Dataset Information

Proteome of silenced minichromosome maintenance protein 2 with siRNA on non-small cell lung cancer H1299 cells

ABSTRACT: Lung cancer is the leading cause of cancer-related mortality. The two main lung cancer types are small cell lung cancer (SCLC) and non-SCLC (NSCLC), where NSCLC comprises about 80-85% of all lung cancer. Despite the introduction of improved treatments, the overall survival rate of lung cancer patients remains low. Further elucidation of the regulatory network perturbations between cancer-related genes and proteins is one promising route to alter this mortality trend. The deregulation of the DNA replication, cell cycle, proliferation and migration are the common factors that are involved in cancer development and progression, and therefore logical targets for analysis. Minichromosome maintenance 2(MCM2) is a DNA replication licensing factor, which belongs to the heterohexameric MCM2-7 complex. MCM2 has been proposed as an excellent proliferation marker in many types of cancer. Our study will establish a global functional distribution of identified proteins in silenced-MCM2 in H1299 NSCLC by the means of iTRAQ. Understanding the molecular basis of MCM2 in lung cancer cells enables us to discover alternative target for lung cancer therapy.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Non-small Cell Lung Cancer Cell, Epithelial Cell

DISEASE(S): Non-small Cell Lung Carcinoma

SUBMITTER: Chantal Hoi Yin Cheung

LAB HEAD: Hsueh-Fen Juan

PROVIDER: PXD003743 | Pride | 2018-10-26

REPOSITORIES: Pride

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MCM2-regulated functional networks in lung cancer by multi-dimensional proteomic approach.

Cheung Chantal Hoi Yin CHY Hsu Chia-Lang CL Chen Kai-Pu KP Chong Siao-Ting ST Wu Chang-Hsun CH Huang Hsuan-Cheng HC Juan Hsueh-Fen HF

Scientific reports 20171016 1

DNA replication control is vital for maintaining genome stability and the cell cycle, perhaps most notably during cell division. Malignancies often exhibit defective minichromosome maintenance protein 2 (MCM2), a cancer proliferation biomarker that serves as a licensing factor in the initiation of DNA replication. MCM2 is also known to be one of the ATPase active sites that facilitates conformational changes and drives DNA unwinding at the origin of DNA replication. However, the biological netwo ...[more]

PMID: 29038488

Similar Datasets

Project description:Minichromosome maintenance protein 2 (MCM2) is a licensing factor for DNA replication. It interacts with other MCM proteins to comprise MCM2-7 complex, which acts as a helicase for DNA unwinding and limits DNA replication to one round per cell cycle. MCM2 has been widely used as a biomarker for proliferation in many types of cancer. However, the molecular regulation underlying MCM2 in lung cancer cells is poorly understood. In this study, we investigated the role of MCM2 in lung adenocarcinoma A549 (wild-type p53) and H1299 (null p53) cells. MCM2 overexpression increased cell proliferation in A549 cells while silencing MCM2 decreased cell proliferation in H1299 cells. We performed global quantitative phosphoproteomic analysis to uncover the important downstream networks regulated by MCM2 in lung cancer cells. We identified 1484 phosphorylation sites in 593 phosphoproteins of MCM2-overexpressed A549 cells. Of these phosphosites, 110 phosphoproteins were significantly changed in response to MCM2 overexpression. In addition, we identified 1599 phosphorylation sites in 592 phosphoproteins of MCM2-silenced H1299 cells. Of these phosphosites, 57 phosphoproteins were significantly changed in response to MCM2 silencing. The differentially regulated phosphoproteins are involved in biological functions such as RNA splicing, cell cycle and cytoskeleton regulation. Functional study demonstrated that MCM2 overexpression promoted cell migration in A549 cells. Moreover, silencing MCM2 inhibits cell migration and induces cell cycle arrest in H1299 cells. Furthermore, we observed a common phosphorylation change at Ser-99 of high mobility group protein HMG-I/HMG-Y (HMGA1) in both MCM2 overexpression and silencing, indicating an important regulatory effect of Ser-99 HMGA1 on lung cancer cells. The phosphoproteomic profiling of MCM2 in lung cancer cells provides new insight about phosphorylation networks regulated by MCM2 and reveals novel targets for lung cancer therapy.

Project description:Introduction: The clinical benefit of EGFR tyrosine kinase inhibitor (TKI) treatment in non-small cell lung cancer (NSCLC) patients with activating EGFR mutations is temporary, as virtually all patients develop acquired EGFR TKI resistance that occurs via diverse mechanisms. Here, we identified increased FGFR1 expression as such a resistance mechanism and using pathways analysis and drug combination testing we identified a novel combination treatment to control growth of these resistant tumors. Methods: Novel erlotinib-resistant NSCLC cell lines were generated and analyzed by mass spectrometry-based proteomics to identify altered pathways associated with erlotinib resistance. The altered pathways were further analyzed in gefitinib and osibenib resistant cell lines. Small molecule inhibitor combinations were used to block the altered pathways and investigate growth reduction in vitro and in two xenograft mouse models. FGFR1 mRNA levels were examined in pre- and (post?)- EGFR TKI treatment clinical tumor samples. Results: Proteomic analysis revealed increased expression of FGFR1 and AXL as well as increased Akt and ERK1/2 activation in a panel of novel erlotinib-resistant HCC827 cell lines. Combined treatment with erlotinib or osimertinib and a panel of small molecule inhibitors targeting AXL/MET, FGFRs, Akt, PI3K/mTOR, MEK or ERK1/2 showed that the most prominent re-sensitization to EGFR TKI occurred with the pan-FGFR inhibitor, PD173074. Interestingly, simultaneous blockade of components of the Akt pathway using specific Akt or dual PI3K-mTOR inhibitors combined with inhibitors targeting the FGFR family exhibited the most efficient growth inhibition of FGFR1 overexpressing EGFR TKI-resistant cell lines. Phosphorylation of proteins downstream of Akt, including PRAS40, FOXO and S6 ribosomal protein, were completely abrogated by PD173074 combined with the Akt inhibitor GSK2141795 . Combination treatment with PD173074 and an Akt inhibitor exhibited synergistic growth inhibition in vivo in two FGFR1 overexpressing NSCLC EGFR TKI-resistant animal models. Conclusion: The significant growth inhibition in vitro and in vivo observed with PD173074 combined with Akt compared to either drug alone imply that inhibition of several key targets may be beneficial in controlling erlotinib-resistant NSCLC. The complete abrogation of PRAS40, FOXO and S6 phosphorylations by PD173074 combined with an Akt inhibitor indicates that the Akt pathway is no longer active.

			Action	DRS
	iTRAQ-10_150608.msf	Msf
	iTRAQ-10_150608.raw	Raw
	iTRAQ-11_150608.msf	Msf
	iTRAQ-11_150608.raw	Raw
	iTRAQ-12_150608.msf	Msf