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22 Activity of the tumor-intrinsic NLRP3 inflammasome pathway predicts for response to checkpoint inhibitor immunotherapy in melanoma patients
  1. Tarek Haykal,
  2. Nagendra Yarla,
  3. Nicholas DeVito,
  4. Georgia Beasley,
  5. April Salama,
  6. Brent Hanks and
  7. Balamayooran Theivanthiran
  1. Duke University, Durham, NC, USA


Background We have previously determined that activation of a novel tumor-intrinsic NOD-, LRR- and pyrin domain-containing protein-3 (NLRP3) inflammasome-heat shock protein-70 (HSP70) signaling axis in response to PD-1 blockade triggers the recruitment of granulocytic myeloid-derived suppressor cells (PMN-MDSCs) into the tumor microenvironment, suppresses anti-tumor immunity and, in select settings, promotes tumor hyperprogression. We, therefore, sought to determine whether the activity of the tumor-intrinsic NLRP3-HSP70 pathway may correlate with anti-PD-1 response by interrogating clinical specimens derived from advanced melanoma patients undergoing anti-PD-1 monotherapy.

Methods Three independent approaches were utilized to measure the activity of the tumor-intrinsic NLRP3-HSP70 signaling pathway in 60 advanced melanoma patients undergoing either pembrolizumab or nivolumab monotherapy: 1. baseline week 0 plasma HSP70 levels were measured by ELISA, 2. germline PCR-based genotyping was performed to detect the single-nucleotide polymorphism (SNP), rs12239046, previously associated with enhanced NLRP3 expression, 3. PCR-based proximity ligation assay (PLA) analysis targeting the NLRP3-ASC proteins in baseline formalin-fixed paraffin-embedded tumor tissue specimens. Detection of the rs12239046 SNP was correlated with progression-free survival (PFS) while plasma HSP70 and NLRP3-ASC PLA levels were correlated with objective response (OR) based on RECIST1.1 assessment of week-12 CT imaging as well as PFS and overall survival (OS).

Results Our studies demonstrate that elevated baseline plasma HSP70 levels (P = 0.0008) and elevated baseline tissue NLRP3-ASC PLA levels (P = 0.0014) independently correlate with resistance to anti-PD-1 immunotherapy (ICI) based on week-12 OR in melanoma patients. Importantly, melanoma patients developing disease hyperprogression (n=5) in response to ICI exhibited elevations in baseline plasma HSP70 levels (P = <0.0001) and baseline tissue NLRP3-ASC PLA levels (P = <0.0001) relative to patients with week-12 disease progression (n=10). Above median baseline tissue NLRP3-ASC PLA levels were determined to correlate with both inferior PFS (HR 0.12, P = 0.0008) and OS (HR 0.16, P = 0.0456) in advanced melanoma patients undergoing ICI. Germline PCR detection of the rs12239046-C SNP was found to be associated with elevated plasma HSP70 levels and trended toward a correlation with inferior PFS (HR 0.50, P = 0.07).

Conclusions Baseline markers of the tumor-intrinsic NLRP3-HSP70 signaling pathway correlate with resistance and disease hyperprogression in melanoma patients undergoing anti-PD-1 immunotherapy. These data strongly support the important role of the tumor-intrinsic NLRP3 inflammasome in regulating responses to anti-PD-1 therapy and verify its relevance as a pharmacologic target to enhance immunotherapy efficacy. Expanded studies are warranted to confirm these findings in a larger patient cohort.

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