Tumor innate immunity primed by specific interferon-stimulated endogenous retroviruses.

Mesenchymal tumor subpopulations secrete pro-tumorigenic cytokines and promote treatment resistance1-4. This phenomenon has been implicated in chemorefractory small cell lung cancer and resistance to targeted therapies5-8, but remains incompletely defined. Here, we identify a subclass of endogenous retroviruses (ERVs) that engages innate immune signaling in these cells. Stimulated 3 prime antisense retroviral coding sequences (SPARCS) are oriented inversely in 3' untranslated regions of specific genes enriched for regulation by STAT1 and EZH2. Derepression of these loci results in double-stranded RNA generation following IFN-γ exposure due to bi-directional transcription from the STAT1-activated gene promoter and the 5' long terminal repeat of the antisense ERV. Engagement of MAVS and STING activates downstream TBK1, IRF3, and STAT1 signaling, sustaining a positive feedback loop. SPARCS induction in human tumors is tightly associated with major histocompatibility complex class 1 expression, mesenchymal markers, and downregulation of chromatin modifying enzymes, including EZH2. Analysis of cell lines with high inducible SPARCS expression reveals strong association with an AXL/MET-positive mesenchymal cell state. While SPARCS-high tumors are immune infiltrated, they also exhibit multiple features of an immune-suppressed microenviroment. Together, these data unveil a subclass of ERVs whose derepression triggers pathologic innate immune signaling in cancer, with important implications for cancer immunotherapy

Characterizing genomic alterations in cancer by complementary functional associations.

Systematic efforts to sequence the cancer genome have identified large numbers of mutations and copy number alterations in human cancers. However, elucidating the functional consequences of these variants, and their interactions to drive or maintain oncogenic states, remains a challenge in cancer research. We developed REVEALER, a computational method that identifies combinations of mutually exclusive genomic alterations correlated with functional phenotypes, such as the activation or gene dependency of oncogenic pathways or sensitivity to a drug treatment. We used REVEALER to uncover complementary genomic alterations associated with the transcriptional activation of β-catenin and NRF2, MEK-inhibitor sensitivity, and KRAS dependency. REVEALER successfully identified both known and new associations, demonstrating the power of combining functional profiles with extensive characterization of genomic alterations in cancer genomes

Whole-exome sequencing and clinical interpretation of FFPE tumor samples to guide precision cancer medicine

Translating whole exome sequencing (WES) for prospective clinical use may impact the care of cancer patients; however, multiple innovations are necessary for clinical implementation. These include: (1) rapid and robust WES from formalin-fixed paraffin embedded (FFPE) tumor tissue, (2) analytical output similar to data from frozen samples, and (3) clinical interpretation of WES data for prospective use. Here, we describe a prospective clinical WES platform for archival FFPE tumor samples. The platform employs computational methods for effective clinical analysis and interpretation of WES data. When applied retrospectively to 511 exomes, the interpretative framework revealed a “long tail” of somatic alterations in clinically important genes. Prospective application of this approach identified clinically relevant alterations in 15/16 patients. In one patient, previously undetected findings guided clinical trial enrollment leading to an objective clinical response. Overall, this methodology may inform the widespread implementation of precision cancer medicine

Pulsed dosing of erlotinib for central nervous system (CNS) progression in EGFR-mutant non-small cell lung cancer (NSCLC).

8116 Background: Lung cancer is the most common cause of CNS metastases. Options for CNS progression are limited, particularly with leptomeningeal metastases (LM). High dose EGFR-TKIs have been used in this setting. This is a retrospective series of our experience with pulsed high dose erlotinib for these patients. Methods: Eligible pts with EGFR-mutant NSCLC were identified through our institutions’ databases and had received pulsed high dose erlotinib for CNS progression. Patients had received erlotinib 1000-1500 mg once weekly. The primary endpoint was CNS response; secondary endpoints included toxicity, systemic response, CNS progression-free survival, and overall survival. Results: Between 10/2010 – 10/2012, 10 eligible pts received pulsed dose erlotinib for CNS progression. The median age was 60 yrs; 8/10 were female, 7/10 were never-smokers, with a median of 2 pack-years. All pts had lung adenocarcinoma, and 9/10 had received prior EGFR-TKI. Median duration of prior TKI was 19 months. 6 received prior erlotinib, 1 received prior dacomitinib, and 2 received prior erlotinib followed by dacomitinib. The overall CNS response rate was 10% (1/10); 2 others achieved CNS stability. Median overall survival was 1.7 months (range 0.6 – 7.0). There was no clear correlation between outcomes and underlying EGFR genotype; type, duration, or dose of prior EGFR-TKI; or extent of CNS involvement. Conclusions: While there has been evidence of higher penetration of EGFR-TKI’s into cerebrospinal fluid with pulsed high doses of EGFR-TKI’s, the clinical efficacy of this strategy remains limited. [Table: see text] </jats:p

Abstract 2181: Genomic correlates of Metastasis in KRAS mutant lung adenocarcinoma

Abstract Background: Lung adenocarcinoma (LUAD) is a disease classified by molecular markers. In KRAS-mutant LUAD, STK11 and KEAP1 mutations are associated with decreased overall survival (OS), but predictors of metastasis have been poorly defined. In this study, we identify clinical and genomic predictors of metastatic KRAS-mutant LUAD. Methods: Patients with KRAS-mutant LUAD profiled by targeted next generation sequencing (OncoPanel) were included. Stage, histology, recurrence-free and overall survival were assessed. Clinical and genomic features between metastatic vs non-metastatic samples were compared. KRAS-mutant LUAD samples profiled using MSK-IMPACT in the AACR GENIE database were used to validate our findings. Continuous variables were compared using the Mann-Whitney U test and categorical variables were compared using the Fisher’s Exact test. Survival analysis was performed using the Cox Proportional Hazards model. WExT was used to identify co-occurring and mutually exclusive genetic alterations. Benjamini-Hochberg was used to correct for multiple comparisons. P-values &amp;lt; 0.05 and q-values &amp;lt; 0.10 were considered significant. Results: In the OncoPanel cohort (metastatic n=290; non-metastatic n=324), tumor mutational burden (TMB) (p = .001) and KEAP1 mutations (q = 0.05) were enriched in metastatic samples, while NFKBIA amplifications (q = 0.07) were enriched in non-metastatic samples. KEAP1/STK11 mutations significantly co-occurred (q &amp;lt; 1e-8). Compared to double wild-type samples: KEAP1/STK11 co-mutations were significantly enriched in metastatic samples (n = 72, p = 0.0002, OR 3.4); KEAP1-mutant samples trended towards enrichment in metastatic samples, (n = 21,p = 0.07, OR 2.47); STK11 mutations did not associate with stage (n = 53, p = 0.88, OR = 0.94). In multivariable survival analysis, metastasis (p &amp;lt; 0.005), KEAP1 mutation (p=0.01), and STK11 mutation (p=0.02) were associated with worse OS. In the MSK-IMPACT validation cohort (metastatic site n=417, primary site n = 781), KEAP1 was the only gene enriched in metastatic samples (q &amp;lt; 0.001) at q &amp;lt; 0.05. Compared to double wild type samples: KEAP/STK11 co-mutations (n=138, p &amp;lt; 0.0001, OR 2.1) and KEAP1 mutations (n=59, p = 0.04, OR 1.77) were enriched in metastatic samples; STK11-mutations did not associate with metastasis (n = 190, p = 0.34, OR 0.83). Other predictors of metastasis included Fraction Genome Altered (FGA) (p &amp;lt; 1e-5), TMB (p &amp;lt; 1e-5), and CDKN2A/B deletions (q &amp;lt; 0.003). Conclusion: While both KEAP1 and STK11 mutations are associated with decreased OS in KRAS-mutant LUAD, we find in two independent cohorts that only KEAP1 mutations and KEAP1/STK11 co-mutations, but not STK11 mutations, are associated with metastasis. We also found that FGA, TMB, CDKN2A/B deletions are strongly associated with metastasis. Further research is necessary to understand the influence of KEAP1 mutations, independent of and in-conjunction with STK11 mutations, on metastasis. Citation Format: Daniel Boiarsky, Christine A. Lydon, Emily Chambers, Pasi A. Janne, Mark M. Awad, Eliezer M. Van Allen, David Barbie, Natalie I. Vokes. Genomic correlates of Metastasis in KRAS mutant lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2181.</jats:p

Phase I/II investigator-initiated study of olaparib and temozolomide in SCLC: Updated analysis and CNS outcomes.

8565 Background: Temozolomide has activity in small-cell lung cancer (SCLC), including patients (pts) with brain metastases (mets; Pietanza M, Clin Cancer Res 2012). Inhibition of Poly (ADP-ribose) polymerase (PARP) is another therapeutic strategy in SCLC. We hypothesized that olaparib plus temozolomide may be safe and effective for pts with relapsed SCLC and clinically active against CNS disease (Farago A, Cancer Discovery 2019). Here, we present an updated analysis of this combination in pts with relapsed SCLC, including a second cohort testing an alternative dosing strategy and an exploratory analysis of CNS-specific outcomes. Methods: In this phase I/II trial of olaparib plus temozolomide in pts with recurrent SCLC, pts were sequentially enrolled into two cohorts defined by dosing schedule. In cohort 1, olaparib was dosed on D1-7 of each 21d cycle. In cohort 2, olaparib was dosed on D1-21. Temozolomide was dosed on D1-7 in both cohorts. Each cohort had a phase I portion (conventional 3+3 dose-escalation) for determination of MTD and RP2D and a phase II portion with primary endpoint of ORR. Per protocol, eligible pts could have untreated asymptomatic brain mets &lt; 1cm and, after mandatory baseline imaging, CNS imaging was performed at investigator’s discretion. A post-hoc exploratory analysis of CNS-specific outcomes was performed using modified RECIST criteria (Long GV, Lancet Oncol 2012) in which brain mets ≥5mm were considered measurable, and intracranial response was independently assessed by an attending radiologist. Results: 66 pts with median of 2 prior lines of therapy (range, 1-7) were enrolled, 50 pts in cohort 1 and 16 pts in cohort 2. 33/66 (50%) pts had history of brain mets, 15/66 (23%) pts had untreated brain mets at enrollment. The confirmed ORR of cohort 2 was 7% (1/14 evaluable pts, 95% CI: 0.2-33.9%), and the updated confirmed ORR of the entire study population was 34% (21/62 evaluable pts, 95% CI: 22.3-47.0%). The most common adverse events were hematologic toxicities (thrombocytopenia, anemia, and neutropenia). 22/50 (44%) of cohort 1 pts and 4/16 (25%) of cohort 2 pts required dose reduction. Of 15 pts with untreated brain mets, best overall intracranial response (including both confirmed and unconfirmed responses) was CR in 6 pts, PR in 4 pts, SD in 3 pts and PD in 1 for a CNS disease control rate of 87% (95% CI: 59.5-98.3%). Of 10 pts with CR/PR as their best intracranial response, 4 responses were confirmed. With non-CNS progression as a competing risk, the probability of CNS progression among the entire study population was 17% (95% CI: 8.8-26.7%) at 6 months and 21% (95% CI: 12.1-32.0%) at 12 months. Conclusions: Olaparib and temozolomide may be an effective therapy for relapsed SCLC, especially for pts with CNS disease where we observed a high rate of intracranial disease control. Ongoing analyses regarding optimal dosing schedule will inform potential for future use of this combination. Clinical trial information: NCT02446704. </jats:p

The prognostic significance of PDL1 expression in locally advanced non-small-cell lung cancer (LA-NSCLC) treated with definitive intent.

150 Background: The PACIFIC trial showed an 11-month (mo) improved median (med) progression-free survival after definitive chemoradiation in LA-NSCLC using the PDL1 inhibitor, durvalumab. This study will likely change practice and puts further emphasis on understanding the significance of immune marker expression in this poor-prognosis disease. Methods: From 1999-2012, pathology specimens from 126 patients (pts) with LA-NSCLC who received combined modality therapy at our institution and had sufficient tissue available were selected for tissue microarrays. Immunohistochemistry for PDL1 (E1L3N), PDL2 (9E5), PD1 (NAT105) and CD3 (F7.2.38) was performed and tumor cells and infiltrating immune cells with moderate or high staining intensity were quantified using Definiens Tissue Studio image analysis software. Univariate and multivariate analyses (MVA) were performed using a Cox proportional hazards model. Results: Distribution by stage was: IIA-3%, IIB-3%, IIIA-65%, IIIB-29%. Sixty-three percent of pts had surgery. All patients received fractionated radiation and chemotherapy with 81% receiving concurrent chemoradiation. Med follow-up from start of therapy was 20 mo (rg 2-168 mo). Median overall survival (OS) was 21 mo (2-yr OS 43%). Median progression-free survival (PFS) was 11 mo (2-yr PFS 26%). Immune marker expression was: PDL1-med 3% (rg 0-58%), PDL2-med 0.03% (rg 0-0.5%), PD1-med 0% (rg 0-76%) and CD3-med 21% (rg 0-78%). PDL2, PD1 and CD3 were not associated with outcomes. However, PDL1 &gt; 8% (chosen as the 75th-percentile) was associated with lower risk of distant metastases (DM) (HR 0.57, 95% CI 0.31-0.98, p = 0.04) and a trend towards longer time to disease progression (HR 0.63, 95% CI 0.37-1.02, p = 0.06). There was no association with local-regional recurrence or OS. On MVA including disease stage, PDL1 &gt; 8% was associated with lower risk of DM (HR 0.59, 95% CI 0.35-0.96, p = 0.03) and overall disease progression (HR 0.53, 95% CI 0.29-0.93), p = 0.02). Conclusions: PDL1 &gt; 8% is associated with lower risk of DM and better PFS in LA-NSCLC. This data suggests that even with traditional cancer therapies, the best outcomes may be seen in patients with higher PDL1 expression. </jats:p

Concurrent osimertinib plus gefitinib for first-line treatment of EGFR-mutated non-small cell lung cancer (NSCLC).

9507 Background: First-line treatment with an EGFR tyrosine kinase inhibitor (TKI) is standard of care for patients (pts) with EGFR-mutated NSCLC. The EGFR TKI osimertinib is active against the acquired gefitinib-resistant mutation EGFR T790M, as is gefitinib against the osimertinib-resistant EGFR C797S. Preclinical evidence suggests dual EGFR inhibition with gefitinib + osimertinib may delay emergence of acquired resistance. Methods: This ongoing phase I/II study enrolled pts with stage IV EGFR-mutated (L858R or del19) NSCLC, without prior therapy for metastatic disease. Treatment in dose escalation (n = 6): concurrent osimertinib 40 mg or 80 mg + gefitinib 250 mg daily. In dose expansion (n = 21): osimertinib + gefitinib at the maximum tolerated dose (MTD). Prior to protocol amendment 6 pts received alternating monthly cycles of TKI monotherapy and were excluded from this analysis. The primary endpoints in the dose escalation and expansion phases were, respectively, identification of the MTD and feasibility, defined as receipt of combination therapy for ≥ 6 four-week cycles. Secondary endpoints included overall response rate (ORR), survival outcomes, plasma EGFR mutation clearance (cell free DNA by droplet digital PCR (ddPCR)), and mechanisms of acquired resistance. Results: From May 2017 to July 2019 27 pts were enrolled and evaluable for the primary endpoints. The MTD was osimertinib 80 mg plus gefitinib 250 mg orally daily. In feasibility analysis, 81.5% completed ≥6 cycles combination therapy (1 pt discontinued for progression, 4 for toxicity). The ORR was 85.2% (95% CI 67.5%-94.1%). Best response: 85.2% partial response, 14.8% stable disease. The most common treatment-related adverse effects (TRAEs) (% any grade, % grade 3) were rash (96.3%, 3.7%), diarrhea (85.2%, 11.1%) and dry skin (70.4%, 0%). Plasma ddPCR (n = 25 pts) detected the driver EGFR mutation at baseline in 68% of pts. In these pts, plasma EGFR cleared to undetectable at 2 weeks treatment in 82.4%. At 14.8 months median follow up the median progression free survival was not yet reached. Conclusions: Combination therapy with osimertinib and gefitinib is tolerable for first-line treatment of EGFR-mutated NSCLC and resulted in rapid plasma clearance of the EGFR mutation. The observed ORR is consistent with previously reported first-line response rates to osimertinib. Analysis of survival outcomes and acquired resistance mechanisms are pending data maturity and will facilitate understanding of the role of first-line dual EGFR TKI therapy for this pt population. Clinical trial information: NCT03122717 . </jats:p