First-generation Tyrosine Kinase Inhibitors Research Articles

Chinese perspectives on clinical efficacy and safety of alectinib in patients with ALK-positive advanced non-small cell lung cancer.

The incidence of lung cancer is increasing in China, in contrast to trends in Western countries, due to the increasing numbers of smokers and high levels of air pollution. Non-small-cell lung cancer (NSCLC) is the most common form of lung cancer, accounting for approximately 85% of lung cancers. Better understanding of the pathogenesis of NSCLC has led to the identification of multiple genetic mutations and chromosomal translocations such as those in the anaplastic lymphoma kinase (ALK) gene. To facilitate the identification of treatment targets, multiple guidelines (European Society for Medical Oncology, National Comprehensive Cancer Network, and American Society of Clinical Oncology) now recommend screening for genetic factors to help guide treatment decisions. In recent years, multiple ALK inhibitors have been developed to treat NSCLC, including the first-generation tyrosine kinase inhibitor (TKI) crizotinib; second-generation TKIs such as ceritinib, ensartinib, brigatinib, and alectinib; the third-generation TKI lorlatinib; and the fourth-generation TKI repotrectinib. These agents differ in structure, potency, and activity, both systemically and their effects on central nervous system (CNS) metastases. Recently, alectinib was approved in China to treat patients with locally advanced or metastatic NSCLC that were ALK+. Alectinib has demonstrated activity against NSCLC, including metastases within the CNS, with better tolerability than crizotinib. These ALK inhibitors represent significant advances in the treatment of NSCLC and yet patients will likely still exhibit disease progression. Alectinib offers greater potency with greater specificity as well as a better toxicity profile than many other TKIs that are currently available. Here, we review the role of ALK as a therapeutic target in NSCLC, the testing methods for identifying ALK-rearranged NSCLC, and the various TKIs currently being used or explored for treatment in this setting, with a focus on alectinib from a Chinese perspective.

Ultra-sensitive EGFR T790M Detection as an Independent Prognostic Marker for Lung Cancer Patients Harboring EGFR del19 Mutations and Treated with First-generation TKIs.

The detection of preexisting EGFR T790M subclones and the assessment of their clinical significance in the pretreatment of patients with EGFR T790M non-small cell lung cancer (NSCLC) remain unclear. A total of 179 tumor samples from patients treated or not with a first-generation tyrosine kinase inhibitor (TKI) was analyzed. The presence of ultra-low levels of preexisting EGFRT790M mutation was evaluated using ultra-sensitive droplet digital PCR (ddPCR) and the clinical implication of these mutations on first-generation TKI efficiency assessed. With a ddPCR linear performance of 0.999 and an analytical sensitivity of approximately 0.001%, we observed a 66% (99/150) overall incidence of ultra-low EGFR T790M mutation. Among 82 patients harboring EGFR activating mutations, the presence of a preexisting EGFR T790M mutation prior to any treatment was significantly associated with a longer progression-free survival (PFS; P = 0.009; log-rank test). Interestingly, longer PFS was linked to concomitant EGFR del19 and ultra-low EGFR T790M mutations. Moreover, the presence of both EGFR del19 and ultra-low EGFR T790M mutations was identified as the best fit for predicting the clinical outcome of patients treated with TKI compared with an ultra-low EGFR T790M mutation status or an activating mutation alone (P = 0.042 and P = 0.0071, respectively). We demonstrate that the detection of the ultra-low EGFR T790M mutation in TKI-naïve patients is not a rare event. We suggest that ddPCR should be used in clinical practice to distinguish patients who may respond to first- or third-generation TKIs.

Zebrafish Xenograft Model of Human Lung Cancer for Evaluating Osimertinib Resistance.

About half of NSCLC patients with EGFR mutation had secondary mutation T790M after treatment with a first-generation tyrosine kinase inhibitor (TKI), Gefitinib. The third-generation of EGFR-TKI Osimertinib is suitable for patients with EGFR mutation and T790M mutation. However, drug screening for NSCLC patients after the emergence of acquired resistance has become a difficult problem for clinicians. In this study, we established drug-resistant cell lines of Gefitinib and Osimertinib to evaluate cell proliferation in vitro. And we investigated the inhibitory effect of different drug concentration gradients on cancer cells. Zebrafish with high homology to human genes were selected as xenotransplantation models to compare the effects of different concentrations of Osimertinib on the proliferation and angiogenesis of zebrafish tumors after transplantation of different lung cancer cell lines. It was confirmed that Osimertinib could inhibit the proliferation of tumor cells with EGFR mutation and T790M resistance mutation in zebrafish, which was consistent with the clinical research conclusion.

The Effect of Next-Generation TKI in Non-Small Cell Lung Cancer after Failure of First-Line Treatment: a Meta-Analysis.

Resistance develops against first-generation tyrosine kinase inhibitors (TKIs), which target the epidermal growth factor receptor (EGFR), after a while for non-small-cell lung cancer (NSCLC). Recently, researchers have developed specific inhibitors against them. Among those inhibitors, next-generation EGFR-TKIs have gained prominence due to the greater efficacy and more favorable tolerability. Today, the efficacy of next-generation EGFR-TKIs in patients with advanced NSCLC after failure on first-generation EGFR-TKIs still remains under investigation. The aim of this meta-analysis was to systematically assess the efficacy and safety profiles of next-generation EGFR-TKIs in advanced NSCLC after failure on first-generation EGFR-TKIs. We performed a comprehensive search of the several electronic databases to September, 2018 to identify clinical trials. The primary endpoint was overall survival (OS), progression-free survival (PFS), disease controlled rate (DCR), objective response rate (ORR), and adverse events (AEs). Severe adverse events (AEs) (grade ≥ 3) based on the EGFR-TKIs were analysed. Odds Ratio (OR) along with 95% confidence interval (95% CI) were utilized for the main outcome analysis. In total, we had 3 randomized controlled trials in this analysis. The group of next-generation EGFR-TKIs was significantly improved PFS (OR = 0.34,95%CI = 0.29-0.40, P < 0.00001), as well with the ORR (OR = 10.48,95%CI = 3.87-28.34, P < 0.00001) and DCR (OR = 6.03,95%CI = 4.41-8.25, P < 0.00001), respectively. However, there is no significant difference in overall survival with next-generation EGFR-TKIs (OR = 1.05,95%CI = 0.85-1.31, P = 0.66). While, the OR for the treatment-related AEs of grade 3 or 4 (diarrhoea, rash/acne, nausea, vomiting, anemia) between the patients who received next-generation EGFR-TKIs and chemotherapy did not show safety benefit (P>0.05). Next-generation EGFR-TKIs was shown to be the better agent to achieve higher response rate and the longer PFS in NSCLC patients as the later-line therapy for previously treated patients with first-generation EGFR-TKIs. While, the benefit of the OS and safety compared with the chemotherapy did not achieved. Further research is needed to develop a database of all EGFR mutations and their individual impact on the differing treatments.

ALK resistance mutations and co-occurring genetic alterations to the ALK tyrosine kinase inhibitors in lung cancer.

e20675 Background: Acquired ALK mutations pose a challenge in multiple ALK tyrosine kinase inhibitors (TKIs) for lung cancer. In our study, we examined the profiles of ALK resistance mutations and co-occurring genetic alterations after targeted therapy. Methods: Using targeted gene capture and next-generation sequencing technologies, we analyzed the somatic mutations from174 patients (pts) with post-TKI samples. Among them, 123 pts received first-generation TKI crizotinib only, 51 pts (34 with second-generation TKI, 17 with third-generation TKI) treated with multiple ALK-TKIs. Results: After the treatment of ALK-TKIs, 29% (50/174) patients developed ALK resistance point mutations, including G1202R (22 pts), G1269A (13 pts), L1196M (8 pts), D1203N (5 pts), F1174L (4 pts), I1171T (4 pts), E1210K (4 pts), G1128A (3 pts), F1174C (3 pts), C1156Y (1 pts), G1123S (1 pts), I1171S (1 pts), L1152R (1 pts), and 10 of them had multi-clone. Specifically, G1269A was found a higher proportion in crizotinib group contrast to multi-TKIs cohort (10/24 vs 3/26, p = 0.024). The recalcitrant G1202R was another common resistance mutation, but there was no significant difference between the two groups (p = 0.052). Other concurrent genetic alterations related to clinical response were usually observed in TP53 mutations (46%), furthermore it seemed to be more frequently detected in post-crizotinb compared with multi-TKIs (P = 0.023). Activated bypass signaling may promote tumor progression. In non-ALK resistance point mutations samples (n = 124), co-occurring genomic alterations in EGFR (32/124, p = 0.004) were significantly more enriched in crizotinib group (n = 99). The driver gene mutation may limit crizotinib response. However, EP300 (24%), CDKN2A (12%), TRIM58 (12%), STK11 (12%) or KRAS (8%) mutations were common in the multiple ALK-TKIs group (n = 25). Conclusions: In lung cancer patients, ALK resistance point mutations G1269A was significantly enriched in post-crizotinib, while patients with multiple ALK-TKIs may frequently found G1202R or L1196M. The co-occurring genetic alterations in TP53 or EGFR after the TKIs therapeutic may offer directions for further research and therapy in lung cancer.

Real-world study of afatinib in first-line or re-challenge settings for patients with EGFR mutant non-small cell lung cancer

Afatinib, a second-generation epidermal growth factor receptor (EGFR)—tyrosine kinase inhibitor (TKI) for mutant non-small cell lung cancer (NSCLC), was approved in Japan in 2014. This study evaluated clinical outcomes of afatinib in real-world practice. Medical records of patients who received afatinib for advanced EGFR-mutant NSCLC were retrospectively reviewed. In total, 128 patients were analyzed. Seventy-six patients received afatinib as the first-line setting and 52 as the re-challenge setting (i.e., after failure of prior first-generation TKI). There was no difference in patient characteristics, such as age, sex, and PS, between the first-line and the re-challenge settings. In the first-line setting, the median progression-free survival (PFS) was 17.8 months (95% confidence interval [CI] 13.7–21.5 months). The overall survival (OS) was 39.5 months (95% CI 34.4- not reached). The response rate (RR) was 64.4%. Subset analysis indicated that patients with dose reduction showed longer PFS than those without dose reduction (18.5 months versus 7.9 months) (P = 0.016). In the re-challenge setting, the median PFS was 8.0 months (95% CI 4.9–9.5 months). The RR was 25%. Most common adverse events leading to dose modification or treatment discontinuation included diarrhea, paronychia, and oral mucositis in both settings. Interstitial lung disease occurred in 5.4% (7/128). In the real-world practice in Japan, afatinib showed comparable or better efficacy compared with that shown in previous clinical trials in both the first-line and the re-challenge settings.

Outcome Differences Between First- and Second-generation EGFR Inhibitors in Advanced EGFR Mutated NSCLC in a Large Population-based Cohort

IntroductionSecond-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) appear superior to first-generation TKIs in clinical trials, but at the cost of greater toxicity. It is unclear whether real-world patients, who often suffer worse outcomes, experience similar survival benefits. Using population-based data, we aim to characterize outcome differences by type of treatment. Patients and MethodsWe reviewed all patients with advanced non–small-cell lung cancer who initiated treatment with an EGFR TKI at BC Cancer between 2010 and 2015. A propensity score was generated to account for imbalances in patient characteristics between treatment groups. A Cox proportional hazards model based on the propensity score was then used to estimate effects of treatment on survival. ResultsA total of 484 patients were identified for analysis. Patients in the second-generation cohort were younger (62 vs. 67 years), had less baseline central nervous system metastases (9% vs. 22%), and more uncommon EGFR mutations (13% vs. 7%). Patients receiving a second-generation TKI had an improved overall survival (hazard ratio, 0.69; P = .05), driven by the subgroup with an EGFR exon 19 deletion. Patients with a L858R mutation did not appear to derive benefit from a second-generation TKI (hazard ratio, 0.91; P = .74). Overall, 40% of patients receiving a second-generation TKI required a dose reduction, but only 1% required discontinuation. ConclusionsSecond-generation TKIs tended to be chosen over first-generation TKIs as frontline therapy in younger patients with uncommon EGFR mutations and without central nervous system metastases. The survival benefit of a second-generation TKI seen in clinical trials appeared to be generalizable to real-world patients and is a reasonable first-line therapy.

Clinical Impact of Rare and Compound Mutations of Epidermal Growth Factor Receptor in Patients With Non–Small-Cell Lung Cancer

BackgroundStandard therapy of advanced non–small-cell lung cancer harboring an activating mutation in the epidermal growth factor receptor (EGFR) gene is treatment with tyrosine kinase inhibitors (TKI). However, for rare and compound mutations of the EGFR gene, the clinical evidence of TKI therapy is still unclear. Patients and MethodsA total of 2906 lung cancer samples were analyzed for EGFR mutations during routine analysis between 2010 and 2017. The samples have been investigated by Sanger sequencing and since 2014 by next-generation sequencing. ResultsWe detected EGFR mutations in 408 specimens (14%). Among these, we found 41 samples with rare and 22 with compound mutations. In these 63 samples, 56 different rare EGFR mutations occurred. Information about the clinical outcome was available for 37. Among those with rare mutations, only one patient harboring the mutation p.G874D had disease that responded to first-generation TKI therapy. In contrast, the disease of all patients with compound mutations responded to first- or second-generation TKI therapy. Furthermore, we collected data on clinical relevance regarding TKI therapy from different databases and from an additional literature search, and only found data for 36 of the 56 detected rare mutations. ConclusionInformation about the clinical outcome of patients with rare and compound EGFR mutations remains limited. At present, second- and third-generation TKIs are available, which may represent new treatment strategies for these patients. Therefore, it is becoming increasingly important to maintain databases concerning rare EGFR mutations.

Brain metastases in resected non-small cell lung cancer: The impact of different tyrosine kinase inhibitors.

ObjectivesThe purpose of this study was to examine the impact of epidermal growth factor receptor (EGFR) mutation status and tyrosine kinase inhibitors (TKIs) on the survival of brain metastases (BM) in patients with surgically resected non-small cell lung cancer (NSCLC).MethodsWe selected the patients who had developed metastatic NSCLC; analyzed the differences between brain metastases and other sites of metastases, including patient characteristics, EGFR status, and survival; and selected the patients who had BM for further investigation. We also compared the treatment effects of first-generation TKIs with those of second-/third-generation TKIs.ResultsA total of 785 cases of stage I-IIIa NSCLC were reviewed. Thirty-six (4.6%) patients were identified as having BM. Among them, 14 patients had a mutated EGFR status. No association between EGFR mutation and the incidence of BM was observed (p = 0.199). Patients with mutated EGFRs had significantly longer overall survival and post-recurrence survival than patients with wild-type EGFR mutation (p = 0.001 for both). However, there was no survival difference between patients with exon 19 and exon 21 mutations (p = 0.426). Furthermore, patients who received the second- and/or third-generation EGFR-TKIs had better survival than patients who only received first-generation EGFR-TKIs (p = 0.031). A multivariate analysis indicated that the next-generation TKIs (HR, 0.007; 95% CI, 0.000 to 0.556; p = 0.026) and a longer interval before BM development (HR, 0.848; 95% CI, 0.733 to 0.980; p = 0.025) were significant factors in longer survival.ConclusionsEGFR-TKIs were effective in treating NSCLC patients with BM after curative pulmonary surgery, especially in those patients harboring EGFR mutations. Furthermore, the second-/third-generation EGFR-TKIs showed more promising results than the first-generation EGFR-TKIs in treating those particular patients, though larger studies needed to further prove the results.

Mutation Profile of Resected EGFR-Mutated Lung Adenocarcinoma by Next-Generation Sequencing.

The efficacy of adjuvant targeted therapy for operable lung cancer is still under debate. Comprehensive genetic profiling is needed for detecting co-mutations in resected epidermal growth factor receptor (EGFR)-mutated lung adenocarcinoma (ADC), which may interfere the efficacy of adjuvant tyrosine kinase inhibitor (TKI) treatment. Mutation profiling of 416 cancer-relevant genes was conducted for 139 resected stage I-IIIa lung ADCs with EGFR mutations using targeted next-generation sequencing. Co-mutation profiles were systematically analyzed. Rare EGFR alterations other than exon 19 deletion and L858R, such as L861Q (∼3%) and G719A (∼2%), were identified at low frequencies. Approximately 10% of patients had mutations in EGFR exon 20 that could confer resistance to first-generation TKIs. Ninety-one percent of patients harbored at least one co-mutation in addition to the major EGFR mutation. TP53 was the top mutated gene and was found more frequently mutated at later stage. Markedly, NF1 mutations were found only in stage II-III ADCs. Conversely, RB1 mutations were more frequent in stage I ADCs, whereas APC mutations were observed exclusively in this group. Thirty-four percent of patients with EGFR TKI-sensitizing mutations had genetic alterations involving EGFR downstream effectors or bypass pathways that could affect the response to EGFR TKIs, such as PIK3CA, BRCA1, and NOTCH1. Operable lung ADCs with EGFR TKI-sensitizing mutations are associated with a high proportion of co-mutations. Mutation profiling of these resected tumors could facilitate in determining the applicability and efficacy of adjuvant EGFR TKI therapeutic strategy. The efficacy of adjuvant epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) therapy for lung cancer harboring EGFR mutation after surgical resection is still under debate. Next-generation sequencing of 416 cancer-relevant genes in 139 resected lung cancers revealed the co-mutational landscape with background EGFR mutation. Notably, the study identified potential EGFR TKI-resistant mutations in 34.71% of patients with a drug-sensitizing EGFR mutation and who were naive in terms of targeted therapy. A comprehensive mutation profiling of these resected tumors could facilitate in determining the applicability and efficacy of adjuvant EGFR TKI therapeutic strategy for these patients.

Repeat biopsy procedures and T790M rates after afatinib, gefitinib, or erlotinib therapy in patients with lung cancer

ObjectivesAfatinib, a second-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), is used for EGFR-mutant non-small cell lung cancer (NSCLC). However, there are few reports about its resistance mechanisms. The aims of this study are to evaluate resistance mechanisms of afatinib compared with other TKIs and analyze the performance of repeat biopsy which is critical for subsequent treatment. Materials and methodsWe screened EGFR-mutant NSCLC patients who started first-line afatinib, gefitinib, or erlotinib from 2014 to 2016, and included patients who acquired resistance. Among those patients, T790 M mutation rates and histologic transformation were compared as an acquired resistance mechanism. ResultsA total of 524 patients started EGFR-TKIs, and 347 experienced disease progression until April 2018. After excluding nine patients with de novo T790 M mutations or who were treated with two TKIs before repeat biopsy, 338 patients were included. Among these patients, 263 (78%) were successfully biopsied and evaluated for EGFR mutations and histologic transformation. T790 M mutation was documented in 35 (41%) of 86 evaluable patients in afatinib group, which is significantly lower than in gefitinib (55%, 73/133) and erlotinib groups (57%, 25/44) (p = 0.026). In multivariate analysis considering both baseline EGFR mutation types (deletion 19 or L858R) and sex, the odds ratio for T790M in afatinib group was 0.45 (95% confidence interval: 0.254-0.795, p = 0.006), compared with gefitinib or erlotinib groups. Five histologic transformations (two small cell, three squamous cell) were detected in afatinib group, while one small cell transformation was detected in gefitinib group, and no transformations were detected in erlotinib group. ConclusionsIn our clinical practice, repeat biopsy was possible in nearly four of five patients. Although T790 M mutation appears to be the main resistance mechanism for afatinib, it affects a lower proportion of patients than observed with first-generation TKIs.

Second-line targeted therapies after nivolumab-ipilimumab failure in metastatic renal cell carcinoma

BackgroundNivolumab-ipilimumab demonstrated a survival benefit over sunitinib in first-line setting for metastatic renal cell carcinomas (mRCCs) and is becoming a new standard of care for naïve patients with intermediate or poor risk prognosis (International mRCC Database Consortium). The efficacy of subsequent vascular endothelial growth factor receptor tyrosine kinase inhibitors (TKIs) after nivolumab-ipilimumab failure remains unclear. MethodsMedical records of mRCC patients treated with nivolumab-ipilimumab, who received subsequent TKI, as part of Checkmate 214 study were reviewed in 13 institutions. Baseline characteristics, outcome data including progression-free survival (PFS), response, overall survival (OS) and toxicities were retrospectively collected. ResultsOverall 33 patients received subsequent TKI after nivolumab-ipilimumab failure. Median follow-up from start of subsequent TKI is 22 months (19–NR). Best response was assessed in 30 patients: 12 partial responses (36%), 13 stable diseases (39%) and five progressive diseases (15%). Median PFS from start of TKI was 8 months [5–13]. Median PFS with first-generation (sunitinib/pazopanib) and second-generation TKI (axitinib/cabozantinib) was 8 months [5–16] and 7 months (5–NA), respectively. PFS in second line was significantly longer in patients with a long first-line duration of response to the double immune checkpoint blockade (≥6 months) with 8 versus 5 months for short responder (<6 months) (p = 0.03). OS rate was 54% at 12 months. Toxicity was as expected: 42% developed at least one toxicity grade ≥3. ConclusionThis is the first report of outcomes with TKI, after first-line nivolumab-ipilimumab failure. Median PFS suggests a sustained benefit of TKI and supports trials investigating the optimal sequence.

Epidermal growth factor receptor tyrosine kinase inhibitors in advanced nonsmall cell lung cancer: what is the preferred first-line therapy?

Epidermal growth factor receptor (EGFR) mt+ nonsmall cell lung cancer (NSCLC) were the first molecularly described NSCLC with an established 'targeted' therapy inhibiting mutated EGFR [EGFR tyrosine kinase inhibitor (TKI)]. EGFR TKI of first and second generation have led to an unprecedented improvement in objective response rate, progression-free survival (PFS) and overall survival (OS) compared with chemotherapy with a significantly reduced toxicity and improved quality of life. Fast elucidation of the most frequent resistance mechanism against first and second-generation TKI, T790M, led to the approval of the third-generation TKI osimertinib in second line. Recently, the FLAURA study showed an impressive PFS benefit and immature OS data for osimertinib against solely first-generation TKI's. Also, the ARCHER study comparing dacomitinib against first-generation TKI showed a PFS and also OS benefit. Two studies combining EGFR TKI and antiangiogenesis showed PFS but no OS benefit. Lately, the combination of TKI and chemotherapy has seen a revival with the NEJ009 study, resulting in an impressive median OS of 55 months. Therefore, potentially four different therapeutic options are available in first-line therapy of EGFR mt+ NSCLC, first, second, third generation, TKI + antiangiogenic agent and TKI + chemotherapy. The purpose of the review is to help to guide physicians to decide in their treatment choice and discuss potential directions of research.

Clinical Relevance of EGFR- or KRAS-mutated Subclones in Patients With Advanced Non–small-cell Lung Cancer Receiving Erlotinib in a French Prospective Cohort (IFCT ERMETIC2 Cohort - Part 2)

IntroductionEvaluation of EGFR Mutation status for the administration of EGFR-TKIs in non-small cell lung Carcinoma (ERMETIC) was a prospective study designed to validate the prognostic value of EGFR/KRAS mutations in patients with advanced non–small-cell lung cancer (NSCLC), all receiving a first-generation tyrosine kinase inhibitor, erlotinib. ERMETIC2 was an ancillary project evaluating the clinical value of common EGFR/KRAS-mutated subclones regarding prognosis using highly sensitive molecular detection methods. Materials and MethodsTumor samples from 228 patients with NSCLC (59% adenocarcinoma, 37% women, and 19% never/former smokers) were available for reanalysis using alternative highly sensitive molecular techniques. A multivariate Cox model was used for prognostic analysis. ResultsUsing alternative highly sensitive techniques, 16 EGFR and 51 KRAS supplementary mutations were newly identified, all still exclusive, leading to an overall rate of 12.3% (n = 28) and 33.3% (n = 76), respectively. Using real-time polymerase chain reaction (hybridization probe), they were significantly associated with progression-free survival (P = .02) and overall survival (OS) (P = .01), which were better for EGFR-mutated patients for progression-free survival (hazard ratio [HR], 0.46; 95% confidence interval [CI], 0.28-0.78) and OS (HR, 0.56; 95% CI, 0.31-1), and worse for KRAS mutations and OS (HR, 1.63; 95% CI, 1.09-2.44). Using the most sensitive technique detection for KRAS–clamp polymerase chain reaction–KRAS mutated subclones did not impact OS. ConclusionsKRAS and EGFR mutations were detected in higher proportions by alternative highly sensitive molecular techniques compared with direct Sanger sequencing. However, minor KRAS-mutated subclones offered no prognostic value when representing less than 1% of the tumor cells.

Lncrna RP1-68D18.2 Promotes K562 Cell Line Resistance to Imatinib through CD44 and Its Downstream Signal Molecules

As the first-generation tyrosine kinase inhibitor specifically targeting the BCR-ABL oncoprotein, imatinib has been successfully used in the therapy for chronic myelogenous leukemia (CML) with a significant response and good compliance. However, imatinib resistance becomes the main reason for failure of the treatment of CML. In addition to the common BCR-ABL dependence of imatinib resistance, e.g. BCR-ABL kinase domain mutations and BCR-ABL gene amplification, BCR-ABL independence also plays important roles, which involves alteration of dynamics of drug import and efflux, persistent or compensatory activation of signaling pathways etc. Recently, several studies have reported the correlation of lncRNA with resistance of chemotherapeutic drugs in many cancers. Here, we report, for the first time, discernible overexpression of lncRNARP1-68D18.2 and CD44 mRNA in the imatinib resistant chronic myelogenous leukemia cell line K562R, using lncRNA and mRNA microarray, and further verify it using qPCR. Knockdown of RP1-68D18.2 in K562R cell lines increased their sensitivity to imatinib, and simultaneously caused a decrease in CD44 and c-Myc protein levels. Correspondingly, overexpression of RP1-68D18.2 in K562, a sensitive cell line, led to a slightly decrease in imatinib sensitivity and partially reverse downregulation of c-Myc, p-ERK and p-MEK caused by imatinib treatment. Analysis of correlations revealed positive correlation between RP1-68D18.2 and CD44 mRNA expression in the primary CML blasts and hematological malignant cell lines. We conclude that overexpression of RP1-68D18.2 could promote imatinib resistance in the K562R cell line through the regulation of CD44 and its downstream signal molecules. DisclosuresNo relevant conflicts of interest to declare.

Multiple configurations of EGFR exon 20 resistance mutations after first- and third-generation EGFR TKI treatment affect treatment options in NSCLC.

After sequential treatment with first- and third-generation EGFR tyrosine kinase inhibitors (TKIs), EGFR-mutant non-small cell lung cancers frequently harbor multiple resistance mutations in exon 20 of EGFR including T790M, mediating resistance to first-generation TKIs, and at codons 792, 796, or 797 mediating resistance to third-generation TKIs. However, whether these resistance mutations are in cis or trans has therapeutic implications for patients. We analyzed a cohort of 29 patients with NSCLC harboring EGFR mutations at codons 792, 796, or 797 to establish the configuration of these mutations. We performed hybrid capture-based, next-generation sequencing on formalin-fixed paraffin-embedded biopsy tissue or liquid biopsy. 27 samples had both a T790M mutation and a mutation at codons 792, 796, or 797. In all of these cases, the mutations were found in the cis configuration; the trans configuration was not observed. Two patients’ samples harbored a mutation at codon 797 but no T790M mutation. In these two cases, longitudinal analysis showed earlier biopsies harbored EGFR T790M, which was undetectable following osimertinib treatment. Treatment of one these patients with both first- and third-generation EGFR TKIs resulted in a mixed response. Here we describe multiple configurations of EGFR T790M and third-generation TKI resistance mutations at codons 792, 796, and 797. These mutations are most commonly found in cis, which confers resistance to all current EGFR TKIs. We also describe two patients that exhibited T790M loss with acquisition of a mutation at codon 797. In addition, one of these patients, with an EGFR C797S in a lung biopsy was subsequently found to have EGFR C797N in a later biopsy of pleural fluid, highlighting the dynamic multiclonal nature of advanced NSCLC.

P2.01-102 Comprehensive Next-Generation Sequencing Guided Targeted Therapies Improve Clinical Outcomes of Lung Cancer Patients

Next-generation sequencing (NGS) has been increasingly involved in the clinical decision-making of cancer care, the primary applications of which are the identification of sensitizing mutations for targeted therapies and drug-resistant mechanisms. Along with more clinical validations and approvals of several commercially available targeted NGS panels by FDA for mutation profiling, it has been debated that whether comprehensive NGS test should be used as a standard procedure in clinical practice. In this study, we recruited 24 patients with stage IV lung cancer. 22 of them are adenocarcinoma and the other 2 are small cell lung cancer and squamous cell carcinoma, respectively. Genomic DNA from 13 tumor tissues, and circulating tumor DNA from 8 pleural effusions and 3 plasma samples in each corresponding patient were collected and subjected to targeted-NGS covering 416 cancer-related genes and 16 genes frequently rearranged in solid tumors. Targeted therapy or chemo/radiochemotherapy was applied in clinical practice with the consideration of patients’ requests and their affordability due to financial barriers. Patients’ clinical outcomes were further evaluated. Of all patients, 54% of them (n=13) were detected with sensitizing mutations that have targeted drugs available, including EGFR exon 19 deletion (n=2), L858R (n=2) and L861Q mutations (n=1), ALK fusion (n=1), ROS1 fusion (n=1), MET exon 14 skipping (n=1), as well as EGFR T790M (n=5) for 4 patients who had progressed on the first-generation tyrosine kinase inhibitors (1st-gen TKI) and 1 TKI-treatment naive patient. 69% of patients (n=9) with actionable mutations were subjected to targeted treatment, while the other 31% patients (n = 4) were treated with chemotherapy or radiochemotherapy. Consistent with clinical validations, the overall survival (OS) of targeted-treatment group is better than the systematic treatment group. On the other hand, for patients without detectable actionable mutations (n=11), 64% of patients (n=7) were underwent chemotherapy, while the rest were treated with 1st-gen EGFR TKI as requested by the patients. As expected, chemotherapy-treatment group had a similar OS as the TKI group. Collectively, patients with sensitizing mutations achieved significantly longer OS from the TKI treatments than those without actionable mutations (p=0.02). Our data demonstrated that the existence of sensitizing mutation is the determining factor for the treatment efficacy of targeted therapies. In the real world, NGS test can not only be involved into the decision-making for the first line treatment, but also be instructive for treatment changes after drug-resistance developed

Malignant peritoneal mesothelioma: a review.

Malignant peritoneal mesothelioma (MPM) is a very rare malignancy of the peritoneum and has a poor prognosis. Of all mesotheliomas, pleural mesothelioma is more common than MPM. In comparison to pleural mesothelioma, the link with asbestos exposure is weaker (33-50% vs. >80%), but it is still the best-defined risk factor. MPM spreads predominantly expansive rather than infiltrative and symptoms are related to tumor spread within the abdominal cavity. Often, MPM is encountered incidentally by diagnostic imaging or by surgery. Computed tomography scan is widely accepted as a first line modality in diagnostic imaging. In diagnostic histopathology, MPM presents some challenges. Firstly, adequate clinical information is of utmost importance to consider the possibility of the diagnosis of MPM. Furthermore, a few morphological subtypes and variants exist. The most sensitive immunohistochemical markers are calretinin (100%), WT1 (94%) and CK5/6 (89%). The malignant character of immunohistochemically demonstrated mesothelial cells is not always obvious. This paradigm somewhat changed with the advent of immunohistochemical demonstration of BAP1 (BRCA-1 associated protein 1). Loss of BAP1 expression supports a diagnosis of malignancy. The gold standard in treatment remains cytoreductive surgery (CRS) combined with hyperthermic intraperitoneal chemotherapy (HIPEC). Targetable molecular pathways in MPM are being identified. An exciting finding was the demonstration of ALK rearrangements in a small subset of patients with MPM and it is hoped for that at least this small subgroup of patients could benefit from treatment with ALK inhibitors. First-generation tyrosine kinase inhibitors against epidermal growth factor receptor (EGFR) did not show any significant activity in MPM. In contrast, nintedanib, an angiokinase inhibitor, improved progression-free survival and bevacizumab, a humanized anti-VEGF antibody increased overall survival in patients with MPM, when administered in combination with cisplatin and pemetrexed. Ongoing immunotherapy trials will offer a possible new treatment.

MA27.10 EGFR-Targeted Therapy Alters the Tumor Microenvironment In EGFR-Driven Lung Tumors: Rationale for Combination Therapies

Non-small cell lung cancer patients harboring EGFR mutations have significant clinical benefit from EGFR-targeted tyrosine kinase inhibitors (TKIs). However, these patients develop resistance eventually. With the promising implementation of immune checkpoint inhibitors targeting the programmed cell death receptor/ligand 1 (PD-1/PD-L1) pathway for the treatment of lung cancer, there is a growing interest in developing combinatorial therapies that could utilize this immune approach in the context of targeted therapies. Although many clinical trials have attempted to study combining EHGR-TKIs with PD-1/PD-L1 inhibitors in NSCLC cases, the clinical benefit is still undefined. Therefore, we carry out this study to investigate the immune response of EGFR-TKIs in EGFR-driven lung tumors, aiming to explore factors may influence the efficacy of this combination strategy. We investigated the early and long-term antitumor effects of first-generation TKI gefitinib and third-generation TKI osimertinib respectively in mice with EGFRL858R andEGFR19DEL/T790M-driven lung tumors.The changes of immune texture in tumors were dynamically tested in different treatment groups by flow cytometry and immunohistochemistry. Upon treatment of gefitinib and osimertinib, we saw significant tumor regression in mice with TKI-sensitive EGFRL858R lung adenocarcinoma. However, mice with EGFR19DEL/T790M-driven tumors did not respond to gefitinib, but did show a significant tumor response to third-generation TKI osimertinib treatment. Accompanied with obvious tumor shrinkage, we saw a significant increase of infiltrating CD11b+ myeloid cells and CD3+ lymphocytes throughout treatment. We further analyzed subpopulation of CD11b+ myeloid cells and CD3+ lymphocytes. Results showed that EGFR-TKIs may demonstrated anti-tumor activity by raising cytotoxic CD8+ T cells, activating dendritic cells, eradicating Foxp3+ Tregs and inhibiting M2-like polarization at early stage. However, these immune benefits occurred temporarily and gradually disappeared with treatment went on. On the other hands, the proportion of myeloid-derived suppressor cells(MDSCs), particular mononuclear-MDSCs were consistently elevated responding to sensitive EGFR-TKIs treatment. Together, results of our study provide novel insights into the immune response to EGFR-TKIs in vivo and provides rationale for potential combinations of EGFR-TKIs and immunotherapies for the treatment of lung carcinomas in the early setting, before the establishment of tumor relapse with long-term EGFR inhibition.And additional therapies aiming to eliminate certain immunosuppressive components should be considered when applying this combination strategy.

EGFR T790M Mutation in TKI-Naïve Clinical Samples: Frequency, Tissue Mosaicism, Predictive Value and Awareness on Artifacts

Background: This study evaluated the distribution of epidermal growth factor receptor (EGFR) T790M mutations in treatment-naïve tumor and normal samples obtained from cancer patients. Methods: We utilized allele-specific PCR (AS-PCR), digital droplet PCR (ddPCR) and next generation sequencing (NGS) to detect EGFR T790M allele in several collections of tumor and normal human tissues. Results: AS-PCR analysis of treatment-naïve tumor samples revealed somatic T790M mutation in 3/394 (1%) non-small cell lung carcinomas (NSCLC) carrying the tyrosine kinase inhibitor (TKI)-sensitizing EGFR mutation, but in none of 334 NSCLC lacking EGFR exon 19 deletions (ex19del) or L858R substitutions and in none of 235 non-lung tumors. Use of highly sensitive and quantitative assays, such as ddPCR and NGS, produced a high number of T790M-specific signals even in presumably T790M-negative DNA specimens. This background noise was evidently higher in degraded DNA isolated from formalin-fixed paraffin-embedded tissues as compared to high molecular weight DNA. A combination of AS-PCR, ddPCR and NGS revealed mosaic EGFR T790M allele in 2/68 (3%) NSCLC treated with the first-generation TKI. Both these tumors produced evident and durable response to gefitinib. Conclusion: Detection of mosaic EGFR T790M mutation in treatment-naïve samples may be compromised by yet unresolved technical issues and may have limited clinical value.