Ye Jin Ha

Feasibility of proposed single-nucleotide polymorphisms as predictive markers for targeted regimens in metastatic colorectal cancer.

Background:Surrogate biomarkers for metastatic colorectal cancer (mCRC) are urgently needed to achieve the best outcomes for targeted therapy.Methods:A clinical association analysis was performed to examine the three single-nucleotide polymorphisms (SNPs) that were previously proposed as markers of chemosensitivity to the cetuximab (124 patients) and bevacizumab regimens (100 patients) in mCRC patients. In addition, biological correlations were examined for the candidate SNPs in terms of their regulatory pathway.Results:For cetuximab regimens, patients homozygous for the wild-type alleles (GG) of LIFR rs3729740 exhibited a 1.9 times greater overall response rate (ORR) and 1.4 months longer progression-free survival (PFS) than those homozygous or heterozygous for the mutant allele (GA and AA; P=0.022 and 0.027, respectively). For bevacizumab regimens, patients homozygous for the minor alleles (TT) of ANXA11 rs1049550 exhibited an ORR twice as high as those homozygous or heterozygous for the ancestral allele (CC and CT; P=0.031). Overall response rate gain was achieved up to 10% in patients with wild-type LIFR rs3729740 patients either with wild-type KRAS or skin toxicity (P=0.001) respectively. Specifically in clones treated with cetuximab and bevacizumab regimens, active p-ERK and MMP-9 expressions were significantly reduced in clones expressing wild-type LIFR rs3729740 (P=0.044) and in those expressing minor-type ANXA11 rs1049550 (P=0.007), respectively.Conclusion:LIFR rs3729740 and possibly ANXA11 rs1049550 may be useful as biomarkers for predicting whether mCRC patients are sensitive to relevant target regimens, although further validation in large cohorts is needed.

Complex Behavior of ALDH1A1 and IGFBP1 in Liver Metastasis from a Colorectal Cancer

Using our data set (GSE50760) previously established by RNA sequencing, the present study aimed to identify upregulated genes associated with colorectal cancer (CRC) liver metastasis (CLM) and verify their biological behavior. The potential roles of candidate genes in tumors were assessed using cell proliferation and invasion assays. Tissue samples were collected from 18 CRC patients with synchronous CLM and two CRC cell lines (SW480 and SW620) were used for transfection and cloning. The roles of the genes identified in CLM were verified using immunohistochemistry in 48 nude mice after intrasplenic transplantation of CRC cells. mRNA and protein expression was determined by quantitative real-time reverse transcription polymerase chain reaction and western blot, respectively. Nine genes were initially selected according to the relevance of their molecular function and biological process and, finally, ALDH1A1 and IGFBP1 were chosen based on differential mRNA expression and a positive correlation with protein expression. The overexpression of ALDH1A1 and IGFBP1 significantly and time-dependently decreased cell proliferation (p ≤ 0.001–0.003) and suppressed invasiveness by ≥3-fold over control cells (p < 0.001) in the SW480 cell line, whereas they had a slight effect on reducing SW620 cell proliferation. The protein expression levels of E-cadherin, N-cadherin, claudin-1, and vimentin were significantly higher in CLM than in primary tumor tissues (p < 0.05). However, the cadherin switch, namely, N-cadherin overexpression with reduced E-cadherin expression, was not observed in CLM tissues and transfected CRC cells. Irrespective of reduced proliferation and invasion found on in vitro cell assays, persistent overexpression of β-catenin, vimentin, and ZO-1 in IGFBP1-overexpressing SW480 cells possibly contributed to CLM development in mice implanted with IGFBP1-overexpressing SW480 cells (CLM occurrences: SW480/IGFBP1-transfected mice vs. SW480/vector- and SW480/ALDH1A1-transfected mice, 4/8 vs. 0/10, p = 0.023). In conclusion, ALDH1A1 and IGFBP1 are differentially overexpressed in CLM and may play a dual role, functioning as both tumor suppressors and metastasis promoters in CRC.

Inhibition of never in mitosis A (NIMA)-related kinase-4 reduces survivin expression and sensitizes cancer cells to TRAIL-induced cell death.

The tumor necrosis factor-related apoptosis inducing ligand (TRAIL) preferentially induces apoptosis in cancer cells. However, many tumors are resistant to TRAIL-induced apoptosis, and resistance mechanisms are not fully understood. To identify novel regulatory molecules of TRAIL resistance, we screened a siRNA library targeting the human kinome, and NEK4 (NIMA-related kinase-4) was identified. Knockdown of NEK4 sensitized TRAIL-resistant cancer cells and in vivo xenografts to cell death. In contrast, over expression of NEK4 suppressed TRAIL-induced cell death in TRAIL-sensitive cancer cells. In addition, loss of NEK4 resulted in decrease of the anti-apoptotic protein survivin, but an increase in apoptotic cell death. Interestingly, NEK4 was highly upregulated in tumor tissues derived from patients with lung cancer and colon cancer. These results suggest that inhibition of NEK4 sensitizes cancer cells to TRAIL-induced apoptosis by regulation of survivin expression.

Abstract 1140: Novel single-nucleotide polymorphism markers predictive of pathologic response to preoperative chemoradiotherapy in rectal cancer patients.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Purpose: Studies aimed at predicting individual responsiveness to preoperative chemoradiotherapy (CRT) are urgently needed, especially considering the risks associated with poorly responsive patients, i.e., delay of curative surgery, immune suppression, and adverse effects. Methods and Materials: A three-step strategy for the determination of CRT sensitivity is proposed that includes: 1) screening of a human genome-wide single-nucleotide polymorphism (SNP) array correlated with histopathological tumor regression grade (TRG), 2) clinical association analysis in 113 patients treated with preoperative CRT, and 3) a cell-based functional assay for biological validation. Genome-wide screening identified nine SNPs associated with preoperative CRT responses. Results: Genome-wide screening identified nine SNPs associated with preoperative CRT responses. The patients carrying the reference allele (C) of the SNP CORO2A rs1985859 exhibited greater positive responses (TRG 1-3) than those with the substitution allele (T) (P=0.01). Down-regulation of CORO2A was associated with significantly reduced early apoptosis in RKO and COLO320DM colorectal cancer cells by 27% (P=0.048) and 39% (P=0.023), respectively, as determined by flow cytometry. Although a specific genotype associated with radiosensitivity was not detected in the clinical association analysis, down-regulation of FAM101A significantly increased the viability of RKO and COLO320DM cells by 25% (P=0.01) and 23% (P=0.023), respectively in the MTT assay. Conclusion: CRT-sensitive SNP markers were identified using a novel three-step process. The candidate marker CORO2A rs1985859 and the putative marker FAM101A rs7955740 may improve the prediction of radiosensitivity to preoperative CRT, although further validation is needed in large cohorts. Citation Format: Jin Cheon Kim, Seon Ae Roh, Dong Hyung Cho, Eun Young Choi, Ye Jin Ha, Tae Won Kim, Jong Hoon Kim, Tae Wook Kang, Seon Young Kim, Yong Sung Kim. Novel single-nucleotide polymorphism markers predictive of pathologic response to preoperative chemoradiotherapy in rectal cancer patients. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1140. doi:10.1158/1538-7445.AM2013-1140

Abstract 255: Identification of a novel TRAIL sensitizer in TRAIL-resistant cancer cells

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL The Tumor Necrosis Factor Related Apoptosis Inducing Ligand (TRAIL), a TNF-family member is a highly promising agent for cancer therapy as it selectively induces apoptosis in various cancer cells but not in normal cells. However, many primary tumors are resistant to TRAIL-induced apoptosis and the resistant mechanisms are not fully understood. To identify novel TRAIL signaling regulatory molecules, we screened a siRNA library that target 719 whole human protein kinases in TRAIL-resistant lung cancer cell line (A549 cell). As a screening result, we identified a member of NIMA-related kinases as a new regulator of TRAIL-induced cell death. NEK family play key role in controlling mitosis but the role in TRAIL signaling was not elucidated. Down-regulation of NEK by siRNA dramatically increased TRAIL-induced apoptosis in TRAIL resistant cancer cells including A549 cells. However it was not effective on both etoposide and TNF/CHX-mediated apoptosis. In addition, inhibition of NEK highly activated caspase-3 in response to TRAIL. On the other hand, over-expression of NEK reduced TRAIL-induced apoptosis in TRAIL sensitive cells. Our findings suggest that NEK is a novel negative modulator of TRAIL-induced apoptosis in cancer cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 255. doi:1538-7445.AM2012-255

Abstract 4590: Genome-wide identification of possible methylation markers chemosensitive to targeted regimens in colorectal cancers

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Purpose Few efficient methylation markers of chemosensitivity have been discovered. The genome-wide analysis of methylation markers is needed to identify chemosensitive candidates to targeted therapy. Methods This study describes a two-step process to select chemosensitive candidates of methylation genes. A genome-wide screening of methylation genes was performed using a beadarray and an in vitro chemosensitivity assay of 119 colorectal cancers (CRCs). Results Ten candidate genes identified during the initial screening were verified by biological utility assessment using cell viability assays of transfected CRC cells. Five methylation genes related to sensitivity to bevacizumab regimens (RASSF1, MMP25, KCNQ1, ESR1, and GALR2) or cetuximab regimens (SCL18A2, GPX7, NID2, IGFBP3, and ALX4) were chosen during the first step. A viability assay revealed that GALR2-overexpressing HCT116 cells were significantly more chemosensitive to bevacizumab regimens than control cells (P = 0.022 and 0.019 for bevacizumab with FOLFIRI and FOLFOX, respectively), concurrently verified on a caspase-3 activity assay. GPX7- or ALX4-overexpressed RKO cells were significantly less viable to cetuximab regimens compared to control cells (GPX7: P = 0.027 each for cetuximab with FOLFIRI and FOLFOX; ALX4: P = 0.049 and 0.003 for cetuximab with FOLFIRI and FOLFOX, respectively), but caspase-3 activity was not prominent in GPX7-overexpressed RKO cells. Conclusions Two novel genes, GALR2 and ALX4, have been identified as chemosensitive methylation candidates to bevacizumab and cetuximab regimens, respectively. As our study did not include a clinical association study, the two candidates should be validated in large clinical cohorts, hopefully predicting responsive patients to targeted regimens. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4590. doi:1538-7445.AM2012-4590

Abstract 3805: Novel chemosensitive single-nucleotide polymorphism markers to targeted regimens in metastatic colorectal cancer

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Purpose: Methods for predicting individual responsiveness to targeted chemotherapy are urgently needed, considering the frequent resistance and extremely high cost of such therapies. Experimental Design: A chemosensitive single-nucleotide polymorphism (SNP) discovery schema is presented that utilizes genome-wide SNP screening with a human SNP array, an in vitro chemosensitivity assay in 118 colorectal cancers, clinical association analysis in 98 patients who had received metastatic chemotherapy, and biological utility assessment using cell viability assays of transfected colorectal cancer (CRC) cells. Results: Eleven SNPs related to bevacizumab and cetuximab sensitivity were initially chosen during screening. Response rates and survival periods revealed that patients carrying specific alleles of the SNPs ANXA11 rs1049550, LINS1 rs11247226, or ITGA3 rs2230392 were chemosensitive to bevacizumab regimens, and patients with DFNB31 rs2274159, LIFR rs3729740, or ISX [rs361863][1] were chemosensitive to cetuximab regimens. Cytotoxicity analyses using MTT and caspase-3 assays showed that all RKO and HCT116 CRC clones transfected with the reference alleles of LIFR rs3729740 and ISX [rs361863][1] (G and C, respectively) were more sensitive to cetuximab regimens than those transfected with the substitution alleles. Conclusions: Chemosensitive SNP markers were identified using a novel three-step process. The chemosensitive markers LIFR rs3729740 and ISX [rs361863][1] will hopefully predict responsive patients to cetuximab regimens, although further validation is needed in large cohorts. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3805. doi:10.1158/1538-7445.AM2011-3805 [1]: /lookup/external-ref?link_type=GEN&access_num=rs361863&atom=%2Fcanres%2F71%2F8_Supplement%2F3805.atom