title: A genotypic test for HIV-1 tropism combining Sanger sequencing with ultradeep sequencing predicts virologic response in treatment-experienced patients.
abstract: A tropism test is required prior to initiation of CCR5 antagonist therapy in HIV-1 infected individuals, as these agents are not effective in patients harboring CXCR4 (X4) coreceptor-using viral variants. We developed a clinical laboratory-based genotypic tropism test for detection of CCR5-using (R5) or X4 variants that utilizes triplicate population sequencing (TPS) followed by ultradeep sequencing (UDS) for samples classified as R5. Tropism was inferred using the bioinformatic algorithms geno2pheno([coreceptor]) and PSSM(x4r5). Virologic response as a function of tropism readout was retrospectively assessed using blinded samples from treatment-experienced subjects who received maraviroc (N = 327) in the MOTIVATE and A4001029 clinical trials. MOTIVATE patients were classified as R5 and A4001029 patients were classified as non-R5 by the original Trofile test. Virologic response was compared between the R5 and non-R5 groups determined by TPS, UDS alone, the reflex strategy and the Trofile Enhanced Sensitivity (TF-ES) test. UDS had greater sensitivity than TPS to detect minority non-R5 variants. The median log(10) viral load change at week 8 was -2.4 for R5 subjects, regardless of the method used for classification; for subjects with non-R5 virus, median changes were -1.2 for TF-ES or the Reflex Test and -1.0 for UDS. The differences between R5 and non-R5 groups were highly significant in all 3 cases (p<0.0001). At week 8, the positive predictive value was 66% for TF-ES and 65% for both the Reflex test and UDS. Negative predictive values were 59% for TF-ES, 58% for the Reflex Test and 61% for UDS. In conclusion, genotypic tropism testing using UDS alone or a reflex strategy separated maraviroc responders and non-responders as well as a sensitive phenotypic test, and both assays showed improved performance compared to TPS alone. Genotypic tropism tests may provide an alternative to phenotypic testing with similar discriminating ability.
authors.affiliation: Department of Infectious Diseases, Quest Diagnostics Nichols Institute, San Juan Capistrano, California, United States of America. Ron.M.Kagan@questdiagnostics.com
authors.forename: Ron M
authors.initials: RM
authors.lastname: Kagan
authors.forename: Erik P
authors.initials: EP
authors.lastname: Johnson
authors.forename: Martin
authors.initials: M
authors.lastname: Siaw
authors.forename: Pinaki
authors.initials: P
authors.lastname: Biswas
authors.forename: Douglass S
authors.initials: DS
authors.lastname: Chapman
authors.forename: Zhaohui
authors.initials: Z
authors.lastname: Su
authors.forename: Jamie L
authors.initials: JL
authors.lastname: Platt
authors.forename: Rick L
authors.initials: RL
authors.lastname: Pesano
mesh_terms: D000328:Adult; D000368:Aged; D000465:Algorithms; D019380:Anti-HIV Agents / Q000378:metabolism / Q000494:pharmacology*; D001681:Biological Assay; D054316:Biomarkers, Pharmacological; D065100:CCR5 Receptor Antagonists; D003510:Cyclohexanes / Q000378:metabolism / Q000494:pharmacology*; D005260:Female; D060005:Genotyping Techniques; D015658:HIV Infections / Q000188:drug therapy* / Q000378:metabolism / Q000821:virology; D015497:HIV-1 / Q000187:drug effects / Q000502:physiology; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008297:Male; D000077592:Maraviroc; D008875:Middle Aged; D011237:Predictive Value of Tests; D019713:Receptors, CCR5 / Q000378:metabolism; D019718:Receptors, CXCR4 / Q000378:metabolism; D012189:Retrospective Studies; D014230:Triazoles / Q000378:metabolism / Q000494:pharmacology*; D056189:Viral Tropism
chemical_list: D019380:Anti-HIV Agents; D054316:Biomarkers, Pharmacological; D065100:CCR5 Receptor Antagonists; C527276:CXCR4 protein, human; D003510:Cyclohexanes; D019713:Receptors, CCR5; D019718:Receptors, CXCR4; D014230:Triazoles; D000077592:Maraviroc