Tseng Y H, Vicent D, Zhu J, Niu Y, Adeyinka A, Moyers J S, Watson P H, Kahn C R
Joslin Diabetes Center, Department of Medicine, Harvard Medical School, Boston, Massachusetts 02215, USA.
Cancer Res. 2001 Mar 1;61(5):2071-9.
Rad is the prototypic member of a family of novel Ras-related GTPases that is normally expressed in heart, skeletal muscle, and lung and that has been shown to exhibit a novel form of bi-directional interaction with the nm23 metastasis suppressor. In the present study, we have investigated the expression of Rad in normal and neoplastic breast tissues by Western blot and immunohistochemistry and the functional effect of altered Rad expression in breast cancer cell lines. We found that, although Rad is frequently expressed in normal breast tissue (23/30 Rad+ve), expression is usually lost in adjacent invasive carcinoma (8/30 Rad+ve; P < 0.0001). However, where Rad expression persists in a small proportion of tumors, it is associated with higher grade, larger size, and extensive axillary nodal involvement (n = 48; P = 0.035, P = 0.016, P = 0.022, respectively). Furthermore, Rad is also highly expressed in a breast cancer cell line with high tumorigenic and metastatic potential (MDA-MB231). To further examine the role of Rad in breast cancer, we stably transfected a Rad-ve breast cancer cell line (MDA-MB435). We observed an increase in growth and marked increased colony formation in soft agar in vitro (P < 0.05) and an increase in tumor growth rate in nude mice (P < 0.05). Moreover, coexpression of nm23 with wild-type Rad inhibited the effect of Rad on growth of these cells in culture and markedly inhibited tumor growth in vivo. Additional transfection studies with mutated Rad cDNAs revealed that the growth-promoting effects of Rad appeared to be mediated through its NH2- and COOH-terminal regions, rather than its GTPase domain, and might involve acceleration of cell cycle transition. These findings suggest that Rad may act as an oncogenic protein in breast tissues and demonstrate a potential mechanism by which interaction between Rad and nm23 may regulate growth and tumorigenicity of breast cancer.
Rad是一类新型Ras相关GTP酶家族的原型成员，通常在心脏、骨骼肌和肺中表达，并且已显示出与nm23转移抑制因子呈现一种新型的双向相互作用形式。在本研究中，我们通过蛋白质印迹法和免疫组织化学研究了Rad在正常和肿瘤性乳腺组织中的表达，以及Rad表达改变对乳腺癌细胞系的功能影响。我们发现，尽管Rad在正常乳腺组织中经常表达（23/30 Rad阳性），但在相邻的浸润性癌中表达通常缺失（8/30 Rad阳性；P<0.0001）。然而，在一小部分肿瘤中Rad表达持续存在的情况下，它与更高的分级、更大的尺寸以及广泛的腋窝淋巴结受累相关（n = 48；P分别为0.035、0.016、0.022）。此外，Rad在具有高致瘤性和转移潜能的乳腺癌细胞系（MDA-MB231）中也高度表达。为了进一步研究Rad在乳腺癌中的作用，我们稳定转染了一个Rad阴性的乳腺癌细胞系（MDA-MB435）。我们观察到体外软琼脂中生长增加且集落形成显著增加（P<0.05），以及裸鼠体内肿瘤生长速率增加（P<0.05）。此外，nm23与野生型Rad共表达抑制了Rad对这些细胞在培养中的生长作用，并显著抑制了体内肿瘤生长。用突变的Rad cDNA进行的额外转染研究表明，Rad的促生长作用似乎是通过其NH2端和COOH端区域介导的，而不是通过其GTP酶结构域，并且可能涉及细胞周期转换的加速。这些发现表明Rad可能在乳腺组织中作为一种致癌蛋白起作用，并证明了Rad与nm23之间的相互作用可能调节乳腺癌生长和致瘤性的潜在机制。
