PIM1 kinase promotes EMT-associated osimertinib resistance via regulating GSK3β signaling pathway in EGFR-mutant non-small cell lung cancer.

Acquired resistance is inevitable in the treatment of non-small cell lung cancer (NSCLC) with osimertinib, and one of the primary mechanisms responsible for this resistance is the epithelial-mesenchymal transition (EMT). We identify upregulation of the proviral integration site for Moloney murine leukemia virus 1 (PIM1) and functional inactivation of glycogen synthase kinase 3β (GSK3β) as drivers of EMT-associated osimertinib resistance. Upregulation of PIM1 promotes the growth, invasion, and resistance of osimertinib-resistant cells and is significantly correlated with EMT molecules expression. Functionally, PIM1 suppresses the ubiquitin-proteasome degradation of snail family transcriptional repressor 1 (SNAIL) and snail family transcriptional repressor 2 (SLUG) by deactivating GSK3β through phosphorylation. The stability and accumulation of SNAIL and SLUG facilitate EMT and encourage osimertinib resistance. Furthermore, treatment with PIM1 inhibitors prevents EMT progression and re-sensitizes osimertinib-resistant NSCLC cells to osimertinib. PIM1/GSK3β signaling is activated in clinical samples of osimertinib-resistant NSCLC, and dual epidermal growth factor receptor (EGFR)/PIM1 blockade synergistically reverse osimertinib-resistant NSCLC in vivo. These data identify PIM1 as a driver of EMT-associated osimertinib-resistant NSCLC cells and predict that PIM1 inhibitors and osimertinib combination therapy will provide clinical benefit in patients with EGFR-mutant NSCLC.

Zhou J ，Wang X ，Li Z ，Wang F ，Cao L ，Chen X ，Huang D ，Jiang R ... -  《Cell Death & Disease》

Targeting the EMT transcription factor Snail overcomes resistance to osimertinib in EGFR-mutant non-small cell lung cancer.

The resistance mechanism of the third generation of epidermal growth factor (EGFR) tyrosine kinase inhibitor (TKI) osimertinib is complex. Epithelial mesenchymal transition (EMT) is a common mechanism of EGFR-TKI acquired resistance. Snail is an important transcription factor related to EMT. Whether targeting Snail can reverse the resistance of osimertinib by downregulating Snail is unknown. The presence of EMT in H1975/OR (osimertinib resistance) cells was confirmed by transwell assay. To explore the EMT role in resistance, the expression levels of EMT markers were detected in both parental cells H1975 and resistant cells H1975OR. We used RNA interference technology to knockdown the key regulator Snail in resistant cells. After the interference efficiency was confirmed, changes in EMT-related molecules of Snail were explicitly downregulated, and changes in sensitivity and migration and invasion ability were also examined. We used CDK4/6 inhibitor to test the ability of reversing drug resistance by downregulating Snail. Compared with the H1975 cell line, the H1975/OR resistant cell line showed increased invasiveness, upregulated expression of vimentin and downregulation of E-cadherin. EMT occurred in the H1975/OR resistant cell line. The expression of Snail was upregulated in the osimertinib-resistant cell line H1975/OR. Knockdown of Snail increased the sensitivity of H1975/OR cells to osimertinib. CDK4/6 inhibitor palbociclib could downregulate the expression of Snail. CDK 4/6 inhibitor palbociclib combined with osimertinib could reverse the resistance of osimertinib in H1975/OR. Snail plays an important role in the third generation of EGFR-TKI osimertinib resistance, which may be reversed by downregulating Snail.

Qin Q ，Li X ，Liang X ，Zeng L ，Wang J ，Sun L ，Zhong D ... -  《-》

Afatinib plus PEM and CBDCA overcome osimertinib resistance in EGFR-mutated NSCLC with high thrombospondin-1 expression.

Osimertinib induces a marked response in non-small-cell lung cancer (NSCLC) patients harboring epidermal growth factor receptor (EGFR) gene mutations. However, acquired resistance to osimertinib remains an inevitable problem. In this study, we aimed to investigate osimertinib-resistant mechanisms and evaluate the combination therapy of afatinib and chemotherapy. We established osimertinib-resistant cell lines (PC-9-OR and H1975-OR) from EGFR-mutant lung adenocarcinoma cell lines PC-9 and H1975 by high exposure and stepwise method. Combination therapy of afatinib plus carboplatin (CBDCA) and pemetrexed (PEM) was effective in both parental and osimertinib-resistant cells. We found that expression of thrombospondin-1 (TSP-1) was upregulated in resistant cells using cDNA microarray analysis. We demonstrated that TSP-1 increases the expression of matrix metalloproteinases through integrin signaling and promotes tumor invasion in both PC-9-OR and H1975-OR, and that epithelial-to-mesenchymal transition (EMT) was involved in H1975-OR. Afatinib plus CBDCA and PEM reversed TSP-1-induced invasion ability and EMT changes in resistant cells. In PC-9-OR xenograft mouse models (five female Balb/c-Nude mice in each group), combination therapy strongly inhibited tumor growth compared with afatinib monotherapy (5 mg/kg, orally, five times per week) or CBDCA (75 mg/kg, intraperitoneally, one time per week) + PEM (100 mg/kg, intraperitoneally, one time per week) over a 28-day period. These results suggest that the combination of afatinib plus CBDCA and PEM, which effectively suppresses TSP-1 expression, may be a promising option in EGFR-mutated NSCLC patients after the acquisition of osimertinib resistance.

Onda N ，Nakamichi S ，Hirao M ，Matsuda K ，Matsumoto M ，Miyanaga A ，Noro R ，Gemma A ，Seike M ... -  《-》

Let-7c regulated epithelial-mesenchymal transition leads to osimertinib resistance in NSCLC cells with EGFR T790M mutations.

Li XF ，Shen WZ ，Jin X ，Ren P ，Zhang J ... -  《Scientific Reports》

Glycogen synthase kinase-3 inhibition overcomes epithelial-mesenchymal transition-associated resistance to osimertinib in EGFR-mutant lung cancer.

A novel epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor, osimertinib, has marked efficacy in patients with EGFR-mutant lung cancer. While epithelial-mesenchymal transition (EMT) plays a role in the resistance to various targeted drugs, its involvement in EGFR-inhibitor resistance remains largely unknown. Preclinical experiments with osimertinib-resistant lung cancer cells showed that EMT was associated with decreased microRNA-200c and increased ZEB1 expression. In several resistant clone cells, pretreatment with the histone deacetylase inhibitor quisinostat helped overcome the resistance by reverting EMT. Furthermore, drug screening from a library of 100 kinase inhibitors indicated that Glycogen synthase kinase-3 (GSK-3) inhibitors, such as LY2090314, markedly inhibited the growth and induced apoptosis of resistant cells, specifically those with a mesenchymal phenotype. These results suggest that GSK-3 inhibition could be useful to circumvent EMT-associated resistance to osimertinib in EGFR-mutant lung cancer.

Fukuda K ，Takeuchi S ，Arai S ，Kita K ，Tanimoto A ，Nishiyama A ，Yano S ... -  《-》