Currently, ara-C is the single most effective agent against acute myelocytic leukemia, (AML) however, drug resistance remains a major obstacle to cure. As a first line chemotherapy, 10% of the leukemic patients are resistant to ara-C and of the approximately 65% that enter remission, 80% will relapse at some point and eventually die with drug resistant leukemia. Therefore, a critical need for new therapy is necessary. Current evidence argues that a major mode of ara-C action in the treatment of AML is directed at nuclear DNA metabolism. However, often ara-C acts in a manner that is more rapid than would be expected at the nuclear level alone. Recently, the applicant has shown that an additional site of ara-C action is on phospholipid (PL) metabolism, and signal transduction events involving protein kinase C (PKC). The function of ara- C's action on PL metabolism is to affect the catalytic activity of choline-phosphotransferase, an enzyme in the CDP-choline pathway of phosphatidylcholine biosynthesis, such that phosphatidylcholine and ara- CMP are catalyzed to diglyceride and ara-CDP-choline. The ara-CDP-choline is thought to be a metabolic end product while the diglyceride produced is assumed, at least in part, to alter PKC activity. The applicant has shown that PKC phosphorylates deoxycytidine (dCyd) kinase, an important, rate limiting enzyme in the anabolism of ara-C and other antitumor drugs as well as normal purine and pyrimidine deoxynucleosides, and increases it's catalytic activity. The metabolic products of araC metabolism may have profound effects on membrane integrity by changing the critical balance of lipids necessary to maintain a functional plasma membrane. This proposal tests the hypothesis that dCyd analog anticancer drugs cause cellular lysis of leukemic blasts by altering lipid metabolism and that this dCyd effect has a major impact on cell homeostasis and viability.
The research aims are: l) To investigate the role of cholinephosphotransferase in the metabolism of dCyd analogs and establish affinity constants of cholinephosphotransferase for the various dCyd analogs. 2) To investigate the activity of PKC in response to dCyd analogs. 3) To examine the effects of dCyd analogs on PL metabolism singly and in combination with 3-deazauridine. The goal of this research project is to identify the mechanism by which ara-C and other dCyd analogs affect membrane homeostasis and signal transduction. The alteration in membrane PLs may account in part for leukemic cell lysis seen with high dose ara-C therapy. Since a number of metabolites derived from membrane PLs are involved in signal transduction, the effects of high dose ara-C therapy may be responsible for changes in signal transduction or membrane integrity, or both. The work described in the current proposal will identify dCyd analog anticancer drugs that act upon cellular membranes and signal transduction mechanisms that involve diglyceride. The results from this work should lead to drug modalities that could be more effective in chemotherapy of leukemia and, possibly, some solid tumors.
