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Fig. S3, related to Fig. 3.

Supplementary Legends, Materials and Methods

Purpose:Stabilization of the transcription factor NRF2 through genomic alterations in KEAP1 and NFE2L2 occurs in a quarter of patients with lung adenocarcinoma and a third of patients with lung squamous cell carcinoma. In lung adenocarcinoma, KEAP1 loss often co-occurs with STK11 loss and KRAS-activating alterations. Despite its prevalence, the impact of NRF2 activation on tumor progression and patient outcomes is not fully defined.Experimental Design:We model NRF2 activation, STK11 loss, and KRAS activation in vivo using novel genetically engineered mouse models. Furthermore, we derive a NRF2 activation signature from human non–small cell lung tumors that we use to dissect how these genomic events impact outcomes and immune contexture of participants in the OAK and IMpower131 immunotherapy trials.Results:Our in vivo data reveal roles for NRF2 activation in (i) promoting rapid-onset, multifocal intrabronchiolar carcinomas, leading to lethal pulmonary dysfunction, and (ii) decreasing elevated redox stress in KRAS-mutant, STK11-null tumors. In patients with nonsquamous tumors, the NRF2 signature is negatively prognostic independently of STK11 loss. Patients with lung squamous cell carcinoma with low NRF2 signature survive longer when receiving anti–PD-L1 treatment.Conclusions:Our in vivo modeling establishes NRF2 activation as a critical oncogenic driver, cooperating with STK11 loss and KRAS activation to promote aggressive lung adenocarcinoma. In patients, oncogenic events alter the tumor immune contexture, possibly having an impact on treatment responses. Importantly, patients with NRF2-activated nonsquamous or squamous tumors have poor prognosis and show limited response to anti–PD-L1 treatment.

Supplementary Tables

BACKGROUND This phase III OAK trial (NCT02008227) subgroup analysis (data cutoff, January 9, 2019) evaluated the predictive value of 2 PD-L1 IHC tests (VENTANA SP142 and Dako 22C3) for benefit from atezolizumab versus docetaxel by programmed death ligand 1 (PD-L1) status in patients with previously treated metastatic non-small cell lung cancer. METHODS PD-L1 expression was assessed prospectively with SP142 on tumor cells (TC) and tumor-infiltrating immune cells (IC) and retrospectively with 22C3 using a tumor proportion score (TPS) based on TC membrane staining. Efficacy was assessed in the 22C3 biomarker-evaluable population (22C3-BEP) (n = 577; 47.1% of SP142-intention-to-treat population) and non-22C3-BEP (n = 648) in PD-L1 subgroups (high, low, and negative) and according to selection by 1 or both assays. RESULTS In the 22C3-BEP, overall survival benefits with atezolizumab versus docetaxel were observed across PD-L1 subgroups; benefits were greatest in SP142-defined PD-L1-high (TC3 or IC3: hazard ratio [HR], 0.39 [95% confidence interval (CI), 0.25-0.63]) and 22C3-defined PD-L1-high (TPS ≥ 50%: HR, 0.56 [95% CI, 0.38-0.82]) and low (TPS, 1% to < 50%: HR, 0.55 [95% CI, 0.37-0.82]) groups. Progression-free survival improved with increasing PD-L1 expression for both assays. SP142 and 22C3 assays identified overlapping and unique patient populations in PD-L1-high, positive, and negative subgroups. Overall survival and progression-free survival benefits favored atezolizumab over docetaxel in double PD-L1-positive and negative groups; patients with both SP142- and 22C3-positive tumors derived the greatest benefit. CONCLUSIONS Despite different scoring algorithms and differing sensitivity levels, the SP142 and 22C3 assays similarly predicted atezolizumab benefit at validated PD-L1 thresholds in patients with non-small cell lung cancer.

Introduction We report the final overall survival (OS) analyses of atezolizumab-carboplatin-paclitaxel (ACP [experimental arm]) and OS data with approximately 39.8 months of median follow-up with atezolizumab-bevacizumab-carboplatin-paclitaxel (ABCP) versus bevacizumab-carboplatin-paclitaxel (BCP) in chemotherapy-naive patients with metastatic nonsquamous NSCLC in the phase 3 IMpower150 study (NCT02366143). Methods In this randomized, open-label study (N = 1202), coprimary end points included investigator-assessed progression-free survival and OS in intention-to-treat (ITT) wild-type (WT; no EGFR or ALK alterations) patients. Secondary and exploratory end points included OS in ITT and programmed death-ligand 1 (PD-L1) subgroups defined by the VENTANA SP142 and SP263 immunohistochemistry assays. Results At the final analysis with ACP versus BCP (data cutoff: September 13, 2019; minimum follow-up: 32.4 mo), ACP had numerical, but not statistically significant, improvements in OS (ITT-WT: median OS = 19.0 versus 14.7 mo; hazard ratio = 0.84; 95% confidence interval: 0.71–1.00). OS benefit was sustained with ABCP versus BCP (ITT-WT: 19.5 versus 14.7 mo; hazard ratio = 0.80; 95% confidence interval: 0.67–0.95). Exploratory analyses in the SP142-defined PD-L1 subgroups revealed longer median OS with ABCP and ACP versus BCP in PD-L1–high and PD-L1–positive subgroups; in the PD-L1–negative subgroups, median OS was similar with ACP and ABCP versus BCP. Safety was consistent with that in earlier analyses (data cutoff: January 22, 2018). Conclusions At the final IMpower150 OS analysis, ACP had numerical, but not statistically significant, OS improvement versus BCP. Updated data with an additional 20 months of follow-up revealed continued OS improvement with ABCP versus BCP in all patients.

The confluence of immunology and oncology has led to a lot of uncertainty and questions about relevant biomarkers. Despite the complexity of the tumour microenvironment, most clinical studies have relied on a single‐parameter immunohistochemical assay to prospectively select patients for checkpoint inhibitor therapy; the results of this strategy have been highly variable and often less than optimal. While great efforts have been made to identify additional or alternative biomarkers, pathologists, drug developers, and clinicians alike have faced technical, logistical, and regulatory challenges on how to implement them successfully. In this review, we will discuss these challenges; we will also highlight recent advances in dissecting the functional diversity of immune cell populations within the tumour microenvironment and their potential for improved, biomarker‐driven therapeutic strategies. The dynamic nature and cellular diversity of the tumour microenvironment may challenge past models of a single biomarker predicting patient response and clinical outcome. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

PURPOSE: IMpower133 (ClinicalTrials.gov identifier: NCT02763579 ), a randomized, double-blind, phase I/III study, demonstrated that adding atezolizumab (anti-programmed death-ligand 1 [PD-L1]) to carboplatin plus etoposide (CP/ET) for first-line (1L) treatment of extensive-stage small-cell lung cancer (ES-SCLC) resulted in significant improvement in overall survival (OS) and progression-free survival (PFS) versus placebo plus CP/ET. Updated OS, disease progression patterns, safety, and exploratory biomarkers (PD-L1, blood-based tumor mutational burden [bTMB]) are reported. PATIENTS AND METHODS: Patients with untreated ES-SCLC were randomly assigned 1:1 to receive four 21-day cycles of CP (area under the curve 5 mg per mL/min intravenously [IV], day 1) plus ET (100 mg/m2 IV, days 1-3) with atezolizumab (1,200 mg IV, day 1) or placebo, and then maintenance atezolizumab or placebo until unacceptable toxicity, disease progression, or loss of clinical benefit. Tumor specimens were collected; PD-L1 testing was not required for enrollment. The two primary end points, investigator-assessed PFS and OS, were statistically significant at the interim analysis. Updated OS and PFS and exploratory biomarker analyses were conducted. RESULTS: Patients received atezolizumab plus CP/ET (n = 201) or placebo plus CP/ET (n = 202). At the updated analysis, median follow-up for OS was 22.9 months; 302 deaths had occurred. Median OS was 12.3 and 10.3 months with atezolizumab plus CP/ET and placebo plus CP/ET, respectively (hazard ratio, 0.76; 95% CI, 0.60 to 0.95; descriptive P = .0154). At 18 months, 34.0% and 21.0% of patients were alive in atezolizumab plus CP/ET and placebo plus CP/ET arms, respectively. Patients derived benefit from the addition of atezolizumab, regardless of PD-L1 immunohistochemistry or bTMB status. CONCLUSION: Adding atezolizumab to CP/ET as 1L treatment for ES-SCLC continued to demonstrate improved OS and a tolerable safety profile at the updated analysis, confirming the regimen as a new standard of care. Exploratory analyses demonstrated treatment benefit independent of biomarker status.

Purpose: Stabilization of the transcription factor NRF2 through genomic alterations in KEAP1 and NFE2L2 occurs in a quarter of patients with lung adenocarcinoma and a third of patients with lung squamous cell carcinoma. In lung adenocarcinoma, KEAP1 loss often co-occurs with STK11 loss and KRAS-activating alterations. Despite its prevalence, the impact of NRF2 activation on tumor progression and patient outcomes is not fully defined. Experimental Design: We model NRF2 activation, STK11 loss, and KRAS activation in vivo using novel genetically engineered mouse models. Furthermore, we derive a NRF2 activation signature from human non–small cell lung tumors that we use to dissect how these genomic events impact outcomes and immune contexture of participants in the OAK and IMpower131 immunotherapy trials. Results: Our in vivo data reveal roles for NRF2 activation in (i) promoting rapid-onset, multifocal intrabronchiolar carcinomas, leading to lethal pulmonary dysfunction, and (ii) decreasing elevated redox stress in KRAS-mutant, STK11-null tumors. In patients with nonsquamous tumors, the NRF2 signature is negatively prognostic independently of STK11 loss. Patients with lung squamous cell carcinoma with low NRF2 signature survive longer when receiving anti–PD-L1 treatment. Conclusions: Our in vivo modeling establishes NRF2 activation as a critical oncogenic driver, cooperating with STK11 loss and KRAS activation to promote aggressive lung adenocarcinoma. In patients, oncogenic events alter the tumor immune contexture, possibly having an impact on treatment responses. Importantly, patients with NRF2-activated nonsquamous or squamous tumors have poor prognosis and show limited response to anti–PD-L1 treatment.

Background The efficacy and safety of the anti–programmed death ligand 1 (PD-L1) monoclonal antibody atezolizumab, as compared with those of platinum-based chemotherapy, as first-line trea...

BackgroundModern machine learning (ML) models based on highly multivariate attribute sets (e.g. unbiased -omics data) can be very successful at generating clinically useful predictions, but at the price of less transparency in how individual attributes are used to make those predictions. In short, ML test algorithms tend to be ”black boxes”. Shapley values (SVs)1 describe the relative importance of the attributes used within a multivariate test to the generation of the test result for an individual patient.2 While typically the calculation of SVs is computationally prohibitive, our ML architecture permits the generation of SVs for large patient cohorts. In this study, we evaluate SVs for the Anti-PD-L1 Response Test (ART), that was shown in independent validation to predict outcomes for patients treated with atezolizumab,3 for the POPLAR Ph2 and OAK Ph3 studies of non-small cell lung cancer (NSCLC) patients .4 5Abstract 831 Figure 1Radar plots illustrating the values of the 10 most important SVs for test classification generation for two samples classified as poor (A and B) and two samples classified as good (C and D). Positive SVs are shown in red and negative SVs are shown in blueAbstract 831 Figure 2Heatmaps of the SVs for samples classified as poor showing two subgroups (top and bottom, separated by horizontal line) with different patterns of SVs for (A) POPLAR and (B) OAKAbstract 831 Figure 3Heatmaps of the SVs for samples classified as good showing three subgroups, top, middle, and bottom, separated by horizontal lines) with different patterns of SVs for (A) POPLAR and (B) OAKMethodsART results, Good or Poor had been produced for 262 patients in POPLAR (NCT01903993) and 786 patients in OAK (NCT02008227). Exact SVs were generated for each pretreatment serum sample for each of the 93 attributes (proteomic features) used in the test. The distribution of SVs across the cohort was investigated to assess the relative importance of each feature to test classification. Subgroups of patients with similar patterns of SVs were identified using t-sne plots and ML methods in the POPLAR cohort and validated in the OAK cohort.ResultsThe SV distributions showed that the features influencing ART classification most were similar in both POPLAR and OAK. The relative importance of features to test classification differed between patients (figure 1), but subgroups of patients within test classification groups showed similar patterns of SVs (figures 2 and 3). Such patient subgroups, identified within POPLAR, were also found in the OAK cohort and were associated with differences in outcome and/or differences in patient characteristics.ConclusionsSVs can explain how complex ML-based tests combine molecular attributes to produce individual patient results. Exact SVs can be obtained for certain ML architectures used in molecular test development, revealing the overall relative importance of attributes used in such molecular tests. Subgrouping of patients with the same test classification by different patterns of SVs is possible. This may reveal different biologies contributing to a Good or Poor phenotype and inform translational studies.Trial RegistrationClinicalTrialsgov NCT01903993 and NCT02008227ReferencesShapley L. A value for n-person games. Contributions to the Theory of Games. 1953;2.28:307–317.Roder J, Maguire L, Georgantas R, Roder H. Explaining multivariate molecular diagnostic tests via Shapley values. BMC Med Inform Decis Mak 2021;21(1):211.Kowanetz M, Leng N, Roder J, et al. Evaluation of immune-related markers in the circulating proteomic and their association with atezolizumab efficacy in patients with 2L+ NSCLC. J Immunother Cancer 2018;6(Suppl1):114.Fehrenbacher L, Spira A, Ballinger M, et al. Atezolizumab versus docetaxel for patients with previously treated non-small-cell lung cancer (POPLAR): a multicentre, open-label, phase 2 randomised controlled trial. Lancet 2016;387(10030):1837–1846.Rittmeyer A, Barlesi F, Waterkamp D, et al. Atezolizumab versus docetaxel in patients with previously treated non-small-cell lung cancer (OAK): a phase 3, open-label, multicenter randomized controlled trial. Lancet 2017;389(10066):255–265.Ethics ApprovalThe OAK study that was done in 194 academic medical centers and community oncology practices across 31 countries worldwide. The study was done in full accordance with the guidelines for Good Clinical Practice and the Declaration of Helsinki. All patients gave written informed consent.The POPLAR trial was done at 61 academic medical centers and community oncology practices across 13 countries in Europe and North America. The study was done in full accordance with the guidelines for Good Clinical Practice and the Declaration of Helsinki. Protocol (and modification) approval was obtained from anindependent ethics committee for each site. Patients gave written informed consent.

The efficacy of atezolizumab (A) and/or bevacizumab (B) with carboplatin/paclitaxel (CP) chemotherapy was explored in the phase III, randomized IMpower150 study in patients with non-squamous non-small cell lung cancer (NSCLC) according toMutation status was determined by circulating tumor DNA next-generation sequencing. Overall survival (OS) and progression-free survival (PFS) were analyzed in a mutation-evaluable intention-to-treat population (MEP; n=920) and SP263 (programmed cell death ligand 1 (PD-L1)) biomarker-evaluable population (n=774).Within the mThese analyses support previous findings of mutation of

Background Novel adjuvant strategies are needed to optimise outcomes after complete surgical resection in patients with early-stage non-small-cell lung cancer (NSCLC). We aimed to evaluate adjuvant atezolizumab versus best supportive care after adjuvant platinum-based chemotherapy in these patients. Methods IMpower010 was a randomised, multicentre, open-label, phase 3 study done at 227 sites in 22 countries and regions. Eligible patients were 18 years or older with completely resected stage IB (tumours ≥4 cm) to IIIA NSCLC per the Union Internationale Contre le Cancer and American Joint Committee on Cancer staging system (7th edition). Patients were randomly assigned (1:1) by a permuted-block method (block size of four) to receive adjuvant atezolizumab (1200 mg every 21 days; for 16 cycles or 1 year) or best supportive care (observation and regular scans for disease recurrence) after adjuvant platinum-based chemotherapy (one to four cycles). The primary endpoint, investigator-assessed disease-free survival, was tested hierarchically first in the stage II-IIIA population subgroup whose tumours expressed PD-L1 on 1% or more of tumour cells (SP263), then all patients in the stage II-IIIA population, and finally the intention-to-treat (ITT) population (stage IB-IIIA). Safety was evaluated in all patients who were randomly assigned and received atezolizumab or best supportive care. IMpower010 is registered with ClinicalTrials.gov, NCT02486718 (active, not recruiting). Findings Between Oct 7, 2015, and Sept 19, 2018, 1280 patients were enrolled after complete resection. 1269 received adjuvant chemotherapy, of whom 1005 patients were eligible for randomisation to atezolizumab (n=507) or best supportive care (n=498); 495 in each group received treatment. After a median follow-up of 32·2 months (IQR 27·4-38·3) in the stage II-IIIA population, atezolizumab treatment improved disease-free survival compared with best supportive care in patients in the stage II-IIIA population whose tumours expressed PD-L1 on 1% or more of tumour cells (HR 0·66; 95% CI 0·50-0·88; p=0·0039) and in all patients in the stage II-IIIA population (0·79; 0·64-0·96; p=0·020). In the ITT population, HR for disease-free survival was 0·81 (0·67-0·99; p=0·040). Atezolizumab-related grade 3 and 4 adverse events occurred in 53 (11%) of 495 patients and grade 5 events in four patients (1%). Interpretation IMpower010 showed a disease-free survival benefit with atezolizumab versus best supportive care after adjuvant chemotherapy in patients with resected stage II-IIIA NSCLC, with pronounced benefit in the subgroup whose tumours expressed PD-L1 on 1% or more of tumour cells, and no new safety signals. Atezolizumab after adjuvant chemotherapy offers a promising treatment option for patients with resected early-stage NSCLC. Funding F Hoffmann-La Roche and Genentech.

Introduction IMpower110 previously revealed significant overall survival (OS) benefit with atezolizumab versus chemotherapy in patients with treatment-naive EGFR- and ALK-negative (wild type [WT]) metastatic NSCLC with high programmed death-ligand 1 (PD-L1) expression (≥50% on tumor cells [TCs] or ≥10% on tumor-infiltrating immune cells [ICs], per SP142 immunohistochemistry assay; p = 0.0106). We present primary OS analyses in lower PD-L1 expression groups and an updated, exploratory analysis in the high PD-L1 expression group. Methods This open-label, phase 3 trial randomized patients with PD-L1 expression on greater than or equal to 1% of TC or IC to receive atezolizumab or platinum-based chemotherapy. The primary end point was OS, hierarchically tested in PD-L1 expression WT subgroups: first the high PD-L1 expression subgroup, then the high-or-intermediate PD-L1 expression subgroup (≥5% on TC or IC), and then the any PD-L1 expression subgroup (≥1% on TC or IC). Results The any PD-L1 expression WT population included 554 patients (excluded 18 EGFR- or ALK-positive patients). With 17 months’ additional follow-up, OS improvement in the atezolizumab versus chemotherapy arm was not statistically significant in high-or-intermediate PD-L1 expression WT patients (n = 328; hazard ratio = 0.87, 95% confidence interval: 0.66–1.14, p = 0.3091; median = 19.9 versus 16.1 mo), precluding formal OS testing in any PD-L1 expression WT patients. Exploratory analysis in high PD-L1 expression WT patients (n = 205) revealed maintained OS benefit in the atezolizumab arm (hazard ratio = 0.76, 95% confidence interval: 0.54–1.09; median = 20.2 versus 14.7 mo). Updated safety data continued to favor atezolizumab. Conclusions Statistical significance for OS was not revealed in the high-or-intermediate expression WT group, and, as a result, OS in the any PD-L1 expression WT group was not formally tested. No new safety signals were found. This updated analysis of IMpower110 supports using atezolizumab in treatment-naive, metastatic WT NSCLC with high PD-L1 expression.

BackgroundCD8+ tissue-resident memory T (TRM) cells, marked by CD103 (ITGAE) expression, are thought to actively suppress cancer progression, leading to the hypothesis that their presence in tumors may predict response to immunotherapy.MethodsHere, we test this by combining high-dimensional single-cell modalities with bulk tumor transcriptomics from 1868 patients enrolled in lung and bladder cancer clinical trials of atezolizumab (anti-programmed cell death ligand 1 (PD-L1)).ResultsITGAE was identified as the most significantly upregulated gene in inflamed tumors. Tumor CD103+ CD8+ TRM cells exhibited a complex phenotype defined by the expression of checkpoint regulators, cytotoxic proteins, and increased clonal expansion.ConclusionsOur analyses indeed demonstrate that the presence of CD103+ CD8+ TRM cells, quantified by tracking intratumoral CD103 expression, can predict treatment outcome, suggesting that patients who respond to PD-1/PD-L1 blockade are those who exhibit an ongoing antitumor T-cell response.

We report the final results of the phase 3 IMpower132 study evaluating atezolizumab plus carboplatin or cisplatin plus pemetrexed (APP) in patients with nonsquamous NSCLC.Chemotherapy-naive patients with stage IV nonsquamous NSCLC without sensitizing EGFR or ALK genetic alterations were randomized in a one-to-one ratio to receive four or six cycles of carboplatin or cisplatin plus pemetrexed (PP) or APP every 3 weeks, followed by maintenance therapy with atezolizumab plus pemetrexed or pemetrexed alone. Co-primary end points were overall survival (OS) and investigator-assessed progression-free survival (PFS).The intention-to-treat population included 578 patients (APP, n = 292; PP, n = 286). At the primary PFS analysis (May 22, 2018; median follow-up, 14.8 mo), APP exhibited significant PFS improvement versus PP (median = 7.6 versus 5.2 mo, stratified hazard ratio [HR] = 0.60, 95% confidence interval [CI]: 0.49-0.72, p0.0001). OS for the APP group was numerically better but not statistically significant at the interim (May 22, 2018; median = 18.1 versus 13.6 mo, stratified HR = 0.81, 95% CI: 0.64-1.03, p = 0.0797) and final analyses (July 18, 2019; median = 17.5 versus 13.6 mo; stratified HR = 0.86, 95% CI: 0.71-1.06, p = 0.1546). The OS and PFS results favored APP versus PP across subgroups. Grade 3 or 4 treatment-related adverse events occurred in 54.6% (APP) and 40.1% (PP) of patients; grade 5 treatment-related events occurred in 3.8% and 2.9%, respectively.IMpower132 met its co-primary PFS end point but not its co-primary OS end point, with numerical improvement for OS in the APP arm. APP had a manageable safety profile, with no new or unexpected safety signals identified.

Immune checkpoint inhibitors targeting the PD-1/PD-L1 axis lead to durable clinical responses in subsets of cancer patients across multiple indications, including non-small cell lung cancer (NSCLC), urothelial carcinoma (UC) and renal cell carcinoma (RCC). Herein, we complement PD-L1 immunohistochemistry (IHC) and tumor mutation burden (TMB) with RNA-seq in 366 patients to identify unifying and indication-specific molecular profiles that can predict response to checkpoint blockade across these tumor types. Multiple machine learning approaches failed to identify a baseline transcriptional signature highly predictive of response across these indications. Signatures described previously for immune checkpoint inhibitors also failed to validate. At the pathway level, significant heterogeneity is observed between indications, in particular within the PD-L1+ tumors. mUC and NSCLC are molecularly aligned, with cell cycle and DNA damage repair genes associated with response in PD-L1- tumors. At the gene level, the CDK4/6 inhibitor CDKN2A is identified as a significant transcriptional correlate of response, highlighting the association of non-immune pathways to the outcome of checkpoint blockade. This cross-indication analysis reveals molecular heterogeneity between mUC, NSCLC and RCC tumors, suggesting that indication-specific molecular approaches should be prioritized to formulate treatment strategies., PD-L1 immune checkpoint inhibition has been used for several tumour types. Here, the authors use immunohistochemistry, tumour mutation burden and RNA-seq data from 366 patients with different indications to identify molecular signatures of response to atezolizumab and reveal pathway heterogeneity and the involvement of non-immune pathways.

Background PD-L1/PD-1 inhibitors (CPI) as monotherapy or in combination with platinum-based doublet chemo (± bevacizumab) are 1L treatment options in metastatic NSCLC, with choice of agent(s) determined by PD-L1 expression. For patients (pts) who may be ineligible for combination therapy, CPI monotherapy remains an attractive treatment choice. IMpower110 evaluated atezo as 1L treatment in PD-L1–selected pts independent of tumor histology. Methods IMpower110 enrolled 572 chemo-naive pts with stage IV nonsquamous (nsq) or squamous (sq) NSCLC, PD-L1 expression ≥ 1% on TC or IC, measurable disease by RECIST 1.1 and ECOG PS 0-1. Pts were randomized 1:1 to receive atezo 1200 mg IV q3w (Arm A) or platinum-based chemo (Arm B; 4 or 6 21-day cycles). Arm B nsq pts received cisplatin (cis) 75 mg/m2 or carboplatin (carbo) AUC 6 + pemetrexed 500 mg/m2 IV q3w; Arm B sq pts received cis 75 mg/m2 + gemcitabine (gem) 1250 mg/m2 or carbo AUC 5 + gem 1000 mg/m2 IV q3w. Stratification factors were sex, ECOG PS, histology and tumor PD-L1 status (TC1/2/3 and any IC vs TC0 and IC1/2/3). The primary endpoint of OS is tested hierarchically in the wild-type (WT; EGFR/ALK-negative) population (TC3 or IC3 then TC2/3 or IC2/3 then TC1/2/3 or IC1/2/3). Results The 3 primary efficacy populations included 554 TC1/2/3 or IC1/2/3 WT pts, 328 TC2/3 or IC2/3 WT pts and 205 TC3 or IC3 WT pts. Median follow-up was 15.7 months (range, 0-35) in TC3 or IC3 WT pts. In the TC3 or IC3 WT population, atezo monotherapy improved median OS by 7.1 months (HR, 0.595; P = 0.0106) compared with chemo (table 1). The safety population comprised 286 pts in Arm A and 263 in Arm B. Treatment-related AEs (TRAEs) and Grade 3-4 TRAEs occurred in 60.5% (Arm A) and 85.2% (Arm B), and 12.9% (Arm A) and 44.1% (Arm B), respectively. Conclusions At this interim analysis, IMpower110 met the primary endpoint of OS with statistically significant and clinically meaningful improvement in the TC3 or IC3 WT population. The safety profile favored Arm A, with no new or unexpected safety signals identified. Trial Registration NCT02409342 Ethics Approval The trial was conducted according to the principles of the Declaration of Helsinki. All patients provided written informed consent. Protocol approval was obtained from independent review boards or ethics committees at each site.

Introduction Cytotoxic agents have immunomodulatory effects, providing a rationale for combining atezolizumab (anti-programmed death-ligand 1 [anti–PD-L1]) with chemotherapy. The randomized phase III IMpower131 study (NCT02367794) evaluated atezolizumab with platinum-based chemotherapy in stage IV squamous NSCLC. Methods A total of 1021 patients were randomized 1:1:1 to receive atezolizumab+carboplatin+paclitaxel (A+CP) (n = 338), atezolizumab+carboplatin+nab-paclitaxel (A+CnP) (n = 343), or carboplatin+nab-paclitaxel (CnP) (n = 340) for four or six 21-day cycles; patients randomized to the A+CP or A+CnP arms received atezolizumab maintenance therapy until progressive disease or loss of clinical benefit. The coprimary end points were investigator-assessed progression-free survival (PFS) and overall survival (OS) in the intention-to-treat (ITT) population. The secondary end points included PFS and OS in PD-L1 subgroups and safety. The primary PFS (January 22, 2018) and final OS (October 3, 2018) for A+CnP versus CnP are reported. Results PFS improvement with A+CnP versus CnP was seen in the ITT population (median, 6.3 versus 5.6 mo; hazard ratio [HR] = 0.71, 95% confidence interval [CI]: 0.60–0.85; p = 0.0001). Median OS in the ITT population was 14.2 and 13.5 months in the A+CnP and CnP arms (HR = 0.88, 95% CI: 0.73–1.05; p = 0.16), not reaching statistical significance. OS improvement with A+CnP versus CnP was observed in the PD-L1–high subgroup (HR = 0.48, 95% CI: 0.29–0.81), despite not being formally tested. Treatment-related grade 3 and 4 adverse events and serious adverse events occurred in 68.0% and 47.9% (A+CnP) and 57.5% and 28.7% (CnP) of patients, respectively. Conclusions Adding atezolizumab to platinum-based chemotherapy significantly improved PFS in patients with first-line squamous NSCLC; OS was similar between the arms.

BackgroundThe efficacy of atezolizumab (A) and/or bevacizumab (B) with carboplatin/paclitaxel (CP) chemotherapy was explored in the phase III, randomized IMpower150 study in patients with non-squamous non-small cell lung cancer (NSCLC) according to KRAS mutations (mKRAS) and co-occurring STK11, KEAP1, or TP53 mutations.MethodsMutation status was determined by circulating tumor DNA next-generation sequencing. Overall survival (OS) and progression-free survival (PFS) were analyzed in a mutation-evaluable intention-to-treat population (MEP; n=920) and SP263 (programmed cell death ligand 1 (PD-L1)) biomarker-evaluable population (n=774).ResultsWithin the mKRAS population (24.5% of MEP), ABCP showed numerical improvements vs BCP in median OS (19.8 vs 9.9 months; HR 0.50; 95% CI 0.34 to 0.72) and PFS (8.1 vs 5.8 months; HR 0.42; 95% CI 0.29 to 0.61)—greater than with ACP (OS: 11.7 vs 9.9 months; HR 0.63; 95% CI 0.43 to 0.91; PFS: 4.8 vs 5.8 months; HR 0.80; 95% CI 0.56 to 1.13) vs BCP. Across PD-L1 subgroups in mKRAS patients, OS and PFS were longer with ABCP vs BCP, but OS with ACP was similar to BCP in PD-L1-low and PD-L1-negative subgroups. Conversely, in KRAS-WT patients, OS was longer with ACP than with ABCP or BCP across PD-L1 subgroups. KRAS was frequently comutated with STK11, KEAP1, and TP53; these subgroups conferred different prognostic outcomes. Within the mKRAS population, STK11 and/or KEAP1 mutations were associated with inferior OS and PFS across treatments compared with STK11-WT and/or KEAP1-WT. In mKRAS patients with co-occurring mSTK11 and/or mKEAP1 (44.9%) or mTP53 (49.3%), survival was longer with ABCP than with ACP or BCP.ConclusionsThese analyses support previous findings of mutation of STK11 and/or KEAP1 as poor prognostic indicators. While clinical efficacy favored ABCP and ACP vs BCP in these mutational subgroups, survival benefits were greater in the mKRAS and KEAP1-WT and STK11-WT population vs mKRAS and mKEAP1 and mSTK11 population, suggesting both prognostic and predictive effects. Overall, these results suggest that atezolizumab combined with bevacizumab and chemotherapy is an efficacious first-line treatment in metastatic NSCLC subgroups with mKRAS and co-occurring STK11 and/or KEAP1 or TP53 mutations and/or high PD-L1 expression.

Introduction: Cancer immunotherapy has revolutionized the treatment of patients with advanced or metastatic non-small cell lung cancer (NSCLC). However, specific patient groups (e.g. patients with activating epidermal growth factor receptor [EGFR] mutations) do not appear to derive benefit from immune checkpoint inhibitor (ICI) monotherapy. Combining ICIs, such as atezolizumab, with chemotherapy and/or targeted therapies may help to address this unmet need.Areas covered: Atezolizumab is an anti-programmed death-ligand 1 therapy for several tumor types. We review its clinical efficacy and safety in the treatment of advanced or metastatic NSCLC, with a specific focus on the combination of atezolizumab with bevacizumab, carboplatin, and paclitaxel (ABCP). Data from IMpower150 show that the ABCP regimen provided clinical benefit to patients with non-squamous NSCLC, including those with EGFR mutations.Expert opinion: Combining ICIs with chemotherapy has proven to be superior to chemotherapy alone. However, tumor resistance to ICIs will likely increase as these drugs enter earlier lines of therapy, underscoring a need for effective treatments when immunotherapy fails. Data suggest that the ABCP regimen may circumvent ICI resistance mechanisms. Continued investigation into the regimen's mechanisms, improved patient profiling/selection, and treatment personalization will drive further development/discoveries.

BackgroundImmunotherapy has become a key element in the arsenal of treatments for advanced non-small cell lung cancer (NSCLC). The anti-PD-L1 Response Test (ART), based on mass spectrometry of pretreatment serum, captures the effect of host biology on outcomes after atezolizumab (A) therapy. It stratified outcomes on A and was predictive of benefit of A over docetaxel (D) in a blinded, retrospective study of 2nd and 3rd line NSCLC patients in the POPLAR Ph2 clinical study.1 Our current work applies the test to the larger OAK NSCLC Ph3 clinical study2 to investigate the interplay between tumor PD-L1 expression and ART classifications in predicting outcomes and benefit from A therapy.MethodsPretreatment serum samples from 823 of the 850 patients in the OAK study (NCT02008227) were analyzed with ART blinded to all clinical data. The ART assigns a result of Good or Poor corresponding to better or worse outcomes on A. Association of test classification with overall survival (OS) within and between treatment arms was investigated using Cox proportion hazards models overall and within PD-L1 subgroups defined by SP142 assay.3ResultsTest classifications were generated for 786 (96%) samples; the remaining samples failed test QC, mainly due to hemolysis. A Good classification was assigned to 359 (46%) samples and a Poor classification to 427 (54%) samples. Overall, OS was better for the Good subgroup than the Poor subgroup within both arms, arm A (hazard ratio (HR)=0.52 (95% Confidence Interval (CI): 0.41–0.66)) and arm D (HR=0.54 (CI:0.43–0.68)). The test was not predictive of benefit of A over D, but was prognostic for both A and D. Patients classified as Good had better outcomes than those classified as Poor in both treatment arms for all PD-L1 subgroups investigated (figure 1). Benefit of A vs D was found in both test classification groups for PD-L1 positive patients (table 1).Abstract 28 Figure 1Kaplan-Meier plots of OS by test classification, Good and Poor, and treatment arm, A and D, within PD-L1 subgroupsAbstract 28 Table 1Hazard ratios between A and D by test classification group and PD-L1 subgroupConclusionsInformation on both tumor and host are essential to predict outcomes of immunotherapy and chemotherapy in NSCLC patients.Trial RegistrationClinicalTrials.gov NCT02008227ReferencesKowanetz M, Leng N, Roder J, et al. Evaluation of immune-related markers in the circulating proteomic and their association with atezolizumab efficacy in patients with 2L+ NSCLC. J Immunother Cancer 2018;6(Suppl1):114.Rittmeyer A, Barlesi F, Waterkamp D, et al. Atezolizumab versus docetaxel in patients with previously treated non-small-cell lung cancer (OAK): a phase 3, open-label, multicenter randomized controlled trial. Lancet 2017;389(10066):255–265.Herbst R, Giaccone G, de Marinis F, et al. Atezolizumab for first-line treatment of PD-L1-selected patients with NSCLC. N Engl J Med 2020;383:1328–1339.Ethics ApprovalThe OAK study (NCT02008227) was done in 194 academic medical centres and community oncology practices across 31 countries worldwide. The study was done in full accordance with the guidelines for Good Clinical Practice and the Declaration of Helsinki. All patients gave written informed consent.

The development of selective ubiquitin-specific protease-7 (USP7) inhibitors GNE-6640 and GNE-6776, which induce tumour cell death and reveal differential kinetics of Lys-48 and Lys-63-linked ubiquitin chain depolymerization by USP7. Deubiquitinating enzymes remove the small modifier protein ubiquitin from target substrates regulating their stability. One such enzyme, USP7, is a potential target for anti-cancer therapy, as its inhibition would result in the degradation of the ubiquitinated oncoprotein MDM2, leading to reactivation of the tumour suppressor protein p53. However, selective inhibitors of USP7 have remained elusive. Here, Ingrid Wertz and team develop two USP7 inhibitors, providing structural insights into the mode of action of these compounds and demonstrating their toxicity towards tumour cells. Elsewhere in this issue, David Komander and colleagues independently report the identification of two small molecules that inhibit USP7 with high affinity and specificity both in vitro and within cells, also demonstrating their ability to inhibit tumour growth. The ubiquitin system regulates essential cellular processes in eukaryotes. Ubiquitin is ligated to substrate proteins as monomers or chains and the topology of ubiquitin modifications regulates substrate interactions with specific proteins. Thus ubiquitination directs a variety of substrate fates including proteasomal degradation1. Deubiquitinase enzymes cleave ubiquitin from substrates and are implicated in disease2; for example, ubiquitin-specific protease-7 (USP7) regulates stability of the p53 tumour suppressor and other proteins critical for tumour cell survival3. However, developing selective deubiquitinase inhibitors has been challenging4 and no co-crystal structures have been solved with small-molecule inhibitors. Here, using nuclear magnetic resonance-based screening and structure-based design, we describe the development of selective USP7 inhibitors GNE-6640 and GNE-6776. These compounds induce tumour cell death and enhance cytotoxicity with chemotherapeutic agents and targeted compounds, including PIM kinase inhibitors. Structural studies reveal that GNE-6640 and GNE-6776 non-covalently target USP7 12 A distant from the catalytic cysteine. The compounds attenuate ubiquitin binding and thus inhibit USP7 deubiquitinase activity. GNE-6640 and GNE-6776 interact with acidic residues that mediate hydrogen-bond interactions with the ubiquitin Lys48 side chain5, suggesting that USP7 preferentially interacts with and cleaves ubiquitin moieties that have free Lys48 side chains. We investigated this idea by engineering di-ubiquitin chains containing differential proximal and distal isotopic labels and measuring USP7 binding by nuclear magnetic resonance. This preferential binding protracted the depolymerization kinetics of Lys48-linked ubiquitin chains relative to Lys63-linked chains. In summary, engineering compounds that inhibit USP7 activity by attenuating ubiquitin binding suggests opportunities for developing other deubiquitinase inhibitors and may be a strategy more broadly applicable to inhibiting proteins that require ubiquitin binding for full functional activity.

Cancer immunotherapy (CIT) has transformed our approach in diagnosis and treatment of cancer. However, durable responses or cures are only seen in a minority of patients, illustrating the need for reliable biomarkers that identify patients most likely to receive meaningful clinical benefit. PD-L1 immunohistochemistry (IHC) has been extensively used in clinical development programs for anti-PD-L1/PD-1 targeted therapies. Notably, four independently developed PD-L1 IHC assays have demonstrated clinically meaningful predictive value in several indications and are approved as companion or complementary diagnostics. PD-L1 IHC is by no means a flawless biomarker or diagnostic. Numerous studies have found that a subset of PD-L1 negative patients do in fact derive clinical benefit from CIT therapy, highlighting the need for more precise diagnostic tools. Gene signatures with emphasis on immune-related biology and tumor mutation burden, a surrogate for neoantigen presentation, have both emerged as new promising CIT biomarkers and have demonstrated predictive value in exploratory clinical studies. As of today, neither of these biomarkers has gained approval as a companion or complementary diagnostic or has shown the capacity to accurately capture all patients that could potentially benefit from CIT. It is likely, based on the complexity of the tumor microenvironment, that more than one biomarker will be required to identify patients that benefit from CIT in the future.

8500 Background: Adjuvant platinum-based chemotherapy (chemo) provides only a modest 5-year survival benefit in fully resected, high-risk early-stage NSCLC. We report the primary disease-free survival (DFS) results from the pre-planned interim analysis of IMpower010, a randomized phase 3 open-label trial of adjuvant atezolizumab (atezo; anti–PD-L1) vs best supportive care (BSC) after adjuvant chemo in patients (pts) with early-stage resected NSCLC. Methods: Eligible pts had completely resected (4-12 weeks prior to enrollment) Stage IB (≥4 cm)-IIIA NSCLC (AJCC/UICC v7) and ECOG PS 0-1. A total of 1280 pts were enrolled, and 1269 pts received up to four 21-day cycles of cisplatin-based chemo (plus pemetrexed, docetaxel, gemcitabine or vinorelbine). Of these pts (n=1269), 1005 were subsequently randomized 1:1 to 16 cycles of atezo 1200 mg Q3W or BSC. The primary endpoint of investigator-assessed DFS and secondary endpoint of overall survival (OS) were tested hierarchically: first DFS in the PD-L1 TC ≥1% (SP263) subgroup with Stage II-IIIA disease, then DFS in all randomized pts with Stage II-IIIA disease, then DFS in the ITT population (Stage IB-IIIA) and finally OS in the ITT population. Efficacy assessments were based on randomized pts. Safety was assessed in the safety-evaluable population, defined as pts who received ≥1 dose of atezo or who had ≥1 post-baseline safety assessment if randomized to the BSC arm. Results: At data cutoff (January 21, 2021), median follow-up was 32.2 months in the ITT population. Baseline characteristics were generally balanced between arms. Atezo showed statistically significant DFS benefit vs BSC in the PD-L1 TC ≥1% Stage II-IIIA and all randomized Stage II-IIIA populations; the significance boundary was not crossed for DFS in the ITT population (Table). OS data were immature and not formally tested. Pts in the atezo arm received a median of 16 (range, 1-16) atezo doses. Any-grade AEs occurred in 92.7% (atezo) and 70.7% (BSC); events were Grade 3/4 in 21.8% and 11.5%, respectively. Grade 5 treatment-related AEs occurred in 0.8% of pts in the atezo arm. AEs leading to atezo discontinuation occurred in 18.2% of atezo-treated pts. Conclusions: IMpower010 met its primary endpoint, showing DFS benefit with adjuvant atezo vs BSC after adjuvant chemo in pts with resected Stage II-IIIA NSCLC, with pronounced benefit in the PD-L1 TC ≥1% subgroup. The safety profile of atezo was consistent with prior experience of atezo monotherapy across indications and lines of therapy. Funding: F. Hoffmann-La Roche Ltd. Clinical trial information: NCT02486718. [Table: see text]

Beginning with promiscuous COT inhibitors, which were found to inhibit CDK8, a series of 6-aza-benzothiophene containing compounds were developed into potent, selective CDK8 inhibitors. When cocrystallized with CDK8 and cyclin C, these compounds exhibit an unusual binding mode, making a single hydrogen bond to the hinge residue A100, a second to K252, and a key cation-π interaction with R356. Structure-based drug design resulted in tool compounds 13 and 32, which are highly potent, kinase selective, permeable compounds with a free fraction >2% and no measurable efflux. Despite these attractive properties, these compounds exhibit weak antiproliferative activity in the HCT-116 colon cancer cell line. Further examination of the activity of 32 in this cell line revealed that the compound reduced phosphorylation of the known CDK8 substrate STAT1 in a manner identical to a CDK8 knockout clone, illustrating the complex effects of inhibition of CDK8 kinase activity in proliferation in these cells.

Colon tumors arise in a stepwise fashion from either discrete genetic perturbations or epigenetic dysregulation. To uncover the key epigenetic regulators that drive colon cancer growth, we used a CRISPR loss-of-function screen and identified a number of essential genes, including the bromodomain and extraterminal (BET) protein BRD4. We found that BRD4 is critical for colon cancer proliferation, and its knockdown led to differentiation effects in vivo. JQ1, a BET inhibitor, preferentially reduced growth in a subset of epigenetically dysregulated colon cancers characterized by the CpG island methylator phenotype (CIMP). Integrated transcriptomic and genomic analyses defined a distinct superenhancer in CIMP + colon cancers that regulates cMYC transcription. We found that the long noncoding RNA colon cancer-associated transcript 1 (CCAT1) is transcribed from this superenhancer and is exquisitely sensitive to BET inhibition. Concordantly, cMYC transcription and cell growth were tightly correlated with the presence of CCAT1 RNA in a variety of tumor types. Taken together, we propose that CCAT1 is a clinically tractable biomarker for identifying patients who are likely to benefit from BET inhibitors.

Background: IMpower133 (NCT02763579), a global Phase I/III, randomized, double-blind, placebo (PBO)-controlled trial, showed that the addition of atezo (anti-PD-L1) to CE for 1L ES-SCLC led to statistically and clinically significant OS and PFS improvement vs CE alone. Here we report updated OS and exploratory analyses. Methods: Pts with untreated ES-SCLC were randomized 1:1 to receive four 21-day cycles of E (100 mg/m2 IV, days 1-3) + C (AUC 5 mg/mL/min IV, day 1) with atezo (1200 mg IV, day 1) or PBO, followed by maintenance therapy with atezo or PBO until intolerable toxicity, progression or loss of clinical benefit. PD-L1 testing was not required for enrollment. Coprimary endpoints were investigator-assessed PFS (RECIST 1.1) and OS. Interim and final OS analyses were planned for ≈240 and ≈306 events, respectively. OS was significant at the interim analysis. Updated OS, exploratory biomarkers and patterns of disease progression were analyzed. Results: 201 and 202 pts were randomized to receive atezo+CE and PBO+CE, respectively. The median follow-up was 22.9 mo and 302 deaths had occurred. Median OS for the atezo and PBO arms was 12.3 and 10.3 mo, respectively (HR, 0.76 [95% CI: 0.60, 0.95]; descriptive P = 0.0154). At the 18-mo landmark, the OS rate was 13% higher with atezo+CE than with PBO+CE (Table). Exploratory analyses showed treatment benefit with atezo+CE regardless of biomarker status. 181 (90.0%) And 194 (96.0%) pts in the atezo+CE and PBO+CE arms, respectively, had RECIST-defined disease progression. Progression at existing, new or existing and new lesions was numerically lower with atezo+CE than with PBO+CE. Common sites of new lesions included the CNS, lung, lymph node and liver, with similar incidences between arms. Conclusion: Adding atezo to CE continued to provide OS improvement for 1L ES-SCLC in an all-comer population. The updated results of IMpower133 further support this regimen for untreated ES-SCLC. Landmark OSAtezo + CE, n = 201PBO + CE, n = 20212 Mo, n (%)93 (51.9)74 (39.0)18 Mo, n (%)61 (34.0)39(21.0)Median OS in biomarker subgroupsAtezo + CEPBO + CEITT (N = 403), mo12.310.3HR (95% CI)0.76 (0.61, 0.96)aITT-BEP (n = 137), mo9.98.9HR (95% CI)0.70 (0.48, 1.02)Non-BEP (n = 266), mo14.611.2HR (95% CI)0.81 (0.61, 1.08)PD-L1 expression, 1% TC or IC< 1% (n = 65), mo10.28.3HR (95% CI)0.51 (0.30, 0.89)≥ 1% (n = 72), mo9.710.6HR (95% CI)0.87 (0.51, 1.49)PD-L1 expression, 5% TC or IC< 5% (n = 108), mo9.28.9HR (95% CI)0.77 (0.51, 1.17)≥ 5% (n = 29), mo21.69.2HR (95% CI)0.60 (0.25, 1.46)Disease progression at sites, n (%)Atezo + CEPBO + CEExisting116 (57.7)131 (64.9)New86 (42.8)99 (49.0)Existing and new42 (20.9)57 (28.2)BEP, biomarker-evaluable population; ITT, intention-to-treat population.a Stratified HR. BEP included pts evaluable by PD-L1 IHC using the VENTANA SP263 assay. Citation Format: Leora Horn, Stephen V. Liu, Aaron S. Mansfield, Tony Mok, Arnaud Scherpereel, Niels Reinmuth, Marina Chiara Garassino, Javier De Castro Carpeno, Raffaele Califano, Makoto Nishio, Francisco Orlandi, Jorge Arturo Alatorre Alexander, Ticiana Leal, Ying Cheng, Jong-Seok Lee, Sivuonthanh Lam, Mark McCleland, Yu Deng, See Phan, Martin Reck. IMpower133: Updated OS and exploratory analyses of first-line (1L) atezolizumab (atezo) + carboplatin (C) + etoposide (E) in extensive-stage SCLC (ES-SCLC) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr CT220.

3130 Background: IMpower150 is a phase 3 study measuring the effect of carboplatin and paclitaxel (CP) combined with atezolizumab (A) and/or bevacizumab (B) in patients with advanced nonsquamous NSCLC, testing the hypothesis that anti-PD-L1 therapy may be enhanced by the blockade of VEGF. Here, we apply a machine-learning based approach to quantify the tumor micro-environment (TME) and vasculature and identify associations with clinical outcome in IMpower150. Methods: Digitized H&E images were registered onto the PathAI research platform (n=1027). Over 200K annotations from 90 pathologists were used to train convolutional neural networks (CNNs) that classify human-interpretable features (HIFs) of cells and tissue structures from images. Blood vessel compression (BVC) indices were calculated using the long versus short axes for each predicted blood vessel. HIFs were clustered to reduce redundancy, and selected features were associated with progression free survival (PFS) within each arm (ABCP, ACP, and BCP) using Cox proportional hazard models. Results: We used the trained CNNs to generate 4,534 features summarizing each patient’s histopathology and TME. After association with survival and correction for multiple comparisons we identified clusters that were significantly associated with survival in at least one arm. Among patients receiving treatments that target PD-L1 (ABCP and ACP), high lymphocyte to fibroblast ratio (LFR) was associated with improved PFS (HR=0.64 (0.51, 0.81), p < 0.001) and showed no significant association with PFS among patients treated with BCP alone (HR=1.13 (0.85, 1.51), p=0.4). Among BCP treated patients, a higher average BVC within the tumor tissue was associated with improved PFS (HR=0.67 (0.50,0.90), p=0.01) and worse PFS among patients treated with ACP (HR=1.50 (1.10,2.06), p=0.009). Conclusions: We developed a deep learning-based assay for quantifying pathology features of the TME and vasculature from H&E images. Application of this system to Impower150 identified an association between high LFR and improved PFS among patients receiving PD-L1 targeting therapy, and between low BVC and improved PFS among patients receiving BCP. These findings support the importance of the TME and vasculature in determining response to PD-L1 and VEGF-targeting therapies.

CDK8 is a dissociable kinase module of the Mediator complex and has been shown to play an important role in transcriptional regulation in organisms as diverse as yeast and humans. Recent studies suggest that CDK8 functions as an oncoprotein in melanoma and colon cancer. Importantly, these studies were conducted using in vitro cell line models and the role of CDK8 in tumourigenesis in vivo has not been explored. We have generated a mouse with a Cdk8 conditional knockout allele and examined the consequences of Cdk8 loss on normal tissue homeostasis and tumour development in vivo. Cdk8 deletion in the young adult mouse did not induce any gross or histopathological abnormalities, implying that Cdk8 is largely dispensable for somatic cellular homeostasis. In contrast, Cdk8 deletion in the Apc(Min) intestinal tumour model shortened the animals' survival and increased tumour burden. Although Cdk8 deletion did not affect tumour initiation, intestinal tumour size and growth rate were significantly increased in Cdk8-null animals. Transcriptome analysis performed on Cdk8-null intestinal cells revealed up-regulation of genes that are governed by the Polycomb group (PcG) complex. In support of these findings, Cdk8-null intestinal cells and tumours displayed a reduction in histone H3K27 trimethylation, both globally and at the promoters of a number of PcG-regulated genes involved in oncogenic signalling. Together, our findings uncover a tumour suppressor function for CDK8 in vivo and suggest that the role of CDK8 activity in driving oncogenesis is context-specific. Sequencing data were deposited at GEO (Accession No. GSE71385).

Purpose: This study is aimed to identify genes within the KRAS genomic amplicon that are both coupregulated and essential for cell proliferation when KRAS is amplified in lung cancer. Experimental Design: We used an integrated genomic approach to identify genes that are coamplified with KRAS in lung adenocarcinomas and subsequently preformed an RNA interference (RNAi) screen to uncover functionally relevant genes. The role of lactate dehydrogenase B (LDHB) was subsequently investigated both in vitro and in vivo by siRNA and short hairpin RNA (shRNA)–mediated knockdown in a panel of lung adenocarcinoma cells lines. LDHB expression was also investigated in patient tumors using microarray and immunohistochemistry analyses. Results: RNAi-mediated depletion of LDHB abrogated cell proliferation both in vitro and in xenografted tumors in vivo. We find that LDHB expression correlates to both KRAS genomic copy number gain and KRAS mutation in lung cancer cell lines and adenocarcinomas. This correlation between LDHB expression and KRAS status is specific for lung cancers and not other tumor types that harbor KRAS mutations. Consistent with a role for LDHB in glycolysis and tumor metabolism, KRAS-mutant lung tumors exhibit elevated expression of a glycolysis gene signature and are more dependent on glycolysis for proliferation compared with KRAS wild-type lung tumors. Finally, high LDHB expression was a significant predictor of shorter survival in patients with lung adenocarcinomas. Conclusion: This study identifies LDHB as a regulator of cell proliferation in a subset of lung adenocarcinoma and may provide a novel therapeutic approach for treating lung cancer. Clin Cancer Res; 19(4); 773–84. ©2012 AACR.

Using Sorafenib as a starting point, a series of potent and selective inhibitors of CDK8 was developed. When cocrystallized with CDK8 and cyclin C, these compounds exhibit a Type-II (DMG-out) binding mode.

CDK8 is a cyclin-dependent kinase that mediates transcriptional control of pathways linked to both cancer and stem cells. In this study, we show that CDK8 is required for both tumor growth and maintenance of tumor dedifferentiation in vivo and uncover a common role for CDK8 in controlling cancer and stem cell function. Acute CDK8 loss in vivo strongly inhibited tumor growth and promoted differentiation. Transcriptional profiling identified a set of embryonic stem cell–related genes that are activated by CDK8 in cancer. Consistent with this, we found that CDK8 expression correlated to the embryonic stem cell pluripotency state and loss of CDK8 caused embryonic stem cells to differentiate. This effect was, at least partially, mediated by the ability of CDK8 to regulate MYC protein and downstream MYC target gene expression. Similar regulation of MYC target genes by CDK8 was observed in colon tumor cells, and increased expression of a CDK8-regulated, embryonic stem cell MYC target gene signature was associated with loss of differentiation and poor outcome in primary human colon cancers. Together, these observations reveal that CDK8 acts, at least in part, through MYC to maintain both tumors and embryonic stem cells in an undifferentiated state. This raises the intriguing possibility that targeting CDK8 therapeutically may specifically inhibit the stem-like properties of cancer cells. Cancer Res; 72(8); 2129–39. ©2012 AACR.

Introduction: Atezo (anti-PD-L1) + bev + chemo prolonged PFS vs bev + chemo in patients (pts) with 1L non-squamous mNSCLC in the randomized Ph 3 IMpower150 study regardless of PD-L1 expression. This analysis aims to further understand the efficacy of atezo + bev + chemo in key subgroups: PD-L1 expression subgroups defined by the SP142 and SP263 IHC assays, patients with EGFR/ALK genetic alterations and patients with liver metastases at baseline. Methods: Pts received atezo 1200 mg + bev 15 mg/kg + carboplatin (C) AUC 6 + paclitaxel (P) 200 mg/m2 (Arm B) or bev + C + P (Arm C) IV q3w. A co-primary endpoint (EP) was PFS in the ITT-WT (EGFR or ALK wild-type pts); a secondary EP was PFS in subgroups defined by PD-L1 expression on tumor cells (TC) and tumor-infiltrating immune cells (IC) with SP142. Retrospective analysis with SP263 was a prespecified exploratory EP. Results: In Arms B and C of the ITT-WT (n = 692), 503 pts had available tumor sections for SP263 testing (BEP). Pt characteristics in the ITT-WT and BEP were similar. A similar PFS benefit was observed for Arm B vs C across all PD-L1 expression subgroups defined by either assay, including pts with PD-L1-negative and PD-L1-low tumors (Table). PFS benefit with Arm B vs C was also observed in patients with EGFR or ALK genomic alterations, including patients with actionable EGFR mutations, and in patients with liver metastases at baseline (Table). Conclusion: PFS benefit with atezo + bev + chemo was observed across all PD-L1 subgroups, regardless of the IHC assay used. Additionally, clinically meaningful PFS benefit was observed in patients with EGFR/ALK genomic alterations, and in patients with liver metastases with this combination. Table. PFS in Biomarker Subgroups and Other Subgroups of Interest in IMpower150Median PFS, moPFSan (%)HRb (95% CI)P ValueArm BArm CITT-WT692 (100%)0.62 (0.52, 0.74) Citation Format: Marcin Kowanetz, Mark A. Socinski, Wei Zou, Mark McCleland, Nancy Yang, Ariel Lopez Chavez, Alexander Spira, Julien Mazières, Fadi Braiteh, David Shames, Alan Sandler, Martin Reck. IMpower150: Efficacy of atezolizumab (atezo) plus bevacizumab (bev) and chemotherapy (chemo) in 1L metastatic nonsquamous NSCLC (mNSCLC) across key subgroups [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT076.