Transmitted and Acquired HIV Drug Resistance in Viet Nam Vu Phuong Thao Kellogg College Clinical Medicine A thesis submitted for the degree of Doctor of Philosophy June 2015 Transmitted and Acquired HIV Drug Resistance in Viet Nam Vu Phuong Thao, Kellogg College, Dphil Thesis, Trinity term 2014 Abstract The roll-out of antiretroviral therapy (ART) in Viet Nam along with limited resources for treatment monitoring are expected to be accompanied by the emergence of transmitted and acquired drug resistance. Drug resistance challenges the success of ART program and the efforts to curb the HIV epidemic in Viet Nam. Understanding factors that impact treatment outcome and prevalence and patterns of drug resistance provides imperative information for strategic and effective management. The first part of this thesis aims to study the prevalence and patterns of transmitted drug resistance (TDR) in ART-naïve patients.
TDR prevalence was detected in 6.4% of ARV-naïve patients with HIV-associated tuberculous meningitis initiating ART in Ho Chi Minh City (HCMC) from 2005-2007. This rate is lower than that in developed countries and is comparable to TDR rates reported in similar resource-limited countries. Pattern of TDR reflected the standard first-line ART regimens with nucleotide and non-nucleotide reverse transcriptase inhibitors in Viet Nam. The second part of this thesis aims to investigate factors that impact treatment outcome and drug resistance in second-line ART in Viet Nam.
In a cohort of adult patients on second-line ART at the Hospital for Tropical Diseases, rate of clinical and/or immunological failure was 18.2% after a median follow up of 29 months. Older age, history of injecting drug use, lower CD4 count at second-line ART initiation, suboptimal ART adherence, and previous protease inhibitor (PI) use independently i predicted treatment failure. Prevalence of virological failure (HIV RNA >1000 copies/mL as recommended by the 2013WHO guidelines) in patients who survived and were in active follow up was 9.5%, and high viral load, non- adherence and previous PI use were independent predictors for virological failure to second-line ART. 64% of patients with virological failure carried major PI mutations.
Cross-resistance to third-line medications was higher than reported in other studies with cross resistance to ETR, TPV, and DRV of 55%, 45%, and 27% patients, respectively. This information informs selection of appropriate third-line ART regimen for patients failing second- line ART in Viet Nam. In conclusion, the work of this thesis provides important data on TDR in the chronically HIV-infected population in Viet Nam, provides, for the first time, data on treatment outcome to lopinavir- based second-line ART in the presence of extensive NRTI drug resistance, and identifies modifying risk factors to improve treatment outcomes in Viet Nam. Strategies to diagnose treatment failure accurately, to switch therapy timely, and to provide targeted adherence support will improve the outcomes of patients.
Continued surveillance of TDR should be performed to assure the effectiveness of ART at the population level. Cost-effectiveness studies should be conducted in order to provide evidence for policy makers to decide whether to apply baseline genotypic testing and viral load monitoring in a resource limited country like Viet Nam. Prospective studies are needed to study the validity of WHO immunological/clinical criteria in defining virological treatment failure in PI-based second-line ART. ii Acknowledgements This thesis would not have been possible without the help and support of many people who in one or another way contributed the completion of this study.
With great pleasure and satisfaction I would like to convey my sincere gratitude and thanks to everyone. I would like to express my utmost gratitude to my principal supervisor, Dr Sarah Dunstan who gave me the opportunity to work in the field of clinical research. I am very grateful to you for your guidance, patience, encouragement and providing me excellent atmosphere of doing research and patiently reviewing my thesis. Thank you for your care and kindness.
This work would not be accomplished without the support and supervision from Dr Thuy Le. Under her guidance and supervision I successfully overcame many difficulties and learned a lot. I am very grateful to her for her knowledge and clinical expertise in HIV treatment, her many insightful discussions and suggestions, and her patience in reviewing the thesis. Chị Thùy ơi, you are like my big sister comforting me and walking me through the ups and downs.
Thank you for everything you have shared with me whether it was academic or personal matter. I would like to thank Prof. Menno de Jong, Dr. Suzanne Jurriaans, Dr.
Nicole Back, and Mr. Ronald Rientsma from Amsterdam Medical Center for HIV genotyping and kind hostage. I am thankful to Dr. Török for her collaboration, giving me access to the HIV-associated tuberculosis meningitis clinical trial cohort, and Dr.
Hue for assistance with sample arrangement. I am grateful to Dr Marcel Wolbers at Oxford iii University Clinical Research Unit for his statistical training using R software and for his assistance with the multiple imputation analyses in chapter 4. I would like to acknowledge the doctors and nurses in the HIV Out-patient Clinic at the Hospital for Tropical Diseases for their assistance with enrolling patients in the second- line ART cohorts. Special thanks to Dr.
Vo Minh Quang, head of the clinic for his support and insightful discussions of the project. Most important is my gratitude to the patients who participated in the tuberculous meningitis clinical trial and the second-line cohorts from the Pham Ngoc Thach hospital and the Hospital for Tropical Diseases. Many thanks to all of my colleagues in Oxford University Clinical Research Unit especially Ms Cam Binh, Ms Thuy Hang, Ms Phuong Tu, Ms Ai Hien, Ms Bich Tram, Ms Hoang Thu and Ms Mai Thu for patiently listening to my troubles. I specially thank my parents, sisters and brothers for their unending support.
My mere expression of thanks does not suffice. Last and not least I would like to thank my dearest husband and children for their unconditional love and support during my good and bad times. These past several years have not been an easy ride yet thanks for being by my side, cheering me up and pulling me out of depression and troubles. Finally, I would like to thank everybody who was important to make this thesis possible, as well as expressing my apology that I could not mention personally one by one.
iv Declaration The work described in this thesis is my own work and was conducted under the supervision of Drs Sarah Dunstan and Thuy Le at the Oxford University Clinical Research Unit (OUCRU) – Vietnam. Thesis result chapter 3: I was in charge of the data management, performed all the sequencing, sequence analysis, and the analysis of correlates of clinical outcomes. Thesis result chapter 4: I designed the data collection form, enrolled all the patients, collected data from patients’ clinic chart, generated the clinical databases and performed the data analysis. Thesis result chapter 5: I enrolled and followed all patients, collected data from patients’ clinic chart, generated the clinical databases, performed viral load measurement, sequencing, sequence analysis, and the analysis of resistance mutations and correlates of clinical outcome.
This thesis has not been submitted for a degree or other qualification to this or any other university. v Table of Content Acknowledgements .v Table of Content. vi List of figures. xii List of tables.
xvi List of Publications .2 Human Immunodeficiency virus.2 HIV Life Cycle .3 Subtypes and diversity .3 Course of HIV infection.1 History of HIV treatment .6 HIV drug resistance .1 How does resistance develop? .2 Types of drug resistance .3 Prevalence of drug resistance.4 Impact of resistance to treatment outcome .5 Mechanisms of HIV-1 drug resistance .6 Subtypes and drug resistance .7 HIV drug resistance testing .7 HIV infection in Viet Nam .2 ARV Treatment and monitoring in Viet Nam .3 HIV drug resistance and treatment outcome in Viet Nam .1 Hospital for Tropical Diseases .2 Pham Ngoc Thach Hospital for tuberculosis and lung diseases .2 Molecular-based methods .1 Quantification of HIV by Abbott Real-Time HIV-1 m2000rt .2 In-house drug resistance testing. 76 3 HIV-1 drug resistance in antiretroviral-naïve individuals with HIV-associated tuberculous meningitis initiating antiretroviral therapy in Viet Nam .1 Study settings and population .2 Statement of ethics .3 HIV-1 drug resistance testing .1 Presence of pre-existing HIV drug resistance mutations in chronically- infected subjects with HIV-associated tuberculous meningitis .2 HIV-1 sub-typing and polymorphisms not associated with drug resistance in Ho Chi Minh City, Viet Nam .3 HIV drug resistance development in subjects with virological failure .105 4 Second-Line Antiretroviral Therapy Outcome in HIV-Infected Adults in Ho Chi Minh City, Viet Nam .4 Therapy adherence measurement and Visual Analogue Scale .1 Study population and baseline characteristics .2 Second-line ART and outcome .3 Predictors of time to second-line ART failure .4 Causes and predictors of death .5 Tuberculosis co-infection on second-line ART .6 Virological outcome in patients with protocol-defined treatment failure .134 ix 5 Virological outcome and drug resistance development in HIV-1 infected adults receiving second-line antiretroviral therapy in Ho Chi Minh City, Viet Nam.1 Study population and data collection .2 HIV viral load measurement and genotype testing .1 Study population and baseline characteristics .2 Drug resistance mutations to first-line ART .3 Virological outcome and factors associated with virological failure.4 Genotypic resistance patterns of patients failing second-line ART .5 Management and follow up of patients with virological failure to second-line ART .1 Case report form for clinical data collection.2 Publications resulting from work conducting during DPhil .241 xi List of figures Figure 1.1: Global prevalence of HIV among adults, 2013.3: HIV life cycle.4: Global distribution of HIV-1 subtypes and recombinants.5: Typical course of HIV infection.6: Time to HIV detection for various generations of diagnostic tests, relative to times of symptom onset and detection of p24 antigen and HIV RNA.7: Evolutionary pathways of viral evolution after transmission .8: Interactive map displaying HIV-1 drug resistance in ARV-naive populations from 287 studies between 2000-2013 .9: Subtype CRF01_AE structure.10: Number of adults and children on ART in Viet Nam from 2006 to 2009.1: Description of the HIV-1 partially double-stranded linear probe.1: Visual analogue scales.2: Flowchart of study population and outcome events .3: Kaplan-Meier estimates of proportion of patients with treatment failure .4: CD4+ T-cell progression after second-line treatment .5: Boxplot of Age in injecting drug users (IDU) and non-IDU group.6: Virological outcome in patients with defined treatment failure.1: Flow-chart of study population .2: Prevalence and patterns of ARV resistance mutations in 173 patients at time of second-line ART initiation in Ho Chi Minh City.3: ARV Drug susceptibility among 22 patients experiencing virological failure to second-line ART in Ho Chi Minh City.4: Two year-follow-up for 22 patients with virological failure to second-line ART. 166 xiii List of tables Table 1.1: WHO clinical staging of HIV/AIDS for adults and adolescents .2: WHO preferred first-line therapy in treatment-naïve adults and adolescents .3: WHO preferred second-line options .4: Guidelines for first- and second-line regimens recommended by WHO and Viet Nam MOH.1: Reagents for reverse transcription PCR .2: Primers for reverse transcription and PCR amplification .3: Reagents for PCR amplification .4: Thermal cycles for nested PCR .5: Primers of PCR amplification for sequencing .1: Prevalence of transmitted drug resistance in Viet Nam over time .2: Pre-existing 2009 WHO surveillance drug resistance mutations in individuals with HIV-associated tuberculous meningitis in Ho Chi Minh City from 2005-2007.3: Non-synonymous polymorphisms at 2009 WHO surveillance drug resistance mutation sites in 218 HIV-1 CRF-01_AE isolates from HCMC .4: Non-synonymous polymorphisms at positions not associated with drug resistance based on 2009 WHO surveillance drug resistance mutation list in 218 HIV-1 CRF-01 AE isolates from HCMC .5: Genotypic drug resistance profile of 5 individuals with virological failure .1: Characteristics of 330 patients starting second-line ART at the Hospital for Tropical Diseases in Ho Chi Minh.2: Impact of covariates on time to treatment failure .3: Impact of covariates on time to treatment failure in sensitivity analysis .4: Impact of covariates on time to death .