L858R Point Mutation In Exon Research Articles

Structural Perspectives in the Development of Novel EGFR Inhibitors for the Treatment of NSCLC.

Non-small cell Lung cancer (NSCLC) is the most common type of lung cancer, which is caused by high consumption of tobacco and smoking. It is an epithelial lung cancer that affects about 2.2 million people across the globe, according to International Agency for Research on Cancer (IARC). Non-small cell lung cancer is a malignant tumor caused by EGFR mutation that occurs in the in-frame deletion of exon 19 and L858R point mutation in exon 21. Presently, clinically available inhibitors of EGFR (including erlotinib, lapatinib, gefitinib, selumetinib, etc.) are not specific and responsible for undesirable adverse effects. Moreover, to solve this problem search for newer EGFR inhibitors is the utmost need for the treatment and/or management of increasing lung cancer burden. The discovery of therapeutic agents that inhibit the specific target in tumorous cells, such as EGFR, is one of the successful strategies in treating many cancer therapies, including lung cancer. The exhaustive literature survey (2018-2023) has shown the importance of medicinally privileged pyrimidine derivatives together, fused and/or clubbed with other heterocyclic rings to design and develop novel EGFR inhibitors. Pyrimidine derivatives substituted with phenylamine, indole, pyrrole, piperazine, pyrazole, thiophene, pyridine and quinazoline derivatives substituted with phenylamine, pyrimidine, morpholine, pyrrole, dioxane, acrylamide, indole, pyridine, furan, pyrimidine, pyrazole etc. are privileged heterocyclic rings shown promising activity by inhibiting EGFR and TKIs. The present review summarizes the structure-activity relationship (SAR) and enzyme inhibitory activity, including IC50 values, percentage inhibition, and kinetic studies of potential compounds from various literature. The review also includes various aspects of molecular docking studies with compounds under clinical trials and patents filed on pyrimidine-based EGFR inhibitors in treating non-small cell lung cancer. The present review may benefit the medicinal chemist for developing novel compounds such as EGFR inhibitors.

Abstract 4476: BI-732, a novel fourth-generation EGFR-TKI, demonstrates promising activities against the C797S-mediated EGFR-TKI resistance

Abstract Introduction: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) is a standard modality of the 1st-line treatments for patients with EGFR-mutated non-small-cell lung cancer (NSCLC). The FDA approved osimertinib, a third-generation EGFR-TKI, is highly selective for EGFR-activating mutations (exon 19 deletion [del19] or L858R point mutation in exon 21 [L858R]) as well as EGFR-T790M mutation. Nevertheless, resistance inevitably emerges and leads to disease progression. Acquired EGFR-C797S mutation is the most common on-target resistance mechanism to osimertinib in first- and second-line settings, and there are currently no approved targeted therapies. We aimed to evaluate the efficacy of BI-732, a fourth-generation EGFR-TKI developed to overcome the C797S mutation. Materials and Methods: We constructed several types of Ba/F3 cells expressing EGFR mutations containing the C797S mutation as follows: EGFRdel19/C797S, EGFRL858R/C797S, EGFRdel19/T790M/C797S, and EGFRL858R/T790M/C797S. We also constructed PC9 cells harboring EGFRdel19/C797S (named PC9-DC cells) using the CRISPR/Cas9 system. Furthermore, we generated two patient-derived cells, YU-1097 with EGFRdel19/T790M/C797S and YU-1182 with EGFRL858R/C797S. The in vitro efficacy of BI-732 was tested in several cell lines harboring the EGFR-C797S mutation mentioned above. We further evaluated in vivo antitumor activity of BI-732 in subcutaneous YU-1097 xenograft model and intracranial YU-1097-tumor model.Results: BI-732 effectively inhibited the viability of Ba/F3 cells expressing EGFRdel19/C797S (IC50, 6.8 nM), EGFRL858R/C797S (IC50, 213.4 nM), EGFRdel19/T790M/C797S (IC50, 3.8nM), and EGFRL858R/T790M/C797S (IC50, 15.2 nM), concomitant with marked reduction in EGFR and downstream signal phosphorylation. Moreover, BI-732 exhibited comparable nanomolar in vitro activity to osimertinib on EGFR activating mutation while sparing wild-type EGFR activity. We further confirmed the superior antiproliferative efficacy of BI-732 in PC9-DC (IC50, 24.9 nM), YU-1182 (IC50, 73 nM) and YU-1097 cells (IC50, 2.9 nM). Oral administration of BI-732 at a concentration of 25 mg/kg as a single agent resulted in significant tumor regression with 183.2% tumor growth inhibition (TGI) in the subcutaneous YU-1097 xenograft model. In a combination treatment strategy with osimertinib, BI-732 showed enhanced antitumor activity at much lower concentrations than monotherapy. BI-732 also demonstrated the ability to penetrate brain-blood barrier (BBB) in the intracranial YU-1097-tumor models.Conclusions: BI-732 is a potent, selective, and orally available fourth-generation EGFR-TKI for EGFR mutations including C797S and has efficient BBB penetration. Our findings suggest that BI-732 can be effective against EGFR mutant NSCLC, especially EGFRDel19/T790M/C797S and EGFRDel19/C797S, that have progressed with previous EGFR inhibitors. Citation Format: Eun Ji Lee, Jiyun Lee, Seung Yeon Oh, You Won Lee, Ju young Kim, Su-Jin Choi, Sewon Park, Mi Ra Yu, Jae Hwan Kim, Kyoung-Ho Pyo, Jii Bum Lee, Min Hee Hong, Sun Min Lim, Anke Baum, Lydia Woelflingseder, Harald Engelhardt, Mark Petronczki, Flavio Solca, Mi Ran Yun, Byoung Chul Cho. BI-732, a novel fourth-generation EGFR-TKI, demonstrates promising activities against the C797S-mediated EGFR-TKI resistance. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4476.

The complexity of EGFR exon 19 deletion and L858R mutant cells as assessed by proteomics, transcriptomics, and metabolomics

Most lung adenocarcinoma-associated EGFR tyrosine kinase mutations are either an exon 19 deletion (19Del) or L858R point mutation in exon 21. Although patients whose tumors contain either of these mutations exhibit increased sensitivity to tyrosine kinase inhibitors, progression-free and overall survival appear to be longer in patients with 19Del than in those with L858R. In mutant-transfected Ba/F3 cells, 19Del and L858R were compared by multi-omics analyses including proteomics, transcriptomics, and metabolomics. Proteome analysis identified increased plastin-2, TKT, PDIA5, and ENO1 expression in L858R cells, and increased EEF1G expression in 19Del cells. RNA sequencing showed significant differences between 19Del and L858R cells in 112 genes. Metabolome analysis showed that amino acids, adenylate, guanylate, NADPH, lactic acid, pyruvic acid glucose 6-phosphate, and ribose 5-phosphate were significantly different between the two mutant cells. Because GSH was increased with L858R, we combined osimertinib with the GSH inhibitor buthionine sulfoximine in L858R cells and observed synergistic effects. The complexity of EGFR 19Del and L858R mutant cells was demonstrated by proteomics, transcriptomics, and metabolomics analyses. Therapeutic strategies for lung cancer with different EGFR mutations could be considered because of their different metabolic phenotypes.

Clinical outcome of Brazilian patients with non-small cell lung cancer in early stage harboring rare mutations in epidermal growth factor receptor.

The common epidermal growth factor receptor (EGFR) mutations, such as the L858R point mutation in exon 21 and the in-frame deletional mutation in exon 19, have been definitively associated with response to EGFR-tyrosine kinase inhibitors (EGFR-TKI). However, the clinical outcome and response to treatment for many other rarer mutations are still unclear. In this study, we report the results of Brazilian patients in stage IB-IIIA non-small cell lung cancer (NSCLC) following complete resection with minimal residual disease and EGFR mutations treated with adjuvant chemotherapy and/or EGFR-TKIs. The frequency of EGFR mutations was investigated in 70 cases of early stage NSCLC. Mutations in exons 18 and 20, uncommon mutations in exons 19 and 21, as well as in exons 3, 7, 14, 16, 22, 27, and 28, and/or the presence of different mutations in a single tumor (complex mutations) are considered rare. EGFR mutations were detected in 23 tumors (32.9%). Fourteen cases carried rare mutations and were treated with platinum-based chemotherapy and two cases were treated with erlotinib. The clinical outcome is described case by case with references to the literature. Notably, we found two rare EGFR mutations and one of them with an unknown response to chemotherapy and/or EGFR-TKIs. We have provided complementary information concerning the clinical outcome and treatment of patients with early stage NSCLC for several rare EGFR mutations not previously or only rarely reported. Description of cases harboring rare mutations can support the decision-making process in this subset of patients.

Efficacy of Icotinib, an EGFR Tyrosine Kinase Inhibitor in Non-Small Cell Lung Cancer Patients with Exon 19 Deletion and Exon 21 L858R: A Retrospective Analysis in China

Introduction: The effect of icotinib on non-small cell lung cancer (NSCLC) patients with EGFR exon 19 deletions (19-Del) or L858R point mutation in exon 21 (21-L858R) remains inconsistent. This study aimed to evaluate the efficacy and safety of icotinib in patients with advanced NSCLC harboring these 2 EGFR mutations. Methods: We retrospectively assessed the clinical effects of first-line icotinib on advanced NSCLC patients with 2 classic EGFR mutations. Kinase activity assays were used to reaffirm the preclinical efficacy. Results: Among 2,757 patients, 2,365 (86%) harbored 19-Del (1,346/2,757, 49%) or 21-L858R (1,019/2,757, 37%) mutation. Patients with 19-Del had a higher response rate (ORR; 67.8 vs. 62.1%; p = 0.0039) and disease control rate (98.5 vs. 97.2%; p = 0.0223) than those with 21-L858R mutation. The median progression-free survival (PFS) in the 19-Del group (22.3 months, 95% confidence interval [CI]: 21.3–23.4) was significantly longer than that in the 21-L858R group (20.4 months, 95% CI: 19.5–21.7) (p = 0.004). In multivariate analysis, mutation types, clinical stage, and smoking history were significant factors for PFS. Additionally, an in vitro study indicated the 50% inhibitory concentrations (IC₅₀) of icotinib was lower for EGFR 19-Del than 21-L858R. Conclusion: These results suggest that EGFR 19-Del confers superior PFS and response to the icotinib treatment compared to 21-L858R.

Epidermal Growth Factor Receptor Gene Mutation Detection in Histology and Cytology Specimens of Primary Lung Adenocarcinoma: Immunohistochemistry Versus the Molecular Method.

Background:Epidermal growth factor receptor (EGFR) gene in lung adenocarcinoma is associated with good clinical response to EGFR-tyrosine kinase therapy. The two most common EGFR gene mutations, representing 80 to 90%, are the E746-A750 deletion in exon 19 and the L858R point mutation in exon 21. Materials and Methods:We have conducted the study to evaluate immunohistochemistry’s performance in detecting the E746-A750 deletion in exon 19 of the EGFR gene in primary lung adenocarcinoma cases. This study examined 133 cases of primary lung adenocarcinoma for three years duration. The selected cases were tested for EGFR gene mutations by real-time PCR by a reference laboratory. Most cases (124) were diagnosed by tissue biopsy, though nine used cell block cytology. We performed an immunohistochemistry test on 75 cases that contained adequate diagnostic material in the paraffin block. Results:The test result was scored as 0 to 3+, based on the staining intensity and percentage of positive tumor cells. We evaluated the immunohistochemistry test’s sensitivity and specificity compared to the EGFR gene mutations by real-time PCR. There was a significant association between gender, smoking status, and the EGFR gene mutations (P < 0.001). The overall sensitivity and specificity of the immunohistochemistry test were 40% and 100%, respectively. The positive predictive value and negative predictive values were 100% and 76.9%, each. Conclusions:The immunohistochemistry has high specificity but low sensitivity in the detection of E746-A750 deletion in exon 19 of the EGFR gene. The mutation-specific antibody used in this study was unable to detect other uncommon variants of exon 19 deletions. With high specificity value, immunohistochemistry may provide an adjunct to molecular testing for detecting the most common EGFR gene mutations in cases of a low cellularity sample, financially-limited situations, or in critically ill cases where urgent targeted therapy is needed.

Treatment of Brain Metastases of Non-Small Cell Lung Carcinoma.

Lung cancer is one of the most common malignant neoplasms. As a result of the disease’s progression, patients may develop metastases to the central nervous system. The prognosis in this location is unfavorable; untreated metastatic lesions may lead to death within one to two months. Existing therapies—neurosurgery and radiation therapy—do not improve the prognosis for every patient. The discovery of Epidermal Growth Factor Receptor (EGFR)—activating mutations and Anaplastic Lymphoma Kinase (ALK) rearrangements in patients with non-small cell lung adenocarcinoma has allowed for the introduction of small-molecule tyrosine kinase inhibitors to the treatment of advanced-stage patients. The Epidermal Growth Factor Receptor (EGFR) is a transmembrane protein with tyrosine kinase-dependent activity. EGFR is present in membranes of all epithelial cells. In physiological conditions, it plays an important role in the process of cell growth and proliferation. Binding the ligand to the EGFR causes its dimerization and the activation of the intracellular signaling cascade. Signal transduction involves the activation of MAPK, AKT, and JNK, resulting in DNA synthesis and cell proliferation. In cancer cells, binding the ligand to the EGFR also leads to its dimerization and transduction of the signal to the cell interior. It has been demonstrated that activating mutations in the gene for EGFR-exon19 (deletion), L858R point mutation in exon 21, and mutation in exon 20 results in cancer cell proliferation. Continuous stimulation of the receptor inhibits apoptosis, stimulates invasion, intensifies angiogenesis, and facilitates the formation of distant metastases. As a consequence, the cancer progresses. These activating gene mutations for the EGFR are present in 10–20% of lung adenocarcinomas. Approximately 3–7% of patients with lung adenocarcinoma have the echinoderm microtubule-associated protein-like 4 (EML4)/ALK fusion gene. The fusion of the two genes EML4 and ALK results in a fusion gene that activates the intracellular signaling pathway, stimulates the proliferation of tumor cells, and inhibits apoptosis. A new group of drugs—small-molecule tyrosine kinase inhibitors—has been developed; the first generation includes gefitinib and erlotinib and the ALK inhibitor crizotinib. These drugs reversibly block the EGFR by stopping the signal transmission to the cell. The second-generation tyrosine kinase inhibitor (TKI) afatinib or ALK inhibitor alectinib block the receptor irreversibly. Clinical trials with TKI in patients with non-small cell lung adenocarcinoma with central nervous system (CNS) metastases have shown prolonged, progression-free survival, a high percentage of objective responses, and improved quality of life. Resistance to treatment with this group of drugs emerging during TKI therapy is the basis for the detection of resistance mutations. The T790M mutation, present in exon 20 of the EGFR gene, is detected in patients treated with first- and second-generation TKI and is overcome by Osimertinib, a third-generation TKI. The I117N resistance mutation in patients with the ALK mutation treated with alectinib is overcome by ceritinib. In this way, sequential therapy ensures the continuity of treatment. In patients with CNS metastases, attempts are made to simultaneously administer radiation therapy and tyrosine kinase inhibitors. Patients with lung adenocarcinoma with CNS metastases, without activating EGFR mutation and without ALK rearrangement, benefit from immunotherapy. This therapeutic option blocks the PD-1 receptor on the surface of T or B lymphocytes or PD-L1 located on cancer cells with an applicable antibody. Based on clinical trials, pembrolizumab and all antibodies are included in the treatment of non-small cell lung carcinoma with CNS metastases.

Osimertinib, an Irreversible Next-Generation EGFR Tyrosine Kinase Inhibitor, Exerts Antitumor Activity in Various Preclinical NSCLC Models Harboring the Uncommon EGFR Mutations G719X or L861Q or S768I.

Osimertinib is an oral, third-generation, irreversible epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) that selectively inhibits both EGFR-TKI-sensitizing and EGFR T790M-resistance mutations with lower activity against wild-type EGFR and has demonstrated efficacy in non-small cell lung cancer (NSCLC) CNS metastases. The sensitizing mutations, the in-frame deletions in exon 19 and the L858R point mutation in exon 21, represent between 80% and 90% of all EGFR mutations. The remaining 10% to 20% are referred to as uncommon activating mutations and are a diverse group of mutations in exons 18 to 21 within the kinase domain of the EGFR gene. Excluding those found as insertion mutations in exon 20, the uncommon mutations involving codons G719, S768, and L861 are the most prevalent.Although the efficacy of EGFR-TKIs for the common EGFR mutations is well established, much less is known about rare EGFR mutations, such as exon 20 insertions, G719X, L861Q, S768I, as most of the data consist of single case reports or small case series.Using available patient-derived xenografts (PDX) and cell lines derived from two of these PDXs that harbor the G719X mutation, we have evaluated in vitro and in vivo the preclinical activity of osimertinib. We report osimertinib inhibits signaling pathways and cellular growth in G719X-mutant cell lines in vitro and demonstrate sustained tumor growth inhibition of PDX harboring the G719X mutation alone or in combination with L861Q and S768I.Together, these data support clinical testing of osimertinib in patients with uncommon EGFR NSCLC.

Impact of neutrophil-to-lymphocyte ratio in patients with EGFR-mutant NSCLC treated with tyrosine kinase inhibitors.

Background Exon 19 deletion and L858R point mutation in exon 21 of the epidermal growth factor receptor (EGFR) are the most commonly encountered mutations in patients with non-small cell lung cancer (NSCLC) and predict better clinical outcomes following treatment with EGFR-tyrosine kinase inhibitors (TKIs). The inflammatory indicator neutrophil-to-lymphocyte ratio (NLR) in peripheral blood serves as a predictive factor for NSCLC patients treated with chemotherapy. Here, we aimed to evaluate the correlation between NLR and clinical efficacy of EGFR-TKIs in NSCLC patients harboring EGFR mutations. Methods We retrospectively collected information of 205 patients with advanced NSCLC harboring exon 19 deletion or L858R point mutation and receiving gefitinib or erlotinib. The clinical outcomes in the NSCLC patients were evaluated based on NLR level before EGFR-TKI therapy. Results The optimal cut-off value for NLR was 3.55. The response rates in the low-NLR and high-NLR groups were 69.2% and 51.5%, respectively. The median progression-free survival (PFS) in the low-NLR and high-NLR groups were 15.7months and 6.7months, respectively. The median overall survival (OS) in the low-NLR and high-NLR groups were 37.6months and 19.2months, respectively. The multivariate analysis identified performance status (PS), NLR, stage, and smoking status as independent predictors of PFS. Moreover, the PS and NLR were identified as independent predictors of OS. Conclusions NLR was a significant predictor of clinical efficacy and OS in NSCLC patients harboring EGFR mutations treated with gefitinib or erlotinib.

Epidermal growth factor receptor mutation status in pulmonary adenocarcinoma: Multi-institutional data discussion at national conference of “Lung Cancer Management in Indian context”

The presence of activating mutations in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR) gene in adenocarcinomas of lung confers sensitivity to tyrosine kinase inhibitor therapy. The prevalence of EGFR mutations varies among different ethnicities and demographic profile. This multi-institutional data was compiled to determine the EGFR mutation status in Indians patients with lung adenocarcinoma. Cohorts represented by 4 tertiary care hospitals participated in data discussion at a national conference entitled as ‘Lung Cancer Management in Indian Context’. The clinicopathologic data and EGFR mutation rate in the patients of lung adenocarcinoma screened in these cohorts were collected and analyzed. The sample types included both surgical and cytological specimens. A variety of methods were used including immunohistochemistry, polymerase chain reaction, Sanger sequencing and next generation sequencing. A total of 3436 cases of treatment naïve lung adenocarcinoma were tested for EGFR mutations. The overall frequency of EGFR mutations observed was 30.03%. The most common baseline mutation detected was exon 19 deletion followed by L858R point mutation in exon 21. Dual mutations were observed in 6.5% of cases and were predominantly combinations of exon 19 deletion and T790M point mutation in exon 20. Incidence of EGFR mutations was higher among females and non-smokers diagnosed with lung adenocarcinomas. The most common histology in EGFR mutant cases was acinar predominant adenocarcinomas. With nearly one-third of Indian patients with lung adenocarcinoma harboring EGFR mutations, routine testing for these mutations is important to get the benefit of targeted therapy.

Abstract 1330: Osimertinib, an irreversible next generation EGFR tyrosine kinase inhibitor, exerts anti-tumor activity in various preclinical NSCLC models harboring G719X mutant-EGFR

Abstract Two first-generation (erlotinib &amp; gefitinib), two second generation (afatinib &amp; dacomitinib) and a third generation (osimertinib) EGFR-TKIs are currently available for the management of EGFR mutation-positive NSCLC. All these TKIs are effective in patients with NSCLC whose tumors harbor the in-frame deletions in exon 19 and the L858R point mutation in exon 21. These two mutations represent 90% of all EGFR mutations. Osimertinib, when used in the front-line setting, has shown more favorable treatment outcomes than first-generation EGFR-TKIs. In approximately 50% of patients, resistance to first and second generation TKI is mediated by the acquisition of the ‘gatekeeper’ mutation T790M. Currently, osimertinib is the only registered EGFR TKI that is active against exon 19 deletions and L858R mutation, regardless of the presence of T790M mutation. While the efficacy of EGFR TKIs for the common EGFR mutations is well established, much less is known about rare EGFR mutations such as exon 20 insertions, G719X, L861Q, S768I, as most of the data consist of single case reports or small case series. This work describes the therapeutic potential of osimertinib in tumors harboring the G719X mutation alone or in combination with L861Q and S768I. Using available patient-derived xenografts (PDX) and cell lines derived from two of these PDX which habor the G719X mutation, we have evaluated in vitro and in vivo the pre-clinical activity of osimertinib. We show that osimertinib inhibits signalling pathways and cellular growth of G719X cell lines in vitro. This translates into sustained tumor growth inhibition in vivo in 3 out of 4 PDX explored (87%, p&amp;lt;0.001, 158% p&amp;lt;0.001 &amp; 181%, p&amp;lt;0.001 respectively at day 14) when compared to the control group. Importantly, in these in vivo models, osimertinib achieved exposure consistent with those observed in patients treated with an 80 mg clinical dose. Moreover, osimertinib demonstrates superior activity to afatinib (145%, p&amp;lt;0.001, &amp; 58% p&amp;lt;0.01 respectively at day 14) in a model harboring the G719A. Out of the 4 PDX models explored in vivo, one model appeared to be inherently resistant to osimertinib. Further genomic characterisation revealed that the model displayed amplification of c-MET, a well know mechanism of resistance to EGFR-TKI. Subsequent in vivo pre-clinical study shows that the model is sensitive to c-MET inhibition. In addition, we performed pharmacodynamic studies to explore the relationship between efficacy and target/pathway modulation. These studies establish a clear relationship between depth and duration of inhibition of the phopshorylation of EGFR and anti-tumor efficacy. The work presented herein demonstrates that osimertinib has the potential to improve upon the current treatment options for NSCLC patients whose tumors harbor a G719X mutation, and warrants further clinical investigation. Citation Format: Nicolas Floch, Sue Bickerton, Matthew J. Martin, Darren A. Cross, Paul D. Smith. Osimertinib, an irreversible next generation EGFR tyrosine kinase inhibitor, exerts anti-tumor activity in various preclinical NSCLC models harboring G719X mutant-EGFR [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1330.

Apparent diffusion coefficient and genetic mutations as effective indexes for radiotherapy time of brain metastases from lung cancer retrospective analysis.

e20641 Background: The potential value of apparent diffusion coefficient (ADC) before chemoradiotherapy and genetic mutation as monitors for radiotherapy time of lung adenocarcinoma patients with brain metastases were retrospective analysed. Methods: Fifty-seven patients of lung adenocarcinoma with brain metastases were enrolled into the clinical trial. Conventional magnetic resonance imaging (MRI) examination and diffusion weighted imaging (DWI) were performed one week before therapy and after two cycles of chemotherapy or after one month targeted therapy. Symptoms such as headache, nausea, vomiting would be alleviated with mannitol and dexamethasone. Chemotherapy,once every 3 weeks, using in these patients with wild gene included pemetrexed, DDP, bevacizumab (pemetrexed 500mg per square meter ; DDP 75mg per square meter and bevacizumab 15mg per kilogram). 27 Patients with genetic mutations includ ed 1 patient with ALK genetic mutations, 2 patients with T790 genetic mutations, the presence of EGFR mutations was determined by the presence of deletions with exons 19 in 9 patients and 20 in 10 patients and L858R point mutations in exon 21 in 5 patients. Patients with genetic mutations were given targeted drug therapy. After these above treatment, if brain metastasis symptoms persist, brain radiotherapy was performed ahead of schedule. Patients with no obvious symptoms or relieved symptoms after targeted therapy or chemotherapy, two-month review was performed with enhanced MRI, radiotherapy was not added until the lesion no longer continues to shrink. Results: In the cohort of patients with wild gene , when ADC values less than 1.01 in 15 patients, the maximum diameter of tumors were significantly decreased after two treatment cycles, While 14 patients with ADC values greater than 1.01, the maximum diameter of the tumors are no difference after two treatment cycles. After targeted therapy for one month, In patients with ALK and T790 genetic mutations, the maximum diameter of tumors were significantly decreased, while no difference in 18 patients and decreased in 6 patients with presence of EGFR. Conclusions: ADC values and genetic mutation may as effective indexes of radiotherapy time for lung adenocarcinoma patients with brain metastases.

Breast cancer and synchronous multiple primary lung adenocarcinomas with heterogeneous mutations: a case report

BackgroundMultiple primary malignant tumors (MPMT) refers to the presence of two or more primary cancers of different organs in the same patient. MPMT is a sparse disease in the past, but there has been a gradual increase in the morbidity. Since multiple primary malignant tumors treatment methods differ, it is essential for clinicians to be able to distinguish between separate primary lesions and metastasis.Case presentationWe present the case of a 57-year-old woman with MPMT presenting with cancer in the left breast and synchronous double primary lung adenocarcinomas. We used IHC and epidermal growth factor receptor(EGFR)mutation to analyze genomic alteration profiles in the patient to validate the difference among the pathological assessments and the clinical differences between double primary lesions of lung and breast. EGFR gene analysis of breast cancer lesion revealed no mutations. The left and right lower lobe lung adenocarcinomas contained EGFR gene mutations: an L858R point mutation in exon 21 in the left lesion and a deletion mutation in exon 19 in the right lesion. The breast cancer and both lung adenocarcinomas were surgically resected. To date, the patient has remained disease-free.ConclusionsBoth pathological and molecular assessment adapted in the current study appeared necessary. Mutational analysis of the EGFR gene provided important information not only in the diagnosis and but also in the treatment of MPMT.

TAS6417, A Novel EGFR Inhibitor Targeting Exon 20 Insertion Mutations.

Activating mutations in the EGFR gene are important targets in cancer therapy because they are key drivers of non-small cell lung cancer (NSCLC). Although almost all common EGFR mutations, such as exon 19 deletions and the L858R point mutation in exon 21, are sensitive to EGFR-tyrosine kinase inhibitor (TKI) therapies, NSCLC driven by EGFR exon 20 insertion mutations is associated with poor clinical outcomes due to dose-limiting toxicity, demonstrating the need for a novel therapy. TAS6417 is a novel EGFR inhibitor that targets EGFR exon 20 insertion mutations while sparing wild-type (WT) EGFR. In cell viability assays using Ba/F3 cells engineered to express human EGFR, TAS6417 inhibited EGFR with various exon 20 insertion mutations more potently than it inhibited the WT. Western blot analysis revealed that TAS6417 inhibited EGFR phosphorylation and downstream molecules in NSCLC cell lines expressing EGFR exon 20 insertions, resulting in caspase activation. These characteristics led to marked tumor regression in vivo in both a genetically engineered model and in a patient-derived xenograft model. Furthermore, TAS6417 provided a survival benefit with good tolerability in a lung orthotopic implantation mouse model. These findings support the clinical evaluation of TAS6417 as an efficacious drug candidate for patients with NSCLC harboring EGFR exon 20 insertion mutations. Mol Cancer Ther; 17(8); 1648-58. ©2018 AACR.

First-generation EGFR tyrosine kinase inhibitor therapy in 106 patients with compound EGFR-mutated lung cancer: a single institution\u2019s clinical practice experience

BackgroundThe antitumour efficacy of tyrosine kinase inhibitors (TKIs) in lung cancer patients with compound epidermal growth factor receptor (EGFR) mutations has not been resolved. Our study summarizes a single institutional experience of first-generation TKI therapy for lung cancers with compound EGFR mutations.MethodsA total of 106 consecutive patients with tumours bearing compound EGFR mutations were identified between January 2012 and May 2016; all patients received first-generation TKI therapy. Deletions in exon 19 and the L858R point mutation in exon 21 were considered common mutations; T790M was considered separately because of its association with TKIs resistances. Any other mutation was defined as a rare mutation. Patients were divided as follows: double common mutations (group A); common plus T790M mutations (group B); common plus rare mutations (group C); double rare mutations (group D); and rare plus T790M mutations (group E). A separate group of 115 consecutive patients with a single common mutation was created for comparative analysis (group F).ResultsThe frequency of patients with compound EGFR was 2.9% (114/3925) and their response rate to first-generation TKIs was 50.9%, which was not significantly different from group F (67.0%, P = 0.088). The progression-free survival (PFS) of the 106 patients receiving TKI therapy was worse than that of group F (median, 9.1 vs. 13.0 months, respectively; P < 0.001). The PFS of the compound mutation group was shorter than that of the single common mutation group (median, 10.1 months in group A, P = 0.240; 9.1 months in group B, P < 0.001; 9.6 months in group C, P = 0.010; 6.5 months in group D, P = 0.048; 5.4 months in group E, P = 0.017). Patients with a co-occurring mutation in exon 20 (excluding T790M) exhibited significantly worse PFS than the patients with other compound mutations or with a single common mutation (median, 6.5 vs. 9.1 vs. 13.0 months, respectively, P = 0.002).ConclusionsThere was significant heterogeneity among the compound EGFR mutations and their response to first-generation TKIs. Individualized treatment in clinical practice should be considered for each case.

Losmapimod Overcomes Gefitinib Resistance in Non-small Cell Lung Cancer by Preventing Tetraploidization

The epidermal growth factor receptor (EGFR) is known to play a critical role in non-small cell lung cancer (NSCLC). Constitutively active EGFR mutations, including in-frame deletion in exon 19 and L858R point mutation in exon 21, contribute about 90% of all EGFR-activating mutations in NSCLC. Although oral EGFR-tyrosine kinase inhibitors (TKIs), gefitinib and erlotinib, show dramatic clinical efficacy with significantly prolonged progression-free survival in patients harboring these EGFR-activating mutations, most of these patients will eventually develop acquired resistance. Researchers have recently named genomic instability as one of the hallmarks of cancer. Genomic instability usually involves a transient phase of polyploidization, in particular tetraploidization. Tetraploid cells can undergo asymmetric cell division or chromosome loss, leading to tumor heterogeneity and multidrug resistance. Therefore, identification of signaling pathways involved in tetraploidization is crucial in overcoming drug resistance. In our present study, we found that gefitinib could activate YAP-MKK3/6-p38 MAPK-STAT3 signaling and induce tetraploidization in gefitinib-resistance cells. Using p38 MAPK inhibitors, SB203580 and losmapimod, we could eliminate gefitinib-induced tetraploidization and overcome gefitinib-resistance. In addition, shRNA approach to knockdown p38α MAPK could prevent tetraploidy formation and showed significant inhibition of cancer cell growth. Finally, in an in vivo study, losmapimod could successfully overcome gefitinib resistance using an in-house established patient-derived xenograft (PDX) mouse model. Overall, these findings suggest that losmapimod could be a potential clinical agent to overcome gefitinib resistance in NSCLC.

P1.02-036 An EGFR Tyrosine Kinase Inhibitor Sensitive Patient-Derived Lung Cancer Xenograft Model without Classical Sensitizing Mutations

Mutations in the tyrosine kinase (TK) domain of EGFR are oncogenic driver in 10-20% of lung adenocarcinoma (AdC) patients in Western countries. Approximately 90% of EGFR-TK inhibitor (TKI) sensitizing mutations occur as small in-frame deletions in exon 19 or L858R point mutations in exon 21. Recently, novel driver mutations in EGFR with oncogenic and TKI sensitizing activity have been reported. We present here an AdC patient-derived xenograft (PDX) model (PDX12) that is highly sensitive to EGFR-TKI, yet failed to demonstrate classical TKI sensitizing mechanisms. Comprehensive genomics profiling was used to characterize the genotype of PDX12, which was established from a resected stage IIIA AdC patient grafted in NGS mouse. The primary human lung cancer cell line (PHLC12) was extracted from its PDX model (PDX12). Aberrant EGFR cell lines used were H3255 (L858R), H2935 (exon 19 deletion), H1975 (L858R and T790M), and H1944 (wild type). Cell viability was assessed after erlotinib treatment at 1nM - 2μM for 72 hours using MTS assay. Levels of EGFR activation in both pre- and post-treatment by Western blot analysis. PDX12 model had no known oncogenic mutations (EGFR wild type) on exons 18-21 by next-generation sequencing, RT-qPCR, and SISH, but was highly sensitive to EGFR-TKI. The IC50 to erlotinib treatment at 72 hr was 67.13 ± 7.63 nM for PHLC12, compared to 9.70 ± 2.64 nM for H3255, 64.88 ± 8.49 nM for HCC2935, > 2 μM for H1975, and > 2 μM for H1944 EGFR mutant or wild type cells, respectively. Western blot analysis demonstrated a relatively higher molecular weight band for EGFR protein with high expression level in PHLC12 when compared to other lung cancer cell lines. Using RT-qPCR, relative expression level of each EGFR domain (extracellular, tyrosine kinase, and c-terminal domain) in PHLC12 showed no difference compared to EGFR wild type. Phosphorylation status of EGFR in PHLC12 was similar in activity as compared to erlotinib sensitive cell lines. PHLC12 represents an enigmatic EGFR TKI sensitive lung PDX model without classical TKI sensitizing aberrations. Additional potential mechanisms of EGFR dependency including exon duplication, or post-translational modification of EGFR protein are being investigated.

P3.02b-016 An Exploration Study of Mechanisms Underlying Primary Resistance to EGFR-TKIs in Patients Harboring TKI-Sensitive EGFR Mutations: Topic: EGFR Biomarkers

Primary resistance to EGFR-TKIs was generally defined as disease progression in less than 3 months without any evidence of objective response. Although possible mechanisms have been investigated in several preclinical and retrospective studies, little is known about the molecular backgrounds of primary resistance. Random Sample of Cases was used to screen TKI-sensitive patients to match with the primary resistant patients on the basis of clinical characteristics (smoking history, EGFR mutations etc.). DNA was extracted from the tumor and their matched normal material. The paired-end whole exome sequencing (WES) of DNA was performed on the Illumina HiSeq 2500 sequencing platform. Totally, five patients exhibiting primary resistance to EGFR-TKIs were enrolled and each was randomly matched with one patient possessing TKI sensitivity (Table 1). The mean depth of the WES was 100x. The mean number of nonsynonymous SNV per sample was 195 (range 97 to 348) in TKI-resistant group versus 84 (range 60 to 101) in TKI-sensitive group (P=0.059). Consistent with the initial result of Sanger sequencing, all 10 patients were found with EGFR sensitizing mutations (exon 19 deletions or L858R point mutation in exon 21), but no T790M mutation; the resistance group present with a lower EGFR mutant allele frequency than the sensitive group. Next generation sequencing of TKI-resistant specimens detected KRAS amplification (CNtumor / CNnormal = 2.6) in one of five patients, and MET amplification (CNtumor / CNnormal = 2.3) in another one. The recurrent mutation genes included FAT4, RBM10, TANC2, ACAN, PPFIA2, UBR4, XIRP2 and PRAMEF1. Next-generation sequencing offers more complete genomic analysis to understand the mechanism of differential responses to EGFR-TKIs, which can lead to more precision therapy. KRAS amplification appears to be a newly mechanism underlying primary resistance to EGFR-TKIs in patients harboring TKI-sensitive EGFR mutations. However, it needs to be validated in a larger cohort.

Final overall survival in JO22903, a phase II, open-label study of first-line erlotinib for Japanese patients with EGFR mutation-positive non-small-cell lung cancer

BackgroundIn Japan, the clinical efficacy of erlotinib monotherapy in epidermal growth factor receptor (EGFR) mutation-positive non-small-cell lung cancer was demonstrated in the phase II JO22903 trial, which reported a median progression-free survival of 11.8 months. Here we report final overall survival data from JO22903.MethodsJO22903 (JapicCTI-101085) was a single-arm, multicenter, phase II, open-label, non-randomized study of first-line erlotinib monotherapy in EGFR mutation-positive non-small-cell lung cancer. Eligible patients (≥20 years) with stage IIIB/IV or recurrent non-small-cell lung cancer and confirmed activating mutations of EGFR (exon 19 deletion or L858R point mutation in exon 21) received oral erlotinib 150 mg/day until disease progression or unacceptable toxicity. The primary endpoints were progression-free survival and safety; overall survival was a secondary endpoint.ResultsAt the final analysis, 102 patients were included in the modified intent-to-treat population and 103 in the safety population. Median follow-up was 32.3 months. Median overall survival was 36.3 months (95 % confidence interval 29.4–not reached). Subgroup analyses of overall survival suggested that the presence of brain metastases was a negative prognostic factor (median overall survival 22.7 months, 95 % confidence interval 19.6–29.4). The impact on overall survival of using versus not using EGFR tyrosine kinase inhibitors in any line of treatment following disease progression was unclear (median 32.8 versus 36.3 months, respectively). No new safety issues were observed.ConclusionIn this survival update, single-agent erlotinib achieved a median overall survival of more than 3 years in patients with EGFR mutation-positive non-small-cell lung cancer.

Standardized uptake value on 18F-FDG-PET/CT is a predictor of EGFR T790M mutation status in patients with acquired resistance to EGFR-TKIs

BackgroundThe development of epidermal growth factor receptor (EGFR) T790M point mutation in exon 20 (T790M) is the most common mechanism of resistance to EGFR tyrosine kinase inhibitors (EGFR-TKIs). The purpose of this study was to determine the association of 18F-2-fluoro-2-deoxyglucose-positron emission tomography/computed tomography (FDG–PET/CT) metrics with T790M status after acquiring resitance to EGFR-TKI resistance. MethodsWe retrospectively reviewed 34 advanced non-small cell lung cancer (NSCLC) patients harboring EGFR mutation who underwent rebiopsy and FDG–PET/CT before rebiopsy. These patients were evaluated for baseline characteristics, initial response to EGFR–TKIs, site of rebiopsy, overall survival, and FDG–PET/CT metrics, such as standardized uptake value (SUV), metabolic tumor volume, and total lesion glycolysis of the rebiopsy site. ResultsThe median age was 67 years (range, 37–86 years); 19 of the patients (56%) were men. Histologic examination revealed adenocarcinoma in all but one patient, with 20 patients (59%) having stage IIIB and IV disease. Upon initial mutational analyses, the types of EGFR mutation included 17 (50%) deletions in exon 19 and 17 (50%) L858R point mutations in exon 21. At the time of acquired resistance to EGFR-TKIs, T790M mutation was identified in 20 (59%) patients. T790M-positive patients had significantly lower levels of median SUVmean and median SUVmax of the rebiopsy site on FDG–PET/CT, compared with T790M-negative patients (SUVmean: 4.57 vs. 9.91, P=0.0069; SUVmax: 7.26 vs. 16.06, P=0.0054). The survival in patients who acquired T790M after failure of EGFR-TKIs was significantly longer than those without T790M (10.2 mos. vs. 18.6 mos., P<0.05). ConclusionT790M status was associated with lower levels of SUVmean and SUVmax on FDG–PET/CT and significantly longer survival.