Jacqulyne P. Robichaux

Mechanisms and clinical activity of an EGFR and HER2 exon 20–selective kinase inhibitor in non–small cell lung cancer

Although most activating mutations of epidermal growth factor receptor (EGFR)-mutant non–small cell lung cancers (NSCLCs) are sensitive to available EGFR tyrosine kinase inhibitors (TKIs), a subset with alterations in exon 20 of EGFR and HER2 are intrinsically resistant and lack an effective therapy. We used in silico, in vitro, and in vivo testing to model structural alterations induced by exon 20 mutations and to identify effective inhibitors. 3D modeling indicated alterations restricted the size of the drug-binding pocket, limiting the binding of large, rigid inhibitors. We found that poziotinib, owing to its small size and flexibility, can circumvent these steric changes and is a potent inhibitor of the most common EGFR and HER2 exon 20 mutants. Poziotinib demonstrated greater activity than approved EGFR TKIs in vitro and in patient-derived xenograft models of EGFR or HER2 exon 20 mutant NSCLC and in genetically engineered mouse models of NSCLC. In a phase 2 trial, the first 11 patients with NSCLC with EGFR exon 20 mutations receiving poziotinib had a confirmed objective response rate of 64%. These data identify poziotinib as a potent, clinically active inhibitor of EGFR and HER2 exon 20 mutations and illuminate the molecular features of TKIs that may circumvent steric changes induced by these mutations.Poziotinib is a candidate inhibitor for a subset of EGFR or HER2 mutant non–small cell lung cancers that lack effective therapy.

Local Consolidation Therapy (LCT) After First Line Tyrosine Kinase Inhibitor (TKI) for Patients With EGFR Mutant Metastatic Non–small-cell Lung Cancer (NSCLC)

Introduction: Although most NSCLC patients with sensitizing epidermal growth factor receptor (EGFR) mutations have an impressive initial response, the vast majority has residual disease and develops acquired resistance after 9 to 14 months of EGFR tyrosine kinase (TKI) therapy. We recently reported a phase II trial showing that, for patients with molecularly unselected oligometastatic NSCLC who did not progress after first‐line systemic therapy, local consolidation therapy (LCT) with surgery or radiation improved progression‐free survival (PFS), compared with maintenance therapy alone. Herein, we report a retrospective analysis of LCT after TKI in patients with metastatic EGFR mutant NSCLC. Patients and Methods: We identified patients with metastatic EGFR mutant NSCLC treated with TKI plus LCT or with TKI alone in the MD Anderson GEMINI (Genomic Marker‐Guided Therapy Initiative) database and in our recently published LCT trial. PFS was compared between LCT plus TKI and TKI only treated patients using the log‐rank test. Results: We identified 129 patients with EGFR mutant NSCLC who were treated with first‐line TKI and 12 that were treated with TKI followed by LCT. Among the 12 patients treated with TKI plus LCT, 8 patients had oligometastatic disease (defined as ≤ 3 metastases), and 4 patients had > 3 metastases. LCT regimens were hypofractionated radiotherapy or stereotactic ablative body radiotherapy for 11 patients and surgery for 1 patient. TKI followed by LCT resulted in a significantly longer PFS (36 months) compared with TKI alone (PFS, 14 months; log‐rank P = .0024). Conclusions: Our data suggests that first‐line TKI plus LCT is a promising therapeutic strategy for patients with EGFR mutant NSCLC that merits further investigation.

Abstract 2061: Non-covalent EGFR T790M targeting TKIs inhibit AZD9291 resistant EGFR C797S mutants

Approximately 10-15% of non-small-cell lung cancers (NSCLC) have epidermal growth factor receptor (EGFR) mutations resulting in increased sensitivity to 1 st generation tyrosine kinase inhibitors (TKIs) such as gefitinib and erlotinib. For common mutations in EGFR, treatment with 1 st generation TKIs results in approximately 70% overall response rate, increased progression free survival, and increased quality of life compared to chemotherapy alone. However, resistance to 1 st generation TKIs typically develops within ~12 months, and approximately 55% of patients acquire a secondary mutation in EGFR Exon 20, T790M. To overcome resistance, 2 nd and 3 rd generation covalently binding TKIs targeting T790M mutations have been developed. Recent studies show that approximately 40% of acquired resistance to 2 nd and 3 rd generation TKIs can also occur via a third acquired EGFR mutation at the site of covalent binding, C797S. To date, there are no standard approved targeted therapies for treating EGFR C797S mutant NSCLC. Moreover, with proposed increased use of covalent inhibitors in the first line setting, C797S mutations are expected to become more prevalent and new strategies to overcome therapeutic resistance will be required. To this end, we have generated stable Ba/F3 and HCC827 NSCLC cell lines expressing C797S mutant EGF receptors with common mutations in EGFR including, L858R/T790M/C797S, and Ex19del/T790M/C797S. EGFR mutant cell lines expressing C797S were screened against 1 st , 2 nd , and 3 rd generation EGFR TKIs and cell viability was determined using Cell Titer Glo. Triple mutant cell lines containing T790M and C797S mutations were not sensitive to any 1 st , 2 nd , or 3 rd generation inhibitors with IC 50 values of >10µM, 7.0µM, and 7.6µM, respectively. However Ba/F3 and HCC827 cell lines transfected with EGFR triple mutants were inhibited by non-covalent inhibitors: CUDC-101, an EGFR, HER2 and HDAC inhibitor, and PKC412, a FLT3 inhibitor. In triple mutant Ba/F3 cells, CUDC-101 and PKC412 had IC 50 values of 470nM and 250nM in Ex19del/T790M/C797S EGFR mutants, respectively, and IC 50 values of 690nM and 270nM in L858R/T790M/C797S EGFR mutants. In HCC827 cells, PKC412 had IC50 values of 610nM and 520nM in L858R/T790M/C797S and Ex19del/T790M/C797S triple mutants, respectively. In addition, western blot analysis of triple mutant Ba/F3 cells showed decreased phosphorylation of EGFR in presence of 50nM PKC412 and 500nM of CUDC-101. Moreover, PKC412 partially decreased p-EGFR expression at 50nM in HCC827 triple mutant cell lines. In conclusion, unlike other EGFR TKIs, the non-covalent EGFR inhibitors CUDC-101 and PKC412 inhibit both T790M and C797S EGFR mutants at low concentrations. Currently, we are establishing additional osimertinib resistant HCC827, H1975, PC9 and H4006 cell lines for further in vitro and in vivo studies. Citation Format: Jacqulyne P. Robichaux, Monique Nilsson, John V. Heymach. Non-covalent EGFR T790M targeting TKIs inhibit AZD9291 resistant EGFR C797S mutants [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2061. doi:10.1158/1538-7445.AM2017-2061

Abstract 4799: Inhibition of HER2 mutant non-small cell lung cancer using 3rd generation EGFR/HER2 inhibitors

HER2 is modified in approximately 20% of lung cancers and is mutated in approximately 2-3% of non-small cell lung cancer (NSCLC). Several HER2 mutations within the tyrosine kinase domain are activating mutations, and irreversible EGFR TKIs such as afatinib are known to be effective at inhibiting HER2 phosphorylation in tumor cells expressing HER2 activating mutations. However, the clinical use of these second generation irreversible EGFR inhibitors, such as afatinib, has been limited by adverse toxicities such as skin rash and diarrhea. Our recent study in lung cancer showed that ibrutinib, a well-tolerated TKI currently FDA approved for B-cell lymphoma, is capable of inhibiting EGFR. Study by others also demonstrated that ibrutinib can inhibit HER2 phosphorylation in HER2 overexpressing breast cancer cell lines. In addition, recent studies in other cancers show that the effects of ibrutinib can be amplified in combination of with other small molecule inhibitors targeting AKT and mTOR. Therefore, we hypothesized that HER2 overexpression or activating mutations render NSCLC tumor cells sensitive to ibrutinib. To investigate this, we generated a panel of Ba/F3 cell lines expressing twelve individual clinically observed HER2 mutations and evaluated the transforming capability of the mutations as indicated by sustained cell viability following IL-3 deprivation. Common HER2 mutations such as A775insYVMA, G776del/insVV, and G776C V777insC as well as others were observed to be activating mutations. Mutant HER2 expressing Ba/F3 cells and the human HER2 mutant NSCLC cell line, H1781, were then screened against first, second, and third generation EGFR/HER2 inhibitors including ibrutinib, and cell viability was determined by the Cell Titer Glo assay. Drugs effective in growth inhibition were verified by Western blot analysis. The results showed that EGFR TKI erlotinib and the EGFR/HER2 TKI lapatinib failed to inhibit cell proliferation. However, ibrutinib and third generation EGFR/HER2 TKIs, EGF816 and AZD9291, inhibited cell proliferation and induced apoptosis at sub-micromolar concentrations. Western blotting analysis showed dose dependent decreases in phosphorylation of HER2 as well as HER2 downstream signaling molecules such as p-AKT, and p-MAPK. In order to determine if targeting downstream effectors of HER2 in combination with inhibition of the receptor would increase inhibition of cell proliferation and induction of apoptosis, we screened ibrutinib, EGF816 and AZD9291 in combination with both mTOR and AKT inhibitors. There was a modest additive effect when ibrutinib was used in combination with everolimus, an mTOR inhibitor, but there was no additive effect of EGF816 or AZD9291 was used in combination with everolimus. These results indicate that EGF816, AZD9291, or ibrutinib alone or in combination with mTOR inhibition may be effective therapeutic strategies for the treatment of HER2-driven NSCLC. Citation Format: Jacqulyne P. Robichaux, Monique Nilsson, Bingliang Fang, John V. Heymach. Inhibition of HER2 mutant non-small cell lung cancer using 3rd generation EGFR/HER2 inhibitors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4799.