Ocular tear growth factor-like protein

Abstract

The present invention relates to a novel lacrimal gland protein (designated lacritin) and the nucleic acid sequences encoding that protein. Lacritin has activity as a growth factor on both human corneal epithelial cells and on the lacrimal acinar cells that produce it Accordingly, one embodiment of the present invention is directed to the use of lacritin to treat Dry Eye and other disorders requiring the wetting of the eye.

1 . A purified polypeptide comprising the amino acid sequence of SEQ ID NO: 4, or an antigenic fragment of SEQ ID NO: 4.  
     
     
         2 . A composition comprising amino acid sequence of SEQ ID NO: 4 and a pharmaceutically acceptable carrier.  
     
     
         3 . The composition of  claim 2 , wherein the composition is a topical ophthalmic formulation.  
     
     
         4 . The composition of  claim 3 , further comprising a pharmaceutically acceptable phospholipid or oil.  
     
     
         5 . A purified nucleic acid sequence that hybridizes to a 100 nucleotide fragment of SEQ ID NO: 1 or its complement under stringent conditions.  
     
     
         6 . A purified nucleic acid sequence comprising a 25 bp nucleic acid sequence that is identical to a contiguous 25 bp sequence of SEQ ID NO: 2 or SEQ ID No: 5.  
     
     
         7 . The nucleic acid sequence of  claim 6  comprising a sequence selected from the group consisting of SEQ ID NO: 3, SEQ ID NO 5 and SEQ ID NO: 6.  
     
     
         8 . A nucleic acid construct comprising a nucleic acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8 operably linked to a heterologous gene.  
     
     
         9 . The nucleic acid construct of  claim 8  wherein said heterologous gene encodes a marker.  
     
     
         10 . The nucleic acid construct of  claim 9 , wherein the marker produces a fluorescent signal.  
     
     
         11 . The nucleic acid construct of  claim 9  wherein said construct is a plasmid.  
     
     
         12 . A transgenic host cell comprising the nucleotide sequence of  claim 8 , 9 or 11.  
     
     
         13 . A nucleic acid construct comprising a nucleic acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8 operably linked to a polylinker.  
     
     
         14 . The nucleic acid construct of  claim 13  wherein said construct is a plasmid.  
     
     
         15 . A purified polypeptide comprising the amino acid sequence of SEQ ID NO: 4, or an amino acid sequence that differs from SEQ ID NO: 4 by one or more conservative amino acid substitutions; or an amino acid sequence that differs from SEQ ID NO: 4 by a single mutation, wherein the single mutation represents a single amino acid deletion, insertion or substitution.  
     
     
         16 . An antibody that binds specifically to the protein of SEQ ID NO: 4.  
     
     
         17 . A method for identifying a test compound capable of modulating lacritin activity, said method comprising: 
 (a) contacting a solution containing lacritin with said test compound; and    (b) measuring lacritin activity in the presence and absence of said test compound, such that if the level obtained in the presence of the test compound differs from that obtained in its absence, then a compound which modulates lacritin gene expression is identified.    
     
     
         18 . The method of  claim 17  wherein the lacritin activity measured is lacritin's ability to promote ductal cell proliferation.  
     
     
         19 . The method of  claim 17  wherein the lacritin activity measured is lacritin's ability to promote tyrosine phosphorylation or calcium signaling.  
     
     
         20 . A method of treating Dry Eye, said method comprising the step of contacting the ocular surface with a composition comprising a polypeptide, wherein the polypeptide comprises the amino acid sequence of SEQ ID NO: 4, or an amino acid sequence that differs from SEQ ID NO: 4 by one or more conservative amino acid substitutions; or an amino acid sequence that differs from SEQ ID NO: 4 by a single mutation, wherein the single mutation represents a single amino acid deletion, insertion or substitution.  
     
     
         21 . A method of enhancing the proliferation of human corneal epithelial cells or lacrimal acinar cells, said method comprising the step of contacting said cells with a composition comprising a polypeptide, wherein the polypeptide comprises the amino acid sequence of SEQ ID NO: 4, or an amino acid sequence that differs from SEQ ID NO: 4 by one or more conservative amino acid substitutions; or an amino acid sequence that differs from SEQ ID NO: 4 by a single mutation, wherein the single mutation represents a single amino acid deletion, insertion or substitution.  
     
     
         22 . A method for detecting a lacritin receptor, said method comprising providing a cell that has been transfected with nucleic acid sequences that encode for potential cell receptors; 
 contacting said transfected cells with lacritin;    detecting cells that display lacritin-dependent calcium signaling;    isolating nucleic acid sequences from the detected cells; and    identifying a nucleic acid sequence that confers lacritin-dependent calcium signaling on cells transfected with that nucleic acid sequence.