Synthesis and Computational Studies on Hiv-1 Integrase Inhibitors

HIV-1 integrase (IN) is essential for viral replication and offers a promising target for the development of anti-retroviral drugs. Two decades of extensive research has lead to the recent approval of raltegravir as the first IN inhibitor. Advancement of drug candidate elvitegravir, which is currently in Phase III clinical trial, has furthermore accelerated efforts against this potential target for combating HIV. However, the emergence of resistance against raltegravir and elvitegravir demands exploration of novel chemical scaffolds that could circumvent resistance against currently used HIV-1 IN inhibitors. With the goal of discovering new agents targeting HIV, a novel structural class of HIV-IN inhibitors have been designed and synthesized. Substantial computational studies were also performed that could aid the design and development of potent HIV IN inhibitors. A part of this dissertation research, covered in Chapter 3, details the design, synthesis, and biological evaluation of 3-keto salicylic acid chalcones as novel HIV-1 IN inhibitors. In the chalcone series, the most active compound, 5-bromo-2-hydroxy-3-[3-(2,3,6-trichloro-phenyl)- acryloyl]-2-hydroxybenzoic acid (96) was selectively active against IN strand transfer (ST) with IC50 of 3.7 µM. While most of the compounds exhibit ST selectivity, a few were nonselective, such as 5-bromo-3-[3-(4-bromo-phenyl)-acryloyl]-2-hydroxybenzoic acid (86), which was active against both 3!-processing (3!-P) and ST with IC50 values of 11 ± 4 and 5 ± 2 M, respectively. The compounds also inhibited HIV-1 replication with potencies comparable to their integrase inhibitory activities. Thus, compounds 96 and 86 inhibited HIV-1 replication with EC50 values of 7.3 and 8.7 M, respectively. Chapter 4 describes the synthesis of structurally related amide derivatives which were designed by modification of the chalcone moiety. In the amide series, the most active compound, 5-bromo-3-[(3-chloro-2,4-difluoro benzyl)- carbamoyl]-2-hydroxybenzoic acid (151), inhibited ST with an IC50 of 4 µM. Chapter 5 discloses the synthesis, and biological studies of halogenated phenanthrene #-diketo acids as novel HIV-1 IN inhibitors. The two most active compounds of the series, 4-(8-chlorophenanthren-3-yl)-2,4-dioxobutanoic acid (179) and 4-(6-chlorophenanthren-2-yl)-2,4-dioxobutanoic acid (177) had ST IC50 values of 1.2 and 1.3 M, respectively, and corresponding 3!-P values of 11.0 and 5.0 M. In the last section of the dissertation detailed in Chapter 6, computational studies were conducted with the aim of exploring the possible binding modes of potent IN inhibitors and evaluating the structural requirements for IN inhibition. To determine the physicochemical parameters important for ligand binding, in the first part of this chapter, a PHASE pharmacophore hypothesis was developed and used for molecular alignments in the initial comparative molecular field analysis (CoMFA) and comparative molecular similarity analysis (CoMSIA) 3D-quantitative structure activity relationship (3D-QSAR) modeling of the chalcone derivatives. A recent breakthrough in the field of anti-HIV research was achieved with the crystallization and 3D structure determination of a complete foamy virus IN-DNA complex. To take advantage of the power of structure-based drug design, in the second part of the computational studies, homology models of HIV-1 IN-DNA were constructed based on the foamy virus IN-DNA complex X-ray crystal structure as template through collaboration with the Oak Ridge National Laboratory. The binding modes of raltegravir and elvitegravir in our homology models were in accordance with their binding modes in their complexes with the foamy virus structure. The homology model was then used for docking and 3D-QSAR studies on our synthesized inhibitors and other integrase inhibitors including the clinically available raltegravir and elvitegravir. Free energy calculations using Molecular Mechanics-Generalized Born Surface Area (MM-GBSA) methods were carried out to rescore and validate the binding modes of HIV-1 integrase inhibitors. 3D-QSAR models derived from this study provided detailed insights into the structural requirements for IN inhibition and established predictive tools to guide further inhibitor design. Linear interaction energy (LIE) calculations were also performed to derive energy parameters contributing to the binding free energies of the IN inhibitors in the data set. These energy parameters were also analyzed to gain insight into the binding modes of raltegravir and elvitegravir as well as to validate the conformations of our synthesized chalcone and amide derivatives. The energy terms were then used as descriptors to develop a linear interaction approximation (LIA) activity model for the inhibition of integration catalytic step. In the next section of the chapter, lead optimization was attempted using structure- and ligand-based drug design tools. RACHEL, a drug optimization software, was used to design an inhibitor with desired binding interactions with the IN active site residues. The hit obtained from RACHEL was used to design a structurally related compound (157), the synthesis and activity testing of which has been described in Chapter 4. Docking studies were also performed on the phenanthrene derivatives synthesized in Chapter 5. The docking studies predominantly revealed two binding poses that were distinct from the possible binding modes of clinically used raltegravir and advanced IN inhibitor elvitegravir and, moreover, do not interact significantly with some of the key amino acids (Q148 and N155) implicated in viral resistance. Therefore, this series of compounds can further be investigated as IN inhibitors to circumvent resistance associated with current clinically used HIV-1 IN inhibitors

Discovery of macrocyclic inhibitors of apurinic/apyrimidinic endonuclease 1

Apurinic/apyrimidinic endonuclease 1 (APE1) is an essential base excision repair enzyme that is upregulated in a number of cancers, contributes to resistance of tumors treated with DNA-alkylating or -oxidizing agents, and has recently been identified as an important therapeutic target. In this work, we identified hot spots for binding of small organic molecules experimentally in high resolution crystal structures of APE1 and computationally through the use of FTMAP analysis (http://ftmap.bu.edu/). Guided by these hot spots, a library of drug-like macrocycles was docked and then screened for inhibition of APE1 endonuclease activity. In an iterative process, hot-spot-guided docking, characterization of inhibition of APE1 endonuclease, and cytotoxicity of cancer cells were used to design next generation macrocycles. To assess target selectivity in cells, selected macrocycles were analyzed for modulation of DNA damage. Taken together, our studies suggest that macrocycles represent a promising class of compounds for inhibition of APE1 in cancer cells.This work was supported by grants from the National Institutes of Health (Grant R01CA205166 to M.R.K. and M.M.G. and Grant R01CA167291 to M.R.K.) and by the Earl and Betty Herr Professor in Pediatric Oncology Research, Jeff Gordon Children's Foundation, and the Riley Children's Foundation (M.R.K.). Work at the BU-CMD (J.A.P., L.E.B., R.T.) is supported by the National Institutes of Health, Grant R24 GM111625. D.B. and S.V. were supported by the National Institutes of Health, Grant R35 GM118078. (R35 GM118078 - National Institutes of Health; R01CA205166 - National Institutes of Health; R01CA167291 - National Institutes of Health; R24 GM111625 - National Institutes of Health; Earl and Betty Herr Professor in Pediatric Oncology Research; Jeff Gordon Children's Foundation; Riley Children's Foundation)Accepted manuscriptSupporting documentatio

Synthetic ion transporters: new analytical approaches for the investigation of ion binding and transport

Synthetic ion transporters: new analytical approaches for the investigation of ion binding and transport by Ionut Alexandru Carasel Doctor of Philosophy in Chemistry Washington University in St. Louis, 2010 Dr. George W. Gokel, co-Chair Dr. Kevin Moeller, co-Chair The work reported in this dissertation focuses on synthetic ion transporters: SATs). SATs have a relatively simple chemical structure but they aggregate, self-assemble and insert in biological membranes much in the same way as their much more complex naturally occurring analogs. This makes SATs valuable tools for the investigation of these supramolecular and membrane related processes with the final goal of developing new therapeutical agents useful in the treatment of conditions stemming from ionic imbalances. Two families of synthetic anion transporters are studied in this dissertation: pyrogallol[4]arene derivatives and dianilides of isophthalic and dipicolininc acids. Experiments aimed at investigating their solution behavior, anion binding properties and the strength of the interactions present in the host*anion adducts employed analytical techniques such as high performance liquid chromatography, electrospray mass spectrometry, UV-vis and NMR spectroscopy. Insights derived from these experiments were instrumental to our understanding of the stability and transport mechanisms pertaining to these two families of compounds

An update on the synthesis and reactivity of spiro-fused β-lactams

Beta-Lactam ring-containing compounds play a pivotal role in drug design and synthetic chemistry. Spirocyclic beta-lactams, representing an important beta-lactam subclass, have recently attracted considerable interest with respect to new synthetic methodologies and pharmacological applications. The aim of this manuscript is to review the recent progress made in this field, covering publications disseminated between 2011 to 2018 concerning the synthesis and application of spirocyclic beta-lactams. In the first part, new approaches to the synthesis of spirocyclic beta-lactams, including Staudinger synthesis, cyclization and transformation reactions, will be presented. The reactivity and biological properties of spiro-beta-lactams will be described in the second and third part, respectively

Synthesis and Evaluation of Unnatural HPAA, Norcoclaurine, and Tyramine Analogues

There are several approaches to obtaining unnatural analogues of natural products. One of these approaches is precursor-directed biosynthesis (PDB), which is the process of using natural biosynthetic machinery to transform unnatural analogues of precursor compounds to produce an unnatural analogue of a target natural product. Berberine and galanthamine are chosen as model systems for the evaluation of PDB for the production of aromatic alkaloids in plants. Several approaches to the synthesis of unnatural 4 hydroxyphenylacetaldehyde (HPAA), norcoclaurine, and tyramine analogues are explored. 3-chloro-, 3-bromo-, and 3-iodoHPAA are all synthesized in good yields via and oxidative decarboxylation of tyrosine. These analogues are evaluated for use as unnatural precursors by monitoring their reaction rates with the enzyme norcoclaurine synthase. 3-chloro-,3-bromo-, and 3-iodonorcoclaurine are all synthesized in good yield from tyrosine using a one-pot biomimetic approach from tyrosine via an oxidative decarboxylation of tyrosine followed by a phosphate catalyzed Pictet-Spengler reaction between the HPAA analogue and dopamine. Additionally, the synthesis of 3-chloro- and 3-bromotyramine analogues from 4-hydroxybenzaldehyde is reported

Investigating Endocrine Disrupting Impacts of Nine Disinfection Byproducts on Human and Zebrafish Estrogen Receptor Alpha

Background: Disinfection byproducts (DBPs) cause endocrine disruption via estrogenic or anti-estrogenic effects on estrogen receptors. However, most studies have focused on human systems, with little experimental data being presented on aquatic biota. This study aimed to compare the effects of nine DBPs on zebrafish and human estrogen receptor alpha (zERα and hERα). Methods: In vitro enzyme response-based tests, including cytotoxicity and reporter gene assays, were performed. Additionally, statistical analysis and molecular docking studies were employed to compare ERα responses. Results: Iodoacetic acid (IAA), chloroacetonitrile (CAN), and bromoacetonitrile (BAN) showed robust estrogenic activity on hERα (maximal induction ratios of 108.7%, 50.3%, and 54.7%, respectively), while IAA strongly inhibited the estrogenic activity induced by 17β-estradiol (E2) in zERα (59.8% induction at the maximum concentration). Chloroacetamide (CAM) and bromoacetamide (BAM) also showed robust anti-estrogen effects in zERα (48.1% and 50.8% induction at the maximum concentration, respectively). These dissimilar endocrine disruption patterns were thoroughly assessed using Pearson correlation and distance-based analyses. Clear differences between the estrogenic responses of the two ERαs were observed, whereas no pattern of anti-estrogenic activities could be established. Some DBPs strongly induced estrogenic endocrine disruption as agonists of hERα, while others inhibited estrogenic activity as antagonists of zERα. Principal coordinate analysis (PCoA) showed similar correlation coefficients for estrogenic and anti-estrogenic responses. Reproducible results were obtained from computational analysis and the reporter gene assay. Conclusions: Overall, the effects of DBPs on both human and zebrafish highlight the importance of controlling their differences in responsiveness for estrogenic activities including the water quality monitoring and endocrine disruption, as DBPs have species-specific ligand-receptor interactions.Peer reviewe

Pioneering a novel class of tetrahydroimidazopyridines with anti-proliferative property study against prostate cancer, 2016

The synthesis of tetrahydroimidazo[1,5-a]pyridine (rIMP) through the transfer hydrogenation of imidazo[1,5-a]pyridine (IMP) was successfully developed. The simple two-step procedure is the most viable and efficient method that results in a unique conjugated system with pyridine at the 1-position and functionalized phenyl group at the 3-position. In the synthesis of imidazo[1,5-a]pyridine, the reaction between di-2-pyridil ketone, substituted benzaldehyde and ammonium acetate in acetic acid underN2 was highly successful, resulting in yields ranging from 35-95%. Highest yields were obtained for compounds that had electron withdrawing group on them. The transfer hydrogenation of IMP using recyclable and inexpensive Pd/C catalyst, hydrazine hydrate in ethanol under N2 was shown to be viable and efficient in the synthesis of rIMP. The reaction conditions for the nitro based IMP were optimized to give singly- or doubly- reduced products, amine-IMP or amine-rIMP, respectively. The optimization of reaction conditions in the synthesis of amine-IMP led to the discovery of 1: 3 mol%: 3 eq ratio of IMP, Pd/C and hydrazine hydrate to obtain high yields. An optimal temperature of 90 oC and a reaction duration of 1.5 hr were established. The optimization of reaction condition for the synthesis of amine-rIMP revealed that of 1: 15 mol%: 15 eq ratio of IMP, Pd/C and hydrazine hydrate lead to high yield. The presence of hydroxy, alkyl and methoxy group was unaffected. The optimized condition for synthesis of amine-rIMP was used on other functionalized IMP. Hydrogenolysis was observed for halogenated (bromine- and chlorine-) IMP to give non-functionalized rIMP. The fluorine atom was not affected by the condition and resulted in fluorine-rIMP. The nitrile group was reduced completely to methyl. The ethoxy and acetaimide groups were unaffected. The thiomethyl group was poisonous to the catalyst. Anti-proliferative assay on human prostate cancer cell lines, PC3 and DU145 revealed that rIMP is more potent at inhibiting growth of PC3 cells and IMP more potent at inhibiting growth of DU145 cells. Of the various treatments amine-rIMP were more potent at inhibiting PC3 cells than Me-rIMP, amine-IMP and OH-IMP. Amine at the ortho-position was most potent compared to meta- and para-position. Cell growth of PC3 cells was inhibited in a dose-dependent manner. KEY TERMS: Tetrahydroimidazopyridine, Imidazopyridine, Catalytic Hydrogenation, Reduction by Hydrazine, Anti-Proliferative Property, Prostate Cancer Cells, Heterocyclic Compounds, Laboratory and Basic Science Research, Medicinal Chemistry and Pharmaceutics, Organic Chemical