Dual MET and ERBB inhibition overcomes intratumor plasticity in osimertinib-resistant-advanced non-small-cell lung cancer (NSCLC).

Third-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) such as osimertinib are the last line of targeted treatment of metastatic non-small-cell lung cancer (NSCLC) EGFR-mutant harboring T790M. Different mechanisms of acquired resistance to third-generation EGFR-TKIs have been proposed. It is therefore crucial to identify new and effective strategies to overcome successive acquired mechanisms of resistance. For Amplicon-seq analysis, samples from the index patient (primary and metastasis lesions at different timepoints) as well as the patient-derived orthotopic xenograft tumors corresponding to the different treatment arms were used. All samples were formalin-fixed paraffin-embedded, selected and evaluated by a pathologist. For droplet digital PCR, 20 patients diagnosed with NSCLC at baseline or progression to different lines of TKI therapies were selected. Formalin-fixed paraffin-embedded blocks corresponding to either primary tumor or metastasis specimens were used for analysis. For single-cell analysis, orthotopically grown metastases were dissected from the brain of an athymic nu/nu mouse and cryopreserved at -80°C. In a brain metastasis lesion from a NSCLC patient presenting an EGFR T790M mutation, we detected MET gene amplification after prolonged treatment with osimertinib. Importantly, the combination of capmatinib (c-MET inhibitor) and afatinib (ErbB-1/2/4 inhibitor) completely suppressed tumor growth in mice orthotopically injected with cells derived from this brain metastasis. In those mice treated with capmatinib or afatinib as monotherapy, we observed the emergence of KRAS G12C clones. Single-cell gene expression analyses also revealed intratumor heterogeneity, indicating the presence of a KRAS-driven subclone. We also detected low-frequent KRAS G12C alleles in patients treated with various EGFR-TKIs. Acquired resistance to subsequent EGFR-TKI treatment lines in EGFR-mutant lung cancer patients may induce genetic plasticity. We assess the biological insights of tumor heterogeneity in an osimertinib-resistant tumor with acquired MET-amplification and propose new treatment strategies in this situation.

Martinez-Marti A ，Felip E ，Matito J ，Mereu E ，Navarro A ，Cedrés S ，Pardo N ，Martinez de Castro A ，Remon J ，Miquel JM ，Guillaumet-Adkins A ，Nadal E ，Rodriguez-Esteban G ，Arqués O ，Fasani R ，Nuciforo P ，Heyn H ，Villanueva A ，Palmer HG ，Vivancos A ... -  《-》

Third generation EGFR inhibitor osimertinib combined with pemetrexed or cisplatin exerts long-lasting anti-tumor effect in EGFR-mutated pre-clinical models of NSCLC.

La Monica S ，Minari R ，Cretella D ，Flammini L ，Fumarola C ，Bonelli M ，Cavazzoni A ，Digiacomo G ，Galetti M ，Madeddu D ，Falco A ，Lagrasta CA ，Squadrilli A ，Barocelli E ，Romanel A ，Quaini F ，Petronini PG ，Tiseo M ，Alfieri R ... -  《JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH》

Preclinical Comparison of Osimertinib with Other EGFR-TKIs in EGFR-Mutant NSCLC Brain Metastases Models, and Early Evidence of Clinical Brain Metastases Activity.

Approximately one-third of patients with non-small cell lung cancer (NSCLC) harboring tumors with EGFR-tyrosine kinase inhibitor (TKI)-sensitizing mutations (EGFRm) experience disease progression during treatment due to brain metastases. Despite anecdotal reports of EGFR-TKIs providing benefit in some patients with EGFRm NSCLC brain metastases, there is a clinical need for novel EGFR-TKIs with improved efficacy against brain lesions. We performed preclinical assessments of brain penetration and activity of osimertinib (AZD9291), an oral, potent, irreversible EGFR-TKI selective for EGFRm and T790M resistance mutations, and other EGFR-TKIs in various animal models of EGFR-mutant NSCLC brain metastases. We also present case reports of previously treated patients with EGFRm-advanced NSCLC and brain metastases who received osimertinib in the phase I/II AURA study (NCT01802632). Osimertinib demonstrated greater penetration of the mouse blood-brain barrier than gefitinib, rociletinib (CO-1686), or afatinib, and at clinically relevant doses induced sustained tumor regression in an EGFRm PC9 mouse brain metastases model; rociletinib did not achieve tumor regression. Under positron emission tomography micro-dosing conditions, [11C]osimertinib showed markedly greater exposure in the cynomolgus monkey brain than [11C]rociletinib and [11C]gefitinib. Early clinical evidence of osimertinib activity in previously treated patients with EGFRm-advanced NSCLC and brain metastases is also reported. Osimertinib may represent a clinically significant treatment option for patients with EGFRm NSCLC and brain metastases. Further investigation of osimertinib in this patient population is ongoing. Clin Cancer Res; 22(20); 5130-40. ©2016 AACR.

Ballard P ，Yates JW ，Yang Z ，Kim DW ，Yang JC ，Cantarini M ，Pickup K ，Jordan A ，Hickey M ，Grist M ，Box M ，Johnström P ，Varnäs K ，Malmquist J ，Thress KS ，Jänne PA ，Cross D ... -  《-》

Audit of Molecular Mechanisms of Primary and Secondary Resistance to Various Generations of Tyrosine Kinase Inhibitors in Known Epidermal Growth Factor Receptor-Mutant Non-small Cell Lung Cancer Patients in a Tertiary Centre.

Presently, three generations of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are approved against oncogene addicted EGFR-mutant non-small cell lung cancer (NSCLC). Patients with actionable EGFR mutations invariably develop resistance. This resistance can be intrinsic (primary) or acquired (secondary). This was a retrospective study carried out between January 2016 and April 2021 analysing 486 samples of NSCLC for primary and secondary resistance to first- (erlotinib, gefitinb), second- (afatinib) and/or third-generation (osimertinib) TKIs in EGFR-mutant NSCLCs by next generation sequencing (NGS). Tissue NGS was carried out using the Thermofischer Ion Torrent™ Oncomine™ Focus 52 gene assay; liquid biopsy NGS was carried out using the Oncomine Lung Cell-Free Total Nucleic Acid assay. All cases were previously tested for a single EGFR gene with the Therascreen® EGFR RGQ PCR kit. The results were divided into four groups: (i) group 1: primary resistance to first- and/or second-generation TKIs. This group, with 21 cases, showed EGFR exon 20 insertions, dual, complex mutations and variant of unknown significance, de novo MET gene amplification besides other mutations. (ii) Group 2: primary resistance to third-generation TKIs. This group showed two cases, with one showing dual EGFR mutation (L858R and E709A) and EGFR gene amplification. (iii) Group 3: secondary resistance to first- and second-generation TKIs. This group had 27 cases, which were previously reported negative for EGFR T790M by single gene testing. Significant findings were MET gene amplification in four cases, with one also showing MET exon 14 skipping mutation. Three cases showed small cell change and one showed loss of primary mutation. (iv) Group 4: secondary resistance to third-generation TKIs. The latter group was further subgrouped into group 4A: secondary resistance to osimertinib (third-generation TKI) when offered as second-line therapy after first- and second-generation TKIs on detection of T790M mutation. This group had 15 cases. EGFR T790M mutation was lost in 10 (10/15; 67%) cases and was retained in five cases. Patients with T790M loss experienced early resistance (6.9 months versus 12.6 months mean, P = 0.0024) compared with cases that retained T790M. Two cases gained MET amplification as the resistance mechanisms. Other mutations that were found when EGFR T790M was lost were in FGFR3, KRAS, PIK3CA, CTNNB1, BRAF genes. One case had EML4-ALK translocation. Two cases showed driver EGFR deletion 19, retained T790M and C797S mutation in Cis form. Group 4B: secondary resistance to osimertinib (when given as first-line therapy) in EGFR-mutant NSCLC. This group had three cases. The duration of osimertinib treatment ranged from 11 to 17 months. Two patients showed additional C797S mutation along with primary EGFR mutation. This study shows the wide spectrum of primary and secondary EGFR resistance mechanisms to first, second and third generation of TKIs and helps us to identify newer therapeutic targets that could carry forward the initial advantage offered by EGFR TKIs.

Suryavanshi M ，Jaipuria J ，Mattoo S ，Dhandha S ，Khatri M ... -  《-》

ERK inhibition effectively overcomes acquired resistance of epidermal growth factor receptor-mutant non-small cell lung cancer cells to osimertinib.

Osimertinib (AZD9291), a third-generation, mutation-selective epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (EGFR-TKI), is an approved drug for patients who have non-small cell lung cancer (NSCLC) with activating EGFR mutations or those harboring a resistant T790M mutation. Unfortunately, all patients eventually relapse and develop resistance to osimertinib. The current study addressed whether ERK inhibition exerts effects similar to those produced by MEK inhibition in overcoming acquired resistance to osimertinib. Drug effects on cell and tumor growth were assessed by measuring cell number alterations and colony formation in vitro and with xenografts in nude mice in vivo. Apoptosis was assessed with annexin V/flow cytometry and protein cleavage. Protein alterations in cells were detected with Western blot analysis. Gene overexpression and knockout were achieved with lentiviral infection and CRISPR/Cas9, respectively. The combination of osimertinib with an ERK inhibitor synergistically decreased the survival of osimertinib-resistant cell lines with enhanced induction of apoptosis and effectively inhibited the growth of osimertinib-resistant xenografts in nude mice. Moreover, the combination of an MEK or ERK inhibitor with a first-generation (eg, erlotinib) or second-generation (eg, afatinib) EGFR-TKI also very effectively inhibited the growth of osimertinib-resistant cells in vitro and of tumors in vivo, although these cell lines were cross-resistant to first-generation or second-generation EGFR-TKIs. The current findings emphasize the importance of targeting MEK/ERK signaling in maintaining the long-term benefit of osimertinib through overcoming acquired resistance to osimertinib, warranting further investigation of this therapeutic strategy to improve the therapeutic efficacy of osimertinib in the clinic.

Li Y ，Zang H ，Qian G ，Owonikoko TK ，Ramalingam SR ，Sun SY ... -  《-》