Validation of digital droplet PCR assays for cell-associated HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 unspliced RNA for clinical studies in HIV-1 cure research
Copyright © 2023. Published by Elsevier B.V..
BACKGROUND: Cell-associated HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 unspliced RNA (usRNA) are important virological parameters for monitoring HIV-1 persistence and activation of latent HIV-1. Assays fully validated by CLIA and/or GCLP standards are needed for future clinical trials that seek to evaluate treatments directed towards HIV-1 cure.
OBJECTIVES: To determine performance characteristics of sensitive, moderate-throughput, digital droplet PCR (ddPCR) assays for cell-associated HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 usRNA that can detect a broad range of HIV-1 M-group subtypes.
STUDY DESIGN: To evaluate linearity, limit of detection, precision, and accuracy of each assay, contrived specimens were analyzed in a background of uninfected PBMC. Detection breadth was evaluated by in silico analysis of primer and probes sets and analysis of material harvested from PBMC infected in vitro with various HIV-1 subtypes. A cohort of clinical specimens from viremic and virologically suppressed individuals was analyzed to demonstrate applicability to clinical research.
RESULTS: The empirically determined limit of detection of these assays was 29, 7, and 60 copies per million PBMC for HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 usRNA, respectively. The assays detect a broad range of HIV-1 M-group subtypes. Finally, analysis of clinical specimens demonstrate that these assays can detect low levels of cell-associated HIV-1 DNA, HIV-1 usRNA, and HIV-1 2-LTR circle and correlate with clinical histories and viral loads of untreated and antiretroviral treated individuals.
CONCLUSIONS: We report the clinical validation of three HIV reservoir assays with broad HIV-1 coverage for future cure studies.
| Medienart: |
E-Artikel |
|---|
| Erscheinungsjahr: |
2024 |
|---|---|
| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:170 |
|---|---|
| Enthalten in: |
Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology - 170(2024) vom: 19. Feb., Seite 105632 |
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Reed, Jonathan [VerfasserIn] |
|---|
| Links: |
|---|
| Anmerkungen: |
Date Completed 22.01.2024 Date Revised 02.04.2024 published: Print-Electronic Citation Status MEDLINE |
|---|
| doi: |
10.1016/j.jcv.2023.105632 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
NLM366043501 |
|---|
| LEADER | 01000caa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLM366043501 | ||
| 003 | DE-627 | ||
| 005 | 20240402235958.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 231227s2024 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.1016/j.jcv.2023.105632 |2 doi | |
| 028 | 5 | 2 | |a pubmed24n1361.xml |
| 035 | |a (DE-627)NLM366043501 | ||
| 035 | |a (NLM)38113685 | ||
| 035 | |a (PII)S1386-6532(23)00255-X | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Reed, Jonathan |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Validation of digital droplet PCR assays for cell-associated HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 unspliced RNA for clinical studies in HIV-1 cure research |
| 264 | 1 | |c 2024 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Completed 22.01.2024 | ||
| 500 | |a Date Revised 02.04.2024 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a Copyright © 2023. Published by Elsevier B.V. | ||
| 520 | |a BACKGROUND: Cell-associated HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 unspliced RNA (usRNA) are important virological parameters for monitoring HIV-1 persistence and activation of latent HIV-1. Assays fully validated by CLIA and/or GCLP standards are needed for future clinical trials that seek to evaluate treatments directed towards HIV-1 cure | ||
| 520 | |a OBJECTIVES: To determine performance characteristics of sensitive, moderate-throughput, digital droplet PCR (ddPCR) assays for cell-associated HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 usRNA that can detect a broad range of HIV-1 M-group subtypes | ||
| 520 | |a STUDY DESIGN: To evaluate linearity, limit of detection, precision, and accuracy of each assay, contrived specimens were analyzed in a background of uninfected PBMC. Detection breadth was evaluated by in silico analysis of primer and probes sets and analysis of material harvested from PBMC infected in vitro with various HIV-1 subtypes. A cohort of clinical specimens from viremic and virologically suppressed individuals was analyzed to demonstrate applicability to clinical research | ||
| 520 | |a RESULTS: The empirically determined limit of detection of these assays was 29, 7, and 60 copies per million PBMC for HIV-1 DNA, HIV-1 2-LTR circle, and HIV-1 usRNA, respectively. The assays detect a broad range of HIV-1 M-group subtypes. Finally, analysis of clinical specimens demonstrate that these assays can detect low levels of cell-associated HIV-1 DNA, HIV-1 usRNA, and HIV-1 2-LTR circle and correlate with clinical histories and viral loads of untreated and antiretroviral treated individuals | ||
| 520 | |a CONCLUSIONS: We report the clinical validation of three HIV reservoir assays with broad HIV-1 coverage for future cure studies | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, N.I.H., Extramural | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 4 | |a 2-LTR circle | |
| 650 | 4 | |a Clinical validation | |
| 650 | 4 | |a HIV-1 DNA | |
| 650 | 4 | |a HIV-1 reservoir | |
| 650 | 4 | |a HIV-1 unspliced RNA | |
| 650 | 4 | |a digital droplet PCR | |
| 650 | 7 | |a DNA, Viral |2 NLM | |
| 650 | 7 | |a RNA, Viral |2 NLM | |
| 700 | 1 | |a Kwak, Ginger |e verfasserin |4 aut | |
| 700 | 1 | |a Piliper, Eli A |e verfasserin |4 aut | |
| 700 | 1 | |a Degli-Angeli, Emily J |e verfasserin |4 aut | |
| 700 | 1 | |a Goecker, Erin A |e verfasserin |4 aut | |
| 700 | 1 | |a Greninger, Alexander L |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology |d 1997 |g 170(2024) vom: 19. Feb., Seite 105632 |w (DE-627)NLM097223964 |x 1873-5967 |7 nnns |
| 773 | 1 | 8 | |g volume:170 |g year:2024 |g day:19 |g month:02 |g pages:105632 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1016/j.jcv.2023.105632 |3 Volltext |
| 912 | |a GBV_USEFLAG_A | ||
| 912 | |a GBV_NLM | ||
| 951 | |a AR | ||
| 952 | |d 170 |j 2024 |b 19 |c 02 |h 105632 | ||