A potential small-molecule synthetic antilymphangiogenic agent norcantharidin inhibits tumor growth and lymphangiogenesis of human colonic adenocarcinomas through blocking VEGF-A,-C,-D/VEGFR-2,-3 ‚Äúmulti-points priming‚Ä?mechanisms in vitro and in vivo
Authors: Wei Sun
Jian-Jun Sun
Wei Zhu
Zhong-Yan Liu
Jing-Tao Zhang
Xin-Ping Li
Yue-Zu Fan
Wei Jing
Journal: BMC Cancer (2015)
Discipline: CLINICAL MEDICINE
Keywords: Tumor lymphangiogenesis; Norcantharidin; Antilymphangiogenic activity; Human colonic adenocarcinomas; In vivo
Affiliations: Department of Surgery, Tongji Hospital, Tongji University School of Medicine, Tongji University, Shanghai, 200065, People's Republic of China. ., Department of Surgery, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, 200072, People's Republic of China. ., Department of Surgery, Dahui Hospital, Postgraduate on-the-job in 2011 Grade, Tongji University School of Medicine, Shanghai, 200237, People's Republic of China. .

Abstract:  Background: Tumor lymphangiogenesis plays an important role in promoting growth and metastasis of tumors, but no antilymphangiogenic agent is used clinically. Based on the effect of norcantharidin (NCTD) on lymphangiogenesis of human lymphatic endothelial cells (LECs), we firstly investigated the antilymphangiogenic activity of NCTD as a tumor lymphangiogenic inhibitor for human colonic adenocarcinomas (HCACs). Methods: In vivo and in vitro experiments to determine the effects of NCTD on tumor growth and lymphangiogenesis of the in-situ colonic xenografts in nude mice, and lymphatic tube formation of the three-dimensional (3-D) of the co-culture system of HCAC HT-29 cells and LECs were done. Proliferation, apoptosis, migration, invasion, Ki-67, Bcl-2 and cell cycle of LECs and the co-culture system in vitro were respectively determined. Streparidin-peroxidase staining, SABC, western blotting and RT-PCR were respectively used to examine the expression of LYVE-1, D2-40, CK20 (including their LMVD), and VEGF-A, VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 in vitro and in vivo. Results: NCTD inhibited tumor growth and lymphangiogenesis of the in-situ colonic xenografts in vivo, and these observations were confirmed by facts that lymphatic tube formation, proliferation, apoptosis, migration, invasion, S-phase cell cycle, and Ki-67 and Bcl-2 expression in vitro, and LYVE-1, D2-40, CK20 expression and their LMVD in vitro and in vivo were inhibited and affected. Furthermore, the expression of VEGF-A, VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 at protein/mRNA levels in the process of lymphatic tube formation in vitro and tumor lymphangiogenesis in vivo was downregulated; NCTD in combination with mF4-31C1 or Sorafenib enhanced these effects. Conclusions: NCTD inhibits tumor growth and lymphangiogenesis of HCACs through "multi-points priming" mechanisms i.e. affecting related malignant phenotypes, inhibiting Ki-67 and Bcl-2 expression, inducing S-phase cell cycle arrest, and directly or indirectly downregulating VEGF-A,-C,-D/VEGFR-2,-3 signaling pathways. The present finding strongly suggests that NCTD could serve as a potential antilymphangiogenic agent for tumor lymphangiogenesis and is of importance to explore NCTD is used for antitumor metastatic comprehensive therapy for HCACs. 

Retracted: No

PubPeer comments:
 Figure 2A. * Red boxes: In the D2-40 row, the NC and NCTD panels appear to show an overlap, albeit shown at a different magnification  , 
 Figure 4A. * Blue boxes: Three panels appear to be showing the same specimen, albeit shown in different orientations and aspect ratios  , 
 Figure 6A. * Boxes of the same color show panels that look more similar than expected, albeit shown in different orientations or exposure.  , 
 Figure 6B. * Purple boxes: Two panels look quite similar.  , 
 Figure 6C. * Green boxes: In the Control column, the HDLECs and Co-Culture panel appear to overlap, if accounting for a 180 degrees rotation  , 
 Figure 7A and 7B: * Boxes of the same color - assigned by ImageTwin - highlight panels that appear to overlap  , 
 Figure 8A and 8B: * Boxes of the same color - assigned by ImageTwin - highlight panels that appear to overlap  , 
 Figure 9B and 10A: * Boxes and arrows of the same color highlight bands that look quite similar, as well as their orientation  This work was supported by a grant from the National Nature Science Foundation of China, No. 81072004. ,