Japanese Journal of Clinical Oncology, 2015, 45(6) 533–540 doi: 10.1093/jjco/hyv037 Advance Access Publication Date: 13 March 2015 Original Article
Original Article
Laminin β3 expression as a prognostic factor and a predictive marker of chemoresistance in colorectal cancer Downloaded from http://jjco.oxfordjournals.org/ at Pennsylvania State University on November 17, 2015
Satomi Fukazawa1,*, Eiji Shinto1, Hitoshi Tsuda2, Hideki Ueno1, Atsushi Shikina1, Yoshiki Kajiwara1, Junji Yamamoto1, and Kazuo Hase1 1
Department of Surgery, National Defense Medical College, Saitama, and 2Department of Pathology, National Defense Medical College, Saitama, Japan *For reprints and all correspondence: Satomi Fukazawa, Department of Surgery, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513, Japan. E-mail: [email protected] Received 25 November 2014; Accepted 22 February 2015
Abstract Objective: Laminin-332, a marker of epithelial–mesenchymal transition, is composed of a heterotrimer of α3, β3 and γ2 chains that regulates cell adhesion and migration. This study aimed to disclose the respective clinical significance of laminin β3 immunoexpression in colorectal cancer as a prognostic factor and a predictive marker of chemoresistance. Methods: Tissue specimens from 323 Stage II and 232 Stage III colorectal cancer patients who underwent curative resection were assessed using laminin β3 immunostaining. Results: Among Stage III colorectal cancer patients, comparisons of 5-year disease-free survival rates revealed a poorer prognosis for the laminin β3-high group than for the laminin β3-low group (52.3 vs. 70.7%, P = 0.038), while there was no significant difference among Stage II patients. Among laminin β3-low Stage III patients, those who received adjuvant chemotherapy showed marginally better disease-free survival than those who did not receive it (75.8 vs. 62.8%; P = 0.096). Furthermore, multivariate analysis corroborated a distinct benefit of adjuvant chemotherapy in laminin β3-low patients (P = 0.035; hazard risk ratio = 1.66). Analyses of the laminin β3-high group, however, failed to show significance. Conclusions: Laminin β3 chain immunoreactivity was a poor prognostic factor for Stage III colorectal cancer patients, and laminin β3-high patients of Stage III colorectal cancer derived no survival benefit from adjuvant chemotherapy. Key words: laminin β3, colorectal cancer, prognostic factor, chemoresistance
Introduction Laminin (LN)-332 (previously known as laminin-5) is a heterotrimer composed of α3, β3 and γ2 chains that is essential for epithelial cell migration and adhesion to the basement membrane (1). Because the γ2 chain is specific to the LN-332 isoform, it is frequently used as a surrogate marker for the LN-332. The expression of the LN γ2 chain is characteristically detected in tumor cells at the invasion front or in budding tumor cells in many types of human cancers (2) and is a poor prognostic
factor for colorectal cancer (CRC) (3–5). However, there are few reports describing expression of the LN β3 chain, specific to the LN-332 isoform as with the LN γ2 chain, in cancer cells. LN-332 formation first requires assembly of β3/γ2 dimers, and then association of the α3 chain to form complete heterotrimers (6). In the absence of the LN α3 chain, the β3 and γ2 chains are secreted as monomers or as a heterodimer (7). Indeed, frequent coexpression of the β3 and γ2 chains in the cytoplasm has been shown in CRC (8,9) and gastric
© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected]
533
534
Clinical implication of laminin β3 in CRC
Patients and methods Patients We retrospectively analyzed the medical records of 555 Stage II or III CRC patients (319 males and 236 females; mean age, 65.1 years; Stage II, n = 323; Stage III, n = 232) who underwent curative resection between January 1997 and December 2005 in a single surgical unit at the National Defense Medical College Hospital (Tokorozawa, Japan). Inclusion criteria were curatively resected and histologically proven Stage II (T3-4N0M0) or III (any TN1-3M0) CRC in accordance with the Japanese Classification of Colorectal Carcinoma (26). All patients who received pre-operative chemotherapy or radiotherapy were excluded from analysis. The mean follow-up period after surgery was 78 months, during which 111 patients developed recurrence. Of the 232 Stage III CRC patients, 140 received adjuvant chemotherapy with 5-FU-based regimens (Mayo, n = 62; oral tegafur-uracil/leucovorin (LV), n = 59; Roswell Park Memorial Institute, n = 12; and simplified bimonthly LV and 5-FU, n = 7). Written informed consent was obtained from each patient in accordance with institutional regulations, and the study protocol was approved by the Ethics Committee of the National Defense Medical College Hospital.
Immunohistochemical analysis Immunohistochemical staining was performed as described elsewhere (5). Briefly, resected tissue containing the tumor invasive front was embedded in paraffin and 4 µm thick sections were immunostained for LN β3 (monoclonal mouse anti-human LN-5 β-3 chain; clone Human LN β3-short arm; dilution, 1:50; BD Oriental Yeast Co., Tokyo, Japan) and LN γ2 (monoclonal mouse anti-human LN-5 γ2 chain; clone D4B5; dilution, 1:200; Chemicon, Temecula, CA, USA). Positive staining for LN β3 and γ2 in the cytoplasm of cancer cells was regarded as immunoreactive and classified as either high (≥30%) or low (
Original Article
Laminin β3 expression as a prognostic factor and a predictive marker of chemoresistance in colorectal cancer Downloaded from http://jjco.oxfordjournals.org/ at Pennsylvania State University on November 17, 2015
Satomi Fukazawa1,*, Eiji Shinto1, Hitoshi Tsuda2, Hideki Ueno1, Atsushi Shikina1, Yoshiki Kajiwara1, Junji Yamamoto1, and Kazuo Hase1 1
Department of Surgery, National Defense Medical College, Saitama, and 2Department of Pathology, National Defense Medical College, Saitama, Japan *For reprints and all correspondence: Satomi Fukazawa, Department of Surgery, National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513, Japan. E-mail: [email protected] Received 25 November 2014; Accepted 22 February 2015
Abstract Objective: Laminin-332, a marker of epithelial–mesenchymal transition, is composed of a heterotrimer of α3, β3 and γ2 chains that regulates cell adhesion and migration. This study aimed to disclose the respective clinical significance of laminin β3 immunoexpression in colorectal cancer as a prognostic factor and a predictive marker of chemoresistance. Methods: Tissue specimens from 323 Stage II and 232 Stage III colorectal cancer patients who underwent curative resection were assessed using laminin β3 immunostaining. Results: Among Stage III colorectal cancer patients, comparisons of 5-year disease-free survival rates revealed a poorer prognosis for the laminin β3-high group than for the laminin β3-low group (52.3 vs. 70.7%, P = 0.038), while there was no significant difference among Stage II patients. Among laminin β3-low Stage III patients, those who received adjuvant chemotherapy showed marginally better disease-free survival than those who did not receive it (75.8 vs. 62.8%; P = 0.096). Furthermore, multivariate analysis corroborated a distinct benefit of adjuvant chemotherapy in laminin β3-low patients (P = 0.035; hazard risk ratio = 1.66). Analyses of the laminin β3-high group, however, failed to show significance. Conclusions: Laminin β3 chain immunoreactivity was a poor prognostic factor for Stage III colorectal cancer patients, and laminin β3-high patients of Stage III colorectal cancer derived no survival benefit from adjuvant chemotherapy. Key words: laminin β3, colorectal cancer, prognostic factor, chemoresistance
Introduction Laminin (LN)-332 (previously known as laminin-5) is a heterotrimer composed of α3, β3 and γ2 chains that is essential for epithelial cell migration and adhesion to the basement membrane (1). Because the γ2 chain is specific to the LN-332 isoform, it is frequently used as a surrogate marker for the LN-332. The expression of the LN γ2 chain is characteristically detected in tumor cells at the invasion front or in budding tumor cells in many types of human cancers (2) and is a poor prognostic
factor for colorectal cancer (CRC) (3–5). However, there are few reports describing expression of the LN β3 chain, specific to the LN-332 isoform as with the LN γ2 chain, in cancer cells. LN-332 formation first requires assembly of β3/γ2 dimers, and then association of the α3 chain to form complete heterotrimers (6). In the absence of the LN α3 chain, the β3 and γ2 chains are secreted as monomers or as a heterodimer (7). Indeed, frequent coexpression of the β3 and γ2 chains in the cytoplasm has been shown in CRC (8,9) and gastric
© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected]
533
534
Clinical implication of laminin β3 in CRC
Patients and methods Patients We retrospectively analyzed the medical records of 555 Stage II or III CRC patients (319 males and 236 females; mean age, 65.1 years; Stage II, n = 323; Stage III, n = 232) who underwent curative resection between January 1997 and December 2005 in a single surgical unit at the National Defense Medical College Hospital (Tokorozawa, Japan). Inclusion criteria were curatively resected and histologically proven Stage II (T3-4N0M0) or III (any TN1-3M0) CRC in accordance with the Japanese Classification of Colorectal Carcinoma (26). All patients who received pre-operative chemotherapy or radiotherapy were excluded from analysis. The mean follow-up period after surgery was 78 months, during which 111 patients developed recurrence. Of the 232 Stage III CRC patients, 140 received adjuvant chemotherapy with 5-FU-based regimens (Mayo, n = 62; oral tegafur-uracil/leucovorin (LV), n = 59; Roswell Park Memorial Institute, n = 12; and simplified bimonthly LV and 5-FU, n = 7). Written informed consent was obtained from each patient in accordance with institutional regulations, and the study protocol was approved by the Ethics Committee of the National Defense Medical College Hospital.
Immunohistochemical analysis Immunohistochemical staining was performed as described elsewhere (5). Briefly, resected tissue containing the tumor invasive front was embedded in paraffin and 4 µm thick sections were immunostained for LN β3 (monoclonal mouse anti-human LN-5 β-3 chain; clone Human LN β3-short arm; dilution, 1:50; BD Oriental Yeast Co., Tokyo, Japan) and LN γ2 (monoclonal mouse anti-human LN-5 γ2 chain; clone D4B5; dilution, 1:200; Chemicon, Temecula, CA, USA). Positive staining for LN β3 and γ2 in the cytoplasm of cancer cells was regarded as immunoreactive and classified as either high (≥30%) or low (
