Comparison between next-generation sequencing and multiplex polymerase chain reaction assays for nonsmall-cell lung cancer molecular diagnosis.

PURPOSE: Genetic mutation detection has become an important step in nonsmall-cell lung cancer (NSCLC) treatment because of the increasing number of drugs that target genomic rearrangements. A multiplex test that can detect multiple gene mutations prior to treatment is thus necessary. Currently, either next-generation sequencing (NGS)-based or polymerase chain reaction (PCR)-based tests are used. We evaluated the performance of the Oncomine Dx Target Test (ODxTT), an NGS-based multiplex biomarker panel test, and the AmoyDx Pan Lung Cancer PCR Panel (AmoyDx PLC panel), a real-time PCR-based multiplex biomarker panel test. MATERIALS AND METHODS: Patients with histologically diagnosed NSCLC and a sufficient sample volume to simultaneously perform the AmoyDx PLC panel and ODxTT-M were included in the study. The success and detection rates of both tests were evaluated. RESULTS: Biopsies revealed 116 cases of malignancies, 100 of which were NSCLC. Of these, 59 met the inclusion criteria and were eligible for analysis. The success rates were 100% and 98% for AmoyDx PLC panel and ODxTT-M, respectively. Nine driver mutations were detected in 35.9% and 37.3% of AmoyDx PLC and ODxTT-M panels, respectively. EGFR mutations were detected in 14% and 12% of samples using the AmoyDx PLC panel and ODxTT-M, respectively. Of the 58 cases in which both NGS and AmoyDx PLC panels were successful, discordant results were observed in seven cases. These differences were mainly due to different sensitivities of the detection methods used and the gene variants targeted in each test. DISCUSSION: The AmoyDx PLC panel, a PCR-based multiplex diagnostic test, exhibits a high success rate. The frequency of the nine genes targeted for treatment detected by the AmoyDx PLC panel was comparable to the frequency of mutations detected by ODxTT-M. Clinicians should understand and use the AmoyDx PLC panel and ODxTT-M with respect to their respective performances and limitations.

Immunotherapy or Chemoimmunotherapy in Older Adults With Advanced Non-Small Cell Lung Cancer.

Importance: Immune checkpoint inhibitor (ICI) plus chemotherapy combination treatment (ICI-chemotherapy) is now a standard treatment for non-small cell lung cancer (NSCLC) without targetable oncogene alterations, but there are few data on ICI-chemotherapy for patients 75 years and older. Objective: To inform the choice of first-line drugs in clinical practice and assess the safety and efficacy of ICI-chemotherapy combination treatment in older adult patients with previously untreated advanced NSCLC. Design, Setting, and Participants: This retrospective cohort study included 58 centers in Japan. The cohort consisted of patients 75 years and older with clinical stage IIIB, IIIC, IV, postoperative or radiotherapy recurrent NSCLC. Patients started first-line systemic therapy between December 2018 and March 2021. Those receiving first-line molecular targeted drugs were excluded. The data were analyzed from February 2022 to October 2022. Exposures: Systemic therapy. Main Outcomes and Measures: The main outcomes were overall survival (OS), progression-free survival (PFS), and safety. Results: A total of 1245 patients (median [range] age, 78 [75-95] years; 967 [78%] male) with NSCLC were included in the cohort. Programmed death ligand-1 (PD-L1) expression of less than 1% occurred in 268 tumors (22%); 1% to 49% in 387 tumors (31%); 50% and higher in 410 tumors (33%), and unknown expression in 180 tumors (14%). Median OS was 20.0 (95% CI, 17.1-23.6) months for the 354 patients receiving ICI-chemotherapy (28%); 19.8 (95% CI, 16.5-23.8) months for the 425 patients receiving ICI alone (34%); 12.8 (95% CI, 10.7-15.6) months for the 311 patients receiving platinum-doublet chemotherapy (25%); and 9.5 (95% CI, 7.4-13.4) months for the 155 patients receiving single-agent chemotherapy (12%). After propensity score matching, no differences in OS and PFS were found between the patients receiving ICI-chemotherapy vs ICI alone. Each group consisted of 118 patients. For PD-L1 expression of 1% and higher the OS hazard ratio (HR) was 0.98 (95% CI, 0.67-1.42; P = .90), and the PFS HR was 0.92 (95% CI, 0.67-1.25; P = .59). Significance was also not reached when separately analyzed for lower or higher PD-L1 expression (1%-49% or ≥50%). However, grade 3 or higher immune-related adverse events occurred in 86 patients (24.3%) treated with ICI-chemotherapy and 76 (17.9%) with ICI alone (P = .03). Conclusions and Relevance: In this study, ICI-chemotherapy combination treatment did not improve survival and increased the incidence of grade 3 and higher immune-related adverse events compared with ICI alone in patients 75 years and older. Based on these results, ICI alone may be recommended for older adult patients with PD-L1-positive NSCLC.

Clinically-meaningful improvements in therapy for unresectable NSCLC.

INTRODUCTION: The ideal management of patients with unresectable non-small-cell lung cancer (NSCLC) is still developing. Unresectable NSCLC has a high mortality rate and poor prognosis. The development of immune checkpoint inhibitors (ICIs) and molecular-targeted therapies has been a breakthrough in the treatment. The correct treatment of this patient population is crucial to maximize the clinical benefits without compromising quality of life (QOL). AREAS COVERED: We review the chemoradiotherapies, cytotoxic chemotherapies, immunotherapies, and molecular-targeted therapies available for unresectable NSCLC, focusing on their effects on overall survival, progression-free survival, and QOL. EXPERT OPINION: Although cure is the ultimate goal of cancer treatment, it is often difficult to achieve in advanced NSCLC. Biomarker surveillance techniques, such as next-generation sequencing, have made it possible to provide the most appropriate treatment for each patient. This has led to clinically-meaningful improvements in therapies for unresectable NSCLC. The development of new molecular-targeted therapies and the establishment of treatment for patients who acquired drug resistance after initial treatment have a positive impact on patients' long-term survival. ICIs lead the long-term survival that can be considered a cure of some patients with advanced NSCLC, but such curative survival is difficult to achieve with cytotoxic chemotherapies and molecular-targeted therapies.

Immunohistochemical markers to diagnose primary squamous cell carcinoma of the lung: a meta-analysis of diagnostic test accuracy.

BACKGROUND: Inconsistent diagnostic test accuracies of immunohistological staining for squamous cell carcinoma (SQC) of the lung have been frequently reported. There have been few meta-analyses of the diagnostic accuracies of the immunohistochemical markers. METHODS: A systematic review and meta-analysis were performed following standard guidelines for systematic reviews of diagnostic test accuracy. Immunohistochemical markers (p40, p63, CK5/6, and DSC3) were evaluated as index tests for SQC. The diagnostic odds ratio (DOR) was obtained by the DerSimonian-Laird variate model. Summary estimates of sensitivity and specificity were calculated using a bivariate model. The protocol registration ID is UMIN000041664. RESULTS: The meta-analysis included 85 of the 1353 first-screened articles. The total number of patients was 17,893, which consisted 6151 SQC cases and 11,742 non-squamous non-small-cell lung cancer cases. The DOR was better for p40 (377, 95% confidence interval (CI) = 213-644, I 2 = 0%) than for CK5/6 (120, 95% CI = 78-184, I 2 = 2.5%), p63 (70, 95% CI = 55-88, I 2 = 9.1%), and DSC3 (94, 95% CI = 35-250, I 2 = 3.7%). Summary estimates of sensitivity and specificity were followings: p40 sensitivity 0.92 (95% CI = 0.89-0.95), specificity 0.94 (95% CI = 0.93-0.96); p63 sensitivity 0.92 (95% CI = 0.90-0.94), specificity 0.83 (95% CI = 0.80-0.86); CK5/6 sensitivity 0.90 (95% CI = 0.87-0.93), specificity 0.91 (95% CI = 0.89-0.93); DSC3 sensitivity 0.81 (95% CI = 0.73-0.88), and specificity 0.95 (95% CI = 0.85-0.98). CONCLUSION: P40 had the best DOR to diagnose SQC in non-small-cell lung carcinoma. Despite its lower sensitivity, DSC3 had the best specificity among the four markers and might be useful to rule-in the diagnosis of SQC.

The best regimens for chemo-naïve incurable non-squamous non-small cell lung cancer with a programmed death-ligand 1, tumor proportion score 1-49%: a network meta-analysis.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death worldwide. There is a rank order of the efficacy and safety of treatment options, including immune checkpoint inhibitors (ICIs), bevacizumab (Bev), and cytotoxic drugs. When patients have low programmed death-ligand 1 (PD-L1) expression, there are multiple options for treatment. In this study, we focused on ICI regimens in patients with non-squamous NSCLC with low PD-L1 expression and no driver alterations and assessed the efficacy of the regimens using network meta-analysis. METHODS: Randomized trials for incurable chemo-naïve non-squamous NSCLC were collected through electronic searches. The data were independently extracted and cross-checked by two investigators. The primary outcome of this analysis was overall survival (OS). A frequentist weighted least-squares approach random-model network meta-analysis was applied. RESULTS: Sixty-eight eligible studies and 22,619 patients were identified. Using a platinum + third-generation cytotoxic agent regimen (platinum regimen) as a reference, the platinum regimen + pembrolizumab (Pemb) [hazard ratio (HR) =0.55, 95% confidence interval (CI): 0.34-0.89, P=0.015] showed the best OS, followed by the platinum regimen + nivolumab (Niv) + ipilimumab (Ipi) (HR =0.61, 95% CI: 0.44-0.84, P=0.003) with no heterogeneity (I2=0%, P=0.348). CONCLUSIONS: The addition of Pemb or Niv/Ipi to platinum-based chemotherapy seems to be a good therapeutic option for non-squamous NSCLC with a PD-L1 tumor proportion score (TPS) of 1-49%.

[Efforts of Lung Cancer Oncologists in the"With COVID-19"Era].

After the identification of the new coronavirus disease(COVID-19)in December 2019, it has spread rapidly around the world. The COVID-19 pandemic has forced medical systems to limit their services, promote telehealth, prolong visits to clinics, and postpone scheduled surgical operations. The mortality rate of COVID-19 in patients with lung cancer appears to be significantly higher than that of patients with other cancers. Medical care offered to patients with lung cancer should take into account the risk of COVID-19 associated with visits to clinics and treatments. This review outlines the efforts of lung cancer oncologists in the"With COVID-19"era based on the current evidence and statements from domestic and foreign academic societies and organizations.

T-cell response to phytohemagglutinin in the interferon-γ release assay as a potential biomarker for the response to immune checkpoint inhibitors in patients with non-small cell lung cancer.

BACKGROUND: Immune checkpoint inhibitors are a standard treatment for advanced lung cancer, although it remains important to identify biomarkers that can accurately predict treatment response. Immune checkpoint inhibitors enhance the antitumor T-cell response, and interferon-γ plays an important role in this process. Therefore, this study evaluated whether the number of interferon-γ-releasing peripheral T cells after phytohemagglutinin stimulation in the interferon-γ release assay might act as a biomarker for the response of non-small cell lung cancer to immune checkpoint inhibitor treatment. METHODS: Data were retrospectively collected regarding 74 patients with non-small cell lung cancer who had received immune checkpoint inhibitors. Pretreatment screening tests had been performed using the T-SPOT.TB assay, which quantifies the number of interferon-γ-releasing T cells (as immunospots) in response to phytohemagglutinin and tuberculosis-specific antigen stimulation. Clinical factors and the number of spots in the T-SPOT fields were evaluated for associations with patient outcomes. The median number of spots was used to categorize patients as having high or low values, and the two groups were compared. RESULTS: Relative to patients with a low ratio, patients with a high ratio of phytohemagglutinin/tuberculosis-specific antigen spots (i.e. more responsive T cells) had significantly better progression-free survival after immune checkpoint inhibitor treatment. When we only considered patients with negative T-SPOT results, a high number of phytohemagglutinin-stimulated spots corresponded to significantly longer progression-free survival. CONCLUSION: The T-SPOT.TB assay can be used to quantify the number of immunospots in response to antigen stimulation, which may predict the response to immune checkpoint inhibitors in patients with non-small cell lung cancer.

Resistance mechanisms of epidermal growth factor receptor tyrosine kinase inhibitors in non-small cell lung cancer patients: A meta-analysis.

BACKGROUND: Differences in the resistance mechanisms of epidermal growth factor receptor tyrosine kinase inhibitors in patients with non-small cell lung cancer (NSCLC) harboring epidermal growth factor receptor mutations are unknown. This meta-analysis aimed to clarify the differences in resistance mechanisms after treatment with various epidermal growth factor receptor tyrosine kinase inhibitors. METHODS: We systematically searched PubMed, Cochrane, and Web of Science on July 29, 2020, for relevant studies on acquired resistance mechanisms against epidermal growth factor receptor tyrosine kinase inhibitors. The primary outcome measure was differences in the resistance mechanism between individual or generations of epidermal growth factor receptor tyrosine kinase inhibitors. RESULTS: In total, 33 trials involving 2418 individuals were included and analyzed. T790M was significantly less frequent after afatinib treatment (40.2%, 95% confidence interval [CI]: 31.7%-48.7%) than after gefitinib and erlotinib treatments (52.5%, 95% CI: 48.7%-56.3%, p = 0.005). There were no significant differences between Asian and non-Asian patients in the incidence of T790M after gefitinib, erlotinib, and afatinib treatments. Regarding epidermal growth factor receptor pathway-independent resistant mechanisms, the incidences of small cell lung cancer transformation (osimertinib: 7.9%, 95% CI: 3.6%-12.2%, others: 2.3%, 95% CI: 0.8%-3.8%) and Kirsten rat sarcoma (KRAS) viral oncogene homolog mutation (osimertinib: 4.6%, 95% CI: 1.5%-7.7%, others: 0.2%, 95% CI: 0.0%-1.7%) were significantly higher following osimertinib treatment than with others. CONCLUSIONS: Significant differences in the incidence of resistance mechanisms among epidermal growth factor receptor tyrosine kinase inhibitors exist, which should be taken into consideration when choosing the treatment strategy.

Identification of a novel biomarker based on lymphocyte count, albumin level, and TBAg/PHA ratio for differentiation between active and latent tuberculosis infection in Japan.

Data from China have shown that the ratio of Mycobacterium tuberculosis-specific antigen (TBAg) spots obtained from the T-SPOT.TB test to the number of positive control phytohemagglutinin (PHA) spots (TBAg/PHA ratio) could help distinguish between active tuberculosis infection (ATBI) and latent tuberculosis infection (LTBI). As the applicability of the T-SPOT.TB test may differ according to region and race, we retrospectively verified the utility of the TBAg/PHA ratio in distinguishing between ATBI and LTBI in Japan. The TBAg/PHA ratio was significantly lower in the LTBI group than in the ATBI group. Area under the receiver operating characteristic curve (AUC) analysis between ATBI and LTBI according to the TBAg/PHA ratio was 0.76, with a sensitivity of 65.8% and a specificity of 75.6%. The best AUC was obtained when the TBAg/PHA ratio was divided by both lymphocyte count and albumin levels. Our results demonstrate that, in Japan, the TBAg/PHA ratio is superior to TBAg alone for distinguishing between ATBI and LTBI. In addition, the sensitivity and specificity were improved by combining the TBAg/PHA ratio with lymphocyte count and albumin levels.

MicroRNA-200b is a potential biomarker of the expression of PD-L1 in patients with lung cancer.

BACKGROUND: Advanced non-small cell lung cancer (NSCLC) has a high mortality rate and poor prognosis. However, outcomes have gradually improved after the introduction of novel immunotherapies, including immune checkpoint inhibitors (ICIs). Although programmed death-ligand 1 (PD-L1) expression in tumor tissues is a known biomarker for guiding ICI treatment of NSCLC, challenges such as difficulty of liquid biopsy and heterogeneous results during treatment persist. This study evaluated the potential of miR200b as a surrogate biomarker for PD-L1 expression. METHODS: We used the human lung cancer cell lines H226, H460, H520, A549, and H1975. miR200b expression in blood and bronchoscopy specimens of NSCLC patients was evaluated using reverse-transcription-quantitative PCR. Using flow cytometry, PD-L1 expression in vitro, as well as in tumor tissues, was evaluated after transfection with a mimic miR200b or siRNA. RESULTS: miR200b expression negatively correlated with PD-L1 expression in all cell lines. The induction or knockdown of miR200b also altered PD-L1 expression in vitro. The patient group with a PD-L1 tumor proportion score ≥ 50% had significantly lower miR200b expression in the bronchoscopy specimens (P = 0.025) and serum-derived exosomes (P = 0.022) than that with PD-L1 tumor proportion score < 50%. CONCLUSIONS: miR200b can regulate PD-L1 expression in lung cancer cells, and miR200b expression in clinical specimens negatively correlated with PD-L1 expression. Thus, miR200b may be a useful surrogate biomarker for PD-L1 expression in lung cancer patients. KEY POINTS: SIGNIFICANT FINDINGS OF THE STUDY: High PD-L1 expression was linked to low miR200b expression, whereas low PD-L1 expression was linked to high miR200b expression in human lung cancer patients. Thus, miR200b overexpression or silencing can control PD-L1 expression in cancer cells. What this study adds We demonstrated the potential of miR200b as a surrogate biomarker for PD-L1 expression in lung cancer patients.

Identification of Biomarkers for Non-small-cell Lung Cancer Patients Treated With an Immune Checkpoint Inhibitor.

BACKGROUND/AIM: Immune checkpoint inhibitors (ICIs) have an important role in lung cancer therapy. Although the programmed cell death protein-1 (PD-L1) tumor proportion score (TPS) and tumor mutational burden are known prognostic factors, they are insufficient to predict clinical outcomes. This study was conducted to identify novel biomarkers for ICI treatment. PATIENTS AND METHODS: We performed univariable and multivariable analyses of 110 patients with advanced non-small-cell lung cancer (NSCLC) who were treated with an ICI to identify novel biomarkers related to prognosis. We assessed their backgrounds, such as performance status (PS), PD-L1 TPS, smoking status, and peripheral white blood cell counts at baseline and on the day the second course of ICI administration. RESULTS: In the multivariable analysis, PS, driver gene, immune-related adverse events, and post-treatment absolute neutrophil counts (post-ANCs) were significantly associated with progression-free survival. CONCLUSION: A high level of post-ANCs was associated with poor outcome in ICI-treated NSCLC patients.

Afatinib + bevacizumab combination therapy in EGFR-mutant NSCLC patients with osimertinib resistance: Protocol of an open-label, phase II, multicenter, single-arm trial.

INTRODUCTION: As most patients with epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer (NSCLC) develop progressive disease after treatment with osimertinib, it is important to develop more effective treatment options. Afatinib has been shown to be more effective in in vitro studies than osimertinib when used in cancer cell lines containing some specific EGFR mutations. Therefore, afatinib may be an effective solution, especially when used in combination with an anti-VEGF agent such as bevacizumab. METHODS: A phase II multicenter, open-label, single-arm trial has been initiated to evaluate the efficacy and safety of afatinib and bevacizumab combination as salvage therapy for EGFR-mutated lung cancer in patients previously treated with osimertinib. The primary endpoint will be the objective response rate (ORR) and secondary endpoints are progression-free survival (PFS), overall survival (OS), disease control rate (DCR), and adverse events (AEs). DISCUSSION: A previous study indicated that afatinib inhibits lung cancer cells with specific EGFR mutations more effectively than other EGFR-TKIs such as osimertinib. Therefore, we expect that combination therapy using afatinib and bevacizumab will be effective in patients previously treated with osimertinib (registration no. jRCTs031190077).

Class A CpG oligodeoxynucleotide inhibits IFN-γ-induced signaling and apoptosis in lung cancer.

BACKGROUND: Currently, anticancer immunotherapy based on PD-1/PD-L1 blockade with immune checkpoint inhibitors (ICIs) is being used as a standard therapy for non-small cell lung cancer (NSCLC). However, more effective treatments are required as these tumors are often resistant and refractory. Here, we aimed to determine the effects of immunomodulatory oligodeoxynucleotides (ODNs) in terms of the presence or absence of CpG motifs and the number of consecutive guanosines. METHODS: Western blots were used to measure the molecules which regulate the expression of PD-L1 in human lung cancer cell lines after incubation with several cytokines and ODNs. The expression of PD-L1 and ß2-microglobulin (ß2-MG) on A549 cells, and IFN-γ-induced apoptosis with ODNs were examined by flow cytometry. The relationship between IFN-γ receptor and ODN was analyzed by ELISA and immunofluorescence chemistry. RESULTS: Our results verified that A-CpG ODNs suppress the upregulation of IFN-γ-induced PD-L1 and ß2-MG expression. In addition, we found that ODNs with six or more consecutive guanosines (ODNs with poly-G sequences) may competitively inhibit the IFN-γ receptor and abolish the effect of IFN-γ, thereby suppressing apoptosis and indoleamine 2,3-dioxygenase 1 expression in human lung cancer cells. The tumor microenvironment regulates whether this action will promote or suppress tumor immunity. Thus, in immunotherapy with CpG ODNs, it is essential to consider the effect of ODNs with poly-G sequences. CONCLUSIONS: This study suggests that ODNs containing six or more consecutive guanosines may inhibit the binding of IFN-γ to IFN-γ receptor. However, it does not directly show that ODNs containing six or more consecutive guanosines competitively inhibit the IFN-γ receptor, and further studies are warranted to confirm this finding. KEY POINTS: Significant findings of the study: Oligodeoxynucleotides with a contiguous sequence of six or more guanosines may competitively inhibit the IFN-γ receptor and abolish the action of IFN-γ. This may suppress IFN-γ-induced apoptosis and indoleamine-2,3-dioxygenase-1 expression in human lung cancer cells. WHAT THIS STUDY ADDS: A-CpG and poly-G ODN may overcome tolerance if the cause of ICI tolerance is high IDO expression. However, IFN-γ also has the effect of suppressing apoptosis of cancer cells, and it is necessary to identify the cause of resistance.

Reproducibility of the T-SPOT.TB test for screening Mycobacterium tuberculosis infection in Japan.

OBJECTIVES: The interferon-gamma release assay (IGRA) is useful for diagnosing Mycobacterium tuberculosis infections, especially in countries where Bacille Calmette-Guérin vaccinations are performed. However, reproducibility of the IGRA is unclear, as recent data suggest high IGRA conversion and reversion rates in serial tests among healthcare workers. This longitudinal study aimed to evaluate reproducibility of T-SPOT.TB for screening M. tuberculosis infections in Japan. METHODS: Results of T-SPOT.TB tests performed between April 2014 and March 2016 at two hospitals in Yokohama, Japan, where the incidence of tuberculosis was 18.0 per 100,000 population in 2014, were analyzed. RESULTS: In total, 3890 T-SPOT.TB tests were included. Overall, positive and negative test rates were 8.4% and 87.6%, respectively. Among 373 serial tests within two years, conversion and reversion rates were only 1.1% and 12.5%, respectively. Almost all patients who were initially negative (98.9%) remained so. There was no statistically significant difference between the outcomes observed at the two hospitals. CONCLUSIONS: The conversion rate of T-SPOT.TB in Japan is as low as that recently reported in other countries where the incidence of tuberculosis is low. These data indicate that T-SPOT.TB is a reproducible tuberculosis screening tool at local hospitals in areas with a moderate incidence of tuberculosis.

Non-small cell lung cancer with mesenchymal-epithelial transition gene exon 14 skipping mutation treated with crizotinib.

We report the case of an 85-year-old man who was surgically diagnosed with lung adenocarcinoma (pT2aN1M0 stage IIA). He was administered platinum combination chemotherapy as first-line treatment for lung cancer recurrence. The patient's pleural fluid sample was obtained and analysed using a next-generation sequencer, which demonstrated the presence of mesenchymal-epithelial transition gene (MET) exon 14 skipping mutations. As the patient developed progressive disease after receiving first-line chemotherapy, crizotinib was administered as the second-line treatment. The treatment was effective, and the patient had a stable disease for 7 months. This case suggests that crizotinib is effective against non-small cell lung cancer with MET exon 14 alterations.