SLDAssay: A software package and web tool for analyzing limiting dilution assays

Trumble, Ilana M.; Allmon, Andrew G.; Archin, Nancie M.; Rigdon, Joseph; Francis, Owen; Baldoni, Pedro L.; Hudgens, Michael G.

doi:10.1016/j.jim.2017.07.004

Cited by 25 publications

(31 citation statements)

References 17 publications

(17 reference statements)

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“…The HIV-1 reservoir in resting CD4+ T cells was measured using quantitative viral outgrowth assays (QVOAs) ( Lee et al, 2017 ; Siliciano and Siliciano, 2005 ; Trumble et al, 2017 ; Ho et al, 2013 ; Massanella and Richman, 2016 ) for all 25 participants ( Figure 1 ). Longitudinal IUPM measurements were obtained for 10 participants ( Supplementary file 1 ).…”

Section: Resultsmentioning

confidence: 99%

“…Only cultures that contained an equivalent or greater level of p24 antigen on day 19 compared with day 15 were scored as positive. A maximum likelihood method was used on day 15 data to estimate the frequency of latent HIV infection, reported as bias-corrected Infectious Units Per Million (IUPM) ( Supplementary file 1 ; Trumble et al, 2017 ). For 10 participants, longitudinal IUPM measurements were available.…”

Section: Methodsmentioning

confidence: 99%

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The HIV-1 latent reservoir is largely sensitive to circulating T cells

Warren

Zhou²,

et al. 2020

eLife

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HIV-1 specific CD8+ T cells are an important component of HIV-1 curative strategies. Viral variants in the HIV-1 reservoir may limit the capacity of T cells to detect and clear virus-infected cells. We investigated the patterns of T cell escape variants in the replication-competent reservoir of 25 persons living with HIV-1 (PLWH) durably suppressed on ART. We identified all reactive T cell epitopes in the HIV-1 proteome for each participant and sequenced HIV-1 outgrowth viruses from resting CD4+ T cells. All non-synonymous mutations in reactive T cell epitopes were tested for their effect on the size of the T cell response, with a ≥50% loss defined as an escape mutation. The majority (68%) of T cell epitopes harbored no detectable escape mutations. These findings suggest that circulating T cells in PLWH on ART could contribute to control of rebound and could be targeted for boosting in curative strategies.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

The HIV-1 latent reservoir is largely sensitive to circulating T cells

Warren

Zhou²,

et al. 2020

eLife

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“…Cultures were scored as positive if p24 was detected at day 15 and was increased in concentration at day 19. The number of resting CD4 + T cells in infected units per million (IUPM) was estimated by a maximum likelihood method 3,38,39 .…”

Section: Methodsmentioning

confidence: 99%

Assessing the impact of AGS-004, a dendritic cell-based immunotherapy, and vorinostat on persistent HIV-1 Infection

Kuruc

Falcinelli

Warren

et al. 2020

Sci Rep

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Approaches to deplete persistent HIV infection are needed. We investigated the combined impact of the latency reversing agent vorinostat (VOR) and AGS-004, an autologous dendritic cell immunotherapeutic, on the HIV reservoir. HIV+, stably treated participants in whom resting CD4 + t cell-associated HIV RNA (rca-RNA) increased after VOR exposure ex vivo and in vivo received 4 doses of AGS-004 every 3 weeks, followed by VOR every 72 hours for 30 days, and then the cycle repeated. Change in VOR-responsive host gene expression, HIV-specific T cell responses, low-level HIV viremia, rca-RNA, and the frequency of resting CD4 + T-cell infection (RCI) was measured at baseline and after each cycle. No serious treatment-related adverse events were observed among five participants. As predicted, VOR-responsive host genes responded uniformly to VOR dosing. Following cycles of AGS-004 and VOR, rca-RNA decreased significantly in only two participants, with a significant decrease in SCA observed in one of these participants. However, unlike other cohorts dosed with AGS-004, no uniform increase in HIV-specific immune responses following vaccination was observed. Finally, no reproducible decline of RCI, defined as a decrease of >50%, was observed. AGS-004 and VOR were safe and well-tolerated, but no substantial impact on RCI was measured. In contrast to previous clinical data, AGS-004 did not induce HIV-specific immune responses greater than those measured at baseline. More efficacious antiviral immune interventions, perhaps paired with more effective latency reversal, must be developed to clear persistent HIV infection.

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“…Code to estimate in the R software environment from an SLDA is provided in Box 1 and in the supporting information. Other widely used software such as the webinterface ELDA (Hu and Smyth, 2009), the popular L-Calc software (Stem Cell Technologies, Vancouver, BC, Canada), the R package SLDAssay (Trumble et al, 2017), which also allows for bias corrected MLE's, can also be used to make these estimates.…”

Section: Estimationmentioning

confidence: 99%

A Two-Stage Experimental Design for Dilution Assays

2019

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Dilution assays to determine solute concentration have found wide use in biomedical research. Many dilution assays return imprecise concentration estimates because they are only done to orders of magnitude. Previous statistical work has focused on how to design efficient experiments that can return more precise estimates, however this work has not considered the practical difficulties of implementing these designs in the laboratory. We developed a two-stage experiment with a first stage that obtains an order of magnitude estimate and a second stage that concentrates effort on the most informative dilution to increase estimator precision. We show using simulations and an empirical example that the best two-stage experimental designs yield estimates that are remarkably more accurate than standard methods with equivalent effort. This work demonstrates how to utilize previous advances in experimental design in a manner consistent with current laboratory practice. We expect that multi-stage designs will prove to be useful for obtaining precise estimates with minimal experimental effort. KEYWORDS efficient experimental design, fisher information, generalized linear model, parameter estimation, single-hit poisson model Biometrics. 2019;1-8 wileyonlinelibrary.com/journal/biom

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SLDAssay: A software package and web tool for analyzing limiting dilution assays

Cited by 25 publications

References 17 publications

The HIV-1 latent reservoir is largely sensitive to circulating T cells

The HIV-1 latent reservoir is largely sensitive to circulating T cells

Assessing the impact of AGS-004, a dendritic cell-based immunotherapy, and vorinostat on persistent HIV-1 Infection

A Two-Stage Experimental Design for Dilution Assays

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