Genomic correlates of response to immune checkpoint therapies in clear cell renal cell carcinoma

PMID: 29301960 · DOI: 10.1126/science.aan5951 · Journal: Science (2018)

TL;DR

Whole-exome sequencing of pre-treatment metastatic clear cell renal cell carcinoma (CCRCC) tumors from 35 patients on a prospective trial of nivolumab (anti-PD-1) showed that loss-of-function (LOF) mutations in PBRM1 — a SWI/SNF (PBAF) complex subunit encoding BAF180 — were enriched in patients with clinical benefit (9/11 vs. 3/13; Fisher p=0.012). The finding was reproduced in an independent validation cohort of 63 ccRCC patients treated with anti-PD-(L)1 ± anti-CTLA-4 (17/27 vs. 4/19; p=0.0071). RNA-seq of PBAF-deficient cell lines and PBRM1-LOF tumors revealed altered transcriptional output in JAK/STAT, hypoxia, and immune signaling pathways, providing a mechanistic basis for the response association.

Cohort & data

• Discovery cohort: 35 patients with metastatic CCRCC on a prospective nivolumab (anti-PD-1) trial; pre-treatment paired tumor/normal whole-exome sequencing (mean target coverage 128× tumor / 91× normal). Dataset: ccrcc_dfci_2019.
• Validation cohort: 63 patients with metastatic CCRCC treated with anti-PD-1 (e.g., nivolumab) or anti-PD-L1 (e.g., atezolizumab) alone or combined with anti-CTLA-4 (ipilimumab); PBRM1 status from whole-exome sequencing (n=49) or panel sequencing (n=14).
• Reference cohorts for expression analysis: TCGA KIRC ccRCC, the Sato (Kyoto University) untreated ccRCC cohort, and on-study tumor RNA-seq.
• Comparator (clinical context): CheckMate 025 compared nivolumab vs. everolimus in advanced RCC.
• Endpoint: Composite response — clinical benefit (CB) = CR/PR by RECIST 1.1 or SD with any objective tumor reduction lasting ≥6 months; no clinical benefit (NCB) = PD by RECIST 1.1 with discontinuation within 3 months; remainder = intermediate benefit (IB).

Key findings

• In the discovery cohort, PBRM1 was the only recurrently mutated gene (by MutSig2CV) in which truncating/LOF mutations were enriched in CB vs. NCB tumors: 9/11 vs. 3/13 (Fisher’s exact p=0.012, q=0.086, OR for CB = 12.93, 95% CI 1.54–190.8).
• All discovery-cohort truncating PBRM1 alterations co-occurred with deletion of the non-mutated allele on chromosome 3p, producing complete biallelic LOF; most mutations were predicted to be clonal by ABSOLUTE.
• Patients with biallelic PBRM1 loss had significantly prolonged overall survival (log-rank p=0.0074) and progression-free survival (p=0.029) compared to PBRM1-intact patients on anti-PD-1.
• Independent validation cohort (n=63): truncating PBRM1 alterations enriched in CB vs. NCB (17/27 vs. 4/19; Fisher p=0.0071, OR=6.10, 95% CI 1.42–32.64).
• IB patients showed intermediate PBRM1 LOF rates in both cohorts (CB/IB/NCB: 82%/64%/23% discovery; 63%/41%/21% validation; Fisher-Freeman-Halton p=0.017 for both).
• Median nonsynonymous tumor mutation burden was modest (82/exome, range 45–157) and did not differ between CB and NCB groups; intratumoral heterogeneity (subclonal/clonal ratio) and antigen-presentation/HLA class I alterations also did not differ.
• PFS benefit conferred by PBRM1 LOF was more pronounced in previously-treated (largely VEGF-inhibitor pre-exposed) patients than in those receiving anti-PD-1 as first-line cancer therapy (p=0.009).
• One of four NCB validation patients with PBRM1 LOF also harbored a B2M alteration affecting antigen presentation, offering a potential explanation for non-response.
• GSEA of A704 ccRCC cell lines with PBAF-complex perturbations (BAF180-null and BRG1-null vs. PBAF-wildtype) showed enrichment of IL6/JAK-STAT3, TNF-α/NF-κB, IL2/STAT5, and hypoxia hallmark gene sets; the KEGG cytokine-cytokine receptor interaction founder gene set was the most strongly enriched (FWER q=0.0020 for BAF180-null vs. WT; q=0.023 for BRG1-null vs. WT).
• GSEA of pre-treatment tumor RNA-seq (n=18 PBRM1-LOF vs. n=14 PBRM1-intact) confirmed up-regulation of hypoxia and IL6/JAK-STAT3 hallmark gene sets in PBRM1-LOF tumors.
• Across TCGA, the Sato cohort, and on-study tumors, PBRM1-LOF tumors expressed lower levels of immune-inhibitory ligands (e.g., CD276, BTLA), though magnitudes were small and possibly confounded by stromal admixture.
• LOF mutations in VHL — the most commonly mutated ccRCC gene — did not correlate with immune-related gene expression, suggesting the immune-expression signal is specific to PBRM1.

Genes & alterations

• PBRM1 — biallelic LOF (truncating mutation + chromosome 3p loss of the WT allele); enriched in anti-PD-(L)1 responders in two ccRCC cohorts; PBRM1-LOF tumors show up-regulated hypoxia and JAK/STAT3 transcriptional programs and reduced expression of immune-inhibitory ligands.
• SMARCA4 (encodes BRG1) — used as an experimental PBAF-complex perturbation in A704 cells; BRG1-null cells phenocopy several BAF180-null transcriptional changes (immune/cytokine signaling enrichment).
• BRD7 and ARID2 — cited as essential PBAF-complex components whose loss (in mouse models, ref. 27) sensitizes tumor cells to T-cell–mediated killing, supporting a PBAF-wide immune-priming mechanism.
• B2M — antigen-presentation gene; one validation-cohort NCB patient had a co-occurring B2M alteration alongside PBRM1 LOF, hypothesized to explain non-response.
• VHL — most commonly mutated ccRCC gene; LOF status did NOT correlate with immune-related gene expression, indicating the immune-transcriptional signal is PBRM1-specific.

Clinical implications

• Truncating/LOF PBRM1 mutation is a candidate biomarker of clinical benefit from anti-PD-(L)1 monotherapy in metastatic CCRCC, independent of tumor mutation burden and PD-L1 IHC (which were not predictive in this cohort).
• The PBRM1-response association was strongest in patients previously treated with VEGF-targeted therapy, motivating prospective study of treatment sequencing (VEGF inhibitor → anti-PD-(L)1) and combinations.
• Concomitant B2M loss-of-function may abrogate PBRM1-associated benefit and warrants assessment in non-responders.
• Given the high prevalence of PBRM1 LOF in ccRCC (~41% per ref. 16) and SWI/SNF alterations across cancer (>20%), the authors propose that PBAF-complex status may be a broadly relevant immunotherapy biomarker beyond ccRCC.
• The authors filed a patent application (Dana-Farber) covering PBRM1 mutational status and immunotherapy response.

Limitations & open questions

• Modest cohort sizes (35 discovery, 63 validation) and retrospective heterogeneity of treatment regimens (anti-PD-1, anti-PD-L1, ± anti-CTLA-4) in the validation cohort.
• Copy-number confirmation of biallelic loss was not feasible for all validation samples, though 3p deletion is near-ubiquitous in ccRCC.
• Mechanistic data are based on a single cell line (A704) with PBAF perturbations and on tumor-bulk RNA-seq; causal in vivo experiments in the immune setting are still needed.
• Immune-inhibitory ligand expression differences (CD276, BTLA) were small and potentially confounded by tumor-stromal admixture.
• Generalizability to other tumor types with SWI/SNF alterations is hypothesized but not tested here.
• The differential PFS benefit of PBRM1 LOF in previously-treated vs. treatment-naive patients hints at an interaction with prior VEGF therapy that requires prospective validation.

Citations from this paper used in the wiki

• “PBRM1 was the only gene in which truncating, or loss-of-function (LOF) … mutations were enriched in tumors from patients in the CB vs. NCB group (9/11 vs. 3/13; Fisher’s exact p=0.012, q=0.086, odds ratio for CB=12.93, 95% C.I. 1.54–190.8)” (Results, discovery cohort).
• “Tumors from CB patients were more likely to harbor truncating alterations in PBRM1 (17/27 vs. 4/19, Fisher’s exact p=0.0071, odds ratio for CB=6.10, 95% C.I. 1.42–32.64)” (Results, validation cohort).
• “Patients whose tumors showed biallelic PBRM1 loss had significantly prolonged OS and PFS compared to patients without PBRM1 LOF (log-rank p=0.0074 and p=0.029, respectively)” (Results).
• “GSEA … revealed five gene sets whose expression was significantly enriched in cell lines that were PBAF-deficient. These included genes linked to IL6/JAK-STAT3 signaling, TNF-α signaling via NF-κB, and IL2/STAT5 signaling” (Results, Fig. 4A).
• “VHL mutation status did not correlate with immune-related gene expression … suggesting that observed differences in immune gene expression in the context of PBRM1 LOF may be specific to the PBRM1 gene” (Results).
• “the progression-free survival benefit conferred by PBRM1 LOF was more prominent in tumors from previously-treated patients compared to those from patients receiving anti-PD-1 therapy as their first cancer therapy (p=0.009)” (Results).

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