Combination anti-PD-1 and anti-CTLA-4 therapy generates waves of clonal responses that include progenitor-exhausted CD8+ T cells

doi:10.1016/j.ccell.2024.08.007

. 2024 Sep 9;42(9):1582-1597.e10.

doi: 10.1016/j.ccell.2024.08.007. Epub 2024 Aug 29.

Combination anti-PD-1 and anti-CTLA-4 therapy generates waves of clonal responses that include progenitor-exhausted CD8⁺ T cells

Kevin Wang¹, Paulina Coutifaris¹, David Brocks², Guanning Wang¹, Tarek Azar¹, Sabrina Solis³, Ajeya Nandi¹, Shaneaka Anderson¹, Nicholas Han¹, Sasikanth Manne⁴, Evgeny Kiner², Chirag Sachar², Minke Lucas⁵, Sangeeth George¹, Patrick K Yan¹, Melanie W Kier¹, Amy I Laughlin¹, Shawn Kothari¹, Josephine Giles⁴, Divij Mathew⁴, Reem Ghinnagow⁶, Cecile Alanio⁷, Ahron Flowers⁸, Wei Xu⁶, Daniel J Tenney⁹, Xiaowei Xu¹⁰, Ravi K Amaravadi¹¹, Giorgos C Karakousis¹², Lynn M Schuchter¹¹, Marcus Buggert¹³, Derek Oldridge¹⁴, Andy J Minn¹⁵, Christian Blank¹⁶, Jeffrey S Weber³, Tara C Mitchell¹¹, Michael D Farwell¹⁷, Ramin S Herati¹⁸, Alexander C Huang¹⁹

Affiliations

¹ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
² Immunai Inc, New York, NY 10016, USA.
³ Department of Medicine, Grossman School of Medicine, New York University, New York, NY 10016, USA.
⁴ Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
⁵ Department of Medical Oncology, Netherlands Cancer Institute, Amsterdam 1066 CX, the Netherlands.
⁶ Department of Radiation Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
⁷ Institut Curie, PSL University, Inserm U932, Immunity and Cancer, 75005 Paris, France; Clinical Immunology and Immunomonitoring Laboratory, Institut Curie, Paris, France.
⁸ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA.
⁹ Bristol Myers Squibb, Princeton, NJ 08540, USA.
¹⁰ Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹¹ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹² Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Surgery, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹³ Institute for Immunology and Immune Health, Philadelphia, PA 19104, USA.
¹⁴ Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Center for Computational and Genomic Medicine, The Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
¹⁵ Department of Radiation Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Institute for Immunology and Immune Health, Philadelphia, PA 19104, USA; Parker Institute for Cancer Immunotherapy, Philadelphia, PA 19104, USA.
¹⁶ Department of Medical Oncology, Netherlands Cancer Institute, Amsterdam 1066 CX, the Netherlands; Department of Medical Oncology, Leiden University Medical Center (LUMC), Leiden 2333 ZA, the Netherlands; Department of Hematology and Oncology, University Clinic of Regensburg (UKR), 93053 Regensburg, Germany.
¹⁷ Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Radiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹⁸ Department of Medicine, Grossman School of Medicine, New York University, New York, NY 10016, USA. Electronic address: ramin.herati@nyulangone.org.
¹⁹ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Institute for Immunology and Immune Health, Philadelphia, PA 19104, USA; Parker Institute for Cancer Immunotherapy, Philadelphia, PA 19104, USA; Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. Electronic address: alexander.huang@pennmedicine.upenn.edu.

PMID: 39214097
PMCID: PMC11387127 (available on 2025-09-09)
DOI: 10.1016/j.ccell.2024.08.007

Combination anti-PD-1 and anti-CTLA-4 therapy generates waves of clonal responses that include progenitor-exhausted CD8⁺ T cells

Kevin Wang et al. Cancer Cell. 2024.

. 2024 Sep 9;42(9):1582-1597.e10.

doi: 10.1016/j.ccell.2024.08.007. Epub 2024 Aug 29.

Authors

Affiliations

¹ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
² Immunai Inc, New York, NY 10016, USA.
³ Department of Medicine, Grossman School of Medicine, New York University, New York, NY 10016, USA.
⁴ Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
⁵ Department of Medical Oncology, Netherlands Cancer Institute, Amsterdam 1066 CX, the Netherlands.
⁶ Department of Radiation Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
⁷ Institut Curie, PSL University, Inserm U932, Immunity and Cancer, 75005 Paris, France; Clinical Immunology and Immunomonitoring Laboratory, Institut Curie, Paris, France.
⁸ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA.
⁹ Bristol Myers Squibb, Princeton, NJ 08540, USA.
¹⁰ Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹¹ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹² Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Surgery, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹³ Institute for Immunology and Immune Health, Philadelphia, PA 19104, USA.
¹⁴ Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Center for Computational and Genomic Medicine, The Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
¹⁵ Department of Radiation Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Institute for Immunology and Immune Health, Philadelphia, PA 19104, USA; Parker Institute for Cancer Immunotherapy, Philadelphia, PA 19104, USA.
¹⁶ Department of Medical Oncology, Netherlands Cancer Institute, Amsterdam 1066 CX, the Netherlands; Department of Medical Oncology, Leiden University Medical Center (LUMC), Leiden 2333 ZA, the Netherlands; Department of Hematology and Oncology, University Clinic of Regensburg (UKR), 93053 Regensburg, Germany.
¹⁷ Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Radiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
¹⁸ Department of Medicine, Grossman School of Medicine, New York University, New York, NY 10016, USA. Electronic address: ramin.herati@nyulangone.org.
¹⁹ Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Tara Miller Melanoma Center, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Institute for Immunology and Immune Health, Philadelphia, PA 19104, USA; Parker Institute for Cancer Immunotherapy, Philadelphia, PA 19104, USA; Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. Electronic address: alexander.huang@pennmedicine.upenn.edu.

PMID: 39214097
PMCID: PMC11387127 (available on 2025-09-09)
DOI: 10.1016/j.ccell.2024.08.007

Abstract

Combination checkpoint blockade with anti-PD-1 and anti-CTLA-4 antibodies has shown promising efficacy in melanoma. However, the underlying mechanism in humans remains unclear. Here, we perform paired single-cell RNA and T cell receptor (TCR) sequencing across time in 36 patients with stage IV melanoma treated with anti-PD-1, anti-CTLA-4, or combination therapy. We develop the algorithm Cyclone to track temporal clonal dynamics and underlying cell states. Checkpoint blockade induces waves of clonal T cell responses that peak at distinct time points. Combination therapy results in greater magnitude of clonal responses at 6 and 9 weeks compared to single-agent therapies, including melanoma-specific CD8⁺ T cells and exhausted CD8⁺ T cell (T_EX) clones. Focused analyses of T_EX identify that anti-CTLA-4 induces robust expansion and proliferation of progenitor T_EX, which synergizes with anti-PD-1 to reinvigorate T_EX during combination therapy. These next generation immune profiling approaches can guide the selection of drugs, schedule, and dosing for novel combination strategies.

Keywords: CTLA-4 blockade; Checkpoint blockade; PD-1 blockade; T cell exhaustion; clonotypic analysis; combination checkpoint blockade; immunotherapy; melanoma; progenitor exhausted CD8(+) T cells; single-cell sequencing.

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Conflict of interest statement

Declaration of interests A.C.H. performed consulting work for Immunai and received research funding from Bristol Myers Squibb and Merck. R.S.H. has performed consulting work for Bristol Myers Squibb (exclusive of the current work). T.C.M. received honorarium for Scientific Advisory Board participation from: BMS, GigaGen, Merck, Pliant, Pfizer. G.C.K. is on the Merck Advisory Board. J.W. consulted for and have received less than $10,000 per annum from Merck, Genentech, AstraZeneca, GSK, Novartis, Nektar, Celldex, Incyte, Biond, Moderna, ImCheck, Sellas, Evaxion, Pfizer, Regeneron, and EMD Serono and received $10–$25,000 from BMS for membership on advisory boards. J.W. also holds equity in Biond, Evaxion, OncoC4, and Instil Bio, and on scientific advisory boards for CytomX, Incyte, ImCheck, Biond, Sellas, Instil Bio, OncoC4, and NexImmune and remunerated between $10,000–$50,000. In addition, J.W. is named on a patent filed by Moffitt Cancer Center on an ipilimumab biomarker and on TIL preparation and also on a PD-1 patent filed by Biodesix; J.W. receives less than $6000 in royalties. D.B., E.K., and C.S. were employed by Immunai when engaged in this project. S.G. and D.T. are employees of BMS. C.A. is a consultant for Biotherapy Partners.

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References

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[1] Larkin J, Chiarion-Sileni V, Gonzalez R, Grob JJ, Rutkowski P, Lao CD, Cowey CL, Schadendorf D, Wagstaff J, Dummer R, et al. (2019). Five-Year Survival with Combined Nivolumab and Ipilimumab in Advanced Melanoma. N Engl J Med 381, 1535–1546. 10.1056/NEJMoa1910836. - DOI - PubMed

[2] Larkin J, Chiarion-Sileni V, Gonzalez R, Grob JJ, Rutkowski P, Lao CD, Cowey CL, Schadendorf D, Wagstaff J, Dummer R, et al. (2019). Five-Year Survival with Combined Nivolumab and Ipilimumab in Advanced Melanoma. N Engl J Med 381, 1535–1546. 10.1056/NEJMoa1910836. - DOI - PubMed

[3] Wei SC, Duffy CR, and Allison JP (2018). Fundamental Mechanisms of Immune Checkpoint Blockade Therapy. Cancer Discov 8, 1069–1086. 10.1158/2159-8290.CD-18-0367. - DOI - PubMed

[4] Wei SC, Duffy CR, and Allison JP (2018). Fundamental Mechanisms of Immune Checkpoint Blockade Therapy. Cancer Discov 8, 1069–1086. 10.1158/2159-8290.CD-18-0367. - DOI - PubMed

[5] Brunner MC, Chambers CA, Chan FK, Hanke J, Winoto A, and Allison JP (1999). CTLA-4-Mediated inhibition of early events of T cell proliferation. J Immunol 162, 5813–5820. - PubMed

[6] Brunner MC, Chambers CA, Chan FK, Hanke J, Winoto A, and Allison JP (1999). CTLA-4-Mediated inhibition of early events of T cell proliferation. J Immunol 162, 5813–5820. - PubMed

[7] Walunas TL, Lenschow DJ, Bakker CY, Linsley PS, Freeman GJ, Green JM, Thompson CB, and Bluestone JA (1994). CTLA-4 can function as a negative regulator of T cell activation. Immunity 1, 405–413. 10.1016/1074-7613(94)90071-x. - DOI - PubMed

[8] Walunas TL, Lenschow DJ, Bakker CY, Linsley PS, Freeman GJ, Green JM, Thompson CB, and Bluestone JA (1994). CTLA-4 can function as a negative regulator of T cell activation. Immunity 1, 405–413. 10.1016/1074-7613(94)90071-x. - DOI - PubMed

[9] Linsley PS, Brady W, Urnes M, Grosmaire LS, Damle NK, and Ledbetter JA (1991). CTLA-4 is a second receptor for the B cell activation antigen B7. J Exp Med 174, 561–569. 10.1084/jem.174.3.561. - DOI - PMC - PubMed

[10] Linsley PS, Brady W, Urnes M, Grosmaire LS, Damle NK, and Ledbetter JA (1991). CTLA-4 is a second receptor for the B cell activation antigen B7. J Exp Med 174, 561–569. 10.1084/jem.174.3.561. - DOI - PMC - PubMed

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Combination anti-PD-1 and anti-CTLA-4 therapy generates waves of clonal responses that include progenitor-exhausted CD8⁺ T cells

Affiliations

Combination anti-PD-1 and anti-CTLA-4 therapy generates waves of clonal responses that include progenitor-exhausted CD8⁺ T cells

Authors

Affiliations

Abstract

Conflict of interest statement

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