Optimization and validation of a robust human T-cell culture method for monitoring phenotypic and polyfunctional antigen-specific CD4 and CD8 T-cell responses

Lin, Yun; Gallardo, Humilidad F.; Ku, Geoffrey Yuyat; Li, Hao; Manukian, Gregor; Rasalan, Teresa; Xu, Yinyan; Terzulli, Stephanie; Old, Lloyd J.; Allison, James P.; Houghton, Alan N.; Wolchok, Jedd D.; Yuan, Jianda

doi:10.3109/14653240903136987

Cited by 33 publications

(15 citation statements)

References 29 publications

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“…PBMCs were obtained with the patient’s consent following resection of the treatment-resistant metastatic lesion and evaluated for reactivity to 7 synthetic in silico predicted neoantigens and their wildtype analogs using standard protocols (Lin et al, 2009; Maeda et al, 2014; Rizvi et al, 2015). Tumor-specific and wild-type peptides were synthesized by GenScript USA Inc.…”

Section: Methodsmentioning

confidence: 99%

Loss of PTEN Is Associated with Resistance to Anti-PD-1 Checkpoint Blockade Therapy in Metastatic Uterine Leiomyosarcoma

George

Miao

Demetri³

et al. 2017

Immunity

421

320

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Summary Response to immune checkpoint blockade in mesenchymal tumors is poorly characterized, but immunogenomic dissection of these cancers may inform immunotherapy mediators. We identified a treatment-naïve patient with metastatic uterine leiomyosarcoma who experienced complete tumor remission for >2 years on anti-PD-1 (pembrolizumab) monotherapy. We analyzed primary tumor, the sole treatment-resistant metastasis, and germline tissue to explore mechanisms of immunotherapy sensitivity and resistance. Both tumors stained diffusely for PD-L2, with sparse PD-L1 staining. PD-1+ cell infiltration significantly decreased in the resistant tumor (p=0.02). Genomically, the treatment-resistant tumor uniquely harbored biallelic PTEN loss and had reduced expression of two neoantigens that demonstrated strong immunoreactivity with patient T cells in vitro, suggesting long-lasting immunological memory. In this near-complete response to PD-1 blockade in a mesenchymal tumor, we identified PTEN mutation and reduced expression of genes encoding neoantigens as potential mediators of resistance to immune checkpoint therapy.

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Section: Methodsmentioning

confidence: 99%

Loss of PTEN Is Associated with Resistance to Anti-PD-1 Checkpoint Blockade Therapy in Metastatic Uterine Leiomyosarcoma

George

Miao

Demetri³

et al. 2017

Immunity

421

320

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“…T-cell analyses were conducted with modifications to published methods (29, 30). Monocytes were isolated using the MACS CD14 isolation kit (Miltenyi Biotec), and cultured in 6-well plates (2×10 6 cell/mL) in RPMI-1640 medium containing 5% pooled human serum (PHS), 50 ng/mL GM-CSF (Miltenyi Biotech) and 50 ng/mL IL-4 (Miltenyi Biotech).…”

Section: Methodsmentioning

confidence: 99%

A Feasibility Study of Cyclophosphamide, Trastuzumab, and an Allogeneic GM-CSF–Secreting Breast Tumor Vaccine for HER2+ Metastatic Breast Cancer

Chen

Gupta

Petrik

et al. 2014

Cancer Immunology Research

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Granulocyte-macrophage colony-stimulating factor (GM-CSF)-secreting tumor vaccines are bioactive, but limited by disease burden and immune tolerance. Cyclophosphamide (CY) augments vaccine activity in tolerant neu mice and metastatic breast cancer (MBC) patients. HER-2-specific monoclonal antibodies (MAb) enhance vaccine activity in neu mice. We hypothesized that CY-modulated vaccination with HER-2-specific MAb safely induces relevant HER-2-specific immunity in neu mice and HER-2+ MBC patients. Adding both CY and the HER-2-specific MAb 7.16.4 to vaccination maximized HER-2-specific CD8+ T-cell immunity and tumor-free survival in neu transgenic mice. We therefore conducted a single arm feasibility study of CY, an allogeneic HER-2+ GM-CSF-secreting breast tumor vaccine, and weekly trastuzumab in 20 HER-2+ MBC patients. Primary clinical trial objectives were safety and clinical benefit (CB), in which CB represents complete response+partial response+stable disease. Secondary study objectives were to assess HER-2-specific T-cell responses by delayed type hypersensitivity (DTH) and intracellular cytokine staining. Subjects received three monthly vaccinations, with a boost 6-8 months from trial entry. This combination immunotherapy was safe, with CB rates at 6 months and 1 year of 55% (95% CI:32-77%, p=0.013) and 40% (95% CI:19-64%) respectively. Median progression-free survival (PFS) and overall survival (OS) were 7 (95% CI:4-16) and 42 months (95% CI:22-70) respectively. Increased HER-2-specific DTH developed in 7/20 subjects (of whom 4 had CB (95% CI:18-90)), with a trend toward longer PFS and OS in DTH responders. Polyfunctional HER-2-specific CD8+ T cells progressively expanded across vaccination cycles. Further investigation of CY-modulated vaccination with trastuzumab is warranted. (Clinicaltrials.gov identifier: NCT00399529)

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“…8,10,[17][18][19] Polyfunctional T cells can be induced in cancer patients after vaccinations. [20][21][22] However, mechanistically how these cells are induced is not understood. Moreover, it is unclear whether these cells are associated with better prognosis, or can be used therapeutically to treat cancer.…”

Section: Introductionmentioning

confidence: 99%

IL-7 signaling imparts polyfunctionality and stemness potential to CD4⁺ T cells

et al. 2016

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The functional status of CD4 C T cells is a critical determinant of antitumor immunity. Polyfunctional CD4 C T cells possess the ability to concomitantly produce multiple Th1-type cytokines, exhibiting a functional attribute desirable for cancer immunotherapy. However, the mechanisms by which these cells are induced are neither defined nor it is clear if these cells can be used therapeutically to treat cancer. Here, we report that CD4C T cells exposed to exogenous IL-7 during antigenic stimulation can acquire a polyfunctional phenotype, characterized by their ability to simultaneously express IFNg, IL-2, TNFa and granzyme B. This IL-7-driven polyfunctional phenotype was associated with increased histone acetylation in the promoters of the effector genes, indicative of increased chromatin accessibility. Moreover, forced expression of a constitutively active (CA) form of STAT5 recapitulated IL-7 in inducing CD4 C T-cell polyfunctionality. Conversely, the expression of a dominant negative (DN) form of STAT5 abolished the ability of IL-7 to induce polyfunctional CD4 C T cells. These in-vitro-generated polyfunctional CD4 C T cells can traffic to tumor and expand intratumorally in response to immunization. Importantly, adoptive transfer of polyfunctional CD4C T cells following lymphodepletive chemotherapy was able to eradicate large established tumors. This beneficial outcome was associated with the occurrence of antigen epitope spreading, activation of the endogenous CD8C T cells and persistence of donor CD4 C T cells exhibiting memory stem cell attributes. These findings indicate that IL-7 signaling can impart polyfunctionality and stemness potential to CD4 C T cells, revealing a previously unknown property of IL-7 that can be exploited in adoptive T-cell immunotherapy.

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Optimization and validation of a robust human T-cell culture method for monitoring phenotypic and polyfunctional antigen-specific CD4 and CD8 T-cell responses

Cited by 33 publications

References 29 publications

Loss of PTEN Is Associated with Resistance to Anti-PD-1 Checkpoint Blockade Therapy in Metastatic Uterine Leiomyosarcoma

Loss of PTEN Is Associated with Resistance to Anti-PD-1 Checkpoint Blockade Therapy in Metastatic Uterine Leiomyosarcoma

A Feasibility Study of Cyclophosphamide, Trastuzumab, and an Allogeneic GM-CSF–Secreting Breast Tumor Vaccine for HER2+ Metastatic Breast Cancer

IL-7 signaling imparts polyfunctionality and stemness potential to CD4⁺ T cells

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Optimization and validation of a robust human T-cell culture method for monitoring phenotypic and polyfunctional antigen-specific CD4 and CD8 T-cell responses

Cited by 33 publications

References 29 publications

Loss of PTEN Is Associated with Resistance to Anti-PD-1 Checkpoint Blockade Therapy in Metastatic Uterine Leiomyosarcoma

Loss of PTEN Is Associated with Resistance to Anti-PD-1 Checkpoint Blockade Therapy in Metastatic Uterine Leiomyosarcoma

A Feasibility Study of Cyclophosphamide, Trastuzumab, and an Allogeneic GM-CSF–Secreting Breast Tumor Vaccine for HER2+ Metastatic Breast Cancer

IL-7 signaling imparts polyfunctionality and stemness potential to CD4+ T cells

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IL-7 signaling imparts polyfunctionality and stemness potential to CD4⁺ T cells