Study on prevalance of drug resistance and genetic mutation for drug resistance HIV on HIV/AIDS patients with first line antiretroviral treatment failure at Dongda hospital Ha Noi

Objectives: To study prevalence of drug resistance and genotype testing for drug resistance HIV on HIV/AIDS patients with first-line antiretroviral treatment failure at Dongda Hospital, Hanoi. Subjects and methods: Cross-sectional, descriptive, prospective study on 47 HIV/AIDS patients with first-line antiretroviral treatment failure at Dongda Hospital, Hanoi during period from 6 - 2011 to 12 - 2016.

Journal of military pharmaco-medicine n08-2017 STUDY ON PREVALANCE OF DRUG RESISTANCE AND GENETIC MUTATION FOR DRUG RESISTANCE HIV ON HIV/AIDS PATIENTS WITH FIRST-LINE ANTIRETROVIRAL TREATMENT FAILURE AT DONGDA HOSPITAL HA NOI Pham Ba Hien*; Do Tuan Anh**; Tran Viet Tien** SUMMARY Objectives: To study prevalence of drug resistance and genotype testing for drug resistance HIV on HIV/AIDS patients with first-line antiretroviral treatment failure at Dongda Hospital, Hanoi Subjects and methods: Cross-sectional, descriptive, prospective study on 47 HIV/AIDS patients with first-line antiretroviral treatment failure at Dongda Hospital, Hanoi during period from - 2011 to 12 - 2016 Results: The prevalence of drug resistance HIV was 90.7%; 97.7% resisted NNRTIs, following NRTIs: 95.3% and PIs group 11.6% Among genetic mutations resistance to NNRTIs: G190A mutation was the highest (51.2%); K103N mutation was 39.5% and Y181C mutation was 34.9% Genetic mutations in NRTIs: M184V mutation was 88.4%; in TAM mutations, K70R mutation was the most common (37.2%); followed D67N, T215F, T69N mutation was 27.9%; 27.9% and 25.6%, respectively Genetic mutations in PIs: M36I and K20R mutation made up 9.3% In NNRTIs, the prevalence of NVP resistance was 95.3% and ETR resitance was 4.7% In NRTIs, lamivudine resistance was 93.0% and AZT resistance was 9.3% No LVP/r resistance was recorded Conclusions: Patients with first-line antiretroviral treatment failure had high prevalence of NNRTIs and NRTIs resistance but still susceptible to PIs * Keywords: HIV; Drug resistance; First-line antiretroviral treatment failure; Genetic mutation for drug resistance INTRODUCTION Thanks to antiretroviral (ARV) drugs that inhibit the growth of HIV, patients treated with ARV can live long and have reasonably heathy lives ARV therapy is recommended due to the capacity assist the immune function and reduces HIVrelated adverse outcomes ARV therapy widely used in patients with indication, contributes to a new pathway in the prevention and treatment of HIV/AIDS by reducing the risk of HIV transmission in the population and improving the quality of life of AIDS patients However, the emergence and widesrpead of drug resistance HIV are inevitable trend due to prolonged treatment Moreover, side effects of ARV are sustainable factors leading to nonadherence to treatment, which contribute to drug resistance HIV and uncontrolled disease in community The development of drug-resistant virus strains predisposes treatment failure and patients who had treatment failure should be changed to use higher-line ARV therapy (higher-level) * Dongda Hospital ** 103 Military Hospital Corresponding author: Pham Ba Hien (phambahien@yahoo.com) Date received: 20/08/2017 Date accepted: 21/09/2017 207 Journal of military pharmaco-medicine n08-2017 In Vietnam, drug-resistance HIV testings are extravagant and are not performed regularly Therefore, to help clinicians optimize the effectiveness of ARV treatment, we performed this study with aims: To determine the proportion of drug resistance and genetic mutation for drug resistance in patients with first-line ARV treatment failure SUBJECTS AND METHODS Subjects 47 HIV patients with fist-line ARV treatment failure in HIV/AIDS outpatient clinic in Dongda Hospital from June, 2011 to December, 2016 were enrolled * Selected criteria: - Age ≥ 18 - Patients were diagnosed HIV positive times, using different commercial kits with different principles and antigens [2] - Patients were treated by first-line ARV (zidovudine + lamivudine + nevirapine or stavudine + lamivudine + nevirapine) and diagnosed treatment failure according to guidelines for diagnosis and treatment HIV/AIDS issue in 2009 [2] - Agree to participate in this study * Exclusion criteria: - Do not come to the examination, test, take medicine by appointment - No cooperation (compliance) in the research process Methods Cross-sectional, descriptive, prospective study * Nucleotid sequencing testing to indentify the drug resistance mutations: 208 Conducted at Molecular Biology Laboratory, National Institute of Hygiene and Epidemiology The study focused on identifying antiretroviral resistance mutations, Pol gene sequences (~ 1,800 bp) including the entire Prot (protease) gene and at least 250 codons of the reverse transcriptase (RT) gene; used Trugene® HIV-1 Genotyping Kit and OpenGene® DNA sequencing system Mutations are recorded when nucleotide mutations occur simultaneously on both downstream and inverted pathways and lead to changes in amino acids Gene sequences were collated with the Stanford University Genomic Database (USA) using the HIVdb (Stanford HIV Drug Resistance Database) software Show each amino acid corresponding to each of Prot and RT sites to detect resistance mutations and estimate resistance levels (potent, low, medium or high) for each drug in the three groups NRTIs, NNRTIs, PIs This drug resistance compiler is based on the International AIDS Society (USA Panel Guidelines) 2010 [6] SPSS 20.0 for Windows program was used for statistical analysis RESULTS AND DISCUSSION The propotion of patients having HIV with genetic mutation for drug resistance Among the 47 patients enrolled, 43 patients had genetic mutation for drug resistance (accounted for 90.7%) Tran Thi Phuong Thuy’s research demonstrated that 5% of patients with HIV-1 treatment Journal of military pharmaco-medicine n08-2017 failure had not resistance mutation [1] The proprotion of these patients was 2.8% in Nguyen Huu Chi’s study [3] Thus, it can be seen that HIV in patients with treatment failure not mean that they resist ARV drugs There are many factors that affect the viral replication, which lead to treatment failure with HAART including poor adherence to ARV therapy, factors related to the pharmacokinetics of drugs such as metabolism for example, absortion, drug interactions; low baseline TCD4 cell count and high pre-treatment viral loads When drug resistance testing are not used, doctor probably determines to unnecessary conversion to second-line regimens Meanwhile, delaying the transition to second-line regimens upon failure of first-line regimen increases the occurrence of new drug-resistant mutations Table 1: Variation in expression of ARV resistance mutation in patients with treatment failure Drug resistance mutations Quantity (n = 43) Percentage % NNRTI resistance mutations 42 97.7 NRTI resistance mutations 41 95.3 PI resistance mutations 11.6 NNRTI + NRTI resistance mutations PI non-resistance mutations 35 81.4 NNRTI + NRTI + PI resistance mutations 11.6 NNRTI resistance mutations NRTI and PI non-resistance mutations 4.7 NRTI resistance mutations NNRTI and PI non-resistance mutations 2.3 Nguyen Huu Chi investigated drugs resistance mutation in 71 treatment failure patients who had drug resistance mutation testing Among them, the highest percentage resistance was seen in NRTI, accounted for 97.2%, followed by NNRTI resistance with 87.3% and PI resistance with 5.6% [3] According to Vietnamese researchers, the rates of PIs resistance were low in our country due to the PI drugs are rarely used and transmitted drug resistance HIV is also rare in Vietnam [3] Adversely, NRTI resistance was the most common as it is the first-choice drug Drug resistance mutations Table 2: The location of NNRTIs resistance mutations Location of mutant gene Quantity The percentage in patiens with drug resistance mutation (n = 43) The percentage in patients with treatment failure (n = 47) A98G 7.0 6.4 M230L 2.3 2.1 K101H 14.0 12.8 K101E 10 23.3 21.3 K101PQ 2.3 2.1 Number 209 Journal of military pharmaco-medicine n08-2017 K101EQ 2.3 2.1 K101HQ 2.3 2.1 K103N 17 39.5 36.2 G190S 2.3 2.1 10 G190A 22 51.2 46.8 11 Y181I 2.3 2.1 12 Y181C 15 34.9 31.9 13 Y181V 2.3 2.1 14 Y188L 7.0 6.4 15 V106I 7.0 6.4 16 V108I 2.3 2.1 17 V179T 7.0 6.4 18 Y179D 2.3 2.1 19 V179E 2.3 2.1 20 P225H 4.7 4.3 G190A is a mutation selected by NVP and EFV G190A is highly resistant to NVP and has a moderate resistance to EFV as well as causing resistance in the NNRTI group This mutation has accumulated over long periods of treatment with all NNRTIs Most of the main resistance, Y181C, G190A, K103N, K101E, K101H were found This is a group with low genetic barriers All mutations were associated with cross-resistance in the NNRTI group The occurrence of these mutations resulted in failure of the NNRTI treatment regimen Therefore, patients with NNRTI resistance mutations should avoid using these agents in the alternative regimen Table 3: The location of NRTIs resistance mutation Quantity The percentage in patiens with drug resistance mutation (n = 43) The percentage in patients with treatment failure (n = 47) D67N* 12 27.9 25.5 D67N* 12 27.9 25.5 V75M 16 37.2 34.0 V75IM 2.3 2.1 V106I 4.7 4.3 V108I 11.6 10.6 V118I 9.3 8.5 K65R 11.6 10.6 10 K70R* 16 37.2 34.0 Number Location of mutant gene 210 Journal of military pharmaco-medicine n08-2017 11 K101Q 4.7 4.3 12 K101H 4.7 4.3 13 K210R 2.3 2.1 14 K219E* 9.3 8.5 15 K219E/Q* 4.7 4.3 16 K219Q* 16.3 14.9 17 T69D 4.7 4.3 18 T69N* 11 25.6 23.4 19 T215Y 4.7 4.3 20 T215F* 12 27.9 25.5 21 T215FS 2.3 2.1 22 T215F/Y 2.3 2.1 23 M41L* 14.0 12.8 24 M184I/V 4.7 4.3 25 M184V 38 88.4 80.9 26 A62V 4.7 4.3 27 A98G 2.3 2.1 28 L74V 7.0 6.4 29 L210W* 11.6 10.6 30 F77L 4.7 4.3 31 F116Y 4.7 4.3 32 Q151M 7.0 6.4 33 E44D 4.7 4.3 34 H208Y 2.3 2.1 * TAM mutation: The rate of M184V mutation was high, accounted for 88.4% This may explain that lamivudine which was a selected drug for this mutation is present in all first-line regimens Lamivudine is a widely used drug in Vietnam for the treatment of chronic hepatitis B, which is a highly prevalent disease in the population This drug is considered to have poor resistance barrier and has a very high rate of drug resistance after initial treatment in chronic hepatitis B [7] In many studies and reports in Vietnam, the incidence of TAM mutations has also been mentioned extensively: in Doan Thu Tra’s study, TAM mutations including the K70R and K219E accounted for 20.25% and 13.95%, respectively [5] According to Huynh Hoang Khanh Thu’s research, the mutation at the D67N site accounted for 37% and was higher than other mutations [4] Tran Thi Phuong Thuy had found that TAM mutation usually occurred at K65R site T215F, K219 E/Q, K70R and 67.4% of patients had both M184V/I and TAM mutations [1] 211 Journal of military pharmaco-medicine n08-2017 Table 4: The location of PIs resistance mutation Number Location of mutant gene Quantity The percentage in drug resistance mutation patients (n = 43) The percentage in treatment failure patients (n = 47) L33F 2.3 2.1 M36I 9.3 8.5 K20R 9.3 8.5 L10I 2.3 2.1 A71V 2.3 2.1 In Vietnam, there was few reports on PIs resistance mutation Studies had shown that PIs resistance mutation has low rate In Doan Thu Tra’s study on patients with L89M, I13V mutation accounted for 6.3% and 5.1%, respectively; Huynh Hoang Khanh Thu indicated that the two main mutants recorded were M46I (0.9%) and V82V (0.9%) [4, 5] Low rates of PIs resistance were a favorable factor in treatment Figure 1: Degree of NNRTIs resistance NVP is a low genetic barrier drug, one mutation in the center of RT (Reverse Transctiptase) activity will occur drug resistance The occurrence of major mutations at high frequency has resulted in the ineffectiveness of NVP (95.3% drug resistance) EFV is substituted for NVP when the patient is allergic to NVP Therefore, the rate of EFV resistance was low with 55.8% due to less common The level of resistance to ETR was lower than that of other drugs in the group because ETR resistance mutations occurred when at least new mutations were accumulated 212 Journal of military pharmaco-medicine n08-2017 Figure 2: Degree of NRTIs resistance The percentage of patients with 3TC resistance was high (93%) Although, there was no case reported with a history using, the similar result was seen in FTC resistance because FTC has similar mechanism to 3TC In patients with combination regimens of d4T or AZT, TAM mutations and secondary mutations occurred, which resulted in multiple drug resistance (41.9% D4T resistance; 9.3% AZT resistance) The rate of patients with TDF resistance was 25.6% Table 5: The degree of PIs resistance (n = 43) Degree of resistance Resistance Ability Non-resistance Drug n % n % n % ATV/r 0 0 43 100 DRV/r 0 0 43 100 FPV/r 0 0 43 100 IDV/r 0 0 43 100 LPV/r 0 0 43 100 NFV 0 0 43 100 SQV/r 0 4.7 41 95.3 TPV/r 0 0 43 100 Among patients with resistant mutation genes to the PI group, however, only two cases were ability to be resistant to SQV/r There were no cases of resistance to other PIs, especially no resistant to LPV/r and ATV/r which were being introduced in second line ARV regimen in Vietnam 213 Journal of military pharmaco-medicine n08-2017 CONCLUSION In 47 patients with first-line ARV therapy failure, 90.7% had HIV with drug resistance genes Among them, the highest resistance rate was seen in NNRTI group with 97.7%, followed by NRTIs (95.3%) and PIs (11.6%) In location of NNRTIs resistance mutation, G190A had the highest rate with 51.2%, followed by K103N, Y181C with 39.5% and 34.9%, respectively In the NRTIresistant mutant genotypes, the highest percentage was seen in M184V accounted for 88.4%; in TAM mutations, K70R mutation was the most common 37.2%, followed by D67N, T215F, T69N with 27.9%; 27.9% and 25.6%, respectively In the location of PIs resistance mutation, M36I and K20R accounted for 9.3% In the NNRTI group, NVP had the highest resistance rate of 95.3% The lowest was the 4.7% ETR In NRTIs, lamivudine had the highest resistance rate (93.0%) and the lowest rate was seen in AZT with 9.3% In PIs, no LVP/r resistance was recorded but might be resistant to SQV/r 214 REFERENCES Tran Thi Phuong Thuy Study on antiretroviral resistance in HIV/AIDS patients treated at the Central Hospital for Tropical Diseases Doctor of Medicine Thesis Institute for Clinical Pharmacomedical Science Research 108 2012 Viet Nam Ministry of Health Guidelines for Diagnosis and Treatment of HIV/AIDS Hanoi Medical Publishing House 2009 Nguyen Huu Chi, Nguyen Tran Chinh, Vo Minh Quang ARV resistance characteristics in AIDS patients failing treatment with HAART at the Tropical Hospital City Medicine Hochiminh 2008, Vol 12 Huynh Hoang Khanh Thu Understanding the antiretroviral resistance (ARV) of HIV in HIV-infected patients Master Thesis in Genetics University of Natural Sciences, Vietnam National University Hochiminh City 2010 Doan Thu Tra Determination of drug resistance of HIV in patients with first-line therapy failure and evaluation of the efficacy of second-line antiretroviral therapy in HIVinfected patients Doctor of Medicine Thesis Institute for Clinical Pharmaco-medical Science Research 108 2016 Johnson V.A, Brun-Vezinet F, Clotet B et al Update of the drug resistance mutations in HIV-1: December 2010 Top HIV Med 2010, 18 (5), pp.156-63 Lok A.S, McMahon B.J Chronic hepatitis B: update 2009 Hepatology 2009, 50 (3), pp.661- 662  ... statistical analysis RESULTS AND DISCUSSION The propotion of patients having HIV with genetic mutation for drug resistance Among the 47 patients enrolled, 43 patients had genetic mutation for drug. .. non -resistance mutations 4.7 NRTI resistance mutations NNRTI and PI non -resistance mutations 2.3 Nguyen Huu Chi investigated drugs resistance mutation in 71 treatment failure patients who had drug resistance. .. expression of ARV resistance mutation in patients with treatment failure Drug resistance mutations Quantity (n = 43) Percentage % NNRTI resistance mutations 42 97.7 NRTI resistance mutations 41