关于阿法替尼（Bibw 2992）耐药的一些理论和解决方案

老马 · 发表于 2013-12-14 02:24:37

An epithelial-mesenchymal transition (EMT) gene signature predicts resistance to EGFR and PI3K inhibitors and identifies Axl as a therapeutic target for overcoming EGFR inhibitor resistance
http://clincancerres.aacrjournal ... 78-0432.CCR-12-1558
Purpose: EMT has been associated with metastatic spread and EGFR inhibitor resistance. We developed and validated a robust 76-gene EMT signature using gene expression profiles from four platforms using NSCLC cell lines and patients treated in the BATTLE study. Methods: We conducted an integrated gene expression, proteomic, and drug response analysis using cell lines and tumors from NSCLC patients. A 76-gene EMT signature was developed and validated using gene expression profiles from four microarray platforms of NSCLC cell lines and patients treated in the BATTLE (Biomarker-integrated Approaches of Targeted Therapy for Lung Cancer Elimination) study, and potential therapeutic targets associated with EMT were identified. Results: Compared with epithelial cells, mesenchymal cells demonstrated significantly greater resistance to EGFR and PI3K/Akt pathway inhibitors, independent of EGFR mutation status, but more sensitivity to certain chemotherapies. Mesenchymal cells also expressed increased levels of the receptor tyrosine kinase Axl and showed a trend towards greater sensitivity to the Axl inhibitor SGI-7079. Furthermore, the combination of SGI-7079 with erlotinib reversed erlotinib resistance in mesenchymal lines expressing Axl and in a xenograft model of mesenchymal NSCLC. In NSCLC patients, the EMT signature predicted 8-week disease control in patients receiving erlotinib, but not other therapies. Conclusion: We have developed a robust EMT signature that predicts resistance to EGFR and PI3K/Akt inhibitors, highlights different patterns of drug responsiveness for epithelial and mesenchymal cells, and identifies Axl as a potential therapeutic target for overcoming EGFR inhibitor resistance associated with the mesenchymal phenotype.

老马 · 发表于 2013-12-14 02:28:53

First Axl inhibitor enters clinical trials
Cormac Sheridan1
Journal name:
Nature Biotechnology
Volume:
31,
Pages:
775–776
Year published:
(2013)
DOI:
doi:10.1038/nbt0913-775a
Published online 10 September 2013
On June 19, BergenBio disclosed that BGB324, which it bills as a first-in-class Axl kinase inhibitor, entered phase 1 clinical trials.

老马 · 发表于 2013-12-17 16:30:01

Clinical Cancer Research Article Demonstrates SGI-7079 Sensitizes Lung Cancer Cells to Clinically Approved Anti-Cancer Agents

Source PRess Release
Company Tolero Pharmaceuticals, The University of Texas MD Anderson Cancer Center
Tags Trial Results, Pre-clinical, Oncology
Date January 07, 2013
A Team of Investigators at MD Anderson Cancer Center Demonstrate Axl Inhibition Reverses the Mesenchymal Phenotype of Cancer Cells and Confers Increased Sensitivity to EGFR Inhibition

SALT LAKE CITY, Utah - January 7, 2013 - Tolero Pharmaceuticals announces that collaborators at the University of Texas MD Anderson Cancer Center in Houston, Texas have developed a robust epithelial-mesenchymal transition (EMT) signature that predicts resistance to EGFR and PI3K/Akt inhibitors and identified  Axl as a potential therapeutic target for overcoming  EGFR inhibitor resistance associated with the mesenchymal phenotype. More importantly, it was demonstrated that Tolero’s  Axl kinase inhibitor,  SGI-7079, sensitized lung cancer cells to erlotinib, particularly in an acquired resistance setting. Published last week in Clinical Cancer Research, the findings show that SGI-7079 may restore sensitivity to EGFR inhibition, particularly in EGFR  mutant lung cancer patients that have acquired resistance to  erlotinib.

This work builds on a general phenomenon described in the scientific literature that Axl is commonly involved in a "kinase switch" as a tumor transitions its dependence from one kinase to another under the survival pressure of a targeted therapeutic agent. More specifically, it provides Tolero with an exciting and concise niche for which a clinical development plan for an  Axl kinase inhibitor could be developed.

"The continued emergence of Axl expression as a resistance mechanism in non-small cell lung cancer and other tumor types provides us with a rational clinical development strategy for our  Axl inhibitor program," said Dr. Steven L. Warner, Tolero’s Vice President of Drug Discovery and Development. "We are moving forward with this strategy in mind and aim to have our  Axl inhibitor in a first-in-man clinical trial in the next 12 months."

Tolero is now looking to broaden these findings to TP-0903, a second generation Axl inhibitor they are moving forward through IND-enabling studies. Preliminary work conducted by the team at MD Anderson and by additional collaborators at the Huntsma

老马 · 发表于 2014-1-5 03:36:01

15th World Conference on Lung Cancer, 27-30 Oct 2013, Sydney, Australia

Oct 10, 2013

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Background:
Somatic activating mutations in the tyrosine kinase (TK) domain of EGFR are harbored by 10-20% of Caucasian NSCLC patients (pts). Reversible TK inhibitors (TKIs), including erlotinib or gefitinib, have demonstrated significantly longer progression-free survival compared to chemotherapy alone in EGFR-mutated pts. Nevertheless, the majority of these tumors develop drug resistance due to an acquired mutation (T790M) in EGFR that determines disease progression. Recent clinical trials have demonstrated interesting activity of the irreversible EGFR-TKI afatinib (BIBW-2992) in advanced NSCLC carrying EGFR mutations and in unselected pts failing previous treatments with reversible TKIs.
The aim of this study was to clarify the mechanisms of acquired resistance to afatinib using in vitro models of resistant cell lines.
Methods:
A dose-escalation study was performed to establish afatinib-resistant (R) clones in NSCLC cell lines harboring different EGFR mutations: H-1650 (exon 19 delE746-A750) and H-1975 (exon 21 L858R/exon 20 T790M). The entire genomes of parental (P) and R cells were screened by array comparative genomic hybridization (aCGH) using a 105 k oligonucleotide microarray. All EGFR and KRAS exons and 10 known hot spots (5 in genes involved in the EGFR signaling cascade and 5 in genes frequently altered in NSCLC) were deep sequenced using an Ion PGM™ Sequencer in P and R cells. The relative expression of 92 genes belonging to the EGF pathway was studied by quantitative polymerase chain reaction (qPCR). The expression of proteins related to the EGF pathway, including EGFR, AKT and ERK (total and activated forms), was investigated by Western blot.
Results:
Genomic analysis indicated that both R cell lines had genomic profiles similar to the P cells. However, H-1975-R showed 3p and 12p loss and gains at 4q and 10q compared to the P cell line. Sequence analyses identified a novel frame-shift mutation within exon 14 of MET and confirmed EGFR mutation status in 100% of H1975-R cells. In contrast, H-1650-R cells showed a single-base deletion 12 bp upstream of exon 8 of PIK3CA within a sequence of nine repeated Ts. Furthermore, a novel missense variant (exon 8 K368E) was found in FGFR2 in both R cell lines compared to the P cell line. Gene expression profiles identified an increase in the FGFR2 and PIK3 regulatory subunits and EGFR ligand silencing in H-1975-R. Notably, H-1975-R cells maintained in afatinib-free medium for over 6 months showed higher EGFR and AKT phosphorylation compared to the P cell lines.

Conclusion:
The lack of novel EGFR mutations suggests the involvement of other mechanisms implicated in afatinib resistance. In particular, the identification of mutations involving MET and FGFR2 in H-1975 and PI3KCA in H-1650 suggests their contribution to resistance against irreversible TKIs, sparing EGFR activation. Furthermore, the different mutation status of the two cell lines indicates that the T790M mutation may be partially responsible for the
mechanism of resistance. Validation studies are ongoing to confirm the genomic results. In conclusion, these preliminary data may help
identify novel therapeutic strategies to delay or reverse resistance to irreversible TKIs in EGFR-mutated NSCLC patients.
http://abstracts.webges.com/wclc2013/myitinerary

老马 · 发表于 2014-1-5 03:36:44

Poster Presentations - Resistance to Molecular Targeted Therapies 3

Abstract 5649: Investigational study of acquired resistance to the EGFR irreversible inhibitor afatinib (BIBW2992) in wild-type and EGFR-mutant NSCLC cell lines.
Angela Alama, Simona Coco, Zita Cavalieri, Anna Truini, Cristina Bruzzo, Mariella Dono, and Francesco Grossi
IRCCS A.O.U. San Martino-IST, Genoa, Italy.

Introduction: Non-small cell lung cancer (NSCLC) represents about 85% of all lung cancers. Almost 20% of NSCLC tumors harbor somatic activating mutations of the tyrosine kinase (TK) encoding domain in EGFR gene (exons 18-21). Patients with EGFR mutations treated with specific inhibitors of EGFR-TKI such as gefitinib and erlotinib demonstrated a significantly longer survival. Nevertheless, the majority of patients acquire resistance to these drugs within one year due to a second-site mutation in EGFR (T790M). The irreversible EGFR-TKI, afatinib, has been found to be effective in inhibiting the growth of NSCLC cells with the T790M mutation of EGFR. In this study, we generated different NSCLC cell lines resistant to afatinib to investigate the biological and molecular mechanisms of acquired resistance.

Methods: A dose-escalation study to establish afatinib-resistant cell lines was performed in 3 NSCLC cell lines with a different EGFR mutational status: A549 (EGFR-wild type); H-1650 (exon19 delE746-A750); H-1975 (exon21 L858R/exon20 T790M). EGFR exons (18-21) from parental and resistant cells were sequenced by Sanger method while the EGF pathway was studied by qPCR using TaqMan® Array EGF Pathway and relative expression values of 92 genes, compared to parental cells, were obtained by 2–ddCT. Concomitantly, protein expression of some relevant genes such as EGFR and AKT, both phosphorylated and unphosphorylated, were investigated by western blot (WB).

Results: All 3 cell lines exhibited different degree of drug resistance and actively proliferated under persistent afatinib pressure (A549: 8.5μM; H-1650: 1.0μM; H-1975: 5.0μM) as compared to parental cultures. WB reported active phosphorylation of EGFR and AKT in resistant cells regardless of EGFR mutational status. This behavior also persisted in absence of EGF stimulation and in cultures maintained in afatinib-free medium for over 2 months. However none of the resistant cell lines harbored new EGFR mutation in the 4 exons (18-21). Gene expression analysis showed an increase of some members of RAS (MRAS, KRAS) and Rho families and also PI3K regulatory subunit 1 (PIK3R1) in A549 and H1975 cells. Notably, SHC3 (Src homology 2 domain containing; transforming protein 3) was up-regulated about 60-fold in H1975 whereas proto-oncogene JUN showed a 6-fold increase in H1650. In addition EGF was down-modulated in both H1650 and H1975.

Conclusion: Silencing of EGF in resistant cells together with the EGFR and AKT phosphorylation, suggest that these cultures acquired a different way to activate the EGF pathway. Furthermore in H-1975 the markedly increase of SHC3, an adaptor immediately downstream of EGFR, leads to hypothesize that it could be implicated in the activation of EGFR. Further investigations of SHC3 involvement in EGF pathway activation as well as sequencing analysis of the whole EGFR gene are ongoing.