High Discordance in Plasma and Genital Tract HIV-1 Drug Resistance in Indian Women Failing First-line Therapy MOPDA0106 S. Saravanan, PhD Session Code: MOPDA01 Molecular Techniques of HIV-1 Analysis 21 July 2014

 It is vital to investigate tissues and compartments other than blood for two important reasons (Cu-Uvin S., et al., 2010). From a patient perspective, it is important to determine whether antiretroviral therapy can reduce viral load in non-blood compartments. From a public health perspective, it is critical to know the factors that contribute to the “infectiousness” of an individual in order to devise strategies to reduce the likelihood of transmission. Background:  Much less work has been directed at HIV in non-blood compartments and those compartments may be the potential sanctuary sites harboring HIV and impacting both the transmission and pathogenesis of HIV infection. 2 of 10

 Sequences were aligned (ClustalX) to an Indian subtype C reference (C.IN.AFo67155) and examined for HIV-1 subtype (REGA v2), nucleotide diversity (Highlighter HIV LANL, SLAC in HyPHY) and drug resistance associated mutations (IAS-USA and Stanford HIV Resistance Database). Materials and Methods:  HIV-infected women (n=200) at YRG CARE in Chennai, India, who were adherent on >6 months of first-line antiretroviral therapy were enrolled.  Genital tract (2 Sno-strips in 500uL of NASBA buffer) RNA levels measured using COBAS® AMPLICOR HIV-1 MONITOR Test, v1.5 & Pol genotyping (Saravanan et al., 2009) was conducted in paired detectable samples from both compartments. 3 of 10

Demographic details of enrolled study subjects Results: Patient Characteristics Women on First-line ART (n=200) Viremic (n=73)Non-Viremic (n=127) Age(Mean) years33.8± ±5.2 PVL (Median) log copies/mL4.6 (3, 5.9)Not Detected CD4(Median) cells/µL246 (15, 832)530 (27, 1182) Duration on HAART (Median) Months 35 (7, 114)34 (6, 122) NVP44 (60%)87 (68.5%) D4T/AZT55 (75%)114 (90%) 4 of 10

STUDY SUBJECTS ENROLLED; n = 200 VIREMIC; n=73 (36.5%)NON-VIREMIC; n=127 (63.5%) DETECTABLE GVL n=30/42 (71%) UNDETECTABLE GVL n=12/42 (29%) GVL >2000 Copies/mL n=21/30 (70%) GVL <2000 Copies/mL n=9/30 (30%) PVL >3000 Copies/mL; n=42/73 (57.5%) PVL <3000 Copies/mL; n=31/73 (42.5%) Results: 5 of 10

Genital and Plasma Sequences (n = 21) Concordant mutation patterns; n = 4/21 (19%) Discordant mutation patterns; n = 17/21 (81%) Total patients with additional mutations in genital tract; n=11/21 (52%) 5 NRTI – T215F, M41L, D67DN, K70T, K219E 7 NNRTI- K103E/N, H221Y, V106M, Y188H, E138A, Y181C Results: 6 of 10

Results: p = <0.005 r 2 = (p=ns) Figure 1a: Pearson’s rank correlation for comparison between PVL and GVL in patients with discordance (n=17). 1b: Fisher’s exact test for comparison of patients with detectable and undetectable viral load in plasma and genital compartments (n=42)  95% have monophyletically clustered with Indian subtype C with one sequence clustered to CRF_02 AE Figure 4: Phylogenetic analyses of RT  Women with detectable PVL tend to shed virus in genital secretions (p<0.005) 7 of 10

Results: Figure 6: Comparison between the presence of intermediate to high-level resistance in plasma and genital secretions in patients with discordant mutations (n=5).  Genital discordant mutations were responsible for an increase to a predicted intermediate or high level drug resistance to at least one drug in 24% of women Figure 5: Prevalence of various NRTI and NNRTI DRMs in plasma and genital tract.  High prevalence of M184V in both compartments followed by TAMs. 8 of 10

 If confirmed, GVL and resistance monitoring may need to be considered to prevent sexual and perinatal HIV-1 resistance transmission in countries like India where sexual transmission is the major mode of HIV infection. Conclusion:  High resistance discordance between plasma and the genital tract among South-Indian women failing first-line antiretroviral therapy, suggesting compartmentalization and independent viral evolution. 9 of 10

Acknowledgement IAS International scholarship (S12858) S. Gomathi, M.Sc S. Sivamalar, M.Sc G. Kausalya, M.Sc P. Selvamuthu, MBBS N. Kumarasamy, MBBS, PhD P. Balakrishnan, PhD Suniti Solomon, MD Susan Cu-Uvin, MD Rami Kantor, MD Sunil S. Solomon, MPH, PhD Indian Council of Medical Research (ICMR) under U.S.-India Collaborative Research Supplement # Indo-US/35/2007-ECD-II 10 of 10