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Radiation and dual checkpoint blockade activate non-redundant immune mechanisms in cancer.

Twyman-Saint Victor C, Rech AJ, Maity A, Rengan R, Pauken KE, Stelekati E, Benci JL, Xu B, Dada H, Odorizzi PM, Herati RS, Mansfield KD, Patsch D, Amaravadi RK, Schuchter LM, Ishwaran H, Mick R, Pryma DA, Xu X, Feldman MD, Gangadhar TC, Hahn SM, Wherry EJ, Vonderheide RH, Minn AJ - Nature (2015)

Bottom Line: Immune checkpoint inhibitors result in impressive clinical responses, but optimal results will require combination with each other and other therapies.Anti-CTLA4 predominantly inhibits T-regulatory cells (Treg cells), thereby increasing the CD8 T-cell to Treg (CD8/Treg) ratio.Addition of PD-L1 blockade reverses T-cell exhaustion to mitigate depression in the CD8/Treg ratio and further encourages oligoclonal T-cell expansion.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA [2] Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

ABSTRACT
Immune checkpoint inhibitors result in impressive clinical responses, but optimal results will require combination with each other and other therapies. This raises fundamental questions about mechanisms of non-redundancy and resistance. Here we report major tumour regressions in a subset of patients with metastatic melanoma treated with an anti-CTLA4 antibody (anti-CTLA4) and radiation, and reproduced this effect in mouse models. Although combined treatment improved responses in irradiated and unirradiated tumours, resistance was common. Unbiased analyses of mice revealed that resistance was due to upregulation of PD-L1 on melanoma cells and associated with T-cell exhaustion. Accordingly, optimal response in melanoma and other cancer types requires radiation, anti-CTLA4 and anti-PD-L1/PD-1. Anti-CTLA4 predominantly inhibits T-regulatory cells (Treg cells), thereby increasing the CD8 T-cell to Treg (CD8/Treg) ratio. Radiation enhances the diversity of the T-cell receptor (TCR) repertoire of intratumoral T cells. Together, anti-CTLA4 promotes expansion of T cells, while radiation shapes the TCR repertoire of the expanded peripheral clones. Addition of PD-L1 blockade reverses T-cell exhaustion to mitigate depression in the CD8/Treg ratio and further encourages oligoclonal T-cell expansion. Similarly to results from mice, patients on our clinical trial with melanoma showing high PD-L1 did not respond to radiation plus anti-CTLA4, demonstrated persistent T-cell exhaustion, and rapidly progressed. Thus, PD-L1 on melanoma cells allows tumours to escape anti-CTLA4-based therapy, and the combination of radiation, anti-CTLA4 and anti-PD-L1 promotes response and immunity through distinct mechanisms.

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RT + anti-CTLA4 promotes regression of irradiated and unirradiated tumors and is inhibited by PD-L1 on tumor cellsa) Waterfall plot of unirradiated tumors after RT to a single index lesion with anti-CTLA4. Dashed lines are thresholds for PD (red) and PR (blue). * Patients with new lesions. ** Clinical progression without imaging. b) PET/CT images of irradiated (white arrows) and unirradiated (yellow arrows) tumors from patient PT-402. c) PFS and OS for all patients (dashed lines: 95% CI). d) B16-F10 tumor growth after RT to the index tumor (n=8), anti-CTLA4 (C4) (n=9), anti-CTLA4 and RT to the index tumor (n=18), or no (control) treatment (n=9). The p-values are comparisons with control. Pie chart shows %CRs (yellow). See Fig. 2d for survival. e) Heat map showing relative abundance of immune cells or their ratios from tumors that are resistant (black hatch) or sensitive to RT + anti-CTLA4. Boxplot shows bootstrap importance scores for each variable. Higher values (red) are more predictive. f) Change in T cell subsets or their ratio after RT + anti-CTLA4 for sensitive parental (Sen) or resistant (Res) tumors. Values are subtracted from average of untreated controls. Red line is mean. g) Heat map of resistance gene signature and PD-L1 across human melanoma. p < 0.001 by gene set enrichment analysis. h) Expression of PD-L1 on Res 499 compared to B16-F10 melanoma cells and of Res 237 compared to TSA breast cancer cells. Isotype control (IgG). i) Total tumor volume from PD-L1 knockout (KO) or control (WT) Res 499 and corresponding survival.
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Figure 1: RT + anti-CTLA4 promotes regression of irradiated and unirradiated tumors and is inhibited by PD-L1 on tumor cellsa) Waterfall plot of unirradiated tumors after RT to a single index lesion with anti-CTLA4. Dashed lines are thresholds for PD (red) and PR (blue). * Patients with new lesions. ** Clinical progression without imaging. b) PET/CT images of irradiated (white arrows) and unirradiated (yellow arrows) tumors from patient PT-402. c) PFS and OS for all patients (dashed lines: 95% CI). d) B16-F10 tumor growth after RT to the index tumor (n=8), anti-CTLA4 (C4) (n=9), anti-CTLA4 and RT to the index tumor (n=18), or no (control) treatment (n=9). The p-values are comparisons with control. Pie chart shows %CRs (yellow). See Fig. 2d for survival. e) Heat map showing relative abundance of immune cells or their ratios from tumors that are resistant (black hatch) or sensitive to RT + anti-CTLA4. Boxplot shows bootstrap importance scores for each variable. Higher values (red) are more predictive. f) Change in T cell subsets or their ratio after RT + anti-CTLA4 for sensitive parental (Sen) or resistant (Res) tumors. Values are subtracted from average of untreated controls. Red line is mean. g) Heat map of resistance gene signature and PD-L1 across human melanoma. p < 0.001 by gene set enrichment analysis. h) Expression of PD-L1 on Res 499 compared to B16-F10 melanoma cells and of Res 237 compared to TSA breast cancer cells. Isotype control (IgG). i) Total tumor volume from PD-L1 knockout (KO) or control (WT) Res 499 and corresponding survival.

Mentions: Anecdotal clinical reports suggest that RT may cooperate with anti-CTLA4 to systemically enhance melanoma response7; however, this combination has not been reported in a clinical trial. To examine the feasibility and efficacy of RT combined with immune checkpoint blockade, we initiated a phase I clinical trial of 22 patients with multiple melanoma metastases (Extended Data Table 1). A single index lesion was irradiated with hypofractionated RT, delivered over two or three fractions, followed by four cycles of the anti-CTLA4 antibody ipilimumab (Extended Data Fig. 1a). Accrual was completed in three out of four RT dose levels, and treatment was well tolerated (Extended Data Table 2). Evaluation of the unirradiated lesions by CT imaging using Response Evaluation Criteria in Solid Tumors (RECIST) demonstrated that 18% of patients had a partial response (PR) as best response, 18% had stable disease (SD), and 64% had progressive disease (PD) (Fig. 1a). For example, patient PT-402 showed a large reduction in sizes of unirradiated tumors and a partial metabolic response by positron emission tomography (PET) (Fig. 1b). None of the 12 patients evaluated by PET had progressive metabolic disease in the irradiated lesion (Extended Data Fig. 1b, Extended Data Table 3). The median progression-free survival (PFS) and overall survival (OS) was 3.8 and 10.7 months with median follow-up of 18.4 and 21.3 months (18.0 and 21.3 for patients without event), respectively (Fig. 1c).


Radiation and dual checkpoint blockade activate non-redundant immune mechanisms in cancer.

Twyman-Saint Victor C, Rech AJ, Maity A, Rengan R, Pauken KE, Stelekati E, Benci JL, Xu B, Dada H, Odorizzi PM, Herati RS, Mansfield KD, Patsch D, Amaravadi RK, Schuchter LM, Ishwaran H, Mick R, Pryma DA, Xu X, Feldman MD, Gangadhar TC, Hahn SM, Wherry EJ, Vonderheide RH, Minn AJ - Nature (2015)

RT + anti-CTLA4 promotes regression of irradiated and unirradiated tumors and is inhibited by PD-L1 on tumor cellsa) Waterfall plot of unirradiated tumors after RT to a single index lesion with anti-CTLA4. Dashed lines are thresholds for PD (red) and PR (blue). * Patients with new lesions. ** Clinical progression without imaging. b) PET/CT images of irradiated (white arrows) and unirradiated (yellow arrows) tumors from patient PT-402. c) PFS and OS for all patients (dashed lines: 95% CI). d) B16-F10 tumor growth after RT to the index tumor (n=8), anti-CTLA4 (C4) (n=9), anti-CTLA4 and RT to the index tumor (n=18), or no (control) treatment (n=9). The p-values are comparisons with control. Pie chart shows %CRs (yellow). See Fig. 2d for survival. e) Heat map showing relative abundance of immune cells or their ratios from tumors that are resistant (black hatch) or sensitive to RT + anti-CTLA4. Boxplot shows bootstrap importance scores for each variable. Higher values (red) are more predictive. f) Change in T cell subsets or their ratio after RT + anti-CTLA4 for sensitive parental (Sen) or resistant (Res) tumors. Values are subtracted from average of untreated controls. Red line is mean. g) Heat map of resistance gene signature and PD-L1 across human melanoma. p < 0.001 by gene set enrichment analysis. h) Expression of PD-L1 on Res 499 compared to B16-F10 melanoma cells and of Res 237 compared to TSA breast cancer cells. Isotype control (IgG). i) Total tumor volume from PD-L1 knockout (KO) or control (WT) Res 499 and corresponding survival.
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Related In: Results  -  Collection

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Figure 1: RT + anti-CTLA4 promotes regression of irradiated and unirradiated tumors and is inhibited by PD-L1 on tumor cellsa) Waterfall plot of unirradiated tumors after RT to a single index lesion with anti-CTLA4. Dashed lines are thresholds for PD (red) and PR (blue). * Patients with new lesions. ** Clinical progression without imaging. b) PET/CT images of irradiated (white arrows) and unirradiated (yellow arrows) tumors from patient PT-402. c) PFS and OS for all patients (dashed lines: 95% CI). d) B16-F10 tumor growth after RT to the index tumor (n=8), anti-CTLA4 (C4) (n=9), anti-CTLA4 and RT to the index tumor (n=18), or no (control) treatment (n=9). The p-values are comparisons with control. Pie chart shows %CRs (yellow). See Fig. 2d for survival. e) Heat map showing relative abundance of immune cells or their ratios from tumors that are resistant (black hatch) or sensitive to RT + anti-CTLA4. Boxplot shows bootstrap importance scores for each variable. Higher values (red) are more predictive. f) Change in T cell subsets or their ratio after RT + anti-CTLA4 for sensitive parental (Sen) or resistant (Res) tumors. Values are subtracted from average of untreated controls. Red line is mean. g) Heat map of resistance gene signature and PD-L1 across human melanoma. p < 0.001 by gene set enrichment analysis. h) Expression of PD-L1 on Res 499 compared to B16-F10 melanoma cells and of Res 237 compared to TSA breast cancer cells. Isotype control (IgG). i) Total tumor volume from PD-L1 knockout (KO) or control (WT) Res 499 and corresponding survival.
Mentions: Anecdotal clinical reports suggest that RT may cooperate with anti-CTLA4 to systemically enhance melanoma response7; however, this combination has not been reported in a clinical trial. To examine the feasibility and efficacy of RT combined with immune checkpoint blockade, we initiated a phase I clinical trial of 22 patients with multiple melanoma metastases (Extended Data Table 1). A single index lesion was irradiated with hypofractionated RT, delivered over two or three fractions, followed by four cycles of the anti-CTLA4 antibody ipilimumab (Extended Data Fig. 1a). Accrual was completed in three out of four RT dose levels, and treatment was well tolerated (Extended Data Table 2). Evaluation of the unirradiated lesions by CT imaging using Response Evaluation Criteria in Solid Tumors (RECIST) demonstrated that 18% of patients had a partial response (PR) as best response, 18% had stable disease (SD), and 64% had progressive disease (PD) (Fig. 1a). For example, patient PT-402 showed a large reduction in sizes of unirradiated tumors and a partial metabolic response by positron emission tomography (PET) (Fig. 1b). None of the 12 patients evaluated by PET had progressive metabolic disease in the irradiated lesion (Extended Data Fig. 1b, Extended Data Table 3). The median progression-free survival (PFS) and overall survival (OS) was 3.8 and 10.7 months with median follow-up of 18.4 and 21.3 months (18.0 and 21.3 for patients without event), respectively (Fig. 1c).

Bottom Line: Immune checkpoint inhibitors result in impressive clinical responses, but optimal results will require combination with each other and other therapies.Anti-CTLA4 predominantly inhibits T-regulatory cells (Treg cells), thereby increasing the CD8 T-cell to Treg (CD8/Treg) ratio.Addition of PD-L1 blockade reverses T-cell exhaustion to mitigate depression in the CD8/Treg ratio and further encourages oligoclonal T-cell expansion.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA [2] Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

ABSTRACT
Immune checkpoint inhibitors result in impressive clinical responses, but optimal results will require combination with each other and other therapies. This raises fundamental questions about mechanisms of non-redundancy and resistance. Here we report major tumour regressions in a subset of patients with metastatic melanoma treated with an anti-CTLA4 antibody (anti-CTLA4) and radiation, and reproduced this effect in mouse models. Although combined treatment improved responses in irradiated and unirradiated tumours, resistance was common. Unbiased analyses of mice revealed that resistance was due to upregulation of PD-L1 on melanoma cells and associated with T-cell exhaustion. Accordingly, optimal response in melanoma and other cancer types requires radiation, anti-CTLA4 and anti-PD-L1/PD-1. Anti-CTLA4 predominantly inhibits T-regulatory cells (Treg cells), thereby increasing the CD8 T-cell to Treg (CD8/Treg) ratio. Radiation enhances the diversity of the T-cell receptor (TCR) repertoire of intratumoral T cells. Together, anti-CTLA4 promotes expansion of T cells, while radiation shapes the TCR repertoire of the expanded peripheral clones. Addition of PD-L1 blockade reverses T-cell exhaustion to mitigate depression in the CD8/Treg ratio and further encourages oligoclonal T-cell expansion. Similarly to results from mice, patients on our clinical trial with melanoma showing high PD-L1 did not respond to radiation plus anti-CTLA4, demonstrated persistent T-cell exhaustion, and rapidly progressed. Thus, PD-L1 on melanoma cells allows tumours to escape anti-CTLA4-based therapy, and the combination of radiation, anti-CTLA4 and anti-PD-L1 promotes response and immunity through distinct mechanisms.

Show MeSH
Related in: MedlinePlus