Integrase strand-transfer inhibitors (InSTIs) are the preferred antiretroviral therapy (ART) regimens for naive HIV-infected individuals [1,2]. However, resistance to InSTIs is not routinely monitored in most centers at baseline. Indeed, recent guidelines suggest performing InSTI baseline resistance test only if transmission from a partner with InSTI resistance is known or suspected .
Although transmission of strains with signature InSTI resistance mutations is extremely rare, previous reports suggest that polymorphic substitutions reducing first generation InSTIs activity may not be infrequent in naïve patients, in studies performed in Europe and USA [3,4]. Indeed, in our institution, 13.89% of patients with acute/recent (less than 6 months) HIV infection were found to carry such substitutions . E157Q has been the one most frequently found in different studies, although with a wide range of prevalence [4–6], sometimes related to specific HIV-1 subtypes .
Rapid ART start is becoming increasingly recognized as a tool to increase adherence, to improve virological outcome and to reduce transmission . Rapid ART is particularly important in acute/recent HIV cases, which frequently have high viral loads . In this context, a rapid decrease of viral load is desirable to control clinical symptoms and decrease secondary cases, as transmission occurs more frequently during acute/recent infection than during chronic infection . Thus, InSTI are ideal drugs, but a fully active ART regimen should be assured, and, at the time of ART initiation, resistance test results are frequently pending .
The aim of this study was to evaluate the prevalence of E157Q substitution in newly diagnosed patients with acute/recent HIV infection, their virological response to ART, and the presence of transmission clusters among patients carrying this substitution.
Genotypic drug resistance tests were performed in all consecutive patients prospectively enrolled with a documented infection of less than 6 months (Hospital Clinic Acute/Recent HIV infection Cohort), from May 2015 (date of initiation of InSTI baseline resistance testing in our institution) to May 2017, and clinical and virological follow-up reported, at least, until 1 year.
Sequences were obtained by ultra-deep sequencing (UDS). UDS sequences obtained were revised to remove error sequencing with quality parameters. Later a single consensus sequence of pol gen (RT, PR, IN) was obtained at threshold 20% using Mesquite, version 2.75 (https://www.mesquiteproject.org). Subtyping was done with REGA, version 3.0 (http://dbpartners.stanford.edu:8080/RegaSubtyping/stanford-hiv/typingtool). All sequences were aligned using ClustalW, the maximum likelihood trees constructed with Mega version 6.06 (https://www.megasoftware.net). We used the best-fit model selected by Find Model: general time-reversible (GTR) with 1.4 gamma-distributed rate across sites model and 1000 bootstrap iterations for this analysis; bootstrap values of 75% or greater were defined for cluster assignment. Ten additional cases from our institution (eight naïve patients, and two patients on virological failure), carrying sequences with E157Q substitution, were also included in the phylogenetic analysis (defined as cases 7–16).
In 16 of 67 (23.9%) cases, at least one substitution was detected (for RT, PROT or INT – including InSTI polymorphisms) in the baseline genotypic tests (a proportion equal to or higher than 20% of sequences). Six of 67 (8.95%, 95% confidence interval = 4.17–18.19%) cases had the E157Q substitution, two patients had another InSTI substitution (163K and 95K), two patients had PROT substitutions (20I and 74S in one case, and 33I and 46L in the other) and nine patients had sequences with at least one substitution for RT including 215S/D in three cases, including one with 41L, the most prevalent substitutions for nonnucleoside reverse transcriptase inhibitor (NRTI); two cases of 138A substitution; two cases of K103N; and 179E and 98G in one case each for nonnucleoside RTI (NNRTI) (see supplementary Table 1, http://links.lww.com/QAD/B481).
As pointed out above, six consecutive patients with acute/recent HIV infection were detected to have strains carrying the E157Q InSTI substitution (cases 1–6), conferring decreased activity of raltegravir and elvitegravir according to Stanford HIV drug resistance data base and French ANRS algorithms (https://hivdb.stanford.edu/ and http://www.hivfrenchresistance.org). All cases were MSM patients with a median age of 35.5 years (31, 40) and infected with subtype B strains. Median viral load was 5.33 [interquartile range (IQR): 4.54–5.71] log10 copies/ml and, in all cases, relative prevalence of the substitution was high (equal to or higher than 95%). None of them presented transmitted drug resistance to other antiretroviral families, or other InSTI substitution. There were no major viral or epidemiological differences when compared with patients without E157Q substitution (data not shown). Three out of six cases were in a cluster of transmission. Patients 3 and 4 had related sequences between them, and patient 2 was in a cluster with one of the naïve patients (chronically infected) included in the tree (patient 16) (Fig. 1 and supplemental Figure 1, http://links.lww.com/QAD/B481).
Three patients initiated dolutegravir-based regimens, and all of them were undetectable at 1 year and at the last follow-up. The other three cases received NNRTI-based regimens (rilpivirine in two cases and efavirenz in the other) one of them switched from elvitegravir/cobicistat when resistance test was available. At 1 year, all cases were undetectable, and all of them remained undetectable in last control (3 years for the longest, 1 year for the shortest follow-up). The main epidemiological and virological characteristics are shown in Table 1.
During the same 2-year period (May 2015–May 2017), E157Q was also detected in 16 additional patients chronically infected, eight cases (50%) in baseline samples of naïve patients. Seven out of these eight cases started a InSTI-based regimen (raltegravir N = 3, elvitegravir N = 2, dolutegravir N = 2), with a median VL = 4.41log (IQR: 3.78–4.95) at ART initiation and all were undetectable at 1 year. Among the eight treatment-experienced cases with E157Q at failure, two patients were on raltegravir-based regimens, and both cases had additional InSTI substitutions (66A, 97A, 74M, 138K in one case and 155H in the other).
Early ART in patients with primary HIV infection controls clinical manifestations, reduces viral reservoir, decreases transmission and optimizes immunological recovery . Indeed, rapid ART start is recommended for all newly diagnosed patients, and particularly for acute/recent infections. The current recommended regimens for rapid ART start (bictegravir, dolutegravir or darunavir-based regimens)  are not affected by these polymorphisms. However, if raltegravir or elvitegravir are the chosen InSTI, activity may be reduced. This might be of particular concern in patients with acute/recent infections, as they frequently have high viral load at diagnosis.
Although there is a lack of large series or cohorts evaluating virological response to different InSTIs when E157Q is present, cases of virological failure with raltegravir  and elvitegravir  have been recently described. In fact, Charpentier et al. reported two out of eight virological failures (25%) in antiretroviral naïve patients starting elvitegravir/cobicistat-based antiretroviral treatment. The Stanford HIV drug resistance database considers E157Q to confer potential low level of resistance to raltegravir and elvitegravir but, according to the results of the previously mentioned French studies, ANRS reports those strains to be fully resistant to raltegravir and elvitegravir, and potentially resistant to dolutegravir. Accordingly, we do not initiate raltegravir or elvitegravir in our clinical practice in acute/recent patients carrying E157Q.
As dolutegravir and bictegravir may not be available in resource limited settings, and as other antiretroviral families do not reduce viral load as fast as InSTIs, what is very relevant in patients with acute/recent infection, our finding may be of concern. If E157Q is confirmed to clinically reduce InSTI activity, and as its prevalence in our study is near 10%, baseline InSTI resistance testing may be justified for this special population. The reasons for the different prevalence of these InSTI substitutions in different studies is unknown; it may be related to different subtype prevalence or to different levels of InSTI use in given clinical contexts, although some studies had reported a high prevalence of these substitutions prior to the widespread use of InSTI in Europe . We avoided using raltegravir or elvitegravir in these acute/recent cases, but the virological response to dolutegravir-based regimens was adequate for a follow-up of 1 year at least, and nearly 3 years for the longest case treated with dolutegravir (case 3).
As expected, and in line with several studies performed in patients with acute/recent infection [9,12], it was not infrequent to find patients from our cohort carrying the E157Q substitution who were involved in transmission clusters. This may be a warning, if a case is detected, that other secondary cases may carry this substitution in a given epidemiological context. Indeed, transmission clusters are more frequently detected among MSM with recent infections [9,12]. It must be highlighted that for cases 3 and 4, involved in a cluster, 8 months had elapsed between them. Considering that all cases have less than 6 months from infection to the beginning of follow-up, the existence of other related cases should be supposed. These two cases had an X4 strain.
Three of the patients with baseline E157Q received dolutegravir-based regimens and all had a good virological response. Among the eight naïve cases with chronic infection, none of them failed (neither on dolutegravir, or on raltegravir or elvitegravir) but, as expected, median viral load was lower than acute/recent cases of viral load (4.41 vs. 5.33). The E157Q substitution was detected in two cases of virological failure, but the baseline InSTI resistance test before InSTI initiation was not available for these two cases, as both started ART before May 2015. Both cases had additional InSTI substitutions added to E157Q.
Although not the objective of our study, when InSTI substitutions are included, we were able to detect a high proportion of transmitted drug resistance, with up to 23.9% patients presenting at least one substitution (for PROT, RT or INT – including InSTI polymorphisms). However, most substitutions were isolated and rarely affecting current front-line recommended regimens.
As a limitation, our study includes only six acute/recent cases and data of eight additional naïve chronic cases with the E157Q substitution. Although this study represents a small sample size, as it is unprovable that more robust data will emerge in the near future, we believe that appropriate surveillance programs on InSTI baseline resistance need to be implemented.
In conclusion, the E157Q substitution, conferring reduced susceptibility to raltegravir and elvitegravir, was frequently found in a baseline genotypic test of acute/recent HIV cases. All cases were infected with subtype B, the most frequent in Europe, and some of these cases were included in transmission clusters. All cases treated with dolutegravir-based regimens were undetectable at 1 year, and remained undetectable at last follow-up. In the context of primary HIV infection with frequently high viral loads, virological response should be carefully monitored in patients carrying InSTIs substitutions and initiating an InSTI-based regimen, particularly, if raltegravir or elvitegravir/cobicistat are used, or if two-drug regimens, such as dolutegravir and 3TC are considered. In this specific setting, InSTI baseline resistance testing may be of particular interest.
This work was supported in part by the ‘Fondo de Investigaciones Sanitarias’ (FIS) grant 04/0363 from the ‘Instituto de Salud Carlos III’, Madrid, Spain awarded to J.M.M. J.M.M. received a personal 80 : 20 research grant from the Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain, during 2017–19. D.N. developed this work in the frame of a postresidency Scholarship Ajuts a la Recerca ‘Josep Font’ 2014, Hospital Clinic, Barcelona, Spain. J.A.F.C.R. was no longer working at Hospital Universitario San Cecilio, Granada, Spain, since 1 March 2018.
J.A., D.N. and J.M.M. attended the patients, collected the information and analyzed the data. M.M.M. and M.A.M. performed resistance tests. E.d.L. performed statistical analyses and data management, J.A.F.-C.R. and F.G. performed phylogenetic analyses. J.A. wrote the first draft. All authors reviewed, commented and approved the last version of the manuscript.
Conflicts of interest
J.M.M. has received consulting honoraria and/or research grants from AbbVie, Bristol-Myers Squibb, Contrafect, Jansen, Genentech, Medtronic, Merck, Novartis, Gilead Sciences, and ViiV Healthcare, outside the submitted work. F.G. has received consulting honoraria and/or research grants from AbbVie, Merck, Roche, Hologic, Gilead Sciences and ViiV Healthcare, outside the submitted work; J.A. has received consulting honoraria from Gilead Sciences and Janssen Pharmaceuticals, outside the submitted work. All other authors have no conflicts of interest.
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