Synthesis and Biological Evaluation of 5'- O -Fatty Acyl Ester Derivatives of 3'-Fluoro-2',3'-dideoxythymidine as Potential Anti-HIV Microbicides.

A number of 5'- O -fatty acyl derivatives of 3'-fluoro-2',3'-dideoxythymidine (FLT, 1 ) were synthesized. These conjugates were evaluated for their potential as topical microbicides with anti-HIV activity against cell-free (X4 and R5), cell-associated, and multidrug-resistant viruses. Compared to FLT and 3'-azido-2',3'-dideoxythymidine (AZT), 5'- O -(12-azidododecanoyl) ( 5 ), 5'- O -myristoyl ( 6 ), and 5'- O -(12-thioethyldodecanoyl) ( 8 ) derivatives of FLT were found to be more active against both cell-free viruses (lymphocytotropic and monocytotropic strains) with EC 50 values of 0.4 μM, 1.1 μM, and <0.2 μM, respectively, as well as cell-associated virus with EC 50 values of 12.6, 6.4, and 2.3 μM, respectively. Conjugates 5 , 6 , and 8 exhibited >4 and >30 times better antiviral index than FLT and AZT, respectively. Conjugates 5 and 8 were significantly more potent than FLT against many multidrug-resistant strains. A comparison of the anti-HIV activity with the corresponding non-hydrolyzable ether conjugates suggested that ester hydrolysis to FLT and fatty acids is critical to enable anti-HIV activity. Cellular uptake studies were conducted using fluorescent derivatives of FLT attached with 5(6)-carboxyfluorescein through either β-alanine ( 23 ) or 12-aminododecanoic acid ( 24 ) spacers. The lipophilic fluorescent analog with a long chain ( 24 ) showed more than 12 times higher cellular uptake profile than the fluorescent analog with a short chain ( 23 ). These studies further confirmed that the attachment of fatty acids improved the cellular uptake of nucleoside conjugates. In addition, 5 , 6 , and 8 were the least cytotoxic and did not alter vaginal cell and sperm viability compared to the positive control, a commercial topical spermicide (N-9), which significantly decreased sperm and vaginal cell viability inducing the generation of proinflammatory cytokines.