Sledge G W, Qulali M, Goulet R, Bone E A, Fife R
Department of Medicine, Indiana University School of Medicine, Indianapolis, USA.
J Natl Cancer Inst. 1995 Oct 18;87(20):1546-50. doi: 10.1093/jnci/87.20.1546.
Matrix metalloproteinases (MMPs) are involved in the invasion and metastasis of human cancers by mediating the degradation of extracellular matrix components. Therefore, these enzymes constitute promising targets in the development of anticancer therapies. Batimastat ([(4-N-hydroxyamino)-2R-isobutyl-3S-(thienyl-thiomethyl)succinyl]-L- phenyl-alanine-N-methylamide) is one of a new class of agents designed to inhibit MMP activity.
We asked whether batimastat, given as adjuvant therapy after primary tumor resection, could inhibit local-regional tumor regrowth and the formation of lung metastases in a human breast cancer xenograft model. We also explored possible effects of batimastat on breast cancer cell viability and on the accumulation of specific messenger RNAs (mRNAs).
Human MDA-MB-435 breast cancer cells were treated in vitro for 6 days with batimastat at concentrations ranging from 0.1 to 10.0 microM, and then viable cell counts were performed. The activity of collagenases, directly associated with cultured MDA-MB-435 cells or released into their culture fluids, was assessed by gelatin zymography after 1 and 3 days of batimastat treatment (drug range, 0.2-2.0 microM). Athymic nude mice were given daily intraperitoneal injections of batimastat (30 mg/kg body weight) after resection of MDA-MB-435 primary tumors grown in their mammary fat pads; the volumes of tumor regrowths and the numbers and volumes of lung metastases were calculated; neovascularization in the regrowths was assessed by immunohistochemical analysis with an antibody directed against CD31, an endothelial cell antigen. The effect of batimastat treatment on the accumulation of mRNAs encoding specific MMPs and the tissue inhibitor of metalloproteinases-2 (TIMP-2) in cultured cells, primary tumors, and tumor regrowths was measured by RNA dot blotting and hybridization with complementary probes. Linear regression analysis, Student's t tests, and chi-squared analysis were used to evaluate the data.
The viability of cultured MDA-MB-435 cells was not affected by treatment with batimastat; however, measured activities for the 72-kd and 92-kd collagenases released by these cells were reduced after batimastat treatment. Intraperitoneal injection of batimastat significantly inhibited the local-regional regrowth of resected MDA-MB-435 tumors in athymic nude mice (in comparison with control mice, P = .035), and it reduced the incidence (P < .05), number (P = .0001), and total volume (P = .0001) of lung metastases. Batimastat treatment did not affect cellular levels of MMP or TIMP-2 mRNAs.
Batimastat inhibits human breast cancer regrowth and metastasis in a nude mouse xenograft model. Potential mechanisms for batimastat's inhibitory activity do not include direct cell toxicity or alteration of MMP or TIMP mRNA levels.
基质金属蛋白酶（MMPs）通过介导细胞外基质成分的降解参与人类癌症的侵袭和转移。因此，这些酶是抗癌治疗开发中很有前景的靶点。batimastat（[(4-N-羟基氨基)-2R-异丁基-3S-(噻吩基-硫甲基)琥珀酰]-L-苯丙氨酸-N-甲基酰胺）是一类旨在抑制MMP活性的新型药物之一。
我们研究了在原发性肿瘤切除后给予batimastat作为辅助治疗，是否能在人乳腺癌异种移植模型中抑制局部区域肿瘤再生和肺转移的形成。我们还探讨了batimastat对乳腺癌细胞活力以及特定信使核糖核酸（mRNAs）积累的可能影响。
将人MDA-MB-435乳腺癌细胞在体外以0.1至10.0微摩尔的浓度用batimastat处理6天，然后进行活细胞计数。在batimastat处理1天和3天后（药物浓度范围为0.2 - 2.0微摩尔），通过明胶酶谱法评估与培养的MDA-MB-435细胞直接相关或释放到其培养液中的胶原酶活性。在切除在无胸腺裸鼠乳腺脂肪垫中生长的MDA-MB-435原发性肿瘤后，每天给无胸腺裸鼠腹腔注射batimastat（30毫克/千克体重）；计算肿瘤再生的体积以及肺转移的数量和体积；用针对内皮细胞抗原CD31的抗体通过免疫组织化学分析评估再生肿瘤中的新生血管形成。通过RNA斑点印迹法并用互补探针杂交，测量batimastat处理对培养细胞、原发性肿瘤和肿瘤再生中编码特定MMPs和金属蛋白酶组织抑制剂-2（TIMP-2）的mRNAs积累的影响。使用线性回归分析、学生t检验和卡方分析来评估数据。
培养的MDA-MB-435细胞的活力不受batimastat处理的影响；然而，batimastat处理后这些细胞释放的72-kd和92-kd胶原酶的测量活性降低。腹腔注射batimastat显著抑制了无胸腺裸鼠中切除的MDA-MB-435肿瘤的局部区域再生（与对照小鼠相比，P = 0.035），并且它降低了肺转移的发生率（P < 0.05）、数量（P = 0.0001）和总体积（P = 0.0001）。batimastat处理不影响细胞中MMP或TIMP-2 mRNAs的水平。
batimastat在裸鼠异种移植模型中抑制人乳腺癌的再生和转移。batimastat抑制活性的潜在机制不包括直接细胞毒性或MMP或TIMP mRNA水平的改变。
