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Innate resistance of PD-1 blockade through loss of function mutations in JAK resulting in inability to express PD-L1 upon interferon exposure

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Journal for ImmunoTherapy of Cancer20153 (Suppl 2) :P311

https://doi.org/10.1186/2051-1426-3-S2-P311

  • Published:

Keywords

  • Melanoma Cell Line
  • Advanced Melanoma
  • Interferon Signaling
  • Innate Resistance
  • pSTAT6 Expression

Introduction

PD-L1-negative tumors assessed by immunohistochemistry often still respond to PD-1 blockade. PD-L1 is inducible by interferon, therefore, absolute negative tumors are the ones unable to up-regulate PD-L1 in response to interferons. Genetic mutations in the interferon receptor signaling pathway leading to loss of PD-L1 up-regulation were hypothesized to exhibit innate resistance to PD-1 blockade.

Experimental procedures

After optimization, 50 primary human melanoma cell lines were exposed to interferons (alpha, beta and gamma) and PD-L1 expression was measured. Interferon signaling was assessed by single cell phospho-proteomics, pSTAT1 (Y701), pSTAT3, pSTAT5, pSTAT6 expression level by flow cytometry (FACS LSRII). Western blot assessed JAK1/JAK2/IRF1 as well as STAT/pSTAT expression and Whole exome sequencing was performed by next generation sequencing for three selected melanoma cell lines and on biopsies from 25 patients with advanced melanoma treated with anti-PD-1 therapy.

Results

Three out of 50 melanoma cell lines were unable to up-regulate PD-L1 in response to interferon gamma; two of them had disruptive mutations in JAK1 or JAK2, and a third one had a defect in expression of IRF1 in response to interferons. Western blot analysis confirmed loss of function for the JAK1/JAK2 mutations and loss of downstream IRF1/STAT1/3/5 phosphorylation events. Whole exome sequencing of biopsies from 15 patients with metastatic melanoma who had objective response to PD-1 blockade (pembrolizumab) showed no homozygous inactivating mutations in interferon signaling pathway genes. Interestingly, one patient with the highest mutational load out of 10 patients without clinical response to PD-1 blockade had an amplified allele of JAK1 with a P429S mutation in the src-homology (SH2) domain. A cell line derived from this patient showed lack of sustained up-regulating of PD-L1 expression in response to interferon gamma by, and Western blot confirmed loss of JAK1 expression. Immunohistochemistry of tumor biopsy for this patient showed few CD8+ T cells.

Conclusions

This study has defined genetic mechanisms of innate resistance to PD-1 blockade which lead to inhibition of adaptive PD-L1 expression in patients with advanced melanoma. This work suggests lack of interferon-gamma induced PD-L1 upregulation has the potential to be a negative selective marker for PD-1 blockade therapy.

Authors’ Affiliations

(1)
UCLA, Los Angeles, CA, USA
(2)
UCLA Molecular and Medical Pharmacology, Los Angeles, CA, USA
(3)
David Geffen School of Medicine at UCLA, Los Angeles, CA, USA
(4)
Division of Hematology - Medical Oncology, UCLA Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA
(5)
UCLA, School of Medicine, LA, CA, USA
(6)
University of California at Los Angeles Medical Center, Los Angeles, CA, USA

Copyright

© Shin et al. 2015

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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