Project description:Tumor cells undergo epithelial-mesenchymal transition (EMT), however, there is a room of disagreement in role of EMT heterogeneity to colorectal cancer metastasis (mCRC) evolution. To uncover new EMT-related metastasis proteins and pathways, we addressed the EMT status in colorectal cancer liver metastasis patient-derived CTCs to identify proteins that promote their distant metastasis. And then, we performed a comparative proteomic analysis in matched pairs of primary tumor tissues, adjacent mucosa tissues and liver metastases tissues. By integrative analysis we show that, unstable Epithelial/Mesenchymal (E/M)-type CTCs had the strongest liver metastases formation ability and the proportion of E/M-type CTCs correlated with distant metastases. Using an optimized proteomic workflow including data independent acquisition (DIA) and parallel reaction monitoring (PRM), we identified novel EMT-related protein cluster (e.g., GNG2, COL6A1, COL6A2, DCN, COL6A3, LAMB2, TNXB, CAVIN1) and well-described (e.g., KRAS and ERBB2) core protein level changes in EMT-related metastasis progression, and the proteomic data indicate KRAS, ERBB2, COL6A1 and CAVIN1 are promising EMT-related metastatic biomarker candidates. This study contributes to catch on EMT in CRC metastasis and in favor of heterogeneous EMT phenotypes as a key piece for tumor outcome and treatment. We further propose that therapies targeting this aggressive subset (E/M-type) of CTCs and related protein may be worthy of exploration as potential suppressors of metastatic evolution
Project description:Using liver metastases from 4 colorectal cancer patients (CRC-LM), we explored one receptor: EGFR whose signaling triggers cell proliferation and is exacerbated in many tumors.
Project description:Anti-EGFR antibodies are effective in therapies for late-stage colorectal cancer (CRC); however, many tumours are unresponsive or develop resistance. We performed genomic analysis of intrinsic and acquired resistance to anti-EGFR therapy in prospectively collected tumour samples from 25 CRC patients receiving cetuximab (an EGFR inhibitor). Of 25 CRC patients, 13 displayed intrinsic resistance to cetuximab; 12 were intrinsically sensitive. We obtained six re-biopsy samples at acquired resistance from the intrinsically sensitive patients. NCOA4–RET and LMNA–NTRK1 fusions and NRG1 and GNAS amplifications were found in intrinsic-resistant patients. In cetuximab-sensitive patients, we found KRAS K117N and A146T mutations in addition to BRAF V600E, AKT1 E17K, PIK3CA E542K, and FGFR1 or ERBB2 amplifications. The comparison between baseline and acquired-resistant tumours revealed an extreme shift in variant allele frequency of somatic variants, suggesting that cetuximab exposure dramatically selected for rare resistant subclones that were initially undetectable. There was also an increase in epithelial-to-mesenchymal transition at acquired resistance, with a reduction in the immune infiltrate. Furthermore, characterization of an acquired-resistant, patient-derived cell line showed that PI3K/mTOR inhibition could rescue cetuximab resistance. Thus, we uncovered novel genomic alterations that elucidate the mechanisms of sensitivity and resistance to anti-EGFR therapy in metastatic CRC patients.
Project description:We showed that a large number of genes and exons were deregulated in colorectal adenomas in comparison with colorectal normal mucosa.
Project description:Intratumor heterogeneity is a major challenge in cancer treatment. To decipher patterns of chromosomal heterogeneity, we analyzed six colorectal cancer cell lines by multiplex interphase FISH. The mismatch repair deficient cell lines DLD-1 and HCT116 had the most stable copy numbers, whereas aneuploid cell lines displayed a higher degree of instability. We subsequently assessed the clonal evolution of a single cell in two aneuploid cell lines, SW480 and HT-29, which both have near-triploid karyotypes but different degrees of chromosomal instability. The clonal compositions of the single cell-derived daughter cell lines, as assessed by multiplex FISH, differed for HT-29 and SW480. Daughters of HT-29 were stable, clonal, and had little heterogeneity. Daughters of SW480 were more heterogeneous, with the single cell-derived daughter cell lines separating into two distinct populations with different ploidy (hyper-diploid and near-triploid), morphology, gene expression and tumorigenicity. To better understand the evolutionary trajectory for the two SW480 populations, we constructed phylogenetic trees which showed ongoing instability in the daughter cell lines.. When analyzing the evolutionary development over time, most single cell-derived daughter cell lines maintained their major clonal pattern, with the exception of one daughter of SW480 that showed a switch involving a loss of APC. Our meticulous analysis of the clonal evolution and composition of these colorectal cancer models shows that all chromosomes are subject to segregation errors, however, specific net genomic imbalances are maintained. Karyotype evolution is driven by the necessity to arrive at and maintain a specific plateau of chromosomal copy numbers as the drivers of carcinogenesis.
Project description:LC-MS/MS glycoproteomic analysis of EGFR N-glycosylation profile. EGFR immunoprecipitated from colorectal cancer cell lines SW48 (wild-type and ST6Gal1 overexpression) and Caco-2 (wild-type and ST6Gal1 knock-out)
