Methods and pharmaceutical compositions for modulating autophagy

Abstract

Autophagy is typically activated by starvation, allowing cells and organisms to mobilize their energy reserves. It is known that pharmacological modulation of autophagy represents a therapeutic potential. Here the inventors report that a protein that is released from cells in an unconventional, autophagy-dependent manner, namely, diazepam binding inhibitor (DBI), regulates autophagy. In particular, the inventors demonstrate that DBI inhibits autophagy and that the supply of recombinant DBI to mice enhanced glycolysis, enhanced lipogenesis, and inhibited fatty acid oxidation. The inventors show that neutralisation of DBI by a monoclonal antibody and an active immunization by means of an immunogenic DBI derivative eliciting autoantibodies induce autophagy and lead to metabolic changes that increase starvation-induced weight loss, reduce food intake upon refeeding, and reduce weight gain in response to hypercaloric diets. Accordingly, the present invention relates to methods and pharmaceutical compositions for modulating autophagy based on the modulation of the activity or expression of DBI.

1 . A method of inhibiting autophagy in a subject in need thereof comprising administering to the subject a therapeutically effective amount of an agent that promotes the activity or expression of DBI diazepam binding protein (DBI). 
     
     
         2 . The method of  claim 1  wherein the subject is underweight. 
     
     
         3 . The method of  claim 2  wherein the subject suffers from a wasting disorder. 
     
     
         4 . The method of  claim 2  wherein the subject suffers from anorexia cachexia, anorexia of the aged, anorexia nervosa, cachexia associated with cancer, cachexia associated with AIDS, cachexia associated with heart failure, cachexia associated with cystic fibrosis, cachexia associated with rheumatoid arthritis, cachexia associated with kidney disease, cachexia associated with chronic obstructive pulmonary disease (COPD), cachexia associated with ALS, cachexia associated with renal failure, cachexia associated with aberrant appetite, cachexia associated with fat mass, cachexia associated with energy balance, and/or cachexia associated with involuntary weight loss. 
     
     
         5 . The method of  claim 1  wherein the subject suffers from a disease selected from the group consisting of cancer diseases, neurodegenerative diseases, cardiovascular diseases, infectious diseases, auto-immune diseases and/or inflammatory diseases. 
     
     
         6 . The method of  claim 1  wherein the agent that promotes the activity of DBI comprises a polypeptide comprising i) an amino acid sequence having at least 80% of identity with SEQ ID NO:1, or ii) an amino acid sequence having at least 80% of identity with the amino acid sequence ranging from the amino acid residue at position 17 to the amino acid residue at position 50 in SEQ ID NO:1, or iii) an amino acid sequence having at least 80% of identity with the amino acid sequence ranging from the amino acid residue at position 33 to the amino acid residue at position 50 in SEQ ID NO:1, or iv) an amino acid sequence having at least 80% identity with the amino acid sequence ranging from the amino acid residue at position 43 to the amino acid residue at position 50 in SEQ ID NO:1. 
     
     
         7 . The method of  claim 1  wherein the agent that promotes the expression of DBI is a nucleic acid molecule that encodes the polypeptide of  claim 6 . 
     
     
         8 . The method of  claim 7  wherein the nucleic acid molecule comprises a nucleic acid sequence having at least 50% identity with SEQ ID NO: 2. 
     
     
         9 . The method of  claim 1  wherein the agent that promotes the activity of DBI is a small organic molecule or peptidomimetic that mimics the activity of DBI. 
     
     
         10 . A method of stimulating autophagy in a subject in need thereof comprising administering to the subject a therapeutically effective amount of an agent that inhibits the activity or expression of DBI. 
     
     
         11 . The method of  claim 10  wherein the subject is overweight. 
     
     
         12 . The method of  claim 11  wherein the subject suffers from obesity. 
     
     
         13 . The method of  claim 10  wherein the subject suffers from type 2 diabetes or metabolic syndrome. 
     
     
         14 . The method of  claim 10  wherein the subject suffers from cancer, neurodegenerative disease, infectious disease, pulmonary disease, cystic fibrosis, liver disease, pancreatitis, or a proteinopathy. 
     
     
         15 . The method of  claim 14  wherein the subject suffers from cancer and the method further comprises, after the step of administering, a step of administering to the subject a therapeutically effective amount of a chemotherapeutic agent. 
     
     
         16 . The method of  claim 10  wherein the agent that inhibits the activity of DBI is an antibody or an aptamer directed against DBI. 
     
     
         17 . The method of  claim 16  wherein the antibody is directed against the peptide fragment ranging from the amino acid residue at position 43 to the amino acid residue at position 50 of DBI. 
     
     
         18 . The method of  claim 16  wherein the antibody is a monoclonal chimeric antibody, a monoclonal humanised antibody, or a monoclonal human antibody. 
     
     
         19 . The method of  claim 10  wherein the agent that inhibits the expression of DBI is siRNA, an endonuclease, an antisense oligonucleotide or a ribozyme. 
     
     
         20 . The method of  claim 10  wherein the agent that inhibits the activity of DBI is a vaccine composition that elicits neutralizing autoantibodies against DBI when administered to the subject. 
     
     
         21 . The method of  claim 20  wherein the vaccine composition comprises an antigen comprising a polypeptide comprising i) an amino acid sequence having at least 80% of identity with SEQ ID NO:1, or ii) an amino acid sequence having at least 80% of identity with the amino acid sequence ranging from the amino acid residue at position 17 to the amino acid residue at position 50 in SEQ ID NO:1, or iii) an amino acid sequence having at least 80% of identity with the amino acid sequence ranging from the amino acid residue at position 33 to the amino acid residue at position 50 in SEQ ID NO:1, or iv) an amino acid sequence having at least 80% of identity with the amino acid sequence ranging from the amino acid residue at position 43 to the amino acid residue at position 50 in SEQ ID NO:1. 
     
     
         22 . The method of  claim 21  wherein the polypeptide is conjugated to a carrier protein. 
     
     
         23 . The method of  claim 20  wherein the vaccine composition comprises an adjuvant. 
     
     
         24 . A method of screening a compound suitable for modulating autophagy comprising i) providing a candidate compound ii) determining whether the candidate compound is capable of modulating the activity or expression of DBI and iii) positively selecting the candidate compound which is capable of modulating the activity or expression of DBI. 
     
     
         25 . A method of determining whether a subject is at risk of weight modulation comprising i) determining the level of DBI in a blood sample obtained from the subject, ii) comparing the level determined at step i) with a predetermined reference value and when a differential between the level determined at step i) and the predetermined reference value is determined, administering to the subject a therapeutically effective amount of an agent that modulates the activity or expression of DBI. 
     
     
         26 . A method of treating a non-alcoholic fatty liver disease (NAFLD) in a subject in need thereof comprising administering to the subject a therapeutically effective amount of an agent that inhibits the activity or expression of DBI. 
     
     
         27 . The method of  claim 26  wherein the NAFLD is nonalcoholic steatohepatitis (NASH).