The integration of linear retrovirus DNA by the viral integrase (IN) into the host chromosome occurs by a concerted mechanism (full-site reaction). IN purified from avian myeloblastosis virus and using retrovirus-like DNA restriction fragments (487 bp in length) as donors and circular DNA (pGEM-3) as the target can efficiently catalyze that reaction. Nonionic detergent lysates of purified human immunodeficiency virus type 1 (HIV-1) virions were also capable of catalyzing the concerted integration reaction. The donor substrates were restriction fragments (469 bp) containing either U3-U5 (H-2 donor) or U5-U5 (H-5 donor) long terminal repeat sequences at their ends. As was shown previously with bacterially expressed HIV-1 IN, the U5 terminus of H-2 was preferred over the U3 terminus by virion-associated IN. The reactions involving two donors per circular target by HIV-1 IN preferred Mg 2؉ over Mn 2؉ . Both metal ions were equally effective for the circular half-site reaction involving only one donor molecule. The linear 3.8-kbp recombinant products produced from two donor insertions into pGEM were genetically selected, and the donor-target junctions of individual recombinants were sequenced. A total of 55% of the 87 sequenced recombinants had host site duplications of between 5 and 7 bp, with the HIV-1 5-bp-specific duplication predominating. The other recombinants that migrated at the linear 3.8-kbp position were mainly small deletions that were grouped into four sets of 17, 27, 40, and 47 bp, each having a periodicity mimicking a turn of the DNA helix. Aprotic solvents (dimethyl sulfoxide and 1,4-dioxane) enhanced both the half-site and the linear 3.8-kbp strand transfer reactions which favored low-salt conditions (30 mM NaCl). The order of addition of the donor and target during preincubation with HIV-1 IN on ice did not affect the quantity of linear 3.8-kbp recombinants relative to that of the circular half-site products that were produced; only the quantity of donor-donor versus donor-target recombinants was affected. The presence of Mg 2؉ in the preincubation mixtures containing donor and target substrates was not necessary for the stability of preintegration complexes on ice or at 22؇C. Comparisons of the avian and HIV-1 concerted integration reactions are discussed.

Eleven A-ring modified hexacyclic analogues of camptothecin (CPT) containing a 1,4-oxazine ring were synthesized from 10-hydroxycamptothecin (11a) and 7-ethyl-10-hydroxycamptothecin (3) (SN-38) in four to five steps and were subjected to the biological tests such as cytotoxicity, topoisomerase I (Topo I) inhibitory activity, acetylcholinesterase (AChE) inhibition, and stability in human plasma. Four compounds 15a, 15b, 16a, and 16c were about 2-fold more potent than topotecan and as potent as CPT toward human cancer cell lines A549, H128, WiDr, MKN45, SK-OV-3, and SK-BR-3 in vitro, even though the most active compound 15b was slightly less potent than SN-38. The potency of Topo I inhibition of these compounds showed relatively good correlation with their cytotoxicity. Most of the compounds exhibited AChE inhibitory activity weaker (9 +/- 2 to 20 +/- 3%) than CPT (23 +/- 5%) or topotecan (20 +/- 4%) and similar to SN-38 (13 +/- 2%), indicating that they might have little effect on causing early diarrhea. The stability of lactone forms of these compounds in human plasma seemed to be much higher than that of CPT and similar to that of topotecan but lower than that of SN-38. Among the new hexacyclic CPT analogues, compound 15b showed higher antitumor activity against human tumor xenograft, WiDr, in nude mice compared to that of SN-38. The most promising compound 15b has been selected for further development.

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Aim: To investigate the pharmacokinetic profile and tissue distribution of a novel phosphodiesterase type 5 inhibitor, 5-ethyl-2-{5-[4-(2-hydroxy-ethyl)-piperazine-1-sulfonyl]-2-propoxy-phenyl}-7-propyl-3,5-dihydro-pyrrolo(3,2-d)pyrimidin-4-one (SK-3530), in rats after administration of the The maximum plasma concentration and area under the curve of the parent SK3530 were 10%-20% compared to those of the total radioactivity. After the oral administration of [ 14 C]SK-3530, the radioactivity was widely distributed in all tissues, and the tissue/plasma ratio of the radioactivity 1 h after administration was calculated as 0.5-2.6 with the exception of excretory organs. A relatively high penetration was shown in the adrenal glands, liver, and lung. In vitro and ex vivo plasma protein binding assay by ultrafiltration showed a considerably high binding rate of more than 97%. Conclusion: SK-3530 was relatively well absorbed in the gastrointestinal tract and showed linear pharmacokinetics over the investigated dose range. SK-3530 had low oral bioavailability due to a high, first-pass metabolism.