphate gene in a sample, comprising a polynucleotide according to  claim 6  and a detection reagent.  
     
     
         50 . A transgenic animal in which sphingosine-1-phosphate lyase activity is reduced compared to a wild-type animal.  
     
     
         51 . A cell line derived from a transgenic animal according to  claim 50.