BCR/ABL inhibition by an escort/phosphatase fusion protein

Cellular transformation by the BCR/ABL oncogene depends on the ABL-encoded tyrosine kinase activity. To block BCR/ABL function, we created a unique tyrosine phosphatase by fusing the catalytic domain of SHP1 (SHP1c) to the ABL binding domain (ABD) of RIN1, an established binding partner and substrate for c-ABL and BCR/ABL. This fusion construct (ABD/SHP1c) binds to BCR/ABL in cells and functions as an active phosphatase. ABD/SHP1c effectively suppressed BCR/ABL function as judged by reductions in transformation of fibroblast cells, growth factor independence of hematopoietic cell lines, and proliferation of primary bone marrow cells. In addition, the leukemogenic properties of BCR/ABL in a murine model system were blocked by coexpression of ABD/SHP1c. Both the “escort” function provided by ABD and the inhibitor function provided by the phosphatase of SHP1c were necessary for effective BCR/ABL interference. Expression of ABD/SHP1c also reversed the transformed phenotype of K562, a human leukemia-derived cell line. These results have direct implications for leukemia therapeutics and suggest an approach to block aberrant signal transduction in other pathologies through the use of appropriately designed escort/inhibitors.

BCR/ABL癌基因所介导的细胞转化，依赖于ABL编码的酪氨酸激酶活性。为阻断BCR/ABL的功能，我们将SHP1的催化结构域（SHP1c）与RIN1的ABL结合结构域（ABD）融合，构建了一种独特的酪氨酸磷酸酶；RIN1是c-ABL与BCR/ABL的公认结合伴侣及底物。该融合构建体（ABD/SHP1c）可在细胞内与BCR/ABL结合，并作为活性磷酸酶发挥功能。通过检测成纤维细胞转化水平降低、造血细胞系的生长因子非依赖性减弱以及原代骨髓细胞增殖受抑，可证实ABD/SHP1c可有效抑制BCR/ABL的功能。此外，在小鼠模型系统中，共表达ABD/SHP1c可阻断BCR/ABL的致白血病特性。ABD所提供的“护送”功能与SHP1c磷酸酶所提供的抑制功能，二者均是有效干扰BCR/ABL功能所必需的。表达ABD/SHP1c还可逆转K562（一种人源白血病来源细胞系）的转化表型。本研究结果为白血病治疗提供了直接的理论依据，并提示可通过设计合适的“护送/抑制”双功能融合蛋白，阻断其他病理状态下的异常信号转导。