Nicholas DeVito

The tumor-intrinsic NOD-, LRR- and pyrin domain-containing protein-3 (NLRP3) inflammasome-heat shock protein 70 (HSP70) signaling axis is triggered by CD8+ T cell cytotoxicity and contributes to the development of adaptive resistance to anti-programmed cell death protein 1 (PD-1) immunotherapy by recruiting granulocytic polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) into the tumor microenvironment. Here, we demonstrate that the tumor NLRP3-HSP70 axis also drives the accumulation of PMN-MDSCs into distant lung tissues in a manner that depends on lung epithelial cell Toll-like receptor 4 (TLR4) signaling, establishing a premetastatic niche that supports disease hyperprogression in response to anti-PD-1 immunotherapy. Lung epithelial HSP70-TLR4 signaling induces the downstream Wnt5a-dependent release of granulocyte colony-stimulating factor (G-CSF) and C-X-C motif chemokine ligand 5 (CXCL5), thus promoting myeloid granulopoiesis and recruitment of PMN-MDSCs into pulmonary tissues. Treatment with anti-PD-1 immunotherapy enhanced the activation of this pathway through immunologic pressure and drove disease progression in the setting of Nlrp3 amplification. Genetic and pharmacologic inhibition of NLRP3 and HSP70 blocked PMN-MDSC accumulation in the lung in response to anti-PD-1 therapy and suppressed metastatic progression in preclinical models of melanoma and breast cancer. Elevated baseline concentrations of plasma HSP70 and evidence of NLRP3 signaling activity in tumor tissue specimens correlated with the development of disease hyperprogression and inferior survival in patients with stage IV melanoma undergoing anti-PD-1 immunotherapy. Together, this work describes a pathogenic mechanism underlying the phenomenon of disease hyperprogression in melanoma and offers candidate targets and markers capable of improving the management of patients with melanoma.

<jats:p> 1091 </jats:p><jats:p> Background: Tumor mutational burden (TMB) has emerged as an imperfect biomarker of immune checkpoint inhibition (ICI) outcomes in solid tumors. Despite the approval for pembrolizumab in all TMB-high (TMB-H) solid tumors, the optimal clinical approach to TMB-H or hypermutated advanced/metastatic breast cancer (MBC) is unknown with sparse prospective data. We hypothesize that TMB-H MBC will have unique genomic alterations compared to TMB-low (TMB-L) breast cancer that could inform novel therapeutic approaches. Methods: Tumor samples (N = 5621) obtained from patients with MBC were analyzed by next-generation sequencing (NGS) of DNA (592-gene panel or whole exome sequencing) and RNA (whole transcriptome sequencing) at Caris Life Sciences (Phoenix, AZ). TMB was calculated based on recommendations from the Friends of Cancer Research TMB Harmonization Project (Merino et al., 2020), with the TMB-H threshold set to ≥ 10 muts/Mb. IHC was performed for PD-L1 (Ventana SP142 ≥1% immune cells). Deficient mismatch repair (dMMR)/high microsatellite instability (MSI-H) was tested by IHC and NGS, respectively. Results: TMB-H was identified in 8.2% (n = 461) of MBC samples, with similar frequencies observed across molecular subtypes (7.8-8.6%, p = 0.85): HR+/HER2- (n = 3087) 7.8%, HR+/HER2+ (n = 266) 8.3%, HR-/HER2+ (n = 179) 7.8%, TNBC (n = 1476) 8.6%. The frequency of TMB-H was significantly increased in lobular (16%) versus ductal (5%) MBC (p &lt; 0.01). TMB-H samples were enriched in genitourinary (42%), soft tissue (20%), and gastrointestinal non-liver (16%) biopsy specimens. Compared to TMB-L tumors, TMB-H tumors exhibited significantly higher mutation rates for TP53 (60 v 52%), PIK3CA (55 vs 31%), ARID1A (34 vs 11%), CDH1 (27 vs 11%), NF1 (22 vs 9%), RB1 (14 vs 5%), KMT2C (12 vs 7%), PTEN (12 vs 7%), ERBB2 (7 vs 2.9%), and PALB2 (3.3 vs 1%) genes (p &lt; 0.05 each). Copy number alteration and fusion rates did not differ between TMB-H and TMB-L breast cancers. PI3K/AKT/MTOR, TP53, Histone/Chromatin remodeling, DNA damage repair (DDR), RAS, and cell cycle pathway alterations were detected in &gt; 25% TMB-H MBCs (p &lt; 0.05 each). dMMR/MSI-High (7.2 vs 0.3%, p &lt; 0.01) and PD-L1 positivity (36 vs 28%, p &lt; 0.05) frequencies were significantly increased in TMB-H tumors. DNA signature analyses including APOBEC and homologous recombination repair deficiency, as well as gene expression profiling to assess immune-related signatures and tumor microenvironment are underway. Conclusions: TMB-H breast cancers contain a unique genomic profile enriched with targetable mutations such as PIK3CA, ARID1A, NF1, PTEN, ERBB2, and PALB2. Concurrent predictive biomarkers of response to immune checkpoint inhibition such as MSI-H and PDL-1 positivity are also more prevalent in TMB-H MBC. These findings suggest novel combination strategies within TMB-H MBC could be explored. </jats:p>

The mechanisms underlying tumor immunosurveillance and their association with the immune-related adverse events (irAEs) associated with checkpoint inhibitor immunotherapies remain poorly understood. We describe a metastatic melanoma patient exhibiting multiple episodes of spontaneous disease regression followed by the development of several irAEs during the course of anti-programmed cell death protein 1 antibody immunotherapy. Whole-exome next-generation sequencing studies revealed this patient to harbor a pyrin inflammasome variant previously described to be associated with an atypical presentation of familial Mediterranean fever. This work highlights a potential role for inflammasomes in the regulation of tumor immunosurveillance and the pathogenesis of irAEs.

AIMS: Use of comprehensive genomic profiling (CGP) in metastatic colorectal cancer (mCRC) is limited. We estimated impacts of expanded 1 L CGP, using the Tempus xT test, on detection of actionable alterations and testing budgets in a modeled US health plan over two-years. MATERIALS AND METHODS: A decision analytic model was developed to estimate the impact of replacing 20% of usual testing (a mix of CGP and non-CGP) with Tempus xT CGP. Actionable alterations for matched treatments or clinical trial included KRAS, NRAS, RAF, BRAF, deficient mismatch repair (dMMR)/microsatellite instability (MSI), NTRK, RET, EGFR, HER2, MET, PIK3CA and POLE1. Costs included initial and repeat testing, physician-associated and administrative costs. RESULTS: In a hypothetical five-million-member plan, 50% Medicare and 50% commercial, 1,112 new cases of mCRC were expected per year. Of these, 566 (51%) would undergo 1 L molecular testing, with 55 re-tested upon progression. Based on current testing rates, there were an expected 521 missed opportunities for genomically informed treatment (47% of new cases), with 442 missed due to lack of testing and 79 due to testing without CGP. Replacing 20% of usual testing with Tempus xT CGP was associated with up to a $0.003 per member per month testing cost increase (net total cost of $202,102 for the five-million-member plan) and 15.5 additional patients with an opportunity for genomically informed care (12.7 patients for treatment and 2.8 for clinical trial). The testing total cost (initial test, repeat test, biopsy and physician services, and administrative cost) to put one additional patient with mCRC on matched therapy or matched clinical trial was estimated to be $13,005. Number needed to test to identify one actionable alteration with Tempus xT CGP versus usual testing was 7.8 patients. LIMITATIONS: Conservative assumptions were made for inputs with limited evidence. Based on high concordance rates with dMMR/MSI status, tumor mutational burden (TMB) status was not calculated separately. CONCLUSIONS: Replacing 20% of usual testing with Tempus xT CGP was associated with a small incremental testing cost and can identify meaningfully more actionable alterations.