Pernasetti Flavia, Nickel Jeffrey, Clark Derek, Baeuerle Patrick A, Van Epps Dennis, Freimark Bruce
Cell-Matrix, Inc., a subsidiary of Micromet Corporation, Carlsbad, CA 92008, USA.
Int J Oncol. 2006 Dec;29(6):1371-9.
Matrix metalloproteases (MMPs) secreted by both tumor and endothelial cells proteolytically degrade collagen during tumor growth and neo-vascularization. This exposes cryptic binding sites on collagen with functional relevance for angiogenesis. In this report, we characterized a novel humanized monoclonal IgG1 antibody, D93. After humanization, the antibody retained the binding specificity of the parental murine IgM antibody for denatured (dn) collagen. D93 recognized dn-collagen but not native (nat) collagen of different species, including mouse, chicken, and human, indicating that its cryptic binding site(s) is conserved across species. In immunohistochemistry (IHC) studies, D93 stained the basement membrane of blood vessels in several xenograft human tumors or in surgically removed tumor tissues from patients with different types of malignancies. D93 staining was rarely or not present in normal blood vessels of healthy tissues. In in vivo experiments, D93 significantly inhibited basic fibroblast growth factor (bFGF)-induced angiogenesis in chick embryo chorioallantoic membrane (CAM) and the tumor growth of pre-established orthotopic human breast (MDA-MB-435) tumors in mice. D93 i.v. administered in mice was subsequently detected in the subendothelial basement membrane of tumor blood vessels but not blood vessels of normal tissues. Inhibition of growth of pre-established orthotopic human breast MDA-MB-435 tumors was more effective when D93 was combined with Taxol, than either treatment alone. In addition, tumors from animals treated with D93 and/or Taxol showed significantly reduced levels of the endothelial cell-marker CD31. Our data suggest that blockade of cryptic epitopes exposed on collagen IV during angiogenesis and tumor growth by a monoclonal antibody may provide a novel therapeutic modality for treatment of cancer and pathogenic neo-vascularization.
肿瘤细胞和内皮细胞分泌的基质金属蛋白酶（MMPs）在肿瘤生长和新血管形成过程中通过蛋白水解作用降解胶原蛋白。这暴露了胶原蛋白上对血管生成具有功能相关性的隐蔽结合位点。在本报告中，我们鉴定了一种新型人源化单克隆IgG1抗体D93。人源化后，该抗体保留了亲本鼠IgM抗体对变性（dn）胶原蛋白的结合特异性。D93识别dn - 胶原蛋白，但不识别包括小鼠、鸡和人在内的不同物种的天然（nat）胶原蛋白，这表明其隐蔽结合位点在物种间是保守的。在免疫组织化学（IHC）研究中，D93在几种异种移植的人类肿瘤或从患有不同类型恶性肿瘤的患者手术切除的肿瘤组织中，对血管基底膜进行了染色。在健康组织的正常血管中，D93染色很少或不存在。在体内实验中，D93显著抑制了鸡胚绒毛尿囊膜（CAM）中碱性成纤维细胞生长因子（bFGF）诱导的血管生成以及小鼠体内预先建立的原位人乳腺（MDA - MB - 435）肿瘤的生长。随后在小鼠静脉注射D93后，在肿瘤血管的内皮下基底膜中检测到了该抗体，但在正常组织的血管中未检测到。当D93与紫杉醇联合使用时，对预先建立的原位人乳腺MDA - MB - 435肿瘤生长的抑制作用比单独使用任何一种治疗方法都更有效。此外，用D93和/或紫杉醇治疗的动物的肿瘤中，内皮细胞标志物CD31的水平显著降低。我们的数据表明，通过单克隆抗体阻断血管生成和肿瘤生长过程中胶原蛋白IV上暴露的隐蔽表位，可能为癌症治疗和病理性新血管形成提供一种新的治疗方式。
