The Type I Insulin-like Growth Factor Receptor (IGF-IR) clearly plays a critical role in breast tumorigenesis through its capacity to facilitate malignant transformation, promote cell proliferation, and protect malignant cells from apoptosis. The human IGF-IR mRNA contains an extraordinarily long (1,038 nucleotides) leader sequence (5'-untranslated RNA, 5'-UTR), which functions as the equivalent of a """"""""promoter"""""""" at the RNA level. We have identified three regulatory proteins (HuR, TIAR, hnRNP C) that bind in a sequence-specific manner to the IGF-IR 5'-UTR and appear to control the efficiency with which the IGF1R mRNA is translated into protein. We hypothesize that alterations in the dynamic interactions between these RNA-binding proteins and the 5'-untranslated sequence of the IGF-IR transcript may be responsible for IGF-IR overexpression in a proportion of human breast tumors, and ultimately contribute to the molecular pathogenesis of this disease. Indeed we have found a marked increase in IGF-IR translational efficiency in association with significant alterations in the activities of these RNA-binding proteins in the human breast carcinoma cell line T47D, which overexpresses IGF-IR. In addition, we propose to test the potential therapeutic utility of the isolated IGF-IR 5'-untranslated RNA, functioning as a dominant negative regulatory RNA, to specifically counteract IGF-IR overexpression and reverse the associated adverse phenotypic consequences in human breast cancer cells. We have established that such a strategy can be used to induce dramatic, favorable phenotypic alterations, including mitotic cell death and loss of tumorigenicity.
The Specific Aims are: 1. Establish the function of the RNA-binding proteins TIAR and hnRNP C in the regulation of IGFIR expression at the translational level. 2. Assay primary human breast tumor specimens for the activities of sequence, specific translation-regulatory proteins binding the 5'-untranslated region of the human IGFIR transcript (including HuR, TIAR, and hnRNP C), and correlate changes in activities of these RNA-binding proteins with IGF-IR overexpression and the malignant phenotype. 3. Ectopically express the isolated IGFIR 5'-untranslated RNA sequence in human breast cancer cells, assess the potential of this non-coding RNA sequence to modulate IGF1R expression, and consequently limit proliferative capacity, enhance apoptotic susceptibility, and reverse tumorigenicity of human breast cancer cells.
