Blood-based Tumor Mutational Burden Research Articles

Predicting therapeutic response to neoadjuvant immunotherapy based on an integration model in resectable stage IIIA (N2) non-small cell lung cancer

ObjectiveAccurately predicting response during neoadjuvant chemoimmunotherapy for resectable non-small cell lung cancer (NSCLC) remains clinically challenging. In this study, we investigate the effectiveness of blood-based tumor mutational burden (bTMB) and a deep learning (DL) model in predicting major pathologic response (MPR) and survival from a phase II trial. MethodsBlood samples were prospectively collected from 45 stage IIIA (N2) NSCLC patients undergoing neoadjuvant chemoimmunotherapy. An integrated model, combining the CT-based DL score, bTMB, and clinical factors, was developed to predict tumor response to neoadjuvant chemoimmunotherapy. ResultsAt baseline, bTMB were detected in 77.8% (35 of 45) of patients. Baseline bTMB ≥11 Muts/Mb was associated with significantly higher MPR rates (77.8% vs. 38.5%, p = 0.042), and longer disease-free survival (DFS, p = 0.043), but not overall survival (p = 0.131), compared to bTMB < 11 Muts/Mb in 35 patients with bTMB available. The developed DL model achieved an area under the curve (AUC) of 0.703 in all patients. Importantly, the predictive performance of the integrated model improved to an AUC of 0.820 when combining the DL score with bTMB and clinical factors. Baseline circulating tumor DNA (ctDNA) status was not associated with pathological response and survival. Compared to ctDNA residual, ctDNA clearance before surgery was associated with significantly higher MPR rates (88.2% vs. 11.1%, p < 0.001) and improved DFS (p = 0.010). ConclusionsThe integrated model shows promise as a predictor of tumor response to neoadjuvant chemoimmunotherapy. Serial ctDNA dynamics provide a reliable tool for monitoring tumor response.

Abstract 6132: Exploring genomic complexity in lung cancer at the Tijuana/California border: A real-world perspective through liquid biopsy comprehensive genomic profiling

Abstract Objective: This study explores the application of liquid biopsy with next-generation sequencing (NGS) in profiling the genomic landscape of lung cancer patients from the Tijuana/California Border region. Specifically, we employ blood-based comprehensive genomic profiling (CGP) to assess the complexity of the genomic profile in these patients. Materials and Methods: From April 2019 to November 2023, 37 lung cancer patients from Tijuana underwent profiling using the FoundationOne®Liquid CDx (F1LCDx®) assay, focusing on circulating tumor DNA (ctDNA). The assay, which spans 324 cancer-related genes and genomic signatures, provides insights into tumor fraction (TF) and blood-based tumor mutational burden (bTMB). Variants were classified based on the ESMO Scale of Clinical Actionability for molecular Targets (ESCAT). Results: A notable 88.6% of patients exhibited at least one detectable alteration in plasma. The most frequently mutated genes were TP53 (51.3%), EGFR (16.2%), DNMT3A (13.5%), and KRAS (10.8%). Tumor mutational burden (TMB) was assessable in 9 out of 37 patients, with values ranging from 0 to 10 mut/Mb. According to ESCAT classification, 63.1% of samples revealed potentially actionable alterations (Tier I-II), while an additional 24.1% harbored alterations for which approved drugs are available in other cancer types (Tier III). Conclusion: Liquid biopsy NGS emerges as a viable and valuable strategy for guiding personalized therapy. Given restricted accessibility to new drugs or clinical trials in Mexico, the use of blood-based CGP may enhance the identification of druggable alterations, thereby increasing the likelihood of accessing targeted drugs or enrolling in clinical trials. Citation Format: Jorge Alberto Guadarrama-Orozco, Jennifer Ramirez-Puente, Elizabeth Dominguez-Gonzalez. Exploring genomic complexity in lung cancer at the Tijuana/California border: A real-world perspective through liquid biopsy comprehensive genomic profiling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6132.

Predictive role of ctDNA in esophageal squamous cell carcinoma receiving definitive chemoradiotherapy combined with toripalimab

The combination of toripalimab (an anti-PD-1 antibody) with definitive chemoradiotherapy (CRT) demonstrated encouraging efficacy against locally advanced esophageal squamous cell carcinoma (ESCC) in the EC-CRT-001 phase II trial (NCT04005170). The primary endpoint of this trial was the clinical complete response rate (cCR), and the secondary endpoints included overall survival (OS), progression-free survival (PFS), duration of response, and quality of life. The exploratory analyses of EC-CRT-001 include exploring the role of circulating tumor DNA (ctDNA) and blood-based tumor mutational burden (bTMB) in predicting the response and survival. In total, 118 blood and 35 tissue samples from 42 enrolled patients were included in the analyses. We found that ctDNA-negative patients achieved a higher cCR compared to those with detectable ctDNA during CRT (83%, 19/23 vs. 39%, 7/18; p = 0.008) or post-CRT (78%, 21/27 vs. 30%, 3/10; p = 0.017). Patients with detectable ctDNA during CRT had shorter PFS (p = 0.014). Similarly, patients with post-CRT detectable ctDNA had a significantly shorter PFS (p = 0.012) and worse OS (p = 0.004). Moreover, patients with high bTMB levels during CRT had prolonged OS (p = 0.027). In conclusion, ctDNA and bTMB have the potential to predict treatment efficacy and survival in ESCC treated with CRT and immunotherapy.

Predictive biomarkers of response and survival following immunotherapy with a PD-L1 inhibitor benmelstobart (TQB2450) and antiangiogenic therapy with a VEGFR inhibitor anlotinib for pretreated advanced triple negative breast cancer

In our phase Ib trial (ClinialTrials.gov Identifier: NCT03855358), benmelstobart (TQB2450), a novel humanized IgG1 antibody against PD-L1, plus antiangiogenic multikinase inhibitor, anlotinib, demonstrated promising antitumor activities in pretreated triple negative breast cancer (TNBC) patients. We conducted explorative analyses of genomic biomarkers to explore the associations with treatment response and survival outcomes. Targeted next generation sequencing (NGS) was undertaken toward circulating tumor DNA (ctDNA) collected from peripheral blood samples prior to the start of treatment and after disease progression. A total of 31 patients received targeted NGS and functional driver mutations in 29 patients were analyzed. The most frequent mutations were TP53 (72%), MLL3 (28%), and PIK3CA (17%). At a blood-based tumor mutational burden (bTMB) cutoff of 6.7 mutations per megabase, patients with low bTMB showed better response to anlotinib plus TQB2450 (50% vs. 7%, P = 0.015) and gained greater PFS benefits (7.3 vs. 4.1 months, P = 0.012) than those with high bTMB. At a maximum somatic allele frequency (MSAF) cutoff of 10%, a low MSAF indicated a better objective response (43% vs. 20%) as well as a significantly longer median PFS (7.9 vs. 2.7 months, P < 0.001). Patients with both low MSAF and low bTMB showed a notably better objective response to anlotinib plus TQB2450 (70% vs. 11%, P < 0.001) and a significantly longer median PFS (11.0 vs. 2.9 months, P < 0.001) than patients with other scenarios. Our findings support future studes and validation of MSAF and the combined bTMB-MSAF classification as predictive biomarkers of immune checkpoint inhibitor-based regimens in advanced TNBC patients.

Improved Prediction of Chemoradiation and Immune Checkpoint Blockade Efficacy with Dynamic bTMB Combined with ctDNA in Unresectable Locally Advanced NSCLC

Consolidation durvalumab after definitive chemoradiation therapy (CRT) has become the new standard of care for patients with unresectable locally advanced non-small-cell lung cancer (LA-NSCLC). However, only a fraction of patients can benefit from it, and current decision-making procedures have limited accuracy. Blood-based tumor mutational burden (bTMB) is a promising biomarker, but whether bTMB alone or combined with circulating tumor DNA (ctDNA) can predict the efficacy of definitive CRT and immune checkpoint inhibitors (ICIs) in patients with LA-NSCLC remains unclear. This cohort study enrolled patients diagnosed with unresectable LA-NSCLC from 2018 to 2022. Patients were assigned to the cohort A receiving definitive CRT alone (intensity modulated radiation therapy or volumetric modulated arc therapy with the prescribed dose of 60 Gy, concurrently or sequentially with two or more cycles of platinum-based doublet chemotherapy), or cohort B undergoing definitive CRT and immunotherapy. Peripheral blood specimens were collected before and after CRT and subjected to next-generation sequencing panel to analyze ctDNA and bTMB. The dynamic change in bTMB (∆bTMB) was calculated as the bTMB level after CRT minus the baseline bTMB level. Potential correlations were identified by Spearman's correlation tests and expressed as R coefficients. Time-dependent receiver operating characteristic curves and areas under the curve (AUCs) were employed to evaluate the predictive power of different models on survival. A total of 73 LA-NSCLC patients were included, with 70 (95.9%) at stage III and 3 at stage II (4.1%). Thirty-six patients (49.3%) assigned to the cohort A were treated with CRT alone and 37 (50.7%) in the cohort B receiving CRT and ICIs. Patients with CRT + ICIs in the cohort B showed significantly improved overall survival (OS; P < 0.01) and progression-free survival (PFS; P = 0.02) than those with CRT alone. Baseline bTMB at cutoff values of 4 to 22 did not predict outcomes (all P > 0.10), while patients with increased bTMB (∆bTMB > 0) after CRT had significantly worse OS and PFS (both P < 0.01) than those with decreased or stable bTMB (∆bTMB ≤ 0). ∆bTMB was independent of the ctDNA level after CRT (P = 0.76, R = -0.04). Patients were further divided into 3 groups based on ∆bTMB and post-CRT ctDNA (∆bTMB > 0/detectable ctDNA vs. ∆bTMB ≤ 0/detectable ctDNA vs. ∆bTMB ≤ 0/undetectable ctDNA), and significant survival differences were observed in the pairwise comparisons of 3 groups (all P < 0.05). The model of ∆bTMB combined with post-CRT ctDNA status exhibited the optimal predictive power on both OS (AUC = 0.80) and PFS (AUC = 0.75) and outperformed each factor, which had been respectively validated in the cohort A and B as well. Dynamic bTMB (∆bTMB) combined with post-CRT ctDNA status is a novel and effective biomarker model of predicting survival outcomes in LA-NSCLC patients treated with CRT ± ICIs, and outperforms each individual feature.

Liquid biopsy comprehensive genomic profiling of lung cancer in the Italian population: A real-world experience

ObjectivesLiquid biopsy with next-generation sequencing (NGS) has emerged as a promising tool for tumor mutation profiling. In this study, we describe the genomic profile of Italian lung cancer patients tested with blood-based comprehensive genomic profiling (CGP) to assess the genomic landscape complexity and its impact on enhancing treatment options for patients. Materials and methodsBetween January 2021 and December 2021, a total of 229 lung cancer patients were profiled by FoundationOne®Liquid CDx (F1LCDx®) assay on circulating tumor DNA (ctDNA). F1LCDx® reports alterations across 324 cancer-related genes and genomic signatures, including tumor fraction (TF) and blood-based tumor mutational burden (bTMB). Detected variants were classified according to the ESMO Scale of Clinical Actionability for molecular Targets (ESCAT). Results90.4% of patients had at least one detectable alteration in plasma. The most frequently mutated genes were TP53 (47.6%), DNMT3A (33.2%), EGFR (20.1%), and KRAS (15.7%). Elevated TF was detected in 18.3% of patients, suggesting high reliability of test results. According to the ESCAT classification, potentially actionable alterations (Tier I-II) were identified in 27.1% of samples. An additional 5.2% harbored an alteration for which an approved drug is available in other cancer types (Tier III). Furthermore, 13.1% of tumors exhibited high bTMB, which may predict response to immunotherapy. Overall, 156 (68.1%) patients were eligible for enrolment in clinical trials. ConclusionLiquid biopsy NGS is a viable and valuable approach to guide personalized therapy. The use of blood-based CGP may help identify a larger number of actionable mutations and increase chances of enrolment in clinical trials.

Predicting role of circulating tumor DNA and blood-based tumor mutational burden in esophageal squamous cell carcinoma receiving chemoradiotherapy combined with toripalimab: Exploratory analyses from a phase II trial (EC-CRT-001).

4056 Background: EC-CRT-001 is a phase II trial investigating the efficacy of toripalimab (an anti-PD1 antibody) combined with definitive chemoradiotherapy (CRT) in locally advanced esophageal squamous cell carcinoma (ESCC) and the results demonstrated encouraging antitumor activity and acceptable toxicity. This exploratory analysis was performed to evaluate the role of circulating tumor DNA (ctDNA) and blood-based tumor mutational burden (bTMB) for efficacy prediction. Methods: Forty-two patients from the phase II trial (NCT04005170) who received toripalimab (240 mg every 3 weeks for up to 1 year, or disease progression, or unacceptable toxicity) combined with definitive CRT (irradiation with 50.4 Gy in 28 fractions concurrent with 5 cycles of weekly paclitaxel in 50 mg/m2 and cisplatin in 25 mg/m2) were enrolled. Plasma samples were collected before, during, and after CRT. Targeted next-generation sequencing was performed on a total of 118 plasma samples and 35 matched tumor samples using a panel covering 474 cancer-associated genes. Results: Twenty-six (62%) of 42 patients achieved clinical complete response (cCR) at three months after CRT. ctDNA was detected in 29 (72.5%) of 40 patients at baseline. The positive rate of ctDNA decreased to 43.9% (20/41) during CRT and continued to descend to 27.0% (10/37) at the completion of CRT. A higher cCR rate was observed in patients with negative during-CRT ctDNA compared to those with detectable ctDNA (82.6% vs. 38.9%, P = 0.008). Patients with post-CRT detectable ctDNA also had poorer cCR rate (30.0% vs. 77.8%, P = 0.017). Within a median follow-up of 24.0 months (IQR 13.7-27.9), the median progression-free survival (PFS) was 12.2 months (95%CI: 8.4-16.0) and median overall survival (OS) was not reached for the whole cohort. Patients with during-CRT detectable ctDNA had shorter PFS compared to ctDNA-negative patients (HR = 2.57, 95%CI: 1.18-5.60, P = 0.014). Similarly, patients with post-CRT detectable ctDNA also had a significantly increased risk of disease progression (HR = 2.88, 95%CI: 1.21-6.83, P = 0.012) and death (HR = 3.67, 95%CI: 1.41-9.55, P = 0.004). Moreover, patients with higher bTMB ( &gt; 1) detected during CRT were associated with a favorable OS (HR = 0.33, 95%CI: 0.13-0.88, P = 0.027). Improved PFS was also observed in those patients with higher post-CRT bTMB ( &gt; 3) (HR = 0.28, 95%CI: 0.08-0.96, P = 0.042). Conclusions: Negative ctDNA status and higher bTMB during or after CRT were associated with better tumor response and favorable survival in ESCC patients who underwent definitive CRT combined with toripalimab. Dynamic ctDNA has great potential in predicting efficacy and risk of disease progression in ESCC. Clinical trial information: NCT04005170 .

Retrospective clinical analysis of circulating tumor DNA (ctDNA)–based molecular response (MR) and baseline blood-based tumor mutational burden (bTMB) for monitoring response in phase 3 trial of bintrafusp alfa vs. pembrolizumab treatment of non-small cell lung cancer (NSCLC).

9098 Background: The clinical utility of ctDNA-based approaches, such as bTMB and MR, to monitor and predict response during chemo-immunotherapy was evaluated using longitudinal cohort samples from a Phase 3 study of bintrafusp alfa versus pembrolizumab as a first-line treatment in patients with PD-L1 expressing advanced NSCLC. Methods: ctDNA was collected at baseline and day 42 after treatment with bintrafusp alfa (Q2W, 1200mg) or pembrolizumab (Q3W, 200mg). The GuardantOMNI liquid biopsy (LBx) assay was used to detect somatic alterations in 497 genes and generate bTMB from baseline, and MR scores from baseline and day 42 (n = 424 samples). bTMB and somatic alterations influencing response were explored. MR scores were calculated using the validated Guardant Response algorithm. Associations between ctDNA metrics and PFS were assessed. Results: 418/424 samples passed sequencing QC; of these, somatic mutations were detected in 236/237 baseline samples and 177/181 day 42 samples. Of 212 NSCLC patients with eligible bTMB results at baseline, 104 (49.1%) patients received bintrafusp alpha and 108 (50.9%) received pembrolizumab. bTMB high (bTMB-H) is defined as &gt;20mut/MB. In the bintrafusp alpha arm, PFS was significantly longer in patients with bTMB-H (median 8.3 months vs 2.7 months, p = 0.00086). In the pembrolizumab arm, PFS was also longer in patients with bTMB-H (median 5.65 months vs 5.5 months, p = 0.0898). Of 132 NSCLC patients with eligible MR results, 67 (50.8%) patients received bintrafusp alpha and 65 (49.2%) patients received pembrolizumab. In the bintrafusp alpha arm, molecular responders had significantly longer PFS (median 7.9 months vs 4.2 months, p = 0.00042). In the pembrolizumab arm, molecular responders also had significantly longer PFS (median 8.95 months vs 4.1 months, p = 0.00054). Conclusions: ctDNA analysis from plasma samples supported MR assessment in patients treated with ICI, indicating its utility as an adjunct to RECIST in monitoring of tumor response. Blood-based TMB-high was associated with immunotherapy treatment benefit. Analysis of b-TMB and MR allowed identification of patients with improved PFS in both treatment arms. Investigation of treatment specific TMB cut off selection and potential clinical utility of defined somatic mutations is ongoing. Clinical trial information: NCT03631706 .

Maximum Somatic Allele Frequency-Adjusted Blood-Based Tumor Mutational Burden Predicts the Efficacy of Immune Checkpoint Inhibitors in Advanced Non-Small Cell Lung Cancer.

Recent studies exhibited the unstable prediction ability of blood-based tumor mutational burden (bTMB) when predicting the response of immune checkpoint inhibitors (ICIs) therapy in patients with non-small cell lung cancer (NSCLC). Circulating tumor DNA (ctDNA) abundance, usually represented by maximum somatic allele frequency (MSAF), was one possible confounding factor influencing bTMB ability in ICIs response prediction. MSAF-adjusted bTMB (Ma-bTMB) was established and validated in patients with advanced NSCLC among Geneplus Cancer Genome Database (GCGD, n = 1679), Zhuo (n = 35), Wang (n = 45), POPLAR (NCT01903993, n = 211) and OAK (NCT02008227, n = 642) cohorts. MSAF demonstrated a modest positive correlation with bTMB and a negative one with survival benefit. Improved survival outcomes of ICIs therapy have been observed among patients with high-Ma-bTMB compared to those with low-Ma-bTMB in Zhuo and Wang cohorts. In addition, compared to low-Ma-bTMB, high-Ma-bTMB was associated with more positive clinical benefits from ICIs therapy than chemotherapy both in POPLAR and OAK cohorts. Further exploration suggested that Ma-bTMB could precisely identify more potential ICIs beneficiaries compared to bTMB and LAF-bTMB, complementary to PD-L1 expression. We developed Ma-bTMB, a convenient, readily available, non-invasive predictive biomarker effectively differentiates beneficiaries of ICIs therapy in advanced NSCLC, warranting future clinical trials.

An exploration of LAF-bTMB as a predictor for the efficacy of immunotherapy combined with chemotherapy in non-small cell lung cancer.

Immune checkpoint inhibitor (ICI) combined with chemotherapy is one of the standards of care for advanced non-small cell lung cancer (NSCLC) without driver mutations. However, the biomarker of combination therapy is still unknown. Although previous studies have confirmed that low allele frequency adjusted blood-based tumor mutational burden (LAF-bTMB) is associated with the efficacy of ICI monotherapy, there has been no report on the correlation between the efficacy of LAF-bTMB and ICI combined chemotherapy. This study aimed to explore whether LAF-bTMB can be used as a predictive biomarker for the efficacy of immunotherapy combined with chemotherapy in advanced NSCLC. This study enrolled patients diagnosed with advanced NSCLC and who received ICI combined with chemotherapy for first-line therapy from May 2020 to December 2021 at Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College. Clinical information, treatment information, survival data, and peripheral blood samples of every patient before treatment were collected. Next-generation sequencing was performed on plasma samples to estimate bTMB and LAF-bTMB. A total of 42 patients with NSCLC were enrolled. In this cohort, 19 patients achieved partial response (PR), and the objective response rate (ORR) was 45.2%. The median progression-free survival (PFS) of all patients was 13.4months (95% CI, 7.49-19.72). Both PFS and the overall survival (OS) were significantly longer in the responder (R) group than in the non-responder (NR) group (median PFS, 16.4months vs. 7.2months, p= 0.028; median OS, NE vs. 9.3months, p= 0.016). There was no significant difference in bTMB and LAF-BTMB between the R and NR group. The ORR of patients with LAF-bTMB≤8muts/Mb was significantly higher than that of patients with LAF-bTMB >8muts/Mb (ORR, 61% vs. 26%, respectively, p= 0.033). When LAF-bTMB ≤8muts/Mb or > 20muts/Mb, ORR was significantly higher than that of patients with LAF-bTMB between 8 and 20muts/Mb (ORR were 57% and 21%, p= 0.047). No correlation has been found between LAF-bTMB and PFS or OS. This study confirmed that neither bTMB nor LAF-bTMB is feasible as a potential predictor of first-line immunochemotherapy for advanced NSCLC. More suitable biomarkers need to be explored to screen patients with better efficacy of immunotherapy combined with chemotherapy in the future.

1092P Clinical potential of circulating tumor DNA (ctDNA)-based molecular response (MR) and baseline blood-based tumor mutational burden (bTMB) for monitoring response to first-line (1L) chemoimmunotherapy in advanced squamous non-small cell lung cancer (sqNSCLC)

The clinical utility of ctDNA-based approaches, such as bTMB and MR, to monitor and predict response during chemo-immunotherapy was evaluated using longitudinal cohort samples from a phase IIa study of patients (pts) with advanced sqNSCLC treated with 1L avelumab in combination with cetuximab and chemotherapy (NCT03717155). Fifty-two plasma samples were obtained from 21 pts treated with 1L avelumab, cetuximab, gemcitabine, and cisplatin for 4 x 3-week cycles followed by avelumab and cetuximab maintenance. The confirmed best overall response (BOR) per RECIST v1.1 was available for 19 pts. The GuardantOMNI liquid biopsy (LBx) assay was used to detect somatic alterations in 497 genes and generate bTMB from baseline, and MR scores from baseline and day 85. bTMB and somatic alterations influencing response were explored. MR scores were calculated using the validated Guardant Response algorithm. Associations between ctDNA metrics and BOR were assessed. We detected somatic mutations in 51 of 52 samples. Biomarker-positive pts were defined as those with high bTMB score (≥20 mut/Mb) and/or STK11, KEAP1, LRP1B, ARID1A/1B/2 mutations. Ten of 18 baseline samples were biomarker-positive; all biomarker-positive pts had a BOR of partial response (PR) or stable disease (SD) and no progressive disease (PD). Eight of 18 baseline samples were biomarker-negative, of which 5 had a BOR of PR or SD and 3 had PD. All 3 pts with PD were TMB-low and 1 had a dual STK11/KRAS alteration, which was previously shown to negatively affect the clinical benefit of immunotherapy. The average ctDNA reduction at the tumor assessment visit following day 85 LBx collection for pts with PR/SD was significantly greater than for pts with PD (82% vs 57%; P=.032). We showed that plasma ctDNA analysis supported MR assessment in pts treated with avelumab combination therapy, which could indicate its clinical utility as an adjunct to RECIST in monitoring tumor response. Plasma TMB-high combined with defined somatic mutations was associated with avelumab combination benefit. Prof. F. Ciardiello and Dr. Z. Feng are acknowledged as first authors and equal contributors to this abstract.

Atezolizumab versus chemotherapy in advanced or metastatic NSCLC with high blood-based tumor mutational burden: primary analysis of BFAST cohort C randomized phase 3 trial

Tumor mutational burden (TMB) is being explored as a predictive biomarker for cancer immunotherapy outcomes in non-small cell lung cancer. BFAST (NCT03178552)—an open-label, global, multicohort trial—evaluated the safety and efficacy of first-line targeted therapies or immunotherapy in patients with unresectable Stage IIIB or IV advanced or metastatic non-small cell lung cancer who were selected for biomarker status using blood-based targeted next-generation sequencing. In the Phase 3 cohort C evaluating blood-based (b)TMB as a biomarker of atezolizumab efficacy, patients with bTMB of ≥10 (N = 471) were randomized 1:1 to receive atezolizumab or platinum-based chemotherapy per local standard of care. Cohort C did not meet its primary endpoint of investigator-assessed progression-free survival in the population with bTMB of ≥16 (hazard ratio, 0.77; 95% confidence interval: 0.59, 1.00; P = 0.053). Adverse events leading to treatment withdrawal occurred in 10% of patients in the atezolizumab arm and 20% in the chemotherapy arm. Adverse events of special interest occurred in 42% of patients in the atezolizumab arm and 26% in the chemotherapy arm. A prespecified exploratory analysis compared the bTMB clinical trial assay with the FoundationOne Liquid Companion Diagnostic assay and showed high concordance between assays. Additional exploration of bTMB to identify optimal cutoffs, confounding factors, assay improvements or cooperative biomarkers is warranted.

Increased blood-based intratumor heterogeneity (bITH) is associated with unfavorable outcomes of immune checkpoint inhibitors plus chemotherapy in non-small cell lung cancer

BackgroundThe combination of immune checkpoint inhibitors (ICIs) and chemotherapy has been the standard first-line treatment for advanced non-small cell lung cancer (NSCLC) patients with driver-gene negative. However, efficacy biomarkers for ICIs-based combination therapy are lacking. We aimed to identify potential factors associated with outcomes of ICIs plus chemotherapy at baseline and dynamic changes in peripheral blood.MethodsWe collected plasma samples of 51 advanced NSCLC patients without EGFR/ALK/ROS1 alteration at baseline and/or after two treatment cycles of ICIs plus chemotherapy. A blood-based intratumor heterogeneity (bITH) score was calculated based on the allele frequencies of somatic mutations using a 520-gene panel. bITH-up was defined as a ≥ 10% increase in bITH score from baseline, with a second confirmatory measurement after treatment.ResultsAt baseline, the number of metastatic organs and lung immune prognostic index (LIPI) were significantly associated with shorter progression-free survival (PFS) of ICIs plus chemotherapy, while bITH and other common molecular biomarkers, including ctDNA level, blood-based tumor mutational burden (bTMB), and PD-L1 expression, had no effect on PFS. LRP1B mutation at baseline was significantly associated with favorable outcomes to ICIs plus chemotherapy. There were 37 patients who had paired samples at baseline and after two cycles of treatment, with the median interval of 53 days. Intriguingly, patients with bITH-up had significant shorter PFS (HR, 4.92; 95% CI, 1.72–14.07; P = 0.001) and a lower durable clinical benefit rate (0 vs 41.38%, P = 0.036) than those with bITH-stable or down. Case studies indicated that bITH was promising to predict disease progression.ConclusionsThe present study is the first to report that increased bITH is associated with unfavorable outcomes of ICIs plus chemotherapy in advanced NSCLC patients.

Maximum somatic allele frequency-adjusted blood-based tumor mutational burden balances the effect of intratumor heterogeneity on response to immune checkpoint Inhibitors in non-small cell lung cancer patients.

e21137 Background: ctDNA abundance can influence the accuracy of blood-based tumor mutational burden (bTMB), driving a paradox of bTMB in predicting response and survival benefit of patients with non-small cell lung cancer (NSCLC) receiving immune checkpoint inhibitors (ICIs) therapy. However, the optimal algorithm of ctDNA adjusted bTMB remains largely unestablished. We aimed to develop a novel approach to optimize the calculation of bTMB by integrating maximum somatic allele frequency (MSAF) to better differentiate the ICI-beneficial NSCLC patients, as well as explore the correlated mechanism. Methods: A 1021-gene panel-based targeted capture sequencing was utilized to evaluate the correlation between MSAF and bTMB based on blood samples in 1679 Chinese NSCLC patients (GCGD database, N = 1679). The establishment of MSAF-adjusted bTMB (Ma-bTMB) was derived from three independent NSCLC cohorts treated with programmed death ligand 1 (PD-L1) or the programmed death 1 (PD-1) inhibitors, including POPLAR cohort (N = 231) as training set, and OAK (N = 642) and Wang (N = 81) cohorts as validation sets separately. PyClone was employed to estimate clonal and subclonal populations. Results: MSAF demonstrated a modest positive correlation between bTMB and MSAF (Spearman’s R = 0.41, P &lt; 0.001, GCGD cohort) and was negatively correlated with survival benefit (Spearman’s rank sum test, overall survival (OS): R = −0.39, P &lt; 0.001; progression-free survival (PFS): R = −0.41, P &lt; 0.001, Wang cohort). Patients with high Ma-bTMB (Ma-bTMB-H) demonstrated improved survival outcomes of anti-PD(L)-1 therapy compared to chemotherapy in POPLAR (PFS, hazard ratio (HR) = 0.63, interaction P = 0.030; OS, HR = 0.44, interaction P = 0.020), which was validated in OAK cohorts (PFS, HR = 0.72, interaction P = 0.040; OS, HR = 0.50, interaction P = 0.010). In addition, Ma-bTMB-H was associated with a better clinical benefit compared to low Ma-bTMB (Ma-bTMB-L) in Wang cohort (PFS, HR = 0.55, P = 0.012; OS, HR = 0.46, P = 0.036). Furthermore, there was a positive correlation between intratumor heterogeneity (ITH) and MSAF (R = 0.37, P &lt; 0.001) in GCGD database, and the effect of ITH on Ma-bTMB (subclones, P = 0.57; subclonal mutations, P = 0.8) was substantially reduced compared to traditional bTMB (subclones, P &lt; 0.001; subclonal mutations, P &lt; 0.001) in Wang cohort. Conclusions: We developed and validated Ma-bTMB, a convenient, readily available, non-invasive predictive biomarker, which reduced the confounding effect of ITH and could effectively differentiate beneficiaries of anti-PD(L)1 therapy, warranting investigation in future clinical trials. Clinical trial information: NCC2018-092.

Impact of cancer susceptibility gene (CSG) mutations in advanced NSCLC (aNSCLC).

9131 Background: Next generation sequencing (NGS) is used widely to identify somatic oncogenic driver mutations in aNSCLC to guide treatment. Comprehensive profiling has led to identification of multiple gene mutations of unclear significance, including germline genes associated with cancer risk, termed CSG. Using data from two randomized trials comparing atezolizumab to docetaxel, we investigated the prognostic and predictive values of CSG in aNSCLC. Methods: We used publicly available data from the OAK (NCT02008227) and POPLAR (NCT01903993) trials. At baseline, plasma was analyzed for cfDNA using FoundationOne CDx NGS assay. We defined CSGs as pathogenic variants of APC, BAP1, BRCA1/2, BRIP1, CDH1, CDKN2A, CHEK2, FH, FLCN, MEN1, MET, MSH2/6, MLH1, PMS2, PALB2, PTCH1, PTEN, RAD51, RB1, SDHB, SMARCA4, STK11, TSC1/2 and VHL. Cox models with treatment covariate, CSG status (mutant [mt] vs wild-type [wt]) and their interaction was used to assess the predictive value by treatment of CSG mt for overall survival (OS) and progression free survival (PFS) using univariate and multivariate models. Similar analysis was performed for objective response rate (ORR). Results: Of 1137 patients, 853 with sufficient tumor content formed the analysis population. In total, 295 (35%) had a known/likely pathogenic CSG mt. The variant allele frequencies (vAF) of CSG mt ≥30%, 10-30%, and &lt;10% were 8% (N=23), 25% (N=73), and 67% (N=199) respectively. Patients with CSG mt were more likely to be smokers (89% vs 81%, P=0.005), had squamous tumors (37% vs 26%, P=0.001) with higher blood-based tumor mutation burden (mean 13.8 vs 10.2 per megabase, P&lt;0.001). CSG mt was not predictive of greater OS benefit with atezolizumab over docetaxel (Table). CSG mt was associated with 35% increase risk of death in univariable analysis (HR 1.35, 95% CI 1.15-1.59). CSG mt was associated with inferior OS in multivariable analysis adjusting for performance status, smoking status, tumor histology, age, sex and number of organ sites of metastasis (HR 1.26, 95% CI 1.07-1.48). Conclusions: Plasma CSG mt is an independent poor prognostic factor in two large aNSCLC clinical trial datasets. Majority of vAF of CSG were low, suggesting that very few were potentially germline in origin, but dedicated sequencing for confirmation will be required. If confirmed, CSG status could be used as a stratification factor in future aNSCLC trials. [Table: see text]

Identification of mutation patterns and circulating tumour DNA-derived prognostic markers in advanced breast cancer patients

BackgroundThe correlations between circulating tumour DNA (ctDNA)-derived genomic markers and treatment response and survival outcome in Chinese patients with advanced breast cancer (ABC) have not been extensively characterized.MethodsBlood samples from 141 ABC patients who underwent first-line standard treatment in Peking University Cancer Hospital were collected. A next-generation sequencing based liquid biopsy assay (PredicineCARE) was used to detect somatic mutations and copy number variations (CNVs) in ctDNA. A subset of matched blood samples and tumour tissue biopsies were compared to evaluate the concordance.ResultsOverall, TP53 (44.0%) and PIK3CA (28.4%) were the top two altered genes. Frequent CNVs included amplifications of ERBB2 (24.8%) and FGFR1 (8.5%) and deletions of CDKN2A (3.5%). PIK3CA/TP53 and FGFR1/2/3 variants were associated with drug resistance in hormone receptor-positive (HR +) and human epidermal growth factor receptor 2-positive (HER2 +) patients. The comparison of genomic variants across matched tumour tissue and ctDNA samples revealed a moderate to high concordance that was gene dependent. Triple-negative breast cancer (TNBC) patients harbouring TP53 or PIK3CA alterations had a shorter overall survival than those without corresponding mutations (P = 0.03 and 0.008). A high ctDNA fraction was correlated with a shorter progression-free survival (PFS) (P = 0.005) in TNBC patients. High blood-based tumor mutation burden (bTMB) was associated with a shorter PFS for HER2 + and TNBC patients (P = 0.009 and 0.05). Moreover, disease monitoring revealed several acquired genomic variants such as ESR1 mutations, CDKN2A deletions, and FGFR1 amplifications.ConclusionsThis study revealed the molecular profiles of Chinese patients with ABC and the clinical validity of ctDNA-derived markers, including the ctDNA fraction and bTMB, for predicting treatment response, prognosis, and disease progression.Trial registration: ClinicalTrials.gov ID: NCT03792529. Registered January 3rd 2019, https://clinicaltrials.gov/ct2/show/NCT03792529.

Blood-based tumor mutational burden as a biomarker for atezolizumab in non-small cell lung cancer: the phase 2 B-F1RST trial

Tumor mutational burden (TMB) in circulating tumor DNA (ctDNA) has shown promise in predicting benefit from PD-L1/PD-1 inhibitors in retrospective studies. Aiming to assess blood TMB (bTMB) prospectively, we conducted B-F1RST (NCT02848651), an open-label, phase 2 trial that evaluated bTMB as a predictive biomarker for first-line atezolizumab monotherapy in locally advanced or metastatic stage IIIB–IVB non-small cell lung cancer (n = 152). The co-primary endpoints were investigator-assessed objective response rate (ORR) per RECIST version 1.1 and investigator-assessed progression-free survival (PFS) between high and low bTMB subgroups at the pre-defined bTMB ≥ 16 (14.5 mutations per megabase) cutoff. Secondary endpoints included investigator-assessed PFS, overall survival (OS) and duration of response at various bTMB cutoffs, as well as safety. Investigator-assessed PFS in the bTMB ≥ 16 versus bTMB < 16 groups was not statistically significant. However, bTMB ≥ 16 was associated with higher ORR, and ORR improved as bTMB cutoffs increased. No new safety signals were seen. In exploratory analyses, patients with maximum somatic allele frequency (MSAF) < 1% had higher ORR than patients with MSAF ≥ 1%. However, further analysis showed that this effect was driven by better baseline prognostics rather than by MSAF itself. At 36.5-month follow-up, an exploratory analysis of OS found that bTMB ≥ 16 was associated with longer OS than bTMB < 16. Further study and assay optimization will be required to develop bTMB as a predictive, standalone biomarker of immunotherapy or for use in conjunction with other biomarkers.

Blood-based tumor mutational burden from circulating tumor DNA and immune checkpoint inhibitors in advanced prostate cancer.

165 Background: A prior study of patients with advanced cancers treated with immune checkpoint inhibitors (ICI) associated improved overall survival with tissue tumor mutational burden above the 80th percentile in each histology (tTMB-H). TMB can also reliably be calculated via liquid biopsy (bTMB), and trends higher than tTMB in analogous tumors. Here, we generate a plasma-informed benchmark for bTMB-H in advanced prostate cancer and report preliminary data from an ongoing multi-institutional case series of patients treated with ICI. Methods: Circulating tumor DNA (ctDNA) next generation sequencing (NGS) results from 3,504 patients with advanced prostate cancer who received a Guardant360 liquid biopsy within a 12-month period were analyzed. The 80th percentile of TMB was defined as the benchmark for bTMB-H in this cohort, and prevalence of microsatellite instability (MSI-H) was determined. Subsequently, clinical data were collected from patients across five institutions who received pembrolizumab based on bTMB scores or other tumor characteristics (i.e. CDK12 mutation). Results: Overall, mean and median bTMB were calculated as 12.34 mut/Mb and 8.61 mut/Mb, respectively (range: 0 – 1164.75 mut/Mb). The 80th percentile of bTMB was 13.4 mut/Mb and the 90th percentile was 16.6 mut/Mb. 2.6% of patients were both MSI-H and bTMB-H. No patients with bTMB &lt; 13.4 mut/Mb (TMB-L) were found to be MSI-H. Ten patients from five institutions received pembrolizumab (8 monotherapy) after a median of 4.5 lines of prior therapy (Table). Four patients were MSI-H. The 3 patients in the cohort with PSA decreases &gt; 50% had both MSI-H and bTMB ≥ 13.4 mut/Mb. Per RECIST criteria, disease was controlled (CR, PR, SD) in 6 of 7 (86%) patients with bTMB scores ≥ 13.4 mut/Mb. Conclusions: These data continue to affirm that ctDNA NGS is a practical and cost-effective means of assessing potential predictors of ICI activity in patients with advanced prostate cancer. Preliminary data from this growing cohort of patients who have received ICI guided by bTMB scores and other molecular parameters identified via ctDNA NGS, including MSI-H, demonstrate a clinical benefit from therapy. Further studies are warranted for contextualizing individual tumor bTMB scores with established, pathology-specific benchmarks.[Table: see text]

Biological and technical factors in the assessment of blood-based tumor mutational burden (bTMB) in patients with NSCLC

BackgroundPatients treated with immunotherapy are at risk of considerable adverse events, and the ongoing struggle is to accurately identify the subset of patients who will benefit. Tumor mutational burden (TMB)...

CCTG BR34: A Randomized Phase 2 Trial of Durvalumab and Tremelimumab With or Without Platinum-Based Chemotherapy in Patients With Metastatic NSCLC

IntroductionFirst-line therapy for patients with metastatic NSCLC includes checkpoint inhibitor monotherapy, dual checkpoint inhibition, or combination with chemotherapy. We compared outcomes with combination chemoimmunotherapy versus dual checkpoint inhibition as first-line treatment for patients with metastatic NSCLC. MethodsThis open-label, randomized clinical trial was conducted at 44 sites in Canada and Australia. Patients with treatment-naive, metastatic NSCLC without sensitizing EGFR or ALK alterations were randomized (1:1) to receive treatment with durvalumab plus tremelimumab with or without platinum-doublet chemotherapy. The primary end point was overall survival (OS). Secondary end points were progression-free survival, overall response rate, and safety. ResultsA total of 301 patients were randomized. Median OS was 16.6 months (95% confidence interval [CI]: 12.6–19.1) with chemotherapy plus immunotherapy and 14.1 months (95% CI: 10.6–18.3) with immunotherapy (hazard ratio = 0.88, 90% CI: 0.67–1.16, p = 0.46). Median progression-free survival with chemotherapy plus immunotherapy was 7.7 months (95% CI: 5.5–8.5) and 3.2 months (95% CI: 2.7–5.1) with immunotherapy (hazard ratio = 0.67, 95% CI: 0.52–0.88). The overall response rate with chemoimmunotherapy was 42.4% and 29.3% with immunotherapy (adjusted OR = 1.69, 95% CI: 1.04–2.76). The percentage of patients with grade 3 or higher adverse events was 82% in the chemotherapy plus immunotherapy group and 70% in the immunotherapy group. Exploratory analyses of programmed death-ligand 1 expression and blood-based tumor mutation burden revealed no differential treatment effect on OS. ConclusionsThe addition of chemotherapy to durvalumab plus tremelimumab in the first-line treatment of stage IV NSCLC did not improve survival compared with durvalumab plus tremelimumab alone. Further study is warranted to identify patients that benefit from initial immunotherapy alone versus combination chemotherapy plus immunotherapy as first-line treatment.