Cancer Chemother Pharmacol DOI 10.1007/s00280-015-2745-4
ORIGINAL ARTICLE
Functional polymorphisms of ITGB1 are associated with clinical outcome of Chinese patients with resected colorectal cancer Feng Zhou1,2 · Xiaojun Huang3 · Zhaohui Zhang2 · Yibing Chen3 · Xiaonan Liu4 · Jinliang Xing3 · Xianli He1
Received: 3 November 2014 / Accepted: 2 April 2015 © Springer-Verlag Berlin Heidelberg 2015
Abstract Purpose Integrin β1 (ITGB1) has been recognized to play a major role in tumor growth, invasion and metastasis. However, effects of single-nucleotide polymorphisms (SNPs) in ITGB1 gene on the prognosis of patients with colorectal cancer (CRC) have not been reported. Methods A total of 372 patients with resected colorectal adenocarcinoma were enrolled in our study. Three functional SNPs (rs2230395, rs1187075 and rs1187076) in ITGB1 were selected and genotyped using the Sequenom iPLEX genotyping system. Results We identified two SNPs (rs2230395 and rs1187075) in ITGB1 gene to be significantly associated with
Feng Zhou and Xiaojun Huang have contributed to this work equally. Electronic supplementary material The online version of this article (doi:10.1007/s00280-015-2745-4) contains supplementary material, which is available to authorized users. * Jinliang Xing [email protected]
CRC overall survival (OS). Compared with the homozygous wild-type (AA) and heterozygous variant (AC), rs2230395 homozygous variant (CC) conferred a 1.55-fold (95 % CI 1.00–2.41, P = 0.049) increased risk of death. Similar result was obtained for homozygous variant (AA) in rs1187075 with a 1.62-fold (95 % CI 1.08–2.42, P = 0.020). In stratified analysis, this association in rs2230395 remained to be significant in patients receiving chemotherapy, but not in those without chemotherapy. We further evaluated the effects of chemotherapy on CRC survival in subgroups stratified by rs2230395 and rs1187075 genotypes. We found that chemotherapy resulted in a significantly better OS in patients with the homozygous wild-type (WW) or heterozygous variant (WV) genotype in both rs2230395 and rs1187075 when compared with patients with homozygous variant (VV) genotype. Conclusions Our data suggest that ITGB1 SNPs might be a prognostic biomarker for CRC patients, especially in those receiving chemotherapy. Our findings warrant validation in larger independent populations. Keywords Single-nucleotide polymorphism · Colorectal cancer · ITGB1 · Prognosis
* Xianli He [email protected] 1
Department of General Surgery, Tangdu Hospital, Fourth Military Medical University, 169 West Changle Street, Xi’an 710032, China
2
Department of General Surgery, Huaihai Hospital, Xuzhou Medical College, Xuzhou 221004, China
3
State Key Laboratory of Cancer Biology, Experimental Teaching Center of Basic Medicine, Fourth Military Medical University, 169 West Changle Street, Xi’an 710032, China
4
Xijing Hospital of Digestive Disease, Fourth Military Medical University, Xi’an 710032, China
Introduction Integrins are heterodimeric cell surface molecules that link the internal signaling components of the cytoskeleton to the extracellular protein microenvironment. There are 18 α and 8 β subunits of integrin, forming 24 unique integrin heterodimers with different extracellular matrix proteins [1]. These integrins transmit signals to promote diverse cellular responses including adhesion, survival and migration [2]. By far the most commonly found subunit in integrin
13
heterodimers is integrin β1 (ITGB1). Several studies have proved that ITGB1 is upregulated in lung [3], prostate [4] and ovarian cancer cells [5]. Later studies have reported that inhibition of ITGB1 expression can reduce cancer cell growth [6, 7], and the deletion of ITGB1 gene in mice dramatically impairs the metastatic dissemination [8, 9]. Furthermore, its overexpression is correlated with poor outcomes in many cancers, including breast cancer [10, 11], lung adenocarcinoma [12], prostate cancer [13] and pancreatic adenocarcinoma [14]. In addition, ITGB1 has been shown to mediate chemoresistance in several human cancers [15]. All these data strongly support the idea that ITGB1 gene is involved in the development and progression of human cancers. Colorectal cancer (CRC) is one of the most common malignancies in the world, and its morbidity and mortality are increasing rapidly in recent years [16]. In China, CRC has become one of the most common malignant tumors. Fortunately, evidences have shown that the mortality rate of CRC has decreased in Asian countries, possibly due to the early screening and detection, as well as the use of more advanced surgical and systemic modalities [17]. However, a considerable proportion of CRC patients develop recurrence or metastasis within 5 years after surgical treatment, highlighting the importance of novel biomarkers for the identification of those patients who are more likely to develop recurrence or metastasis and thus should receive more aggressive therapies. Single-nucleotide polymorphism (SNP) is the most common type of genetic variation, and numerous previous studies have shown that SNPs may be used as surrogates of patients’ genetic background to predict therapeutic response and prognosis. Our previous studies also demonstrate that some SNPs are significantly associated with OS in CRC patients [18–20]. Previous studies have demonstrated that SNPs in genes encoding integrins are associated with the susceptibility and prognosis of several types of malignancies [21, 22]. However, whether functional SNPs in ITGB1 gene have any influence on CRC patients’ clinical outcomes remains unclear. In this study, we assessed the effects of functional SNPs in ITGB1 gene on recurrence and survival in a cohort of 372 Chinese CRC patients.
Subjects and methods Study population Between February 2006 and March 2010, CRC patients were enrolled at Xijing and Tangdu Hospitals affiliated to the Fourth Military Medical University in Xi’an, China.
13
Cancer Chemother Pharmacol
The enrollment was based on the following criteria: (1) histologically confirmed with primary colorectal adenocarcinoma and no history of other cancers; (2) received curative surgical resection treatment, but without any preoperative anticancer treatment; and (3) with complete clinical and follow-up data, as well as common epidemiological data. Our study had a power of 80 % to detect a hazard ratio (HR) of 1.5 for SNP on an assumed 5-year OS rate of 70 % in the high-risk group and 65 % in the low-risk group with a two-sided log-rank test at a significance level of 0.05. The sample size was assessed as previously reported, and at least 357 patients were needed [23]. Finally, 372 patients were included in the present study. Before surgical resection, 5 mL of peripheral blood sample was collected from each patient for DNA preparation. This study was approved by the Ethic Committee of the Fourth Military Medical University, and the signed informed consent was obtained from each participant. Epidemiologic and clinical data collection Demographic and personal data were collected through in-person interview using a standardized epidemiological questionnaire, including gender, ethnicity and residential region. Detailed clinical information was collected through medical chart review or consultation with treating physicians, including time of diagnosis, time of surgery and/ or chemotherapy, time of recurrence and/or death, tumor stage, differentiation, location site, lymph node invasiveness, treatment protocol and serum carcinoembryonic antigen (CEA). A standard follow-up was performed at 6-month intervals by a trained clinical specialist through on-site interview, direct calling or medical chart review. The latest follow-up data in this analysis were obtained in January 2014. SNP selection and genotyping Functional SNPs in ITGB1 gene were selected using a set of web-based SNP selection tools (freely available at http://snpinfo.niehs.nih.gov/snpfunc.htm), by which one can select SNPs based on linkage disequilibrium (LD) and predicted functional characteristics of both coding and noncoding SNPs. The 5′ and 3′ flanking regions were arbitrarily set at 2000 bp for all genes. Only validated SNPs were selected, and SNPs with minor allele frequency (MAF) 0.85), only one SNP was included. Finally, a total of three SNPs in ITGB1 gene were selected for further genotyping, including two SNPs in transcription factor binding site of
Cancer Chemother Pharmacol Table 1 Distribution of patients’ characteristics and prognosis analysis Parameter
All patients, n (%) n = 372
OS
RFS a
Death, n (%) n = 130
HR
95 % CI
P value
Recurrence, n (%) n = 144
HRa
95 % CI
P value
Gender Female Male
164 (44.1) 208 (55.9)
55 (42.3) 75 (57.7)
Ref. 1.29
0.91–1.84
0.158
60 (41.7) 84 (58.3)
Ref. 1.35
0.96–1.88
0.083
Age ≤60 >60
180 (48.4) 192 (51.6)
68 (52.3) 62 (47.7)
Ref. 0.74
0.52–1.05
0.091
80 (55.6) 64 (44.4)
Ref. 0.65
0.47–0.91
0.013
Tumor position Colon Rectum
173 (46.5) 199 (53.5)
62 (47.7) 68 (52.3)
Ref. 1.06
0.75–1.50
0.754
66 (45.8) 78 (54.2)
Ref. 0.99
0.71–1.38
0.957
TNM stage I + II III + IV
249 (66.9) 123 (33.1)
67 (51.5) 63 (48.5)
Ref. 2.93
2.02–4.27
ORIGINAL ARTICLE
Functional polymorphisms of ITGB1 are associated with clinical outcome of Chinese patients with resected colorectal cancer Feng Zhou1,2 · Xiaojun Huang3 · Zhaohui Zhang2 · Yibing Chen3 · Xiaonan Liu4 · Jinliang Xing3 · Xianli He1
Received: 3 November 2014 / Accepted: 2 April 2015 © Springer-Verlag Berlin Heidelberg 2015
Abstract Purpose Integrin β1 (ITGB1) has been recognized to play a major role in tumor growth, invasion and metastasis. However, effects of single-nucleotide polymorphisms (SNPs) in ITGB1 gene on the prognosis of patients with colorectal cancer (CRC) have not been reported. Methods A total of 372 patients with resected colorectal adenocarcinoma were enrolled in our study. Three functional SNPs (rs2230395, rs1187075 and rs1187076) in ITGB1 were selected and genotyped using the Sequenom iPLEX genotyping system. Results We identified two SNPs (rs2230395 and rs1187075) in ITGB1 gene to be significantly associated with
Feng Zhou and Xiaojun Huang have contributed to this work equally. Electronic supplementary material The online version of this article (doi:10.1007/s00280-015-2745-4) contains supplementary material, which is available to authorized users. * Jinliang Xing [email protected]
CRC overall survival (OS). Compared with the homozygous wild-type (AA) and heterozygous variant (AC), rs2230395 homozygous variant (CC) conferred a 1.55-fold (95 % CI 1.00–2.41, P = 0.049) increased risk of death. Similar result was obtained for homozygous variant (AA) in rs1187075 with a 1.62-fold (95 % CI 1.08–2.42, P = 0.020). In stratified analysis, this association in rs2230395 remained to be significant in patients receiving chemotherapy, but not in those without chemotherapy. We further evaluated the effects of chemotherapy on CRC survival in subgroups stratified by rs2230395 and rs1187075 genotypes. We found that chemotherapy resulted in a significantly better OS in patients with the homozygous wild-type (WW) or heterozygous variant (WV) genotype in both rs2230395 and rs1187075 when compared with patients with homozygous variant (VV) genotype. Conclusions Our data suggest that ITGB1 SNPs might be a prognostic biomarker for CRC patients, especially in those receiving chemotherapy. Our findings warrant validation in larger independent populations. Keywords Single-nucleotide polymorphism · Colorectal cancer · ITGB1 · Prognosis
* Xianli He [email protected] 1
Department of General Surgery, Tangdu Hospital, Fourth Military Medical University, 169 West Changle Street, Xi’an 710032, China
2
Department of General Surgery, Huaihai Hospital, Xuzhou Medical College, Xuzhou 221004, China
3
State Key Laboratory of Cancer Biology, Experimental Teaching Center of Basic Medicine, Fourth Military Medical University, 169 West Changle Street, Xi’an 710032, China
4
Xijing Hospital of Digestive Disease, Fourth Military Medical University, Xi’an 710032, China
Introduction Integrins are heterodimeric cell surface molecules that link the internal signaling components of the cytoskeleton to the extracellular protein microenvironment. There are 18 α and 8 β subunits of integrin, forming 24 unique integrin heterodimers with different extracellular matrix proteins [1]. These integrins transmit signals to promote diverse cellular responses including adhesion, survival and migration [2]. By far the most commonly found subunit in integrin
13
heterodimers is integrin β1 (ITGB1). Several studies have proved that ITGB1 is upregulated in lung [3], prostate [4] and ovarian cancer cells [5]. Later studies have reported that inhibition of ITGB1 expression can reduce cancer cell growth [6, 7], and the deletion of ITGB1 gene in mice dramatically impairs the metastatic dissemination [8, 9]. Furthermore, its overexpression is correlated with poor outcomes in many cancers, including breast cancer [10, 11], lung adenocarcinoma [12], prostate cancer [13] and pancreatic adenocarcinoma [14]. In addition, ITGB1 has been shown to mediate chemoresistance in several human cancers [15]. All these data strongly support the idea that ITGB1 gene is involved in the development and progression of human cancers. Colorectal cancer (CRC) is one of the most common malignancies in the world, and its morbidity and mortality are increasing rapidly in recent years [16]. In China, CRC has become one of the most common malignant tumors. Fortunately, evidences have shown that the mortality rate of CRC has decreased in Asian countries, possibly due to the early screening and detection, as well as the use of more advanced surgical and systemic modalities [17]. However, a considerable proportion of CRC patients develop recurrence or metastasis within 5 years after surgical treatment, highlighting the importance of novel biomarkers for the identification of those patients who are more likely to develop recurrence or metastasis and thus should receive more aggressive therapies. Single-nucleotide polymorphism (SNP) is the most common type of genetic variation, and numerous previous studies have shown that SNPs may be used as surrogates of patients’ genetic background to predict therapeutic response and prognosis. Our previous studies also demonstrate that some SNPs are significantly associated with OS in CRC patients [18–20]. Previous studies have demonstrated that SNPs in genes encoding integrins are associated with the susceptibility and prognosis of several types of malignancies [21, 22]. However, whether functional SNPs in ITGB1 gene have any influence on CRC patients’ clinical outcomes remains unclear. In this study, we assessed the effects of functional SNPs in ITGB1 gene on recurrence and survival in a cohort of 372 Chinese CRC patients.
Subjects and methods Study population Between February 2006 and March 2010, CRC patients were enrolled at Xijing and Tangdu Hospitals affiliated to the Fourth Military Medical University in Xi’an, China.
13
Cancer Chemother Pharmacol
The enrollment was based on the following criteria: (1) histologically confirmed with primary colorectal adenocarcinoma and no history of other cancers; (2) received curative surgical resection treatment, but without any preoperative anticancer treatment; and (3) with complete clinical and follow-up data, as well as common epidemiological data. Our study had a power of 80 % to detect a hazard ratio (HR) of 1.5 for SNP on an assumed 5-year OS rate of 70 % in the high-risk group and 65 % in the low-risk group with a two-sided log-rank test at a significance level of 0.05. The sample size was assessed as previously reported, and at least 357 patients were needed [23]. Finally, 372 patients were included in the present study. Before surgical resection, 5 mL of peripheral blood sample was collected from each patient for DNA preparation. This study was approved by the Ethic Committee of the Fourth Military Medical University, and the signed informed consent was obtained from each participant. Epidemiologic and clinical data collection Demographic and personal data were collected through in-person interview using a standardized epidemiological questionnaire, including gender, ethnicity and residential region. Detailed clinical information was collected through medical chart review or consultation with treating physicians, including time of diagnosis, time of surgery and/ or chemotherapy, time of recurrence and/or death, tumor stage, differentiation, location site, lymph node invasiveness, treatment protocol and serum carcinoembryonic antigen (CEA). A standard follow-up was performed at 6-month intervals by a trained clinical specialist through on-site interview, direct calling or medical chart review. The latest follow-up data in this analysis were obtained in January 2014. SNP selection and genotyping Functional SNPs in ITGB1 gene were selected using a set of web-based SNP selection tools (freely available at http://snpinfo.niehs.nih.gov/snpfunc.htm), by which one can select SNPs based on linkage disequilibrium (LD) and predicted functional characteristics of both coding and noncoding SNPs. The 5′ and 3′ flanking regions were arbitrarily set at 2000 bp for all genes. Only validated SNPs were selected, and SNPs with minor allele frequency (MAF) 0.85), only one SNP was included. Finally, a total of three SNPs in ITGB1 gene were selected for further genotyping, including two SNPs in transcription factor binding site of
Cancer Chemother Pharmacol Table 1 Distribution of patients’ characteristics and prognosis analysis Parameter
All patients, n (%) n = 372
OS
RFS a
Death, n (%) n = 130
HR
95 % CI
P value
Recurrence, n (%) n = 144
HRa
95 % CI
P value
Gender Female Male
164 (44.1) 208 (55.9)
55 (42.3) 75 (57.7)
Ref. 1.29
0.91–1.84
0.158
60 (41.7) 84 (58.3)
Ref. 1.35
0.96–1.88
0.083
Age ≤60 >60
180 (48.4) 192 (51.6)
68 (52.3) 62 (47.7)
Ref. 0.74
0.52–1.05
0.091
80 (55.6) 64 (44.4)
Ref. 0.65
0.47–0.91
0.013
Tumor position Colon Rectum
173 (46.5) 199 (53.5)
62 (47.7) 68 (52.3)
Ref. 1.06
0.75–1.50
0.754
66 (45.8) 78 (54.2)
Ref. 0.99
0.71–1.38
0.957
TNM stage I + II III + IV
249 (66.9) 123 (33.1)
67 (51.5) 63 (48.5)
Ref. 2.93
2.02–4.27
