The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug tox... more The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug toxic side effects as well as therapeutic costs. HIV-1 integrase (IN) and reverse transcriptase-associated ribonuclease H (RNase H) are both selective targets for HIV-1 chemotherapy and the identification of dual IN/RNase H inhibitors is an attractive strategy for new drug development. We synthesized newly pyrrolyl derivatives that exhibited good potency against IN and a moderate inhibition of the RNase H function of RT, confirming the possibility to develop dual HIV-1 IN/RNase H inhibitors and obtaining new information for the further development of more effective dual HIV-1 inhibitors.
The increasing efficiency of HAART has helped to transform HIV/AIDS into a chronic disease. Still... more The increasing efficiency of HAART has helped to transform HIV/AIDS into a chronic disease. Still, resistance and drug-drug interactions warrant the development of new anti-HIV agents. We previously discovered hit 6, active against HIV-1 replication and targeting RNase H in vitro. Because of its diketo-acid moiety, we speculated that this chemotype could serve to develop dual inhibitors of both RNase H and integrase. Here, we describe a new series of 1-benzyl-pyrrolyl diketohexenoic derivatives, 7a-y and 8a-y, synthesized following a parallel solution-phase approach. Those 50 analogues have been tested on recombinant enzymes (RNase H and integrase) and in cell-based assays. Approximately half (22) exibited inhibition of HIV replication. Compounds 7b, 7u, and 8g were the most active against the RNase H activity of reverse-transcriptase, with IC50 values of 3, 3, and 2.5 μM, respectively. Compound 8g was also the most potent integrase inhibitor with an IC50 value of 26 nM.
The DNA polymerase and ribonuclease H (RNase H) activities of human immunodeficiency virus type 1... more The DNA polymerase and ribonuclease H (RNase H) activities of human immunodeficiency virus type 1 (HIV-1) are needed for the replication of the viral genome and are validated drug targets. However, there are no approved drugs inhibiting RNase H and the efficiency of DNA polymerase inhibitors can be diminished by the presence of drug resistance mutations. In this context, drugs inhibiting both activities could represent a significant advance towards better anti-HIV therapies. We report on the mechanisms of allosteric inhibition of a newly synthesized isatin-based compound designated as RMNC6 that showed IC 50 values of 1.4 and 9.8 μM on HIV-1 RT-associated RNase H and polymerase activities, respectively. Blind docking studies predict that RMNC6 could bind two different pockets in the RT: one in the DNA polymerase domain (partially overlapping the non-nucleoside RT inhibitor [NNRTI] binding pocket), and a second one close to the RNase H active site. Enzymatic studies showed that RMNC6 interferes with efavirenz (an approved NNRTI) in its binding to the RT polymerase domain, although NNRTI resistance-associated mutations such as K103N, Y181C and Y188L had a minor impact on RT susceptibility to RMNC6. In addition, despite being naturally resistant to NNRTIs, the polymerase activity of HIV-1 group O RT was efficiently inhibited by RMNC6. The compound was also an inhibitor of the RNase H activity of wild-type HIV-1 group O RT, although we observed a 6.5-fold increase in the IC 50 in comparison with the prototypic HIV-1 group M subtype B enzyme. Mutagenesis studies showed that RT RNase H domain residues Asn474 and Tyr501, and in a lesser extent Ala502 and Ala508, are critical for RMNC6 inhibition of the endonuclease activity of the RT, without affecting its DNA polymerization activity. Our results show that RMNC6 acts as a dual inhibi-tor with allosteric sites in the DNA polymerase and the RNase H domains of HIV-1 RT.
The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug tox... more The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug toxic side effects as well as therapeutic costs. HIV-1 integrase (IN) and reverse transcriptase-associated ribonuclease H (RNase H) are both selective targets for HIV-1 chemotherapy and the identification of dual IN/RNase H inhibitors is an attractive strategy for new drug development. We synthesized newly pyrrolyl derivatives that exhibited good potency against IN and a moderate inhibition of the RNase H function of RT, confirming the possibility to develop dual HIV-1 IN/RNase H inhibitors and obtaining new information for the further development of more effective dual HIV-1 inhibitors.
HIV-1 reverse transcriptase (RT) is still an extremely attractive pharmaceutical target for the i... more HIV-1 reverse transcriptase (RT) is still an extremely attractive pharmaceutical target for the identification of new inhibitors possibly active on drug resistant strains. Medicinal plants are a rich source of chemical diversity and can be used to identify novel scaffolds to be further developed by chemical modifications. We investigated the ability of the main lignans from Schisandra chinensis (Turcz.) Baill. fruits, commonly used in Traditional Chinese Medicine, to affect HIV-1 RT functions. We purified 6 lignans from Schisandra chinensis fruits and assayed their effects on HIV-1 RT and viral replication. Among the S. chinensis fruit lignans, Schisandrin B and Deoxyschizandrin selectively inhibited the HIV-1 RT-associated DNA polymerase activity. Structure activity relationship revealed the importance of cyclooctadiene ring substituents for efficacy. In addition, Schisandrin B was also able to impair HIV-1 RT drug resistant mutants and the early phases of viral replication. We identified Schisandrin B and Deoxyschizandrin as new scaffold for the further development of novel HIV-1 RT inhibitors.
Chembiochem : a European journal of chemical biology, Jan 11, 2015
HIV-1 integrase (IN) active site inhibitors are the latest class of drugs approved for the HIV tr... more HIV-1 integrase (IN) active site inhibitors are the latest class of drugs approved for the HIV treatment. The selection of IN strand transfer drug-resistant HIV strains in patients supports the development of new agents active as allosteric IN inhibitors. Herein, a docking-based virtual screening was applied on a small library of natural ligands to identify novel allosteric IN inhibitors targeting the sucrose binding pocket. From theoretical studies Kuwanon-L emerged as the most promising binder and was thus selected for biological studies. Biochemical studies showed that Kuwanon-L is able to inhibit the HIV-1 IN catalytic activity in absence and in presence of LEDGF/p75 protein, the IN dimerization and the IN/LEDGF binding.Kuwanon-L also inhibited the HIV-1 replication in cell culture. Overall, docking and biochemical results suggest that Kuwanon-L binds to an allosteric binding pocket and can be considered an attactive lead for the development of new allosteric IN antiviral agents.
The degradative activity of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase... more The degradative activity of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT), termed ribonuclease H (RNase H), which hydrolyzes the RNA component of the heteroduplex RNA:DNA replication intermediate, is an excellent target for drug discovery. Anthraquinones (AQs) and their derivatives, which are common secondary metabolites occurring in bacteria, fungi, lichens and a large number of families in higher plants, have been reported to have several biological activities including that of inhibiting HIV-1 RT activities in biochemical assays. We have assayed new AQ derivatives on HIV-1 RNase H activities in biochemical assays. Six series of new AQ derivatives with various substituents at positions 1, 2, 3 and 4 of the AQ ring were tested, and new analogs able to inhibit HIV-1 RT-associated RNase H activity in the low micromolar range were found. Our results demonstrate that AQ derivatives are promising anti-RNase H inhibitors.
During Ebola virus (EBOV) infection, the type I interferon α/β (IFN-α/β) innate immune response i... more During Ebola virus (EBOV) infection, the type I interferon α/β (IFN-α/β) innate immune response is suppressed by EBOV viral protein 35 (VP35), a validated drug target. Identification of EBOV VP35 inhibitors requires a cellular system able to assess the VP35-based inhibitory functions of viral double-stranded RNA (dsRNA) IFN-β induction. We established a miniaturized luciferase gene reporter assay in A549 cells that measures IFN-β induction by viral dsRNA and is dose-dependently inhibited by VP35 expression. When compared to influenza A virus NS1 protein, EBOV VP35 showed improved inhibition of viral dsRNA-based IFN-β induction. This assay can be used to screen for EBOV VP35 inhibitors.
The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug tox... more The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug toxic side effects as well as therapeutic costs. HIV-1 integrase (IN) and reverse transcriptase-associated ribonuclease H (RNase H) are both selective targets for HIV-1 chemotherapy and the identification of dual IN/RNase H inhibitors is an attractive strategy for new drug development. We synthesized newly pyrrolyl derivatives that exhibited good potency against IN and a moderate inhibition of the RNase H function of RT, confirming the possibility to develop dual HIV-1 IN/RNase H inhibitors and obtaining new information for the further development of more effective dual HIV-1 inhibitors.
The increasing efficiency of HAART has helped to transform HIV/AIDS into a chronic disease. Still... more The increasing efficiency of HAART has helped to transform HIV/AIDS into a chronic disease. Still, resistance and drug-drug interactions warrant the development of new anti-HIV agents. We previously discovered hit 6, active against HIV-1 replication and targeting RNase H in vitro. Because of its diketo-acid moiety, we speculated that this chemotype could serve to develop dual inhibitors of both RNase H and integrase. Here, we describe a new series of 1-benzyl-pyrrolyl diketohexenoic derivatives, 7a-y and 8a-y, synthesized following a parallel solution-phase approach. Those 50 analogues have been tested on recombinant enzymes (RNase H and integrase) and in cell-based assays. Approximately half (22) exibited inhibition of HIV replication. Compounds 7b, 7u, and 8g were the most active against the RNase H activity of reverse-transcriptase, with IC50 values of 3, 3, and 2.5 μM, respectively. Compound 8g was also the most potent integrase inhibitor with an IC50 value of 26 nM.
The DNA polymerase and ribonuclease H (RNase H) activities of human immunodeficiency virus type 1... more The DNA polymerase and ribonuclease H (RNase H) activities of human immunodeficiency virus type 1 (HIV-1) are needed for the replication of the viral genome and are validated drug targets. However, there are no approved drugs inhibiting RNase H and the efficiency of DNA polymerase inhibitors can be diminished by the presence of drug resistance mutations. In this context, drugs inhibiting both activities could represent a significant advance towards better anti-HIV therapies. We report on the mechanisms of allosteric inhibition of a newly synthesized isatin-based compound designated as RMNC6 that showed IC 50 values of 1.4 and 9.8 μM on HIV-1 RT-associated RNase H and polymerase activities, respectively. Blind docking studies predict that RMNC6 could bind two different pockets in the RT: one in the DNA polymerase domain (partially overlapping the non-nucleoside RT inhibitor [NNRTI] binding pocket), and a second one close to the RNase H active site. Enzymatic studies showed that RMNC6 interferes with efavirenz (an approved NNRTI) in its binding to the RT polymerase domain, although NNRTI resistance-associated mutations such as K103N, Y181C and Y188L had a minor impact on RT susceptibility to RMNC6. In addition, despite being naturally resistant to NNRTIs, the polymerase activity of HIV-1 group O RT was efficiently inhibited by RMNC6. The compound was also an inhibitor of the RNase H activity of wild-type HIV-1 group O RT, although we observed a 6.5-fold increase in the IC 50 in comparison with the prototypic HIV-1 group M subtype B enzyme. Mutagenesis studies showed that RT RNase H domain residues Asn474 and Tyr501, and in a lesser extent Ala502 and Ala508, are critical for RMNC6 inhibition of the endonuclease activity of the RT, without affecting its DNA polymerization activity. Our results show that RMNC6 acts as a dual inhibi-tor with allosteric sites in the DNA polymerase and the RNase H domains of HIV-1 RT.
The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug tox... more The development of HIV-1 dual inhibitors is a highly innovative approach aimed to reduce drug toxic side effects as well as therapeutic costs. HIV-1 integrase (IN) and reverse transcriptase-associated ribonuclease H (RNase H) are both selective targets for HIV-1 chemotherapy and the identification of dual IN/RNase H inhibitors is an attractive strategy for new drug development. We synthesized newly pyrrolyl derivatives that exhibited good potency against IN and a moderate inhibition of the RNase H function of RT, confirming the possibility to develop dual HIV-1 IN/RNase H inhibitors and obtaining new information for the further development of more effective dual HIV-1 inhibitors.
HIV-1 reverse transcriptase (RT) is still an extremely attractive pharmaceutical target for the i... more HIV-1 reverse transcriptase (RT) is still an extremely attractive pharmaceutical target for the identification of new inhibitors possibly active on drug resistant strains. Medicinal plants are a rich source of chemical diversity and can be used to identify novel scaffolds to be further developed by chemical modifications. We investigated the ability of the main lignans from Schisandra chinensis (Turcz.) Baill. fruits, commonly used in Traditional Chinese Medicine, to affect HIV-1 RT functions. We purified 6 lignans from Schisandra chinensis fruits and assayed their effects on HIV-1 RT and viral replication. Among the S. chinensis fruit lignans, Schisandrin B and Deoxyschizandrin selectively inhibited the HIV-1 RT-associated DNA polymerase activity. Structure activity relationship revealed the importance of cyclooctadiene ring substituents for efficacy. In addition, Schisandrin B was also able to impair HIV-1 RT drug resistant mutants and the early phases of viral replication. We identified Schisandrin B and Deoxyschizandrin as new scaffold for the further development of novel HIV-1 RT inhibitors.
Chembiochem : a European journal of chemical biology, Jan 11, 2015
HIV-1 integrase (IN) active site inhibitors are the latest class of drugs approved for the HIV tr... more HIV-1 integrase (IN) active site inhibitors are the latest class of drugs approved for the HIV treatment. The selection of IN strand transfer drug-resistant HIV strains in patients supports the development of new agents active as allosteric IN inhibitors. Herein, a docking-based virtual screening was applied on a small library of natural ligands to identify novel allosteric IN inhibitors targeting the sucrose binding pocket. From theoretical studies Kuwanon-L emerged as the most promising binder and was thus selected for biological studies. Biochemical studies showed that Kuwanon-L is able to inhibit the HIV-1 IN catalytic activity in absence and in presence of LEDGF/p75 protein, the IN dimerization and the IN/LEDGF binding.Kuwanon-L also inhibited the HIV-1 replication in cell culture. Overall, docking and biochemical results suggest that Kuwanon-L binds to an allosteric binding pocket and can be considered an attactive lead for the development of new allosteric IN antiviral agents.
The degradative activity of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase... more The degradative activity of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT), termed ribonuclease H (RNase H), which hydrolyzes the RNA component of the heteroduplex RNA:DNA replication intermediate, is an excellent target for drug discovery. Anthraquinones (AQs) and their derivatives, which are common secondary metabolites occurring in bacteria, fungi, lichens and a large number of families in higher plants, have been reported to have several biological activities including that of inhibiting HIV-1 RT activities in biochemical assays. We have assayed new AQ derivatives on HIV-1 RNase H activities in biochemical assays. Six series of new AQ derivatives with various substituents at positions 1, 2, 3 and 4 of the AQ ring were tested, and new analogs able to inhibit HIV-1 RT-associated RNase H activity in the low micromolar range were found. Our results demonstrate that AQ derivatives are promising anti-RNase H inhibitors.
During Ebola virus (EBOV) infection, the type I interferon α/β (IFN-α/β) innate immune response i... more During Ebola virus (EBOV) infection, the type I interferon α/β (IFN-α/β) innate immune response is suppressed by EBOV viral protein 35 (VP35), a validated drug target. Identification of EBOV VP35 inhibitors requires a cellular system able to assess the VP35-based inhibitory functions of viral double-stranded RNA (dsRNA) IFN-β induction. We established a miniaturized luciferase gene reporter assay in A549 cells that measures IFN-β induction by viral dsRNA and is dose-dependently inhibited by VP35 expression. When compared to influenza A virus NS1 protein, EBOV VP35 showed improved inhibition of viral dsRNA-based IFN-β induction. This assay can be used to screen for EBOV VP35 inhibitors.
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Papers by Francesca Esposito