M. P. Kallis1,3, C. Tzanavaris1,3, S. Wang1,3, C. Maloney1,3, M. Symons1,3, B. M. Steinberg1,3, S. Z. Soffer2,3 1The Elmezzi Graduate School of Molecular Medicine,Northwell Health,Manhasset, NY, USA 2Cohen Children’s Medical Center, Northwell Health,Division Of Pediatric Surgery,New Hyde Park, NY, USA 3The Feinstein Institute for Medical Research,Center For Oncology And Cell Biology,Manhasset, NY, USA
Introduction:
Excision of the primary tumor is the mainstay of treatment for osteosarcoma (OS), but may result in a significant increase in pulmonary metastasis. Previous studies in our lab have shown that gefitinib decreases surgically accelerated pulmonary metastasis in an animal model of OS via its effect on tumor associated macrophages. Angiogenesis is essential for tumor growth and metastasis, but also increases following systemic inflammation and surgery. The purpose of this study is to determine whether vascular endothelial growth factor receptor (VEGFR), used as a marker of a pro-angiogenic tumor environment, specifically increases in response to removal of the primary tumor or is a general response to surgical stress.
Methods:
K7M2 OS cells were injected into the tibia of syngeneic BALB/c mice. One week post implantation, mice were randomly assigned to 4 groups: control, amputation of primary tumor, amputation of contralateral limb, amputation of primary tumor plus perioperative gefitinib. Mice were sacrificed 3 weeks later, and lungs were harvested to assess pulmonary metastatic nodules. Lung sections (10 high-power fields per mouse) were analyzed by immunohistochemistry for the presence of VEGFR, quantified by ImageJ photo processing software.
Results:
Mice that underwent amputation of the primary tumor had increased pulmonary metastases compared to tumor bearing controls (17 vs 9.5, p<0.05). Histological lung sections from these two groups were analyzed for VEGFR positivity. Lung sections from mice that underwent amputation of tumor showed an increase in the number of VEGF+ cells (105.6 vs 74.8 per field, p<0.05). Surgical stress, represented by amputation of the contralateral limb, had no effect on metastasis with metastatic nodules akin to controls (7 vs 9.5, p<0.05). Lung sections from the contralateral amputation group showed a marked reduction in VEGFR+ cells (58.4 per field, p<0.05). Gefitinib suppressed both the increase of gross metastatic nodules and the increase of VEGFR+ cells induced by amputation of the primary tumor (6 vs 17, p<0.05; 74 vs 105.6 per field, p<0.05).
Conclusions:
Post-surgical metastatic enhancement reflects distinct microenvironmental changes that occur upon removal of the primary tumor. Among these is a pro-angiogenic tumor environment. These changes are independent of intrinsic surgical stress. Gefitinib prevents this pro-angiogenic switch, further supporting the use of gefitinib as a clinically important anti-metastatic strategy.