Drug Resistance Mutations In The Connection Subdomain Of The HIV-1 Reverse Transcriptase
Funder
National Health and Medical Research Council
Funding Amount
$376,710.00
Summary
Human immunodeficiency virus type 1 (HIV-1) infections can be controlled with antiretroviral drugs. In the majority of patients on antiretroviral therapy the virus mutates and is no longer inhibited by the drug. The emergence of drug-resistant HIV-1 is one of the major factors that lead to loss of drug efficacy in patients. Mutations that confer drug resistance have been defined and are specific for different drug classes. Genotype assays that are used to predict drug resistance are routinely us ....Human immunodeficiency virus type 1 (HIV-1) infections can be controlled with antiretroviral drugs. In the majority of patients on antiretroviral therapy the virus mutates and is no longer inhibited by the drug. The emergence of drug-resistant HIV-1 is one of the major factors that lead to loss of drug efficacy in patients. Mutations that confer drug resistance have been defined and are specific for different drug classes. Genotype assays that are used to predict drug resistance are routinely used to guide therapeutic decisions in the treatment of HIV-1 infected individuals. For drugs that target the HIV-1 reverse transcriptase (RT), commonly used genotype kits normally analyse mutations in the first 240 out of 560 amino acids of the reverse transcriptase. This ignores the impact of mutations in other regions of the enzyme, which are potentially important in drug resistance. Recently, mutations that inhibit ribonuclease H function of the HIV-1 RT have been shown to confer high-level resistance to zidovudine, providing the precendent that mutations beyond codon 240 can confer drug resistance. Our analysis of a different region to ribonuclease H called the connection subdomain has demonstrated the presence of mutations that are highly prevalent in drug-treated versus drug naive patients. In this study we will use in vitro assays to define the effect of these mutations on drug resistance and viral fitness . We will also determine the mechanism by which these mutations confer drug resistance. Finally, using our unique database consisting of over 20,000 genotyped samples , we will establish the role of these mutations in the patient. This study is anticipated to identify clinically significant mutations that are present in the RT connection subdomain. Additionally, this study will lead to the development of more accurate genotype assays which will improve the clinical management of HIV infected individuals.Read moreRead less
Molecular Interactions Between The Subunits Of The HIV-1 Reverse Transcriptase
Funder
National Health and Medical Research Council
Funding Amount
$347,625.00
Summary
Human immunodeficiency virus type 1 (HIV-1) infection is a serious public health problem affecting millions of individuals world-wide. HIV-1 infection can be controlled or prevented by antiretroviral drugs. However, these drugs eventually become ineffective because the virus can mutate to become resistant to them. Therefore, it is important to identify new targets in the virus life-cycle for chemotherapeutic intervention. A successful target for anti-HIV-1 drugs has been the reverse transcriptas ....Human immunodeficiency virus type 1 (HIV-1) infection is a serious public health problem affecting millions of individuals world-wide. HIV-1 infection can be controlled or prevented by antiretroviral drugs. However, these drugs eventually become ineffective because the virus can mutate to become resistant to them. Therefore, it is important to identify new targets in the virus life-cycle for chemotherapeutic intervention. A successful target for anti-HIV-1 drugs has been the reverse transcriptase (RT) enzyme, which converts the viral RNA genome into a proviral DNA precursor, and is absolutely essential for virus replication. The RT is a dimer consisting of two polypeptides of 66 and 51kDa, and formation of this heterodimer is required for its enzymatic functions. We believe that destabilisation or enhancement of the RT subunits represents a potential target for chemotherapeutic intervention. We have developed a novel system in yeast which we can use to study this interaction and determine any mutations that either enhance or decrease the interaction between the two RT subunits. These mutations will then be assessed for their effects on RT activity and HIV-1 replication. Using this system we have made the novel discovery that certain types of anti-HIV drugs called nonnucleoside reverse transcriptase inhibitors (NNRTIs) can increase the interaction between the two RT subunits. This suggests that increased subunit interaction may, in part, explain the inhibitory activity of RT dimerisation enhancing NNRTIs. Elucidation of the mechanism and structural basis of this phenomenon is of interest to provide insight into the actions of NNRTIs and into the dimerisation process, neither of which is well understood.Read moreRead less
Worldwide there are approximately 40 million people living with HIV-AIDS. An effective HIV vaccine does not exist at present. Therefore, current strategies to control the HIV pandemic include the use of life saving antiretroviral drugs. While the current drugs are successful in controlling infections, new and more effective agents are needed that inhibit HIV replication by distinct mechanisms due to the inevitable development of drug resistant strains of HIV. The HIV reverse transcriptase enzyme ....Worldwide there are approximately 40 million people living with HIV-AIDS. An effective HIV vaccine does not exist at present. Therefore, current strategies to control the HIV pandemic include the use of life saving antiretroviral drugs. While the current drugs are successful in controlling infections, new and more effective agents are needed that inhibit HIV replication by distinct mechanisms due to the inevitable development of drug resistant strains of HIV. The HIV reverse transcriptase enzyme is essential for HIV replication and has been a successful target for nucleoside reverse transcriptase inhibitors (NRTIs) and nonnucleoside reverse transcriptase inhibitors (NNRTIs). NNRTIs act in part by stabilizing the reverse transcriptase enzyme, thus blocking enzyme function. However, no drugs have been developed that can specifically prevent formation of the reverse transcriptase enzyme, which would result in the production of noninfectious viral particles. We propose that formation of the active reverse transcriptase enzyme, from a large polyprotein called Gag-Pol, proceeds through a homodimer intermediate, which represents an ideal target for blocking reverse transcriptase formation in HIV infected cells. This homodimer intermediate is an attractive target with greater potential for disruption with small molecule inhibitors compared to the mature reverse transcriptase enzyme as it is less stable than the reverse transcriptase found in viruses. This study will determine whether formation of the active RT enzyme is dependent on this intermediate. In addition, we will examine how the reverse transcriptase encoded on Gag-Pol regulates activation of the HIV protease, which is also critical for the formation of infectious virus particles. These studies will increase our understanding of how the virus produces infectious particles and will identify new approaches for targeting the HIV reverse transcriptase enzyme.Read moreRead less