Cyclosporine A sensitizes lung cancer cells to crizotinib through inhibition of the Ca2+/calcineurin/Erk pathway

Abstract

Background Crizotinib has potent anti-tumor activity in patients with advanced MET-amplified non-small cell lung cancer (NSCLC). However, the therapeutic effect is still not satisfying. Thus, developing approaches that improve the efficacy of crizotinib remains a significant challenge. Methods MET-amplified NSCLC cell lines were treated with crizotinib and cyclosporine A (CsA). Cell viability was determined by MTS assay. The changes of apoptosis, cell cycle and calcineurin-Erk pathways were assessed by western blot. Xenograft mouse model, primary human NSCLC cells and hollow fiber assays were utilized to confirm the effects of CsA. Findings We demonstrated that CsA significantly increased the anti-tumor effect of crizotinib on multiple MET-amplified NSCLC cells in vitro and in vivo . Mechanistically, crizotinib treatment led to the activation of Ca 2+ -calcineurin (CaN)-Kinase suppressor of Ras 2 (KSR2) signaling, resulting in Erk1/2 activation and enhanced survival of cancer cells. CsA effectively blocked CaN-KSR2-Erk1/2 signaling, promoting crizotinib-induced apoptosis and G2/M arrest. Similarly, pharmacologic or genetic inhibition of Erk1/2 also enhanced crizotinib-induced growth inhibition in vitro . Xenograft studies further confirmed that CsA or Erk1/2 inhibitor PD98059 enhanced the anti-cancer activity of crizotinib through inhibition of CaN-Erk1/2 axis. The results were also validated by primary human NSCLC cells in vitro and hollow fiber assays in vivo . Interpretation This study provides preclinical evidences that combination therapy of CsA and crizotinib is a promising approach for targeted treatment of MET-amplified lung cancer patients. Fund This work was supported by the National Natural Science Foundation of China, the Key Projects of Natural Foundation of Zhejiang Province, the Ten thousand plan youth talent support program of Zhejiang Province, the Zhejiang Natural Sciences Foundation Grant, and the Zhejiang medical innovative discipline construction project-2016.

Calcineurin in development and disease

“…A recent study revealed that when non-small cell lung cancer (NSCLC) cells were treated with crizotinib and CsA, apoptosis was promoted, and G2/M arrest was induced compared with crizotinib-only treatment. 148 In contrast, CsA and FK506 increased the risk of cancers in organ-transplant patients due to suppression of tumor immunosurveillance mechanisms. 49 For example, CsA treatment promotes the rapid growth and survival of renal cancer cells via activating Ras and inducing the expression of cytoprotective molecule heme oxygenase-1 (HO-1).…”

RCAN1-mediated calcineurin inhibition as a target for cancer therapy

“…Preclinical studies show that the tumor inhibitory mechanism of CsA involves the sensitization of lung cancer cells to crizotinib via suppression of the Ca2 + /CN/Erk pathway. Treatment with a combination of CsA and crizotinib might have potential in MET-amplified lung cancer (Liu et al 2019 ). CsA increases pancreatic cancer cell response to phospho-sulindac (P-S) treatment by overcoming NFATc1-mediated resistance (Murray et al 2014 ).…”

Strategies for Targeting Serine/Threonine Protein Phosphatases with Small Molecules in Cancer

“…In the process of exploring their mechanisms, PP3 is identified as a target. , Before combining with PP3, CsA and FK-506 first form a binary complex with immunophilins, their specific carrier protein. Immunophilins are highly conserved and ubiquitously expressed with peptidylprolyl isomerase activity, and they are mainly divided into two types: cyclophilins (Cyp, for CsA binding) and FK-506 binding proteins (FKBPs). − Recently, they were found to exhibit antitumor effects in cancers, including colorectal cancer, breast cancer, bladder cancer, lung cancer, , oral squamous cell carcinoma, and other cancers. ,− …”

“…In addition, several tumor suppressor genes, such as p53, p21, and p27, are upregulated, and some oncogenic genes are downregulated, including cyclin D1, cyclin D3, and cyclin E. , CsA and FK-506 retard tumor growth by inhibiting proliferation, migration, and invasion and promoting apoptosis. In addition to evidence from the Masuo group, other evidence also revealed the antitumor activities of CsA and FK-506 on other cancers. ,− In a breast cancer model, pyruvate kinase M2 (PKM2) is overexpressed which is closely related to the usage of glycolytic intermediates for macromolecular biosynthesis and tumor growth. Treatment with CsA can significantly downregulate the expression of PKM2 and reduce ATP synthesis, which induces cancer cell necrosis .…”

“…Treatment with CsA can significantly downregulate the expression of PKM2 and reduce ATP synthesis, which induces cancer cell necrosis . Other cancers, including lung cancer and oral squamous cell carcinoma, are also response to CsA. − …”