Characterization of the Immune Response to PD-1 Blockade during Chemoradiotherapy for Head and Neck Squamous Cell Carcinoma

Background: Chemoradiotherapy is a standard treatment for HNSCC. Blockade of the PD-1/L1-2 interaction may represent a target to overcome immune escape during this treatment. Methods: Utilizing a HNSCC mEERL C57BL/6 mouse model, we evaluated a PD-1 blockade alone or in combination with cisplatin-based chemoradiotherapy. Next, we evaluated peripheral blood mononuclear cells (PBMCs) with relative PD-1, TIM-3, and LAG-3 expression, and myeloid-derived suppressor-like (MDSC-like) populations from a clinical trial evaluating PD-1 blockade with chemoradiotherapy in HNSCC. Finally, we analyzed the effect of therapy on human T-cell clonality through T-cell Receptor (TCR) sequencing. Results: Anti-PD-1 monotherapy induced no response in the mEERL model; however, combination with chemoradiotherapy improved tumor clearance and survival. PBMCs from patients treated with this combination therapy demonstrate a decline in circulating T-cell populations with knockdown of PD-1 expressing CD3+CD4+ and CD3+CD8+ T cells during treatment. However, TIM-3, LAG-3 expressing T-cell and MDSC-like populations concordantly rose. During treatment, the TCR repertoire demonstrates overall clonal expansion, with both unique and previously reported T-cell clones. Conclusions: Our murine HNSCC model demonstrates efficacy of PD-1 blockade during chemoradiotherapy. However, while PD-1-expressing T cells decreased with this therapy, human PBMC findings also identified an increase in populations contributing to immune exhaustion. These findings further characterize PD-1 blockade during chemoradiotherapy for HNSCC and highlight potential competing mechanisms of immune evasion

CD4+ T Cells Expressing PD-1, TIGIT and LAG-3 Contribute to HIV Persistence during ART.

HIV persists in a small pool of latently infected cells despite antiretroviral therapy (ART). Identifying cellular markers expressed at the surface of these cells may lead to novel therapeutic strategies to reduce the size of the HIV reservoir. We hypothesized that CD4+ T cells expressing immune checkpoint molecules would be enriched in HIV-infected cells in individuals receiving suppressive ART. Expression levels of 7 immune checkpoint molecules (PD-1, CTLA-4, LAG-3, TIGIT, TIM-3, CD160 and 2B4) as well as 4 markers of HIV persistence (integrated and total HIV DNA, 2-LTR circles and cell-associated unspliced HIV RNA) were measured in PBMCs from 48 virally suppressed individuals. Using negative binomial regression models, we identified PD-1, TIGIT and LAG-3 as immune checkpoint molecules positively associated with the frequency of CD4+ T cells harboring integrated HIV DNA. The frequency of CD4+ T cells co-expressing PD-1, TIGIT and LAG-3 independently predicted the frequency of cells harboring integrated HIV DNA. Quantification of HIV genomes in highly purified cell subsets from blood further revealed that expressions of PD-1, TIGIT and LAG-3 were associated with HIV-infected cells in distinct memory CD4+ T cell subsets. CD4+ T cells co-expressing the three markers were highly enriched for integrated viral genomes (median of 8.2 fold compared to total CD4+ T cells). Importantly, most cells carrying inducible HIV genomes expressed at least one of these markers (median contribution of cells expressing LAG-3, PD-1 or TIGIT to the inducible reservoir = 76%). Our data provide evidence that CD4+ T cells expressing PD-1, TIGIT and LAG-3 alone or in combination are enriched for persistent HIV during ART and suggest that immune checkpoint blockers directed against these receptors may represent valuable tools to target latently infected cells in virally suppressed individuals

CD4⁺ T Cells Expressing PD-1, TIGIT and LAG-3 Contribute to HIV Persistence during ART

<div><p>HIV persists in a small pool of latently infected cells despite antiretroviral therapy (ART). Identifying cellular markers expressed at the surface of these cells may lead to novel therapeutic strategies to reduce the size of the HIV reservoir. We hypothesized that CD4⁺ T cells expressing immune checkpoint molecules would be enriched in HIV-infected cells in individuals receiving suppressive ART. Expression levels of 7 immune checkpoint molecules (PD-1, CTLA-4, LAG-3, TIGIT, TIM-3, CD160 and 2B4) as well as 4 markers of HIV persistence (integrated and total HIV DNA, 2-LTR circles and cell-associated unspliced HIV RNA) were measured in PBMCs from 48 virally suppressed individuals. Using negative binomial regression models, we identified PD-1, TIGIT and LAG-3 as immune checkpoint molecules positively associated with the frequency of CD4⁺ T cells harboring integrated HIV DNA. The frequency of CD4⁺ T cells co-expressing PD-1, TIGIT and LAG-3 independently predicted the frequency of cells harboring integrated HIV DNA. Quantification of HIV genomes in highly purified cell subsets from blood further revealed that expressions of PD-1, TIGIT and LAG-3 were associated with HIV-infected cells in distinct memory CD4⁺ T cell subsets. CD4⁺ T cells co-expressing the three markers were highly enriched for integrated viral genomes (median of 8.2 fold compared to total CD4⁺ T cells). Importantly, most cells carrying inducible HIV genomes expressed at least one of these markers (median contribution of cells expressing LAG-3, PD-1 or TIGIT to the inducible reservoir = 76%). Our data provide evidence that CD4⁺ T cells expressing PD-1, TIGIT and LAG-3 alone or in combination are enriched for persistent HIV during ART and suggest that immune checkpoint blockers directed against these receptors may represent valuable tools to target latently infected cells in virally suppressed individuals.</p></div

HIV persistence and T-cell activation in blood, rectal and lymph node tissue in HIV-infected individuals receiving suppressive ART.

Background. Immune activation and inflammation remain elevated in human immunodeficiency virus (HIV)-infected individuals receiving antiretroviral therapy (ART) and may contribute to HIV persistence. Methods. Using flow cytometry expression of CD38, HLA-DR and PD-1 were measured in blood (n = 48), lymph node (LN; n = 9), and rectal tissue (n = 17) from virally suppressed individuals. Total and integrated HIV DNA, 2-LTR circles, and cell-associated unspliced HIV RNA were quantified. Results. CD4+ T cells from rectal tissue had a higher frequency of integrated HIV DNA compared with blood (4.26 fold-change in DNA; 95% confidence interval [CI] = 2.61-7.00; P &lt;.001) and LN (2.32 fold-change in DNA; 95% CI = 1.22-4.41; P =.01). In rectal tissue, there were positive associations between integrated HIV DNA with PD-1+ CD4+ T-cells (1.44 fold-change in integrated HIV DNA per 10-unit increase in PD-1+ CD4+ T cells; 95% CI = 1.01-2.05; P =.045) and CD38+HLA-DR+ CD8+ T cells (1.40 foldchange in integrated HIV DNA per 1-unit increase in CD38+HLA-DR+ CD8+ T cells; 95% CI = 1.05-1.86; P =.02). Both associations were independent of current and nadir CD4+ T-cell counts. Conclusions. During ART, rectal tissue is an important reservoir for HIV persistence with a high frequency of activated CD4+ and CD8+ T cells. PD-1 may represent a marker of HIV persistence in rectal tissue

Growth hormone enhances thymic function in HIV-1–infected adults

Growth hormone (GH) is an underappreciated but important regulator of T cell development that can reverse age-related declines in thymopoiesis in rodents. Here, we report findings of a prospective randomized study examining the effects of GH on the immune system of HIV-1–infected adults. GH treatment was associated with increased thymic mass. In addition, GH treatment enhanced thymic output, as measured by both the frequency of T cell receptor rearrangement excision circles in circulating T cells and the numbers of circulating naive and total CD4+ T cells. These findings provide compelling evidence that GH induces de novo T cell production and may, accordingly, facilitate CD4+ T cell recovery in HIV-1–infected adults. Further, these randomized, prospective data have shown that thymic involution can be pharmacologically reversed in humans, suggesting that immune-based therapies could be used to enhance thymopoiesis in immunodeficient individuals

Influenza A(H3N2) Virus in Swine at Agricultural Fairs and Transmission to Humans, Michigan and Ohio, USA, 2016

In 2016, a total of 18 human infections with influenza A(H3N2) virus occurred after exposure to influenza-infected swine at 7 agricultural fairs. Sixteen of these cases were the result of infection by a reassorted virus with increasing prevalence among US swine containing a hemagglutinin gene from 2010–11 human seasonal H3N2 strains

Negative binomial regression models to assess the relationship between integrated HIV DNA and IC expression on CD4⁺ T cells.

<p>Negative binomial regression models to assess the relationship between integrated HIV DNA and IC expression on CD4⁺ T cells.</p

Co-expression of PD-1, TIGIT and LAG-3 identifies HIV-infected cells during ART.

<p>(A) Pie chart representing the frequencies of memory CD4⁺ T cells co-expressing PD-1 and/or TIGIT and/or LAG-3 in cohort 1 (n = 48). Coloured bars on the right side designate categories of ICs expressing cells: Triple -: PD-1/TIGIT/LAG-3 triple—in blue; single +: PD-1 single +, TIGIT single +, LAG-3 single + in green; double +: PD-1/TIGIT double +, TIGIT/LAG-3 double +, PD-1/LAG-3 double + in orange and triple +: PD-1/TIGIT/LAG-3 triple + in red. (B) Frequency of memory CD4⁺ T cells harboring integrated HIV DNA represented as a fold change over frequency in total CD4⁺ T cells. Mean values and standard deviations from 5 independent donors are represented (n = 5). (C) Frequency of cells harboring inducible msRNA measured by TILDA in memory CD4⁺ T cells expressing any (i.e. at least one) versus none of PD-1, TIGIT or LAG-3 (mLPT+ and mLPT- respectively). P value and fold-difference were obtained from a maximum likelihood model.</p

PD-1, TIGIT and LAG-3 identify HIV-infected cells in distinct memory CD4⁺ T-cell subsets during ART.

<p>(A), (B), (C) Frequencies of memory CD4⁺ T-cell subsets (naïve (T_N), central memory (T_CM), transitional memory (T_TM), effector memory (T_EM) and terminally differentiated (T_D)) expressing PD-1, TIGIT or LAG-3, respectively. Horizontal bars indicate median values with interquartile ranges. (D), (E), (F) Frequencies of cells harboring integrated
HIV DNA in T_CM, T_TM and T_EM CD4⁺ T-cell subsets sorted based on their expression of PD-1 (n = 12), TIGIT (n = 9) or LAG-3 (n = 7), respectively. Results are expressed as the HIV copy number in million cells of a
given subset. P values were obtained from negative binomial regression analysis. Significant differences (p<0.05) are designated by a p value in bold. Open circles represent the limit of detection in the negative samples (based on cell input).</p