Design, Synthesis And Biological Evaluation Of New Non-peptide Small Molecules Targeting Gp41As HIV-1Fusion Inhibitors

AIDS is one of the most severe epidmics worldwide. Currently, antiretroviraldrugs in combination have significantly reduced the morbidity and mortality ofHIV/AIDS. However, an increasing number of patients fail to respond to currentantiretrovirual therapeutics due to the emergence of drug resistance to current RTIsand PIs drugs and serious adverse side effects. So development novel anti-HIV drugtargeting different steps of HIV replication cycle is necessary, particularly forinhibiting HIV fusion and entry process.HIV-1envelop glycoprotein transmembrane subunit gp41plays a crucial role inmediate virus fusion and entry into the host cell. During the entry process, three CHRof gp41anti-parallelly fold back with the inner NHR trimer to form a stable six-helixbundle core structure, bringing the viral and host cell membranes into sufficientproximity and fusion. Licensed by FDA in2003, peptide T-20is the first HIV fusioninhibitor for treatment RTIs and PIs failed HIV/AIDS patients. The success of T-20strongly supports the idea that gp41could be effective target for developing newanti-HIV drug. However, the clinical use of peptide drug T-20is limited by its lack ofbioavailability, metabolic instability in vivo and high product cost, therefore, it isessential to develop orally available, non-peptide small-molecule fusion inhibitors.A deep conserved hydrophobic pocket on the surface of gp41inner NHR trimerhas been identified as an attractive drug target, a small molecule binding to this cavityis expected to block the formation of gp416-HB and despair its structural stability,thus inhibit the HIV-1entry and replication. Based on the structural modification of aknown compound NB-64, a new lead N-(3-(1H-tetrazol-5-yl)phenyl)-2,5-dimethylpyrrole A14have been discovered in our previous work, docking studies showed thatA14only partially occupied the hydrophobic pocket, and thus leaving moremodifiable space for developing new inhibitors.This work focused on expanding the molecular size to provide morecomplementary shape and more interaction with the binding pocket, especially withthe critical residue K574, thus enhancing the binding affinity of lead A14.Consequently, various electronegative groups and fragments were successivelyintroduced into the3position on the2,5-dimethyl pyrrole ring according to themolecular modeling and preliminary SAR analysis, Thus, four kinds of targetcompounds with different skeletons were designed and synthesized. The SAR haveinvestigated that:(1) open chain and linear four-ring skeleton structure;(2) negatively charged tetrazolyl and its bioisotere carboxyl on the phenyl ring (A ring);(3) replaceor cut2,5-dimethyl pyrrole (B ring);(4) electronegative rhodanine ring or otherhetero-five-member rings (C ring);(5) N-phenyl rings (D ring) with differentsubstituent, and the CH2linker between C and D ring.Total113synthetic target compounds were synthesized and evaluated againstHIV replication in MT-2cell line and gp416-HB formation. The results indicated thatmany compounds exhibited improved inhibitory activity against6-HB formation andHIV-1replication at low μM level. Among of them, the most active compounds are (1)24a,24j containing a tetrazolyl moiety with low IC50values of4.4and4.6μMagainst gp416-HB formation and low EC50values of3.2and2.2μM, against HIV-1replication in the MT-2cells, respectively;(2)32p,32u containing a carboxyl groupwith low IC50values of1.8and2.6μM for inhibiting gp416-HB and EC50values of0.3and0.8μM against HIV-1replication respectively. Four new active compounds allshowed higher potency than prior lead A14in both assays, thus providing a newstarting point for further structural modifications.Current SAR studies reveal as below:1. A negatively charged tetrazolyl moiety or a carboxylic group on the A ring isnecessary for anti-HIV activity.2.2,5-Dimethyl pyrrole moiety as B ring is favorable for potency. Areplacement of a fluorophenyl moiety or cut the B-ring resulted in the loss ofanti-HIV activity.3. For C ring, the rhodanine moiety prefers to other five-member rings.4. A linear multi-ring skeleton constructed by a double bond between B and Cring is suitable for anti-HIV activity.5. A linker between the C and D rings obviously affected molecular anti-HIVactivity, such as n=0is favorable in the tetrazolyl series, whereas n=1isbetter for the series of compounds containing a carboxylic group.6. Substituents and positions on the D ring are crucial for enhancing potency.These results indicated the direction for our further studies.