NOVEL ACID-LABILE COL1 TRITYL-LINKED DIFLUORONUCLEOSIDE IMMUNOCONJUGATES - SYNTHESIS, CHARACTERIZATION, AND BIOLOGICAL-ACTIVITY

LY207702 (1) is a difluorinated purine nucleoside that exhibits impres sive antitumor activity in preclinical models. This agent, however, al so possesses cardiotoxicity which limits the potential clinical utilit y of this novel drug candidate. We therefore developed linker chemistr y whereby regioselective NG-tritylation of LY207702 (1) allowed this d rug to be coupled to epsilon-lysine amino groups of mAb's reactive wit h human tumor-associated antigens. The resulting immunoconjugates 3 po ssessed conjugation ratios ranging from 5 to 7 mol of LY207702/mol of mAb, minimal aggregate content (5-10%), and good immunoreactivity. The electronic nature of substituents on the aromatic rings of the trityl group dictated the degree of acid lability of the trityl linker. Incr eased electronic stabilization of the transient trityl carbocation led to increase in the release rate of free drug, i.e., m-DMT 10a = p-DMT 10b > p-MMT 10d > p-T 10f. Consequently, the more acid labile DMT con jugates 3a and 3b proved to be the most potent cytotoxic agents, and t he most stable p-T conjugate 3f exhibited the least antitumor activity when evaluated in vitro and in vivo. p-MeT-linked conjugate 3e, the m ost stable construct that retained excellent in vivo antitumor activit y, was selected for more extensive evaluation. No detectable free drug or metabolite was observed in mouse plasma at a single intravenous do se of p-MeT conjugate 3e, which was consistent with its predicted stab ility under physiological conditions. This construct did, however, exh ibit significant antigen-mediated antitumor activity in vivo. No cardi otoxicity was detected in mice dosed with conjugate 3e (6 mg/kg free d rug content per day for 21 days) equivalent to similar to 8 times the total dose required for complete regression of well-established (simil ar to 1 g) HC1 human colon tumor xenografts in nude mice. Cardiotoxici ty was induced in 20% of free drug 1 treated group at the equivalent d ose. Cardiomyopathy was, however, observed when the dose of conjugate 3e was increased to 8 mg/kg per day for 21 days. These data suggest th at antitumor activity of LY207702 (1) was maintained and its cardiotox ic potential reduced when this agent was administered to human tumor x enograft bearing nude mice as COL1-N6-p-MeT-207702 conjugate 3e.