Oropharyngeal squamous cell carcinoma (OPSCC) is the most common neoplasm originating at the base of the tongue or in the tonsils or soft palate. In this study, we investigated the prognostic value ...of FOXP3+ regulatory T cells in OPSCC. Tumor tissues of patients with locally advanced OPSCC were analyzed using quantitative multiplex immunohistochemistry. Staining of CD8+ T cells, conventional CD4+FOXP3- T cells (Tconv cells), CD4+FOXP3+ regulatory T cells (Treg cells), CD20+ B cells, and CD68+ macrophages was performed, and cell density was evaluated in both the tumor and its stroma. Among the 71 patients included in this study, males constituted 93.0% of the cohort, and the median age was 59 years (range: 42-80 years). A total of 56 patients (78.9%) had a smoking history, and 53 (74.6%) patients were positive for human papillomavirus (HPV). The most frequent site of OPSCC was the tonsils (70.4%), followed by the base of the tongue (25.4%). The proportion of Treg cells was lower in the tumors of patients with HPV than in those of patients without HPV. Patients with OPSCC whose tumor Treg cell levels were above the median had longer relapse-free survival (RFS) periods than those with tumor Treg cell levels below the median (HR, 0.12; 95% CI, 0.03-0.46; p = 0.02). Our multivariate analysis identified high Treg levels (HR, 0.13; 95% CI, 0.02-1.00; p = 0.05) as an RFS factor that predicted a good prognosis. Our results demonstrated that high Treg cell density in locally advanced OPSCC tumors was correlated with longer RFS.
Advanced metastatic cancer poses utmost clinical challenges and may present molecular and cellular features distinct from an early-stage cancer. Herein, we present single-cell transcriptome profiling ...of metastatic lung adenocarcinoma, the most prevalent histological lung cancer type diagnosed at stage IV in over 40% of all cases. From 208,506 cells populating the normal tissues or early to metastatic stage cancer in 44 patients, we identify a cancer cell subtype deviating from the normal differentiation trajectory and dominating the metastatic stage. In all stages, the stromal and immune cell dynamics reveal ontological and functional changes that create a pro-tumoral and immunosuppressive microenvironment. Normal resident myeloid cell populations are gradually replaced with monocyte-derived macrophages and dendritic cells, along with T-cell exhaustion. This extensive single-cell analysis enhances our understanding of molecular and cellular dynamics in metastatic lung cancer and reveals potential diagnostic and therapeutic targets in cancer-microenvironment interactions.
Background
Because of the growing number of actionable biomarkers in non–small cell lung cancer (NSCLC), sufficient tissue availability for testing is becoming a greater challenge. Liquid biopsy ...offers a potential solution by complementing standard tissue‐based methods. In this study, the authors analyzed the concordance of actionable genomic alterations sequenced from circulating tumor DNA (ctDNA; Guardant360) and tissue (Oncomine Focus Assay).
Methods
From September 2015 to May 2018, 421 paired plasma and tissue samples from patients with advanced NSCLC who had previously undergone tissue testing by standard methods were collected. Both types of samples were available for 287 patients (262 in cohort 1 treatment‐naive and 25 in cohort 2 treatment failure), and only 1 sample type was available for 134 patients (50 in cohort 3 plasma only and 84 in cohort 4 tissue only).
Results
In cohort 1, 198 samples (77.6%) showed concordance between tissue and plasma next‐generation sequencing (NGS). Among the discordant cases, plasma testing detected additional genomic alterations in 11 patients (4.2%). In 50 patients without tissue‐based NGS results (cohort 3), the ctDNA‐based test detected genomic alterations in 20 samples (40.0%). The median allele frequency (AF) of mutations identified with ctDNA‐based NGS (0.74%) was lower than that identified with the tissue‐based NGS test (13.90%). Clinical responses to matched targeted therapy occurred, regardless of the ctDNA AF. Upfront ctDNA‐based testing identified 60.4% of patients with genomic alterations. In addition, ctDNA‐based testing uncovered 12.0% more actionable alterations when it was performed after tissue‐based NGS testing.
Conclusions
The results indicate that a ctDNA‐based test identifies additional patients with actionable genomic alterations and could, therefore, be used to complement traditional tissue‐based testing for NSCLC.
Lay Summary
Circulating tumor DNA (ctDNA)–based next‐generation sequencing (NGS) testing is becoming essential as the number of actionable genomic biomarker increases for the treatment selection of non–small cell lung cancer.
This study demonstrates the additive value of ctDNA‐based testing in addition to tissue‐based NGS and standard of care–based biomarker testing for detecting additional patients with actionable genomic alterations.
Clinical responses have also been observed in patients with a low allele frequency detected by ctDNA‐based NGS testing.
The results of this study support the benefit of circulating tumor DNA–based next‐generation sequencing (NGS) when it is used along with the tissue‐based standard of care and NGS to prevent undergenotyping in patients with non–small cell lung cancer at the time of the initial diagnosis and during later disease progression.
BackgroundOropharyngeal squamous cell carcinoma (OPSCC) is the most common neoplasm originating at the base of the tongue or in the tonsils or soft palate. In this study, we investigated the ...prognostic value of FOXP3+ regulatory T cells in OPSCC.MethodsTumor tissues of patients with locally advanced OPSCC were analyzed using quantitative multiplex immunohistochemistry. Staining of CD8+ T cells, conventional CD4+FOXP3- T cells (Tconv cells), CD4+FOXP3+ regulatory T cells (Treg cells), CD20+ B cells, and CD68+ macrophages was performed, and cell density was evaluated in both the tumor and its stroma.ResultsAmong the 71 patients included in this study, males constituted 93.0% of the cohort, and the median age was 59 years (range: 42-80 years). A total of 56 patients (78.9%) had a smoking history, and 53 (74.6%) patients were positive for human papillomavirus (HPV). The most frequent site of OPSCC was the tonsils (70.4%), followed by the base of the tongue (25.4%). The proportion of Treg cells was lower in the tumors of patients with HPV than in those of patients without HPV. Patients with OPSCC whose tumor Treg cell levels were above the median had longer relapse-free survival (RFS) periods than those with tumor Treg cell levels below the median (HR, 0.12; 95% CI, 0.03-0.46; p = 0.02). Our multivariate analysis identified high Treg levels (HR, 0.13; 95% CI, 0.02-1.00; p = 0.05) as an RFS factor that predicted a good prognosis.ConclusionsOur results demonstrated that high Treg cell density in locally advanced OPSCC tumors was correlated with longer RFS.
Summary
Entrectinib is a pan-tyrosine-kinase inhibitor that targets oncogenic rearrangements in
NTRK
,
ROS1
and
ALK
. The combined results of two clinical trials demonstrated the efficacy of ...entrectinib in
ROS1
-rearranged NSCLC. Because the development of drug resistance is inevitable, it would be helpful to determine the mechanisms of entrectinib resistance in a
ROS1
-rearranged tumor model so that future therapeutic strategies can be developed. Here, we characterized the molecular basis of resistance in entrectinib-resistant
ROS1
-rearranged HCC78 cells (HCC78ER cells). These cells were analyzed by next-generation sequencing and genetic profiling, which revealed the acquisition of
KRAS
G12C and the amplification of
KRAS
and
FGF3
. However, there were no secondary mutations in the ROS1 kinase domain. We also found that sustained ERK activation was involved in entrectinib resistance, and that combined treatment with selumetinib resensitized HCC78ER cells to entrectinib in cell viability and colony formation assays. Our data suggest that activation of the RAS signaling pathway can cause entrectinib resistance in
ROS1
-rearranged NSCLC, and is unlikely to be overcome by sequential single agent ROS1-targeting strategies against such tumors. Instead, co-targeting ROS1 and MEK may be an effective strategy for overcoming entrectinib resistance in
ROS1
-rearranged NSCLC.
AZD9291 (osimertinib) is approved for standard care in patients with EGFR T790M-positive non-small cell lung cancer (NSCLC) after prior EGFR TKI progression. Furthermore, AZD9291 is now being ...evaluated as a first-line treatment for NSCLC patients with activation EGFR mutations. Based on previous experiments, resistance to AZD9291 as a first-line treatment may also emerge. Thus, identification and understanding of resistance mechanisms to AZD9291 as a first-line treatment can help direct development of future therapies. AZD9291-resistant cells (PC9/AZDR) were established using EGFR inhibitor-naïve PC9 cells. Resistance mechanisms were analyzed using next-generation sequencing (NGS) and a proteome profiler array. Resistance to AZD9291 developed through aberrant activation of ERK signaling by an EGFR-independent mechanism. The combination of a MEK inhibitor with AZD9291 restored the sensitivity of PC9/AZDR cells in vitro and in vivo. PC9/AZDR cells also showed increased MET expression and an HRAS G13R mutation. In addition, maspin expression was higher after AZD9291 treatment in PC9/AZDR cells. Sustained ERK activation confers resistance to AZD9291 as a first-line therapy. Thus, co-targeting EGFR and MEK may be an effective strategy to overcome resistance to AZD9291.
Since the first discovery of rearranged during transfection (RET) fusion in lung adenocarcinoma in 2011, two tyrosine kinase inhibitors, namely vandetanib and cabozantinib, are currently available. ...Despite favorable outcomes in systemic control, the intracranial therapeutic response remains insufficient. In this study, the clinical characteristics and outcomes of non-small cell lung cancer (NSCLC) patients with RET rearrangements were analyzed.
Patients with NSCLC harboring RET fusion who received treatment between January 2006 and January 2018 were analyzed. RET rearrangement was identified by FISH or NGS.
A total of 59 patients were identified. About half of the patients were female (47.5%) and never smokers (50.9%). Most patients had adenocarcinoma (89.8%). A total of 17 patients (28.8%) had an intracranial lesion at the initial diagnosis of stage IV disease, and 11 additional patients (18.6%) developed intracranial metastases during follow-up. The median time to development of intracranial metastases was 19.0 months (95% CI: 9.6-28.5), resulting in a >60% cumulative incidence of brain metastasis at 24 months. The systemic efficacy of pemetrexed-based regimens was favorable with progression-free survival of 9.0 (95% CI: 6.9-11.2) and OS of 24.1 (95% CI: 15.2-33.0) months. The median progression-free survival for vandetanib and immunotherapy was 2.9 (95% CI: 2.0-3.8) and 2.1 (95% CI: 1.6-2.6) months, respectively.
Given the likelihood of RET-rearranged NSCLC progressing to intracranial metastases and the absence of apparent clinical benefit of currently available targeted or immunotherapeutic agents, development of novel treatment with higher selectivity and better penetration of the blood-brain barrier remains a priority.
Although anti-programmed death-1 (PD-1) treatment has shown remarkable anti-tumor efficacy, immune-related adverse events (irAEs) develop with heterogeneous clinical manifestations. However, the ...immunological understanding of irAEs is currently limited. In the present study, we analyzed peripheral blood T cells obtained from cancer patients who received anti-PD-1 treatment to determine the immunological characteristics of irAEs. This study included 31 patients with refractory thymic epithelial tumor (TET) who were enrolled in a phase II trial of pembrolizumab (NCT02607631) and 60 patients with metastatic non-small cell lung cancer (NSCLC) who received pembrolizumab or nivolumab. T-cell profiling was performed by multicolor flow cytometry using peripheral blood obtained before treatment and 7 days after the first dose of anti-PD-1 antibodies. irAEs developed in 21 TET patients and 24 NSCLC patients. Severe (≥ grade 3) irAEs occurred in 7 TET patients (22.6%) and 6 NSCLC patients (10.0%). Patients with severe irAEs exhibited a significantly lower fold increase in the frequency of effector regulatory T (eTreg) cells after anti-PD-1 treatment, a higher ratio of T helper-17 (Th17) and T helper-1 cells at baseline, and a higher percentage of Ki-67
+
cells among PD-1
+
CD8
+
T cells posttreatment. In clustering analysis using the T-cell parameters, patients with irAEs were grouped into four distinct subtypes: Th17-related, TNF-related, CD8-related Treg-compensated, and CD8-related Treg-uncompensated. The T-cell parameters showed a predictive value for the development of each subtype of severe irAEs. In conclusion, severe irAEs after anti-PD-1 treatment were clustered into four immunological subtypes, and potential biomarkers for early prediction of severe irAEs were proposed.
Owing to rapid advancements in NGS (next generation sequencing), genomic alteration is now considered an essential predictive biomarkers that impact the treatment decision in many cases of cancer. ...Among the various predictive biomarkers, tumor mutation burden (TMB) was identified by NGS and was considered to be useful in predicting a clinical response in cancer cases treated by immunotherapy. In this study, we directly compared the lab-developed-test (LDT) results by target sequencing panel, K-MASTER panel v3.0 and whole-exome sequencing (WES) to evaluate the concordance of TMB. As an initial step, the reference materials (n = 3) with known TMB status were used as an exploratory test. To validate and evaluate TMB, we used one hundred samples that were acquired from surgically resected tissues of non-small cell lung cancer (NSCLC) patients. The TMB of each sample was tested by using both LDT and WES methods, which extracted the DNA from samples at the same time. In addition, we evaluated the impact of capture region, which might lead to different values of TMB; the evaluation of capture region was based on the size of NGS and target sequencing panels. In this pilot study, TMB was evaluated by LDT and WES by using duplicated reference samples; the results of TMB showed high concordance rate (R 2 = 0.887). This was also reflected in clinical samples (n = 100), which showed R 2 of 0.71. The difference between the coding sequence ratio (3.49%) and the ratio of mutations (4.8%) indicated that the LDT panel identified a relatively higher number of mutations. It was feasible to calculate TMB with LDT panel, which can be useful in clinical practice. Furthermore, a customized approach must be developed for calculating TMB, which differs according to cancer types and specific clinical settings. BMB Reports 2021; 54(7): 386-391
Owing to rapid advancements in NGS (next generation sequencing), genomic alteration is now considered an essential predictive biomarkers that impact the treatment decision in many cases of cancer. ...Among the various predictive biomarkers, tumor mutation burden (TMB) was identified by NGS and was considered to be useful in predicting a clinical response in cancer cases treated by immunotherapy. In this study, we directly compared the lab-developed-test (LDT) results by target sequencing panel, K-MASTER panel v3.0 and whole-exome sequencing (WES) to evaluate the concordance of TMB. As an initial step, the reference materials (n = 3) with known TMB status were used as an exploratory test. To validate and evaluate TMB, we used one hundred samples that were acquired from surgically resected tissues of non-small cell lung cancer (NSCLC) patients. The TMB of each sample was tested by using both LDT and WES methods, which extracted the DNA from samples at the same time. In addition, we evaluated the impact of capture region, which might lead to different values of TMB; the evaluation of capture region was based on the size of NGS and target sequencing panels. In this pilot study, TMB was evaluated by LDT and WES by using duplicated reference samples; the results of TMB showed high concordance rate (R2 = 0.887). This was also reflected in clinical samples (n = 100), which showed R2 of 0.71. The difference between the coding sequence ratio (3.49%) and the ratio of mutations (4.8%) indicated that the LDT panel identified a relatively higher number of mutations. It was feasible to calculate TMB with LDT panel, which can be useful in clinical practice. Furthermore, a customized approach must be developed for calculating TMB, which differs according to cancer types and specific clinical settings. BMB Reports 2021; 54(7): 386-391.
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