Oncogene-driven non-small cell lung cancers in patients with a history of smoking lack smoking-induced mutations.

Non-small cell lung cancers (NSCLCs) in non-smokers are mostly driven by mutations in the oncogenes EGFR, ERBB2, and MET and fusions involving ALK and RET. In addition to occurring in non-smokers, alterations in these "non-smoking-related oncogenes" (NSROs) also occur in smokers. To better understand the clonal architecture and genomic landscape of NSRO-driven tumors in smokers compared to typical-smoking NSCLCs, we investigated genomic and transcriptomic alterations in 173 tumor sectors from 48 NSCLC patients. NSRO-driven NSCLCs in smokers and non-smokers had similar genomic landscapes. Surprisingly, even in patients with prominent smoking histories, the mutational signature caused by tobacco smoking was essentially absent in NSRO-driven NSCLCs, which was confirmed in two large NSCLC datasets from other geographic regions. However, NSRO-driven NSCLCs in smokers had higher transcriptomic activities related to regulation of the cell cycle. These findings suggest that, while the genomic landscape is similar between NSRO-driven NSCLC in smokers and non-smokers, smoking still affects the tumor phenotype independently of genomic alterations.

Detection of circulating tumor DNA with ultradeep sequencing of plasma cell-free DNA for monitoring minimal residual disease and early detection of recurrence in early-stage lung cancer.

BACKGROUND: In early-stage non-small cell lung cancer (NSCLC), recurrence is frequently observed. Circulating tumor DNA (ctDNA) has emerged as a noninvasive tool to risk stratify patients for recurrence after curative intent therapy. This study aimed to risk stratify patients with early-stage NSCLC via a personalized, tumor-informed multiplex polymerase chain reaction (mPCR) next-generation sequencing assay. METHODS: This retrospective cohort study included patients with stage I-III NSCLC. Recruited patients received standard-of-care management (surgical resection with or without adjuvant chemotherapy, followed by surveillance). Whole-exome sequencing of NSCLC resected tissue and matched germline DNA was used to design patient-specific mPCR assays (Signatera, Natera, Inc) to track up to 16 single-nucleotide variants in plasma samples. RESULTS: The overall cohort with analyzed plasma samples consisted of 57 patients. Stage distribution was 68% for stage I and 16% each for stages II and III. Presurgery (i.e., at baseline), ctDNA was detected in 15 of 57 patients (26%). ctDNA detection presurgery was significantly associated with shorter recurrence-free survival (RFS; hazard ratio [HR], 3.54; 95% confidence interval [CI], 1.00-12.62; p = .009). In the postsurgery setting, ctDNA was detected in seven patients, of whom 100% experienced radiological recurrence. ctDNA positivity preceded radiological findings by a median lead time of 2.8 months (range, 0-12.9 months). Longitudinally, ctDNA detection at any time point was associated with shorter RFS (HR, 16.1; 95% CI, 1.63-158.9; p < .0001). CONCLUSIONS: ctDNA detection before surgical resection was strongly associated with a high risk of relapse in early-stage NSCLC in a large unique Asian cohort. Prospective studies are needed to assess the clinical utility of ctDNA status in this setting.

Combinatorial Hypofractionated Radiotherapy and Pembrolizumab in Anaplastic Thyroid Cancer.

Objectives Anaplastic thyroid cancer (ATC) is an aggressive disease associated with poor outcomes and resistance to therapies. Our study aim was to evaluate the activity of a combinatorial regimen of sandwich sequencing of pembrolizumab immunotherapy and hypofractionated radiotherapy (RT). Methods In this case series, patients with ATC received hypofractionated RT (QUAD-shot) and intravenous pembrolizumab 200mg every 3-4 weeks. Pembrolizumab was continued until disease progression or up till 24 months. Concurrent Lenvatinib treatment was allowed. Primary endpoint was best overall response (BOR) and progression-free survival (PFS). Additionally, we performed immune profiling of circulating T cells in a responder to investigate the immune response to our combinatorial treatment. Results At median follow-up of 32.6 months (IQR: 26.4-38.8), of a cohort of 5 patients, BOR was 80%; with 2 complete responses (CR) and 2 partial responses (PR). Patients who achieved CR remained disease-free at last follow-up. Median PFS was 7.6 months (IQR: 6.2-NR), and 1-year PFS and overall survival rate was 40% (95% CI: 13.7-100) for both. Treatment was well-tolerated, with mostly grade 1-2 adverse events. Immune profiling of one partial responder revealed an increase in activated CD4 and CD8 T cells post-QUAD-shot RT, which was further enhanced during the maintenance phase of pembrolizumab. Conclusions Herein, we reported a case series of 5 patients with ATC, with 2 long-term survivors who were treated with surgical debulking followed by QUAD-shot RT and pembrolizumab, possibly due to synergy of local and systemic treatments in activating anti-tumour immunogenic cytotoxicity. This regimen warrants further investigation in a larger cohort of patients.

Longitudinal post-radiotherapy plasma Epstein-Barr virus DNA trends inform on optimal risk stratification in endemic nasopharyngeal carcinoma.

OBJECTIVES: To characterize longitudinal changes in Epstein-Barr virus (EBV) DNA post-radiotherapy in nasopharyngeal carcinoma (NPC) patients, and investigate whether an early (0-2 weeks) or delayed (8-12 weeks) EBV DNA result better predicts for disease-free survival (DFS). MATERIALS AND METHODS: Histologically-confirmed NPC patients with ≥1 EBV DNA test quantified using the harmonized BamHI-W polymerase chain reaction-based assay at 0-2 and 8-12 weeks post-radiotherapy were included. RESULTS: We identified 302 patients with EBV DNA measured at 0-2 weeks post-radiotherapy; of which, 110 (36.4 %) underwent a repeat test at 8-12 weeks post-treatment. Patients harboring a detectable EBV DNA at 0-2 weeks experienced an inferior DFS (adjusted HR1-264 copies 1.72 [95 %CI: 1.05-2.83], P = 0.031; AHR≥265 copies 4.39 [95 %CI: 1.68-11.44], P = 0.002 relative to 0 copies/mL). At 8-12 weeks, we observed substantial shifts in EBV DNA readings from 0 to 2 weeks; 76/110 (69.1 %) and 34/110 (30.9 %) patients at 0-2 weeks versus 90/110 (81.8 %) and 20/110 (18.2 %) at 8-12 weeks recorded undetectable and detectable EBV DNA, respectively. Positive EBV DNA at 8-12 weeks was strongly associated with relapse (73.3 % [11/15] for 1-264; 80.0 % [4/5] for ≥265 subgroups had relapses versus 15.6 % [14/90] for 0 copies/mL). Area under receiver operating curve values for 2-year relapse rates were 0.817 (95 %CI: 0.725-0.909) for stage + EBV DNA8-12w versus 0.654 (95 %CI: 0.542-0.765) for stage + EBV DNA0-2w. CONCLUSION: EBV DNA is dynamic post-radiotherapy, and delayed EBV DNA testing better enriched for higher-risk NPC patients. This implicates trials investigating adjuvant chemotherapy intensification based on early EBV DNA testing.

Boosted abscopal effect from radiotherapy and pembrolizumab in anaplastic thyroid cancer: a mini-review and case report.

BACKGROUND: The abscopal effect, in which radiation induces a systemic anti-tumour immune response, has been demonstrated with radiotherapy. Immunotherapy boosts the abscopal effect by facilitating the immune response to radiation. Radiotherapy and programmed cell death protein 1/programmed cell death ligand 1 (PD-1/PD-L1) blockade has resulted in the boosted abscopal effect in solid cancers, but its role in anaplastic thyroid cancer (ATC) is unknown. In this mini-review, we describe the abscopal effect and summarise its proposed underlying mechanisms. We then present a potential case of boosted abscopal effect in ATC. CASE DESCRIPTION: In our case presentation, we describe a 51-year-old female who presented with 3 weeks of rapidly enlarging thyroid mass. Examination revealed a 3-cm thyroid nodule which was Bethesda V on fine needle aspiration cytology (FNAC). Intraoperatively, there was a gross extrathyroidal extension into the cricoid cartilage. After total thyroidectomy, post-operative histopathology showed widely invasive follicular thyroid cancer with anaplastic transformation (>50%). Immunohistochemistry showed high PD-L1 expression [combined positive score (CPS) >70%]. Due to residual cricoid cartilage disease and several peri-hilar and lung metastases on positron emission tomography-computed tomography (PET-CT) scan, she underwent post-operative palliative radiotherapy and pembrolizumab. After two cycles of pembrolizumab, repeat PET-CT scan showed complete response (CR) of local and distant disease. She remained well for 32 months, before recent discovery of a right mandible bony metastasis planned for radiotherapy. CONCLUSIONS: This case demonstrates exceptional response to radiotherapy and anti-PD-1 immunotherapy in ATC, potentially illustrating the first known abscopal effect in ATC with this treatment.

Clinical efficacy and biomarker analysis of dual PD-1/CTLA-4 blockade in recurrent/metastatic EBV-associated nasopharyngeal carcinoma.

Single-agent checkpoint inhibitor (CPI) activity in Epstein-Barr Virus (EBV) related nasopharyngeal carcinoma (NPC) is limited. Dual CPI shows increased activity in solid cancers. In this single-arm phase II trial (NCT03097939), 40 patients with recurrent/metastatic EBV-positive NPC who failed prior chemotherapy receive nivolumab 3 mg/kg every 2 weeks and ipilimumab 1 mg/kg every 6 weeks. Primary outcome of best overall response rate (BOR) and secondary outcomes (progression-free survival [PFS], clinical benefit rate, adverse events, duration of response, time to progression, overall survival [OS]) are reported. The BOR is 38% with median PFS and OS of 5.3 and 19.5 months, respectively. This regimen is well-tolerated and treatment-related adverse events requiring discontinuation are low. Biomarker analysis shows no correlation of outcomes to PD-L1 expression or tumor mutation burden. While the BOR does not meet pre-planned estimates, patients with low plasma EBV-DNA titre (<7800 IU/ml) trend to better response and PFS. Deep immunophenotyping of pre- and on-treatment tumor biopsies demonstrate early activation of the adaptive immune response, with T-cell cytotoxicity seen in responders prior to any clinically evident response. Immune-subpopulation profiling also identifies specific PD-1 and CTLA-4 expressing CD8 subpopulations that predict for response to combined immune checkpoint blockade in NPC.

Real world efficacy of osimertinib in second line/beyond in patients with metastatic EGFR+ non-small cell lung cancer and role of paired tumour-plasma T790M testing at tyrosine kinase inhibitor resistance.

Background: Osimertinib is a third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) approved for use in EGFR-mutant lung cancer. We examined its performance in the second/subsequent line after resistance to first- and second-generation (1/2G) EGFR-TKI. Methods: We reviewed electronic records of 202 patients who received osimertinib from July 2015 to January 2019 in the second/subsequent line after progression on prior EGFR-TKI. Of these, complete data from 193 patients were available. Clinical data including patient characteristics, primary EGFR mutation, T790M mutation status, presence of baseline brain metastases (BM), first-line EGFR-TKI use, and survival outcomes were extracted, and results retrospectively analyzed. Results: Of 193 evaluable patients, 151 (78.2%) were T790M+ (T790M positive) with 96 (49.2%) tissue confirmed; 52% of patients received osimertinib in the second line setting. After median follow up of 37 months, median progression-free survival (PFS) of the entire cohort was 10.3 [95% confidence interval (CI): 8.64-11.50] months and median overall survival (OS) was 20 (95% CI: 15.61-23.13) months. Overall response rate (ORR) to osimertinib was 43% (95% CI: 35.9-50.3%); 48.3% in T790M+ vs. 20% in T790M- (T790M negative) patients. OS in T790M+ patients was 22.6 vs. 7.9 months in T790M- patients (HR 0.43, P=0.001), and PFS was 11.2 vs. 3.1 months respectively (HR 0.52, P=0.01). Tumour T790M+ was significantly associated with longer PFS (P=0.007) and OS (P=0.01) compared to tumour T790M- patients, however this association was not seen with plasma T790M+. Of the 22 patients with paired tumor/plasma T790M testing, response rate (RR) to osimertinib was 30% for those plasma T790M+/tumour T790M-, compared to 63% and 67% for those who were plasma T790M+/tumour T790M+ and plasma T790M-/tumour T790M+, respectively. By multivariable analysis (MVA), Eastern Cooperative Oncology Group (ECOG) performance status ≥2 was associated with shorter OS (HR 2.53, P<0.001) and PFS (HR 2.10, P<0.001), whereas presence of T790M+ was associated with longer OS (HR 0.50, P=0.008) and PFS (HR 0.57, P=0.027). Conclusions: This cohort demonstrated the efficacy of osimertinib in second line/beyond for EGFR+ (EGFR mutation-positive) non-small cell lung cancer (NSCLC). Tissue T790M result appeared more predictive of osimertinib efficacy compared to plasma, highlighting potential T790M heterogeneity and the advantage with paired tumor-plasma T790M testing at TKI resistance. T790M- disease at resistance remains an unmet treatment need.

Efficacy of Anti-PD1 Blockade in Treating Recurrent or Metastatic Nasopharyngeal Cancer: A Systematic Review and Meta-analysis.

OBJECTIVES: Anti-PD1 antibody has emerged as a promising immunotherapeutic option in patients with recurrent and/or metastatic nasopharyngeal cancers (RM-NPC). We aim to summarise existing evidence on the use of anti-PD1 antibodies in the treatment of these patients and compare its effectiveness with standard-of-care palliative chemotherapy. Our secondary aim is to explore potential combination therapies with anti-PD1 antibodies. MATERIALS AND METHODS: PubMed, Embase and Cochrane databases were systematically searched for studies comparing the efficacy of various anti-PD1 antibodies in the treatment of RM-NPC (either as first or second line treatment) from inception to 2 September 2022. Meta-analyses were performed to correlate the various anti-PD1 antibodies with primary endpoints including overall response rate disease control rate (DCR), progression free survival (PFS) and overall survival (OS). RESULTS: Eighteen studies with 1,887 patients met the inclusion criteria. The use of anti-PD1 antibody monotherapy as second-line treatment of RM-NPC revealed an ORR of 23 % (95 % CI = 19 %-28 %) and DCR of 51 % (95 % CI = 42 %-60 %). The ORRs for first-line as well as a combination of first and second-line treatments were 21 % (95 % CI = 15 % - 30 %) and 22 % (95 % CI = 6 % - 56 %, I2 = 75 %) respectively. The 12-month PFS and 12-month OS was also 27 % (95 % CI = 21 %-33 %) and 63 % (95 % CI = 53 %-72 %) respectively. ORR was much higher at 73 % (95 % CI = 32 %-94 %) when anti-PD1 antibodies were combined with Gemcitabine plus Cisplatin. CONCLUSION: Anti-PD1 antibody demonstrate considerable activity in previously treated RM-NPC patients. Combining anti-PD1 antibodies with gemcitabine and cisplatin chemotherapy enhanced the efficacy of treatment.

PD-L1 score as a prognostic biomarker in asian early-stage epidermal growth factor receptor-mutated lung cancer.

AIM: To determine the prognostic value of programmed death-ligand 1 (PD-L1) score in early-stage epidermal growth factor receptor (EGFR)-mutated non-small cell lung cancer (NSCLC), contrasted against EGFR-wildtype NSCLC. METHODS: Consecutive patients with Stage IA-IIIA NSCLC diagnosed 1st January 2010-31st December 2019 at National Cancer Centre Singapore with evaluable EGFR and PD-L1 status were included. Co-primary end-points were 2-year disease-free survival (DFS) and 5-year overall survival (OS) by Kaplan-Meier method. RESULTS: 455 patients were included (267 EGFR-mutated, EGFR-M+; 188 EGFR-wildtype, wt). Median age at diagnosis was 65 years, 52.3% (238/455) of patients were males, 62.9% (286/455) of patients were never-smokers and 92.5% (421/455) of patients had R0 resection. Stage IA comprised 42.4% (193/455) of patients, Stage IB comprised 23.1% (105/455) of patients, Stage IIA comprised 10.8% of patients (49/455), Stage IIB comprised 5.1% of patients (23/455) and Stage IIIA comprised 18.7% (85/455) of patients. Among EGFR-M+, 45.3% (121/267) were Ex19del and 41.9% (112/267) were L858R. PD-L1 ≥1% among EGFR-M+ and EGFR-wt was 45.3% (121/267) and 54.8% (103/188) respectively (p = 0.047). At median follow-up of 47 months, 178 patients had relapsed. Among EGFR-M+, 2-year DFS comparing PD-L1 <1% and PD-L1 ≥1% was 78.1% and 67.6% (p = 0.007) while 5-year OS was 59.5% and 42.8% (p = 0.001), respectively. Controlling for age, gender, lymphovascular invasion, adjuvant therapy and resection margin status, PD-L1 ≥1% (hazard ratio, HR 2.18, 95% CI 1.04-4.54, p = 0.038), stage IIB (HR 7.78, 95% CI 1.72-35.27, p = 0.008) and stage IIIA (HR 4.45, 95% CI 1.44-13.80, p = 0.01) emerged as independent predictors of inferior OS on multivariable analysis. In exploratory analysis, genomic analysis of 81 EGFR-M+ tumours was performed. PD-L1 ≥1% tumours had significantly higher rates of TP53 mutations (36.1% versus 15.6%, p = 0.04), with predominantly missense mutations. CONCLUSION: PD-L1 ≥1% is an independent predictor of worse OS among early-stage EGFR-mutated NSCLC and is associated with inferior DFS regardless of EGFR status. PD-L1 score as a risk stratification factor should be evaluated in prospective adjuvant studies among EGFR-mutated NSCLC.

Adjuvant Immunotherapy in Patients with Early-Stage Non-small Cell Lung Cancer and Future Directions.

OPINION STATEMENT: While cisplatin-based adjuvant chemotherapy has been the standard of care for the past two decades, the recent introduction of immunotherapy has heralded an important milestone in the adjuvant landscape of early-stage non-small cell lung cancer (NSCLC). The landmark approval of adjuvant atezolizumab based on disease-free survival (DFS) benefit in IMpower010 was swiftly followed by the recent data for use of adjuvant pembrolizumab in PEARLS/KEYNOTE-091, and similar trials involving other immune checkpoint inhibitors are eagerly anticipated. Although both atezolizumab and pembrolizumab demonstrated a significant DFS benefit in the intention-to-treat population, key subgroup analyses have raised questions about the role of predictive biomarkers such as PD-L1 expression and EGFR-mutation status. In this review, we examine the data from the two important trials (IMpower010 and PEARLS/KEYNOTE-091), discuss the controversies surrounding adjuvant immunotherapy including appropriate endpoints, biomarker selection and highlight key considerations in oncogene-driven NSCLC. Finally, we propose future directions including the impact of neoadjuvant therapy on developments in the adjuvant immunotherapy paradigm and role of minimal residual disease (MRD).

A Randomized Phase 2 Trial of Nivolumab Versus Nivolumab-Ipilimumab Combination in EGFR-Mutant NSCLC.

Introduction: Although immune checkpoint inhibitors (ICIs) have dramatically improved outcomes for nononcogene-addicted NSCLC, monotherapy with programmed cell death protein-1 (PD1) inhibition has been associated with low efficacy in the EGFR-mutant setting. Given the potential for synergism with combination checkpoint blockade, we designed a trial to test the activity of combination nivolumab (N)-ipilimumab (NI) in EGFR-mutant NSCLC. Methods: This is a randomized phase 2 study (NCT03091491) of N versus NI combination in EGFR tyrosine kinase inhibitor (TKI)-resistant NSCLC, with crossover permitted on disease progression. The primary end point was the objective response rate, and the secondary end points included progression-free survival, overall survival, and safety of ICI after EGFR TKI. Results: Recruitment ceased owing to futility after 31 of 184 planned patients were treated. A total of 15 patients received N and 16 received NI combination. There were 16 patients (51.6%) who had programmed death-ligand (PDL1) 1 greater than or equal to 1%, and 15 (45.2%) harbored EGFR T790M. Five patients derived clinical benefits from ICI with one objective response (objective response rate 3.2%), and median progression-free survival was 1.22 months (95% confidence interval: 1.15-1.35) for the overall cohort. None of the four patients who crossed over achieved salvage response by NI. PDL1 and tumor mutational burden (TMB) were not able to predict ICI response. Rates of all grade immune-related adverse events were similar (80% versus 75%), with only two grade 3 events. Conclusions: Immune checkpoint inhibition is ineffective in EGFR TKI-resistant NSCLC. Whereas a small subgroup of EGFR-mutant NSCLC may be immunogenic and responsive to ICI, better biomarkers are needed to select appropriate patients.

Clinical and Genomic Features of HER2 Exon 20 Insertion Mutations and Characterization of HER2 Expression by Immunohistochemistry in East Asian Non-Small-Cell Lung Cancer.

PURPOSE: HER2-altered non-small-cell lung cancer (NSCLC) represents a diverse subgroup, including mutations, amplifications, and overexpression. However, HER2 exon 20 insertion mutations are emerging as a distinct molecular subtype with expanding therapeutic options. We describe the molecular epidemiology and genomic features of HER2-altered NSCLC in an Asian tertiary cancer center. METHODS: We identified patients with HER2-mutated NSCLC in our institutional database, collating clinicopathological features and treatment outcomes. The genomic landscape of human epidermal growth factor receptor 2 (HER2)-mutated NSCLC was further evaluated using whole-exome sequencing (WES) data from combined local and publicly available data sets. HER2 amplification and overexpression as selection biomarkers in NSCLC were further interrogated using HER2 immunohistochemistry and correlations with WES and RNA sequencing data. RESULTS: Among 1,252 patients with consecutive lung adenocarcinoma undergoing routine next-generation sequencing, the prevalence of HER2 mutations was 3.1%-exon 20 insertion mutations comprised 2.7%. We examined the clinicopathological features in 55 patients with HER2-mutated NSCLC comprising 40 exon 20 insertion and 15 nonexon 20 insertion mutations. The most common exon 20 insertion mutation was HER2Y772_A775dup in 30 (75%), followed by HER2G776delinsVC in five patients (13%). There were limited responses to HER2-directed therapies apart from trastuzumab-deruxtecan, and no responses were seen with immunotherapy monotherapy. Evaluating the genomics features of HER2 exon 20 insertion mutations using WES data revealed low tumor mutational burden (TMB), low incidence of cancer driver comutations, and a predominance of aging mutational signature-similar to EGFR-mutated tumors. In contrast, uncommon (or nonexon 20 insertion) HER2-mutated tumors resembled EGFR wild-type tumors with higher TMB, higher frequency of cancer driver comutations, and greater presence of smoking and APOBEC mutational signature. Finally, in evaluating HER2 immunohistochemistry in all lung adenocarcinoma, there was significant discordance comparing different scoring systems and poor correlation with HER2 RNA expression and HER2 amplification. CONCLUSION: The incidence of HER2 mutations is 3.1% in East Asian nonsquamous NSCLC. HER2 exon 20 insertion-mutated tumors appear genomically distinct from uncommon (nonexon 20 insertion) HER2 mutations, the latter demonstrating higher TMB, co-occurring drivers, and predominant nonaging mutational signature. The therapeutic implications of the genomic and clinical features of HER2-mutated NSCLC warrant further investigation.

Association of Clinicopathologic and Molecular Tumor Features With Recurrence in Resected Early-Stage Epidermal Growth Factor Receptor-Positive Non-Small Cell Lung Cancer.

Importance: The recently published ADAURA study has posed a significant dilemma for clinicians in selecting patients for adjuvant osimertinib. Risk factors for recurrence in early-stage epidermal growth factor receptor (EGFR)-positive non-small cell lung cancer (NSCLC) also remain undefined. Objective: To determine clinicopathologic characteristics and recurrence patterns of resected early-stage EGFR-positive NSCLC, using wildtype EGFR as a comparator cohort, and identify features associated with recurrence. Design, Setting, and Participants: This is a cohort study including patients diagnosed with AJCC7 Stage IA to IIIA NSCLC between January 1, 2010, and June 30, 2018, who underwent curative surgical procedures at a specialist cancer center in Singapore. The cutoff for data analysis was October 15, 2020. Patient demographic characteristics, treatment history, and survival data were collated. In exploratory analysis, whole-exome sequencing was performed in a subset of 86 patients. Data were analyzed from September 3, 2020, to June 6, 2021. Exposures: Adjuvant treatment was administered per investigator's discretion. Main Outcomes and Measures: The main outcome was 2-year disease-free survival (DFS). Results: A total of 723 patients were included (389 patients with EGFR-positive NSCLC; 334 patients with wildtype EGFR NSCLC). There were 366 women (50.6%) and 357 men (49.4%), and the median (range) age was 64 (22-88) years. A total of 299 patients (41.4%) had stage IA NSCLC, 155 patients (21.4%) had stage IB NSCLC, 141 patients (19.5%) had stage II NSCLC, and 125 patients (17.3%) had stage IIIA NSCLC. Compared with patients with wildtype EGFR NSCLC, patients with EGFR-positive NSCLC were more likely to be women (106 women [31.7%] vs 251 women [64.5%]) and never smokers (121 never smokers [36.2%] vs 317 never smokers [81.5%]). At median (range) follow up of 46 (0-123) months, 299 patients (41.4%) had cancer recurrence. There was no statistically significant difference in 2-year DFS for EGFR-positive and wildtype EGFR NSCLC (70.2% [95% CI, 65.3%-74.5%] vs 67.6% [95% CI, 62.2%-72.4%]; P = .70), although patients with EGFR-positive NSCLC had significantly better 5-year overall survival (77.7% [95% CI, 72.4%-82.1%] vs 66.6% [95% CI, 60.5%-72.0%]; P = .004). Among patients with EGFR-positive NSCLC, 2-year DFS was 81.0% (95% CI, 74.0%-86.3%) for stage IA, 78.4% (95% CI, 68.2%-85.6%) for stage IB, 57.1% (95% CI, 43.7%-68.4%) for stage II, and 46.6% (95% CI, 34.7%-57.7%) for stage IIIA. Overall, 5-year DFS among patients with stage IB through IIIA was 37.2% (95% CI, 30.1%-44.3%). Sites of disease at recurrence were similar between EGFR-positive and wildtype EGFR NSCLC, with locoregional (64 patients [16.5%] vs 56 patients [16.8%]), lung (41 patients [10.5%] vs 40 patients [12.0%]), and intracranial (37 patients [9.5%] vs 22 patients [6.6%]) metastases being the most common. A risk estimation model incorporating genomic data and an individual patient nomogram using clinicopathologic features for stage I EGFR-positive NSCLC was developed to improve risk stratification. Conclusions and Relevance: This cohort study found that recurrence rates were high in early-stage EGFR-positive NSCLC including stage IA, yet 37.2% of patients with stage IB through IIIA were cured without adjuvant osimertinib. Further studies are needed to elucidate individualized surveillance and adjuvant treatment strategies for early-stage EGFR-positive NSCLC.

Complementary Sequential Circulating Tumor Cell (CTC) and Cell-Free Tumor DNA (ctDNA) Profiling Reveals Metastatic Heterogeneity and Genomic Changes in Lung Cancer and Breast Cancer.

INTRODUCTION: Circulating tumor cells (CTCs) and cell-free tumor DNA (ctDNA) are tumor components present in circulation. Due to the limited access to both CTC enrichment platforms and ctDNA sequencing in most laboratories, they are rarely analyzed together. METHODS: Concurrent isolation of ctDNA and single CTCs were isolated from lung cancer and breast cancer patients using the combination of size-based and CD45-negative selection method via DropCell platform. We performed targeted amplicon sequencing to evaluate the genomic heterogeneity of CTCs and ctDNA in lung cancer and breast cancer patients. RESULTS: Higher degrees of genomic heterogeneity were observed in CTCs as compared to ctDNA. Several shared alterations present in CTCs and ctDNA were undetected in the primary tumor, highlighting the intra-tumoral heterogeneity of tumor components that were shed into systemic circulation. Accordingly, CTCs and ctDNA displayed higher degree of concordance with the metastatic tumor than the primary tumor. The alterations detected in circulation correlated with worse survival outcome for both lung and breast cancer patients emphasizing the impact of the metastatic phenotype. Notably, evolving genetic signatures were detected in the CTCs and ctDNA samples during the course of treatment and disease progression. CONCLUSIONS: A standardized sample processing and data analysis workflow for concurrent analysis of CTCs and ctDNA successfully dissected the heterogeneity of metastatic tumor in circulation as well as the progressive genomic changes that may potentially guide the selection of appropriate therapy against evolving tumor clonality.

Integrative Profiling of T790M-Negative EGFR-Mutated NSCLC Reveals Pervasive Lineage Transition and Therapeutic Opportunities.

PURPOSE: Despite the established role of EGFR tyrosine kinase inhibitors (TKIs) in EGFR-mutated NSCLC, drug resistance inevitably ensues, with a paucity of treatment options especially in EGFR T790M-negative resistance. EXPERIMENTAL DESIGN: We performed whole-exome and transcriptome analysis of 59 patients with first- and second-generation EGFR TKI-resistant metastatic EGFR-mutated NSCLC to characterize and compare molecular alterations mediating resistance in T790M-positive (T790M+) and -negative (T790M-) disease. RESULTS: Transcriptomic analysis revealed ubiquitous loss of adenocarcinoma lineage gene expression in T790M- tumors, orthogonally validated using multiplex IHC. There was enrichment of genomic features such as TP53 alterations, 3q chromosomal amplifications, whole-genome doubling and nonaging mutational signatures in T790M- tumors. Almost half of resistant tumors were further classified as immunehot, with clinical outcomes conditional on immune cell-infiltration state and T790M status. Finally, using a Bayesian statistical approach, we explored how T790M- and T790M+ disease might be predicted using comprehensive genomic and transcriptomic profiles of treatment-naïve patients. CONCLUSIONS: Our results illustrate the interplay between genetic alterations, cell lineage plasticity, and immune microenvironment in shaping divergent TKI resistance and outcome trajectories in EGFR-mutated NSCLC. Genomic and transcriptomic profiling may facilitate the design of bespoke therapeutic approaches tailored to a tumor's adaptive potential.

Individualized Molecular Profiling for Allocation to Clinical Trials Singapore Study-An Asian Tertiary Cancer Center Experience.

PURPOSE: Precision oncology has transformed the management of advanced cancers through implementation of advanced molecular profiling technologies to identify increasingly defined subsets of patients and match them to appropriate therapy. We report outcomes of a prospective molecular profiling study in a high-volume Asian tertiary cancer center. PATIENTS AND METHODS: Patients with advanced cancer were enrolled onto a prospective protocol for genomic profiling, the Individualized Molecular Profiling for Allocation to Clinical Trials Singapore study, at the National Cancer Center Singapore. Primary objective was to identify molecular biomarkers in patient's tumors for allocation to clinical trials. The study commenced in February 2012 and is ongoing, with the results of all patients who underwent multiplex next-generation sequencing (NGS) testing until December 2018 presented here. The results were discussed at a molecular tumor board where recommendations for allocation to biomarker-directed trials or targeted therapies were made. RESULTS: One thousand fifteen patients were enrolled with a median age of 58 years (range 20-83 years). Most common tumor types were lung adenocarcinoma (26%), colorectal cancer (15%), and breast cancer (12%). A total of 1,064 NGS assays were performed, on fresh tumor tissue for 369 (35%) and archival tumor tissue for 687 (65%) assays. TP53 (39%) alterations were most common, followed by EGFR (21%), KRAS (14%), and PIK3CA (10%). Of 405 NGS assays with potentially actionable alterations, 111 (27%) were allocated to a clinical trial after molecular tumor board and 20 (4.9%) were enrolled on a molecularly matched clinical trial. Gene fusions were detected in 23 of 311 (7%) patients tested, including rare fusions in new tumor types and known fusions in rare tumors. CONCLUSION: Individualized Molecular Profiling for Allocation to Clinical Trials Singapore demonstrates the feasibility of a prospective broad molecular profiling program in an Asian tertiary cancer center, with the ability to develop and adapt to a dynamic landscape of precision oncology.

Phase II study of nimotuzumab (TheraCim-hR3) concurrent with cisplatin/radiotherapy in patients with locally advanced head and neck squamous cell carcinoma.

BACKGROUND: The efficacy of a combination of nimotuzumab, a humanized monoclonal antibody to the epidermal growth factor receptor, with chemoradiation in locally advanced head and neck squamous cell carcinoma (HNSCC) was evaluated in a phase II study. METHODS: Patients with stage III/IV HNSCC received 3-weekly cisplatin 100 mg/m2 for three cycles and weekly nimotuzumab 200 mg for 8 weeks concurrently with radiotherapy. Primary endpoint was best overall response (BOR) and secondary endpoint was progression-free survival (PFS). RESULTS: Thirty-seven patients were included; the majority were Chinese (76%), male (89%), and had stage IVA/IVB HNSCC (92%). BOR of complete and partial response was seen in 22/37 (59%) and 10/37 (27%) patients, respectively. Median PFS was 17.5 months (95% CI: 11.1-54.5) and 3-year PFS was 40.4% (95% CI: 24.3-55.9). The frequency and type of adverse events observed were similar to standard chemoradiation. CONCLUSION: The combination of nimotuzumab with cisplatin and radiotherapy was safe and achieved high response rates in HNSCC.

Pairing a prognostic target with potential therapeutic strategy for head and neck cancer.

OBJECTIVES: We have previously identified and validated a panel of molecular prognostic markers (ATP13A3, SSR3, and ANO1) for Head and Neck Squamous Cell Carcinoma (HNSCC). The aim of this study was to investigate the consequence of ATP13A3 dysregulation on signaling pathways, to aid in formulating a therapeutic strategy targeting ATP13A3-overexpressing HNSCC. MATERIALS AND METHODS: Gene Set Enrichment Analysis (GSEA) was performed on HNSCC microarray expression data (Internal local dataset [n = 92], TCGA [n = 232], EMBL [n = 81]) to identify pathways associated with high expression of ATP13A3. Validation was performed using immunohistochemistry (IHC) on tissue microarrays (TMAs) of head and neck cancers (n = 333), staining for ATP13A3 and phosphorylated Aurora kinase A (phospho-T288). Short interfering RNA was used to knockdown ATP13A3 expression in patient derived HNSCC cell lines. Protein expression of ATP13A3 and Aurora kinase A was then assessed by immunoblotting. RESULTS: GSEA identified Aurora kinase pathway to be associated with high expression of ATP13A3 (p = 0.026). The Aurora kinase pathway was also associated with a trend towards poor prognosis and tumor aggressiveness (p = 0.086, 0.094, respectively). Furthermore, the immunohistochemical staining results revealed a significant association between Aurora kinase activity and high ATP13A3 expression (p < 0.001). Knockdown of ATP13A3 in human head and neck cell lines showed decrease in Aurora kinase A levels. CONCLUSION: Tumors with high ATP13A3 are associated with high Aurora kinase activity. This suggests a potential therapeutic role of Aurora kinase inhibitors in a subset of poor prognosis HNSCC patients with overexpression of ATP13A3.