Growth Factor Receptor Mutations Research Articles

Effects of reduced platelet count on the prognosis for patients with non-small cell lung cancer treated with EGFR-TKI: a retrospective study

BackgroundProgressive lung cancer is associated with abnormal coagulation. Platelets play a vital part in evading immune surveillance and angiogenesis in the case of tumor metastasis. The study aimed to analyze the predictive and prognostic effects of platelet count on non-small cell lung cancer (NSCLC) patients treated with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs).MethodsThis study retrospectively analyzed the prognostic effects of platelets on 52 NSCLC patients with epidermal growth factor receptor (EGFR) mutant following EGFR-TKI treatment. Related data, together with the progression-free survival (PFS) and overall survival (OS) were collected before and after 2 cycles of treatments (60 days).ResultsThe anti-EGFR treatment markedly reduced the platelet count in 33 (63.5%) patients after 2 cycles of treatment. Multivariate Cox analysis revealed that, the decreased platelet count was closely correlated with the longer OS (HR = 0.293; 95%CI: 0.107-0.799; p = 0.017). Besides, the median OS was 326 days in the decreased platelet count group and 241 days in the increased platelet count group (HR = 0.311; 95%CI: 0.118-0.818; P = 0.018), as obtained from the independent baseline platelet levels and other clinical features.ConclusionsThe platelet count may predict the prognosis for EGFR-TKI treatment without additional costs. Besides, changes in platelet count may serve as a meaningful parameter to establish the prognostic model for NSCLC patients receiving anti-EGFR targeted therapy.

Predictive and Prognostic Potential of TP53 in Patients With Advanced Non–Small-Cell Lung Cancer Treated With EGFR-TKI: Analysis of a Phase III Randomized Clinical Trial (CTONG 0901)

BackgroundMutations in TP53 are commonly found in patients with epidermal growth factor receptor (EGFR) mutant advanced non–small-cell lung cancer (NSCLC). In this study, we determined the predictive and prognostic potential of different subtypes of TP53 using data from a phase III randomized trial (CTONG 0901). Patients and MethodsThe trial enrolled 195 patients who had undergone next-generation sequencing of 168 genes before treatment with EGFR tyrosine kinase inhibitors. Mutations in TP53 (exon 4 or 7, other mutations, and wild-type) were analyzed based on the therapeutic response and survival. A Cox proportional hazards model was used to determine the potential of the predictive and prognostic factors. ResultsAll 195 patients harbored activating EGFR mutations: the most common concomitant mutations were TP53 (134/195, 68.7%), CTNNB1 (20/195, 10.3%), and RB1 (16/195, 8.2%). The genetic profiles between patient subgroups administered first-line (132, 67.7%) or later-line (63, 32.3%) treatments did not significantly differ. The median progression-free survival in patients with mutations in exon 4 or 7 of TP53, other TP53 mutations, and wild-type TP53 were 9.4, 11.0, and 14.5 months (P = .009), respectively. Overall survival times were 15.8, 20.0, and 26.1 months (P = .004), respectively. Mutations in exon 4 or 7 of TP53 served as independent prognostic factors for progression-free (P = .001) and overall survival (P = .004) in patients. ConclusionMutations in exon 4 and/or 7 in TP53 are promising predictive and prognostic indicators in EGFR-mutated NSCLC.

Survival outcomes and symptomatic central nervous system (CNS) metastasis in EGFR-mutant advanced non-small cell lung cancer without baseline CNS metastasis: Osimertinib vs. first-generation EGFR tyrosine kinase inhibitors

ObjectivesCentral nervous system (CNS) metastases are common complications in epidermal growth factor receptor (EGFR) mutant non-small-cell lung cancer (NSCLC) treated with EGFR-tyrosine kinase inhibitors (TKIs). However, for patients without baseline CNS metastasis, data regarding the incidence of symptomatic CNS metastasis with EGFR-TKI treatment and its risk factors are still rare. Materials and MethodsPatients with EGFR-mutant advanced NSCLC without baseline CNS metastasis who are receiving first- and/or third-generation EGFR-TKIs were included. Overall survival (OS), cumulative incidence of symptomatic CNS metastasis upon treatment failure, and their risk factors were evaluated. ResultsThere were 813 patients enrolled, with 562, 106, and 32 received first-line gefitinib, erlotinib, and osimertinib, respectively, while 113 received second-line osimertinib. At a median follow-up of 18.1 months, the median OS was 45.5 months. There were 38 patients developed symptomatic CNS metastases. Osimertinib-treated patients tended to have a lower risk of CNS metastases compared with those treated with first-generation EGFR-TKIs (p = 0.059). However, the cumulative incidence curves of symptomatic CNS metastasis tended to reach a plateau after approximately 3 years regardless of which generation was used, and incidences beyond that period were similar in the two groups. Patients with L858R mutation exhibited a higher risk of developing CNS metastasis than patients with 19del mutation (p = 0.001). Interestingly, the presence of baseline neuroimaging was not associated with the risk of developing CNS metastasis or OS. ConclusionCompared with first-generation EGFR-TKIs, osimertinib can delay but not prevent the development of symptomatic CNS metastasis. L858R mutation is an independent risk factor for CNS metastasis.

Immunotherapy in EGFR-Mutant and ALK-Positive Lung Cancer: Implications for Oncogene-Driven Lung Cancer.

Non-small cell lung cancer (NSCLC) is a heterogeneous disease, commonly defined by genetic alterations in oncogenic drivers. Targeted therapies have transformed the management of oncogene-driven lung cancers, with targeted agents now approved in the United States for 7 distinct molecular alterations. Nonetheless, acquired resistance remains an ongoing challenge, underscoring the need for alternative therapeutic approaches. Immune checkpoint inhibitors targeting the programmed cell death 1 (PD-1) axis have emerged as important therapies in the management of advanced NSCLC, but the role of these agents in patients with oncogenic driver mutations remains unclear. Here, we focus on epidermal growth factor receptor-mutant and anaplastic lymphoma kinase-rearranged NSCLC as paradigms to explore the role of immune checkpoint inhibitors in oncogene-driven NSCLC. We provide an overview of the clinical data examining programmed death ligand 1 (PD-L1) inhibitor monotherapy, PD-(L)1 inhibitors, and tyrosine kinase inhibitor combinations, as well as combinations of PD-(L)1 inhibitors and chemotherapy.

Superior efficacy of immunotherapy-based combinations over monotherapy for EGFR-mutant non-small cell lung cancer acquired resistance to EGFR-TKIs.

BackgroundWhile prospective clinical studies on immunotherapy in epidermal growth factor receptor (EGFR) mutant non‐small‐cell lung cancer (NSCLC) with acquired resistance to EGFR tyrosine kinase inhibitors (TKIs) are ongoing, this study aimed to investigate the outcomes of immunotherapy combinations in such a population in a real‐world setting.MethodsThe clinical data of pretreated EGFR‐mutated NSCLC patients who acquired EGFR‐TKI resistance and received immunotherapy were retrospectively analyzed in this study. Progression‐free survival (PFS) was assessed using the Kaplan‐Meier log‐rank test, and univariate and multivariate analysis were performed.ResultsA total of 31 patients were analyzed in this study. A total of 25 (80.6%) patients received combination immunotherapy. In the univariate analysis, patients who received combination immunotherapy seemingly acquired longer PFS than those who received monotherapy, although there was no significant difference (3.42 months vs. 1.61; P = 0.078; hazard ratio (HR) 0.43, 95% CI: 0.16–1.13). Patients who received antiangiogenic drugs prior to immunotherapy acquired better PFS (3.42 months vs. 1.58; P = 0.027; HR 0.37, 95% CI: 0.15–0.93), while patients with liver metastasis had inferior PFS (2.04 months vs. 3.42; P = 0.031; HR 2.83, 95% CI: 1.05–7.60). Furthermore, multivariate analysis confirmed that the above three factors had independent prognostic value.ConclusionsThe study revealed that immunotherapy combinations are better choices than single‐agent regimens in previously treated and EGFR‐mutant NSCLC patients with progressive disease. In addition, antiangiogenic drugs administered before immunotherapy might be a favorable prognostic factor, while liver metastasis was associated with a short PFS in this setting. In future, more robust and prospective clinical trial results are expected to guide clinical practice.Key pointsSignificant study findings Immunotherapy‐based combination therapies are better choices than single‐agent regimens in heavily treated EGFR‐mutant NSCLC patients. What this study adds Patients without liver metastasis and with prior antiangiogenic drugs obtained more benefit from immunotherapy in this setting.

Classical versus non-classical EGFR mutations: Erlotinib response and impact of renal insufficiency.

Erlotinib is an effective treatment option for EGFR-mutant non-small cell lung cancer. It is important to predict patients who will respond better to erlotinib. We designed this study to investigate the effect of renal insufficiency (RI) on erlotinib treatment outcomes. All patients receiving erlotinib were stratified into 3 groups. Group 1 consisted of non-RI subjects with classical epidermal growth factor receptor (EGFR) mutations, Group 2 consisted of those with RI (Estimated glomerular filtration rate <60 mL/min) and classical EGFR mutations, and Group 3 consisted of those with non-classical EGFR mutations. 82 patients were included in the study. Median progression-free survival (PFS) in patients with classical mutation was approximately 6 months shorter in those with RI, although not statistically significant. Median overall survival (OS) in Group 1, 2 and 3 was 34.1 months, 35.2 months, and 15 months, respectively and although not statistically significant, median OS was 20 months shorter in Group 3. Univariate and multivariate cox-regression analysis revealed shorter PFS and OS in males and those with ECOG ≥2 while PFS and OS were longer in those with recurrent lung tumors and generating rash during erlotinib treatment. There was no difference between RI and non-RI patients in terms of adverse events except for fatigue and appetite loss. This research showed OS in patients with and without RI was comparable. Although not statistically significant, PFS in patients with classical mutation was approximately 6 months shorter in those with RI patients.

Treatment and Outcomes of Metastatic Non-Small-Cell Lung Cancer Harboring Uncommon EGFR Mutations: Are They Different from Those with Common EGFR Mutations?

Simple SummaryThe present study showed the comprehensive analysis of disease characteristics and treatment patterns in uncommon EGFR mutation-positive NSCLC at a major cancer center. This study showed the efficacy of 1G or 2G EGFR-TKIs as the 1L treatment, and subsequent therapy including 3G EGFR-TKIs in the real-world setting.Approximately 10% of the epidermal growth factor receptor (EGFR) mutations in non-small-cell lung cancer (NSCLC) are uncommon EGFR mutations. Although the efficacy of second (2G) or third generation (3G) EGFR tyrosine kinase inhibitors (EGFR-TKIs) in the patients with uncommon EGFR mutation has been proven, further studies are warranted to define the optimal treatment approach for uncommon EGFR mutation-positive NSCLC. This study retrospectively investigated the treatment patterns and outcomes of patients with uncommon EGFR mutation-positive NSCLC from January 2011 to December 2019 at the Samsung Medical Center, Seoul, Korea. During the study, 2121 patients with EGFR mutation-positive NSCLC received first-generation (1G, gefitinib or erlotinib) or 2G EGFR-TKI (afatinib) as the first-line (1L) systemic therapy. Of this, 135 (6.4%) patients harbored uncommon EGFR mutations. Of 135, 54 (40%, 54/135) patients had overlapping mutations with major EGFR mutations. The objective response rate (ORR) for the 1L EGFR-TKI was 63.3%. The median progression-free survivals (PFSs) were 8.6 months (95% CI: 3.8–13.5), 11.7 months (95% CI: 6.6–16.7), 7.7 months (95% CI: 4.9–17.4), and 5.0 months (95% CI: 3.7–6.1) for major uncommon EGFR mutation (G719X, L861Q), compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), other compound mutation, and other uncommon mutations, respectively. The median overall survivals (OSs) were 25.6 months (16.9–34.2), 28.8 (95% CI: 24.4–33.4), 13.5 months (95% CI: 7.4–27.8), and 9.4 months (95% CI: 3.4–10.5) for major uncommon EGFR mutation (G719X), compound mutation with major EGFR mutation (Del 19 or EGFR exon 21 p.L858R), other compound mutation, and other uncommon mutations, respectively. The response rate, median PFS, and OS were 63.3%, 16.3 months (95% CI: 15.6–16.9), and 37.5 months (95% CI: 35.4–39.6) for common EGFR mutation-positive NSCLC. After failing 1L EGFR-TKI, repeated tissue or liquid biopsy were carried out on 44.9% (35/78) of patients with T790M detected in 10/35 (28.6%) patients. With subsequent 3G EGFR-TKI after failing the first-line EGFR-TKI, the ORR and PFS for 3G EGFR-TKI were 80% and 8.9 months (95% CI: 8.0–9.8). These patients showed a median OS of 34.6 months (95% CI: 29.8–39.4). The ORR, PFS and OS were poorer in patients with uncommon (especially other compound and other uncommon mutation) than those with common EGFR mutations. T790M was detected in 28.6% of the uncommon EGFR mutation-positive patients for whom prior 1G/2G EGFR-TKIs failed and underwent repeat biopsy at the time of progression.

Abstract IA18: The PI3K isoforms in myeloid leukemia and hematopoietic stem cells

Abstract The Class IA PI3 kinase isoforms (p110α, β, and δ) transduce many growth factor signals that are important for the proliferation, differentiation, and self-renewal of hematopoietic stem cells (HSCs). Mutations in growth factor receptors or RAS proteins are commonly observed in patients with acute myeloid leukemia (AML), leading to activation of the PI3K/AKT pathway. Therefore, PI3K inhibition is an attractive therapeutic strategy for a large subset of AML patients. We previously reported that p110α is dispensable for HSC function, suggesting that redundancy exists between the Class IA isoforms in HSCs (Gritsman et al., J Clin Invest 2014). However, we have identified a specific role for p110α in RAS-mutated myeloid leukemia. Furthermore, we found that the p110α-selective inhibitor BYL-719 can sensitize RAS-mutated leukemic cells to the MEK inhibitor MEK-162. While PI3K inhibitors have multiple clinical indications, including in hematologic malignancies, it is still unclear whether PI3K plays an important role in normal HSC function. Upon environmental stresses like chemotherapy or infection, quiescent HSCs must enter the cell cycle to re-establish homeostasis and promote emergency myelopoiesis. To test for redundancy between p110α and p110δ in HSCs, we generated p110α and p110δ double knockout (DKO) mice, with germline deletion of p110δ and conditional deletion of p110α. DKO mice have leukopenia, anemia, and decreased numbers of lymphoid-primed multipotent progenitors (LMPPs) in the bone marrow, though HSC numbers are unchanged. In competitive repopulation assays, B-cell maintenance was severely impaired, while the myeloid and T-cell lineages were relatively preserved. This suggests that HSC function is not impaired at steady state. We performed transcriptome analysis of DKO, p110δ, and p110α KO HSCs and LMPPs after bone marrow transplantation. Gene set enrichment analysis revealed downregulation of gene sets associated with cell cycle progression. Interestingly, we also observed negative enrichment of inflammatory response gene sets in both p110δ KO and DKO HSCs and LMPPs. To examine the roles of p110δ and α in the hematopoietic stress response, we injected p110δ, DKO, and WT mice with 5-fluorouracil. We observed significantly decreased survival of 5-FU-treated DKO mice due to impaired hematopoietic recovery, associated with the failure of HSCs to enter the cell cycle. Consistent with this finding, we also observed a decrease in myeloid reconstitution by DKO HSCs in secondary competitive transplantation, suggesting that p110α and p110δ play redundant roles in emergency myelopoiesis. We also observed defects in downstream signaling in DKO HSCs and progenitors in response to IL1β or TNFα stimulation. Our results suggest that p110α and δ act in a redundant fashion to transduce specific inflammatory signals in HSPCs in response to hematopoietic stress. Our results have important implications for the use of PI3K inhibitors in combination with chemotherapy and in other hematopoietic stress conditions. Citation Format: Shayda Hemmati, Taneisha Sinclair, Meng Tong, Boris Bartholdy, Rachel Okabe, Kristina Ames, Leanne Ostrodka, Tamanna Haque, Imit Kaur, Anupriya Agarwal, Jean Zhao, Thomas Roberts, Kira Gritsman. The PI3K isoforms in myeloid leukemia and hematopoietic stem cells [abstract]. In: Proceedings of the AACR Special Conference on Targeting PI3K/mTOR Signaling; 2018 Nov 30-Dec 8; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(10_Suppl):Abstract nr IA18.

Cost-effectiveness analysis of osimertinib for first-line treatment of locally advanced or metastatic EGFR mutation positive non-small cell lung cancer in Singapore

Objective Non-small cell lung cancer (NSCLC) accounts for 80–90% of all lung cancer cases and is usually associated with a poor prognosis. However, targeted therapy with first and second generation tyrosine kinase inhibitors (TKIs) has so far improved progression-free survival of epidermal growth factor receptor (EGFR) mutant NSCLC patients. Osimertinib, a third generation EGFR TKI has recently shown improved overall survival of 6.8 months in previously untreated EGFR mutant NSCLC patients. We assessed the cost-effectiveness of osimertinib versus standard EGFR TKIs (gefitinib or erlotinib) as first-line treatment for advanced or metastatic EGFR mutant NSCLC patients in Singapore. Methods A partitioned survival model with three health states (progression-free, progressive disease, and death) was developed from the Singapore healthcare payer perspective. Survival curves based on the overall trial population from the FLAURA trial were extrapolated beyond trial period over a 10-year time horizon to estimate the underlying progression-free survival and overall survival parametric distributions. Health state utilities were derived from the literature and direct costs were sourced from public healthcare institutions in Singapore. Deterministic and probabilistic sensitivity analyses were conducted to explore the impact of uncertainties and assumptions on cost-effectiveness results. Results Compared with first or second generation TKI, osimertinib had a base-case incremental cost-effectiveness ratio (ICER) of SG$418,839 (US$304,277) per quality-adjusted life year gained. One-way sensitivity analysis showed the ICER was most sensitive to time horizon and variations in progression-free utility values. Scenario analyses showed that a 50% reduction in the cost of osimertinib was still associated with a high ICER that was unlikely to be deemed cost effective. Conclusions Osimertinib is not cost effective as a first-line treatment compared to standard EGFR TKIs in advanced EGFR mutant NSCLC patients in Singapore. The findings from our evaluation, alongside other considerations including the lack of survival benefit in the Asian subgroup of the FLAURA trial, will be useful to inform policy makers on funding decisions for NSCLC treatments in Singapore.

Poor prognosis of NSCLC located in lower lobe is partly mediated by lower frequency of EGFR mutations

It is controversial whether a tumor located in the lower lobe is related with worse outcome of non-small cell lung cancer (NSCLC). This study aimed to clarify the prognostic role of primary tumor location in NSCLC. Patients newly diagnosed with NSCLC in a tertiary referral hospital from January 2011 to December 2014 were followed up for 5 years. Of the 2,289 NSCLC cases, 911 (39.8%) cases pertained to lower lobe cancers. Patients with lower lobe cancer showed a higher all-cause mortality rate than those with non-lower lobe cancer (48.6% vs. 40.3%, p < 0.001). Patients with lower lobe cancer had a lower proportion of adenocarcinoma histology and epidermal growth factor receptor (EGFR) mutations. Furthermore, compared to patients with non-lower lobe cancer, those with lower lobe cancer had a higher level of tumor markers (neuron-specific enolase and cytokeratin fragment 21-1). Mediation analysis revealed that the association between lower lobe cancer and higher all-cause mortality could be explained by an indirect pathway through EGFR mutations (percent mediated = 17.3%, p = 0.005). The sensitivity analysis for adenocarcinoma patients showed similar results (percent mediated = 18.8%, p = 0.021). Lower lobe cancer is associated with a higher all-cause mortality risk in patients with NSCLC, which is partly mediated by a lower proportion of EGFR mutations.

ORAL AND NAIL BED ULCERS CAUSED BY AFATINIB (MONOCLONAL ANTIBODY) IN LUNG CANCER

Medicated oral ulcers are often observed in patients who use antineoplastic drugs. Afatinib is an orally administered, irreversible inhibitor of the ErbB family of tyrosine kinases. It is the first-line treatment of patients with non–small cell lung cancer (NSCLC) with activating epidermal growth factor receptor (EGFR) mutations. During the treatment, some adverse effects such as epistaxis, diarrhea, nausea, vomiting, aphthous stomatitis, rash, acneiform dermatitis, pruritus, and xeroderma can be observed. We present a case of a 63-year-old female patient with advanced EGFR-mutated NSCLC and started treatment with oral afatinib 40 mg daily. On clinical examination there were atopic lesions located on the mucosa of the soft palate, lips, and tongue as well as involvement of the nails. This case demonstrates aphthous stomatitis as an adverse effect of antineoplastic medication that should be placed as a differential diagnosis in patients who use afatinib.

PLCγ1‑dependent invasion and migration of cells expressing NSCLC‑associated EGFR mutants.

The increased tyrosine kinase activity of non‑small cell lung cancer (NSCLC)‑associated epidermal growth factor receptor (EGFR) mutants results in deregulated pathways that contribute to malignant cell survival, tumor progression and metastasis. Previous studies investigating lung cancer‑associated EGFR have focused on the prognostic implications of receptor kinase mutations in patients with NSCLC; however, the role of EGFR mutations in tumor cell invasion and migration remains undetermined. The present study was designed to investigate the role of NSCLC‑associated mutant EGFR‑driven signaling pathways in cell proliferation and invasion. Non‑endogenous EGFR‑expressing 293cells stably expressing EGFR mutants that are sensitive or resistant to Food and Drug Administration(FDA)‑approved EGFR‑targeted tyrosine kinase inhibitors(TKIs) were used in the present study. The experiments demonstrated an increased phosphorylation of phospholipase (PLC)γ1, c‑Cbl, signal transducer and activator of transcription(Stat), extracellular regulated kinase (Erk)1/2, Akt, Shc and Gab1 proteins in cells expressing a mutant form, rather than the wild‑type receptor. As PLCγ1 is a known regulator of metastatic development, mutant receptor‑mediated PLCγ1 activation was further evaluated. To examine the effects of EGFR and PLCγ1 phosphorylation, the metastatic potential of cells expressing mutants was investigated using wound healing, Transwell cell migration and invasion assays. The inhibition of receptor phosphorylation with the 1st, 2nd and 3rdgeneration TKIs, gefitinib, afatinib, osimertinib, respectively, reduced PLCγ1 phosphorylation, and reduced the invasive and migratory potential of 293cells, confirming PLCγ1 as one of the probable downstream effectors of mutant EGFR signaling. However, the PLC inhibitor, U73122, inhibited cell migration and invasion without affecting EGFR signaling and PLCγ1 phosphorylation. Notably, U73122 reduced Akt and Erk1/2 phosphorylation within 25min of its application; however, 100%cell viability was recorded even after 48h. Upon further investigation, proliferative signaling pathways remained active at 48h, in accordance with cell viability. Therefore, the present study concludes that mutant receptor‑mediated PLCγ1 activation may play a significant role in the migration and invasion of NSCLC tumors; however, its regulatory role in tumor cell proliferation warrants further investigation and validation in lung tumor cell lines harboring EGFR mutations.

Abstract 697: Improvement in quality and length of life following treatment with mifepristone in women with stage IV non-small cell lung cancer, positive for the epidermal growth factor receptor (EGFR) mutation, that previously progressed on targeted therapy

Abstract Treatment with the oral single agent progesterone receptor modulator, mifepristone, has been demonstrated to significantly increase the quality and length of life in a patient with stage IV non-small cell lung cancer (NSCLC) that progressed despite multiple rounds of chemotherapy, when no markers existed for specific targeted therapy. The purpose of the present study was to determine if mifepristone could similarly provide palliative benefits to patients with NSCLC, that were positive for EGFR mutation, who had progressed despite targeted therapies. Patient 1, a 59 year old woman presented with 6 months of upper back pain and dyspnea, was found to have innumerable tiny lung nodules (miliary pattern), bone metastases, and a single brain metastasis (left occipital lobe). The tumor was an adenocarcinoma, with an EGFR exon leu858 arg identified. The brain metastasis was treated with palliative radiation therapy and erlotinib was started. She remained stable for 1 ½ years, but then her lung nodules started to increase in size. Erolotinib was stopped, and osimertinib was started. However, she progressed 6 months later, with continued growth of pulmonary nodules bilaterally and a new brain metastasis, in the right lateral cuneus. Her dyspnea, which had improved on erlotinib, returned to pre-treatment status. She was started on oral single agent mifepristone, 200mg per day. Patient 2, a 46 year old woman, had presented with headaches, difficulty with executive function, and memory loss. She was diagnosed with stage IV NSCLC, with extensive lung lesions and brain metastases. A MRI of the brain revealed innumerable supratentorial rim enhancing lesions, with one dominant left parietal lobe lesion measuring 3 × 2.1cm (treated with palliative radiation). CT scans showed innumerable bilateral pulmonary nodules with evidence of lymphangitic spread and multiple bone lesions. Because the tumor was positive for the EGFR1 exon 19 mutations (E746/751 mutation), she was started on afatinib, which was stopped 8 months later. She then had a second course of brain radiation and 5 months after stopping afatinib she began osimertinib. After 10 months of osimertinib, it seemed to be losing effectiveness, and so 200mg oral mifepristone was added. Both women have had stable disease, with no growth of previous nodules and no new nodules, or new brain metastases after 6 months and 14 months of mifepristone therapy, respectively. More importantly they both feel much better, with no dyspnea, or pain, and remain ECOG-0. Thus, similar to small cell lung cancer and NSCLC devoid of tumor markers, or with the PD-L1 mutation, patients with advanced NSCLC with EGFR mutation also seem to respond to the progesterone receptor modulator, mifepristone. Citation Format: Jerome H. Check, Diane Check, Trina Poretta, James Aikins, Carrie Wilson. Improvement in quality and length of life following treatment with mifepristone in women with stage IV non-small cell lung cancer, positive for the epidermal growth factor receptor (EGFR) mutation, that previously progressed on targeted therapy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 697.

Abstract 4852: CBLC functions as a novel therapeutic target to enhance the efficacy of paclitaxel on EGFR wild-type lung adenocarcinoma by downregulating AURKA

Abstract Patients with lung adenocarcinoma harboring driver genes can be treated with target therapies, such as tyrosine kinase inhibitors for epithelial growth factor receptor (EGFR) mutations, anaplastic lymphoma kinase (ALK) inhibitors for ALK gene rearrangements, and so on. However, a large portion of lung adenocarcinoma patients, especially for those with wild-type EGFR, still do not have valid target therapies in clinics. The CBL family proteins (CBL, CBLB, and CBLC) are E3 ubiquitin ligases of protein tyrosine kinases (PTKs), including EGFR, hepatocyte growth factor receptor (MET), and SRC proto-oncogene (SRC). It has been reported that the K63-linkaged ubiquitination mediated by CBL can negatively regulate activated EGFR (aEGFR) signaling through lysosomal degradation. Previously, we discovered that CBLC, unlike CBL, possesses an oncogenic function and positively regulates aEGFR stability through polyubiquitination via K6 and K11 linkages. The CBLC-mediated conjugation of polyubiquitin promotes aEGFR preferentially recycled back to the plasma membrane, or trafficked to the cell nucleus, leading to the sustained activation of EGFR signaling in lung adenocarcinoma. Aside from aEGFR dysregulation, we recently found that CBLC assumes a novel role in positively regulating the expression of Aurora kinase A (AURKA), which contributes to tumor development by promoting cell proliferation, metastasis, epithelial-mesenchymal transition, and drug resistance. In lung adenocarcinoma cells with wild-type EGFR, CBLC knockdown significantly reduced cell viability, delayed cell cycle progression and increased apoptotic death. Our study also showed that CBLC knockdown in EGFR wild type cells was associated with increased sensitivity to paclitaxel, probably through downregulating AURKA. Taken together, these findings indicated that CBLC may play a role in promoting cell cycle progression and may be a novel therapeutic target to enhance the efficacy of paclitaxel on EGFR wild-type lung adenocarcinoma. Citation Format: Shiao-Ya Hong, Yu-Rung Kao, Yi-Ping Lin, Meng-Hsuan Lee, Jyun-Yi Wu, Cheng-Wen Wu. CBLC functions as a novel therapeutic target to enhance the efficacy of paclitaxel on EGFR wild-type lung adenocarcinoma by downregulating AURKA [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4852.

Abstract 6645: Anti-EGF antibodies significantly improve the activity of RET, BRAF, MEK and PI3K kinase inhibitors in preclinical models

Abstract Background: We have recently shown that antibodies generated by vaccination (anti-EGF VacAbs) potentiate the effects of tyrosine kinase inhibitors (TKIs) in epidermal growth factor receptor mutant (EGFR-mut) cell lines (1) and a Phase I/II clinical trial of an anti-EGF vaccine in combination with afatinib has been initiated. In this study we aimed to determine the efficacy of anti-EGF VacAbs to improve the antitumor activity of RET, BRAF, MEK and PI3K inhibitors in non-small cell lung cancer (NSCLC) and colorectal cancer (CRC) cell lines. Methods: Cell lines with RET translocations (LC-2/ad) and BRAF, KRAS and PIK3CA mutations (HT29, DLD1, LS174T and H508) were used. Anti-EGF VacAbs were obtained by immunizing rabbits with recombinant human EGF. Cell lines were treated with anti-EGF VacAbs alone and in combination with RET, BRAF, MEK and PI3K inhibitors. Cell viability was determined by MTT, cell cycle was analyzed by flow cytometry, changes of total and phosphorylated proteins by Western blot and emergence of resistance by direct microscopic examination in low density cultures. Results: Anti-EGF VacAbs suppressed EGF-induced cell proliferation and blocked EGFR transduction signaling pathway in all cell lines tested. In combination, the anti-EGF VacAbs significantly enhanced the antitumor activity of BLU667 in LC-2/ad, trametinib and encorafenib in HT29, trametinib in DLD1 and LS174T and trametinib, taselisib, alpelisib and copanlisib in H508 cells. In these cell lines, anti-EGF VacAbs in combination with kinase inhibitors suppressed EGFR, Akt and Erk 1/2 phosphorylation. Cell cycle experiments revealed that anti-EGF VacAbs significantly increased the antiproliferative effects of the kinase inhibitors, measured as the percentage of cells in G2+M. Finally, the addition of the anti-EGF VacAbs to the culture medium significantly delayed the emergence resistant clones to trametinib in DLD1 cells. Conclusions: Anti-EGF VacAbs potentiate the antitumor effects of RET, MEK, BRAF and PI3K inhibitors in tumor cell lines. Our data provide a rationale for clinical trials testing the combination of anti-EGF VacAbs with kinase inhibitors in RET-translocated, KRAS, BRAF and PIK3CA mutant NSCLC and CRC patients.(1)“Anti-Epidermal Growth Factor Vaccine Antibodies Enhance the Efficacy of Tyrosine Kinase Inhibitors and Delay the Emergence of Resistance in EGFR Mutant Lung Cancer Cells” Codony-Servat J, García-Roman S, Molina-Vila MA, et al. J Thorac Oncol. 2018. Citation Format: Silvia Garcia Roman, Jordi Codony Servat, Miguel Angel Molina Vila, Jordi Bertran Alamillo, Monica Garzon, Alejandro Martínez Bueno, Santiago Viteri, María González Cao, Andrés Aguilar, Juan José García, Erik d'Hondt, Rafael Rosell. Anti-EGF antibodies significantly improve the activity of RET, BRAF, MEK and PI3K kinase inhibitors in preclinical models [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6645.

Simultaneous Single Cell Gene Expression and EGFR Mutation Analysis of Circulating Tumor Cells Reveals Distinct Phenotypes in NSCLC.

While cancer cell populations are known to be highly heterogeneous within a tumor, the current gold standard of tumor profiling is through a tumor biopsy. These biopsies are invasive and prone to missing these clones due to spatial heterogeneity, and this bulk analysis approach can miss information from rare subpopulations. To noninvasively investigate tumor cell heterogeneity, a streamlined workflow is developed to scrutinize rare cells, such as circulating tumor cells (CTCs), for simultaneous analysis of mutations and gene expression profiles at the single cell level. This powerful workflow overcomes low-input limitations of single cell analysis techniques. The utility of this multiplexed workflow to unravel inter- and intra-patient heterogeneity is demonstrated using non-small-cell lung cancer (NSCLC) CTCs (n = 58) from six epidermal growth factor receptor (EGFR) mutant positive NSCLC patients. CTCs are isolated using a high-throughput microfluidic technology, the Labyrinth, and their EGFR mutation status and gene expression profiles are characterized.

Non-small cell lung cancer patients with ex19del or exon 21 L858R mutation: distinct mechanisms, different efficacies to treatments.

With the development of antitumor therapies, different treatment methods including monotherapy and combined therapy have achieved clinical efficacy in advanced epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC) patients. Exon 19 deletion (ex19del) and exon 21 L858R mutation are common sensitive subtypes of EGFR mutation. However, potential distinct mechanisms are found from several dimensions including molecular structures, biological behaviors, concomitant mutations, resistance mechanisms and tumor mutation burdens. More evidence indicates the prognostic difference of EGFR subgroups. This review focused on the progress of potential distinct mechanisms and outcomes in clinical trials of advanced NSCLC patients with ex19del or exon 21 L858R mutation.

Abstract B36: Identifying single-cell gene expression and EGFR mutation profile heterogeneity in NSCLC patients’ CTCs

Abstract The efficacy of targeted therapies was first shown in patients with non-small cell lung cancer (NSCLC) who possess activating epidermal growth factor receptor (EGFR) mutations that responded to tyrosine kinase inhibitors (TKIs). Current standard of care is to screen metastatic patients for the presence of EGFR mutations to determine if they qualify for TKI treatment. Due to the invasiveness of a tumor biopsy, this characterization is typically only done at the time of diagnosis. This single time-point determines the entire treatment plan. Despite strong initial response to TKI therapy, most patients develop resistance within months, most commonly by acquiring a secondary EGFR mutation. Identifying resistance through current monitoring techniques, such as measuring tumor volume, can lead to a delay in detection. Alternatively, circulating tumor cells (CTCs) are cancer cells present in the blood that provide access to tumor cells without the requirement of a biopsy. Shed from the tumor into the vasculature, CTCs circulate throughout the body before a fraction extravasate, leading to metastatic sites. CTCs have been shown to carry tumor-matched characteristics in both genotype and phenotype. Due to their ease of access, they can be used to serially track patients’ conditions to detect the early emergence of new tumor clones and, potentially, therapeutic resistance. One of the major limitations of CTCs’ clinical utility is their low abundance in the blood. Our group previously developed the Labyrinth, a high-throughput, label-free microfluidic technology, which enables rapid and efficient CTC enrichment. These CTCs are used for enumeration or single-cell transcriptomic analysis. Initial single-cell analysis was performed through highly multiplexed RT-qPCR that compared 96-gene expression profiles to unravel inter- and intrapatient CTC heterogeneity. We recently developed a complimentary single-cell analysis approach using digital PCR (dPCR) to detect the presence of sensitizing EGFR mutations, exon 19 deletion and L858R, and resistance mutation, T790M. dPCR is an ultrasensitive approach with single-molecule resolution, which enables detection and quantification at a single-cell level by partitioning the sample into individual PCR reaction droplets. This resolution can distinguish if a cell is homo- or heterozygous for a mutation and each allele’s relative expression level. We validated this system for single-cell analysis using lung cancer cell lines and demonstrated its agreement with qPCR results. We present initial findings of EGFR mutation screening in CTCs from a small cohort of NSCLC patients with known EGFR status. We highlight the single-cell heterogeneity of CTCs within a patient at both the gene expression and EGFR mutation level. By tracking these patients’ CTCs over time, we may be able to identify the emergence of resistant clones sooner than can be done in the clinic. This may allow for more rapid treatment modification and lead to improved patient outcomes. Citation Format: Sarah Owen, Ting-Wen Lo, Shamileh Fouladdel, Mina Zeinali, Evan Keller, Ebrahim Azizi, Nithya Ramnath, Sunitha Nagrath. Identifying single-cell gene expression and EGFR mutation profile heterogeneity in NSCLC patients’ CTCs [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B36.

Evaluation of the fully automated Idylla EGFR Mutation Assay in Chinese patients with lung adenocarcinoma.

e21716 Background: Epidermal growth factor receptor ( EGFR) mutation testing is nowadays an essential part of lung adenocarcinoma diagnosis. Its rapid detection is of utmost significance in order to ensure that patients receive timely and appropriate treatment. However, the current techniques for EGFR examination in China, ARMS-PCR and NGS, often take 3~7 days to deliver the final results. In contrast, the fully automated Idylla EGFR assay could detect 51 EGFR mutations directly from formalin-fixed, paraffin-embedded (FFPE) samples within 2.5 hours with &lt; 2 minutes of hands-on time. The test has proven both accurate and robust among Caucasian lung adenocarcinoma patients but has never been clinically validated in Chinese patients. Methods: FFPE samples were retrospectively collected from 94 lung adenocarcinoma patients followed by EGFR testing using the Idylla system. Seventy-eight samples were also previously assessed with ARMS-PCR, 9 with NGS and 7 with both. In case of discordance, repeat testing was performed on the same tissue block with a third method. The sensitivity and specificity of the Idylla EGFR assay were evaluated against ARMS-PCR or NGS. Mutations beyond the scope of detection by the Idylla EGFR assay were not included in the analyses. Results: Of the 94 tumors enrolled, 77 were stage I-II. The Idylla system was consistent with ARMS-PCR or NGS for the EGFR mutational profiles of 92 tumors: 80 positive and 12 negative. Among the 2 cases of discrepancy, one was wild-type using Idylla but were found to harbor G719X/S768I using ARMS-PCR (and confirmed by NGS), while the other one carried L858R according to Idylla, as confirmed by ARMS-PCR, but was discovered to have an additional mutation L861Q using NGS. The Idylla EGFR test had a sensitivity of 97.6%, a specificity of 100% and an overall concordance of 97.9%. Conclusions: The Idylla system provides a rapid, accurate and user-friendly solution to EGFR characterization, especially in time-sensitive scenarios where special expertise and staff training could be a challenge.

CONCORDANCE study: An observational, multicenter, prospective study to evaluate concordance of detecting EGFR mutations by circulating tumor-free DNA versus tissues biopsy in NSCLC.

e21615 Background: Epidermal growth factor receptor (EGFR) mutations on circulating tumour free DNA (ctDNA) by liquid biopsy is suitable option in those with difficulty in obtaining tissue samples. Correlation in tissue and plasma results of EFGRm has not been established in the Indian population. 1,2,3 This study was done to investigate the utility of ctDNA for EGFRm testing with Next-Generation Sequencing (NGS) in a real-world diagnostic setting. Methods: This is a multicentre, prospective, diagnostic observational study in 245 newly diagnosed treatment naïve, histologically confirmed, advanced lung adenocarcinoma patients. This was a single visit study. The primary objective of the study was to determine the level of concordance between EGFR mutation status obtained by tissue and ctDNA from blood (plasma) based testing in terms of overall concordance, sensitivity &amp; specificity. Study was registered at Clinicaltrials.gov NCT03562819 &amp; CTRI/2018/08/015290. Results: Seventy-five (30.6%) and eighty-four (34.3%) subjects showed positive mutation status by plasma &amp; tissue testing respectively. The overall concordance of 82.9% was observed between tissue and ctDNA (Plasma) based testing. Sensitivity of EGFR mutation status between ctDNA and tissue was observed for EGFR mutation subtypes was observed to be 100% while specificity was observed to be 90.1%. Plasma and tissue sample testing detected 1.2% (n = 3) and 2.4% (n = 6) positive in exon 20 T790M EGFR mutation, respectively. In terms of secondary outcomes, plasma sample testing detected 16.7% (n = 41) positive for Exon 19 deletions type EGFR mutation followed Exon 21 L858R [11.4% (n = 28)]. Conclusions: CONCORDANCE, a real-world study in Indian patients suggest that ctDNA testing for EGFR mutation analysis is a diagnostic option in newly diagnosed lung adenocarcinoma patients. EGFR Mutation testing in ctDNA, being non-invasive and especially in patients without available/evaluable tumor sample may enable more patients to receive appropriate targeted therapies. [Table: see text]