Overexpression of the amplified Pip4k2β gene from 17q11–12 in breast cancer cells confers proliferation advantage

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Abstract
Gene amplification is common in solid tumors and is associated with adverse prognosis, disease progression, and development of drug resistance. A small segment from chromosome 17q11-12 containing the HER-2/Neu gene is amplified in about 25% of breast cancer. HER-2/Neu amplification is associated with adverse prognosis and may predict response to chemotherapy and hormonal manipulation. Moreover, HER-2/Neu amplification may select patients for anti-HER-2/Neu-based therapy with Herceptin. We and others recently described a common sequence element from the HER-2/Neu region that was amplified in breast cancer cells. In addition, most, if not all, of the amplified genes from this region display overexpression. This raises the intriguing possibility that genes immediately adjacent to HER-2/Neu may influence the biological behavior of breast cancer carrying HER-2/Neu amplification and serve as rational targets for therapy. By extracting sequence information from public databases, we have constructed a contig in bacterial artificial chromosomes (BACs) that extends from HER-2/Neu to a phosphotidylinositol phosphate kinase (PIPK), Pip4k2beta from 17q11-12. Although a role of PI-3-kinase and AKT in cancer biology has been previously described, PIPK has not been previously implicated. We show that Pip4k2beta, initially known as Pip5k2beta, is amplified in a subset of breast cancer cell lines and primary breast cancer samples that carry HER-2/Neu amplification. Out of eight breast cancer cell lines with HER-2/Neu amplification, three have concomitant amplification of the Pip4k2beta gene--UACC-812, BT-474 and ZR-75-30. Similarly, two out of four primary breast tumors with HER-2/Neu amplification carry Pip4k2beta gene amplification. Intriguingly, one tumor displays an increase in the gene copy number of Pip4k2beta that is significantly more than that of HER-2/Neu. Moreover, dual color FISH reveals that amplified Pip4k2beta gene may exist in a distinct structure from that of HER-2/Neu in ZR-75-30 cell line. These studies suggest that Pip4k2beta may reside on an amplification maximum distinct from that of HER-2/Neu and serve as an independent target for amplification and selective retention. Pip4k2beta amplification is associated with overexpression at the RNA and protein level in breast cancer cell lines. Stable expression of Pip4k2beta in breast cancer cell lines with and without HER-2/Neu amplification increases cell proliferation and anchorage-independent growth. The above observations implicate Pip4k2beta in the development and/or progression of breast cancer. Our study suggests that Pip4k2beta may be a distinct target for gene amplification and selective retention from 17q11-12.