The Role of PI3K Beta in Breast Cancer Metastasis

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Tumor metastasis is the major cause of mortality in human breast cancer. Previous studies have shown that breast cancer metastasis is driven by paracrine signaling between tumor cells and stromal cells, which promotes invasion, intravasation, extravasation and tumor growth at secondary sites. This paracrine signaling is dependent on the reciprocal production of growth factors, cytokines and chemokines produced by stromal cells and tumor cells, many of which signal via G-protein-coupled receptors (GPCRs). We now present extensive preliminary data showing that GPCR signaling to PI3Kβ is critical for tumor cell invasion, intravasation and extravasation. Importantly, loss of GPCR signaling to PI3Kβ has a more severe phenotype on tumor intravasation and extravasation in vivo than loss of kinase activity, suggesting that inhibition of p110β-Gβγ binding might provide an alternative therapeutic approach for the prevention of breast cancer metastasis. This proposal examines the role of PI3Kβ in breast cancer metastasis, using both in vitro and in vivo approaches. The first aim comprises mechanistic studies to evaluate the role of PI3Kβ in the formation of invadopodia, which allow tumor cells to invade into surrounding tissue. We will focus on two models of p110β function in invadopodia maturation: (a) as a local source of PI[3,4,5]P3, whose metabolism to PI[3,4]P2 recruits the critical invadopodia protein Tks5; and (b) as a regulator of integrin signaling, which is important for invadopodia maturation and MMP secretion. Aim 2 examines how p110β integrates upstream signals from GPCRs, RTKs and Rac1, and examines the role of Rac1 signaling to PI3Kβ in breast cancer metastasis. In particular, we find that mutation of the Gβγ binding site in p110β has no effect on Rac1GTP binding and activation of p110β in vitro, but blocks PI3Kβ activation by constitutively active Rac1 in cells. Similarly, inhibitors that block the binding of the p85 regulatory subunit to tyrosine-phosphorylated proteins also inhibit PI3Kβ activation by CA-Rac. We will explore two hypotheses to explain these data: first, that Rac binding to PI3Kβ in cells requires the targeting of PI3Kβ to the membrane, and second, that activation of PI3Kβ by Gβγ or SH2-mediated interactions sensitizes PI3Kβ to Rac. We will also directly test the role of Rac binding to p110β in breast cancer metastasis using in vitro and in vivo xenograft models. Finally, we will study the role of PI3Kβ in breast cancer metastasis using an established genetic mouse model, MMTV-PyMT, which develops mammary epithelial tumors with high penetrance and has an intact immune system. We will cross PyMT mice to a knock-in mouse expressing the GPCR-uncoupled mutant of p110β, to definitively establish the role of GPCR signaling to PI3Kβ in tumor progression and metastasis. Altogether, these studies will lead to important new insights into the basic biology of PI3Kβ, and the role of this complex signaling enzyme in breast cancer metastasis.