Efavirenz, a drug that is still inaccessible to millions of people worldwide, is potent non nucleoside reverse transcriptase inhibitor (NNRTI), is one of the preferred agents used in combination therapy for first-line treatment of the human immunodeficiency virus (HIV). NNRTIs attach to and block an HIV enzyme called reverse transcriptase, by blocking reverse transcriptase; NNRTIs prevent HIV from multiplying and can reduce the amount of HIV in the body. Efavirenz can't cure HIV/AIDS, but taken in combination with other HIV medicines (called an HIV regimen) every day helps people with HIV live longer healthier lives. Efavirenz also reduces the risk of HIV transmission and can be used by children who are suffering from HIV/AIDS. All the above therapeutic uses of efavirenz prompted us to identify the novel and hopefully cost efficient synthetic methodology for the preparation of efavirenz. In this thesis a new synthetic method for asymmetric synthesis of efavirenz is described. This route started from commercially available starting materials and it is first established in traditional batch chemistry and further the parameters transferred to a semi continuous flow protocol for optimization.

Translocation of viral integrase into nucleus is a critical precondition of integration during the life cycle of HIV, a causative agent of Acquired Immunodeficiency Syndromes (AIDS). It has been considered as an important target for the drug development to treat AIDS. In order to understand the detailed mechanisms of integrase-host cell protein complex interactions, we cloned HIV-1 integrase-EGFP into pTRE2hyg as visible tag to monitor the translocation process. When transiently transfected this vector into Tet-off ready HeLa cells, the EGFP: integrase is mainly localized in the nucleus. It has been hypothesized that any drugs that can inhibit the translocation process are novel class of drugs for AIDS treatment. More than 30000 synthetic compounds and 80000 natural products were screened by virtual screening. A total of 34 compounds were obtained and screened for their ability to block the nuclear entry of HIV-1 integrase by monitoring the EGFP fluorescence in the cells by high-throughput live cell imaging. Eight synthetic compounds (DW-IN4, DW-IN5, DW-IN6, DW-IN9, DW-IN15, DW-IN16, DW-IN17, DW-IN21) and one natural product (DW-IN719) were found to block integrase translocation significantly. According to our screening result, six compounds (INNB-1, INNB-2, INNB-3, INNB-4, INNB-5, INNB-6) were designed and synthesized. INNB-1 and INNB-2 had significant inhibition on integrase nuclear translocation. DW-IN6, DWIN719, INNB-1, INNB-2, INNB-3 and INNB-4, showed significant inhibition on P24 production in live virus assay. DW-IN6, INNB-1, INNB-2, INNB-3 and INNB-4 showed significant syncitia formation inhibition in live virus assay. Six compounds (KM7, KM8, KM14, KM30, KM37, KM79) from Kunming were screened as integrase nuclear translocation inhibitors. Using similar cell imaging techniques, we have cloned the GFP-tagged chemokine receptor CXCR4 using the lentivirus transfection system. CXCR4 receptor is a critical co-receptor in CD4 positive lymphocytes mediating the fusion of HIV into the CD4 positive cells. CXCR4-GFP was over-expressed in 293T cells and the results showed that GFP:CXCR4 receptor is expressed at the plasma membrane of the cells. These cells have been used to monitor the blockage of CXCR4 receptor internalization for drug development. Four compounds (KX128, KX166, KX171, KX180) from Kunming showed CXCR4 internalization blockage in imaging assay. The interaction of these compounds with CXCR4 was predicted by molecular docking. KX128 showed significant HIV inhibition in live virus assays. / Gu, Wangang. / Advisers: Pang Chui Shaw; David Chi Cheong Wan. / Source: Dissertation Abstracts International, Volume: 73-06, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 165-179). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Submiltted in fulfillment of the requirements for the degree of Master of Science in the faculty of Science, University of the Witwatersrand, Johannesburg • Johannesburg 1993. / The roles of the highly conserved aspartic acid residues found at positions 443 and 498 within the RNase H domain of Human Immunodeficiency Virus type-1 reverse transcription were investigated by the defined substitution of these residues using site-directed mutagenesis. [Abbreviated Abstract. Open document to view full version] / MT2016

According to the 2012 UNAIDS global report, sub-Saharan Africa hosts 69% of the world’s total population living with HIV, South Africa being the most affected with a reported 24% incidence rate. To date, extensive research is being conducted globally, particularly involving anti-HIV treatment that targets the retroviral enzymes: reverse transcriptase, integrase and protease. The discovery of inhibitors to HIV protease which disrupts virion protein assembly has made this enzyme a prime target of anti-retroviral therapies, thus there exists a concerted research initiative to identify compounds with HIV protease inactivation potential. This study employs HIV protease that is isolated and purified from a genetically modified HIV protease overexpressing Escherichia coli strain to monitor the inhibitory capacity of new lead compounds. Optimized growth conditions for HIV protease production displayed that the use of chemically defined media resulted in higher yields of the enzyme. Recent research studies have shown that peptide-based cage and glycosylated compounds displayed HIV protease inhibitor activity in cell free enzymatic reactions that are comparable to commercially available HIV protease inhibitors. However, in contrast it has also been reported that these inhibitors are inactive in whole T-cell assays, when employing HIV infected CD4 cells. It is a well-known fact that potential new chemical entities that do not possess oral bioavailability, in terms of their absorption properties, are not successful candidates within the drug discovery industry. Following this, the current study was designed to determine if inefficient membrane permeability of these promising anti-HIV protease lead compounds could result in their inactivity in whole T-cell assays. Two different methods were considered, a cell-based method using the Madin Darby Canine Kidney strain I (MDCKI) cell line and a non-cell based method, the parallel artificial membrane permeability assay (PAMPA). MDCKI cells have been extensively used to form monolayers that mimic human intestinal membranes whilst the PAMPA utilizes an artificial lipid membrane composition on a filter support. Data from permeability assays using the novel chemically synthesized inhibitors have been compared to commercially available drugs, antipyrine, metoprolol and caffeine, which displayed efficient membrane permeability characteristics, thereby validating the assay. The results indicated that novel cage-derived and glycosylated peptide inhibitors do not possess sufficient passive diffusion properties which may explain their inactivity in whole T-cell assays. / M.Sc. University of KwaZulu-Natal, Durban 2014.

Indiana University-Purdue University Indianapolis (IUPUI) / The antiretroviral drug efavirenz remains a cornerstone for treatment-naïve HIV patients. Subsequent to the demonstration that efavirenz is a substrate of cytochrome P450 (CYP) 2B6, a number of clinical studies found that the CYP2B6*6 allele is significantly associated with higher efavirenz exposure and/or adverse reactions. However, the mechanism of reduced efavirenz metabolism by this genetic variant is not fully understood and whether this variant exhibits differential susceptibility to metabolic inhibition is also unknown. Ths use of efavirenz is further complicated by the drug interactions associated with it. Therefore, I hypothezised that 1) the CYP2B6*6 allele reduces efavirenz metabolism by altering catalytic properties of CYP2B6; 2) efavirenz alters the pharmacokinetics of co-administered drugs by inhibiting drug metabolizing enzymes. A series of studies was carried out in hepatic microsomal preparations to determine the functional consequences of the CYP2B6*6 allele and to assess inhibition potency of efavirenz on 8 CYPs. The major findings for these studies include: 1) the CYP2B6*6 allele reduces efavirenz metabolism by decreasing substrate binding and catalytic efficiency; 2) functional consequences of the CYP2B6*6 allele appear to be substrate- and cytochrome b5-dependent; 3) the CYP2B6*6 allele confers increased susceptibility to metabolic inhibition; and 4) efavirenz inhibits the activities of CYP2B6, 2C8, 2C9 and 2C19 at therapeutically relevant concentrations. In addition, I explored the hypothesis that the incorporation of in vitro mechanism by which the CYP2B6*6 allele reduced efavirenz metabolism predicts the genetic effect of this allele on efavirenz clearance after a single oral dose by modeling approach. A pharmacogenetics-based in vitro-in vivo extrapolation (IVIVE) model was developed to predict human efavirenz clearance. Taken together, results from this dissertation provide new mechanistic information on how the CYP2B6*6 allale alters substrate metabolism and drug interactions; demonstrate new mechanisms of efavirenz-mediated inhibition interactions; and demonstrate the utility of a pharmacogenetics-based predictive model that can serve as a basis for future studies with efavirenz and other CYP2B6 substrates. Overall these data provide improved understanding of genetic and non-genetic determinant of efavirenz disposition and drug interactions associated with it.