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Project 1: Inhibition of PI3 kinase as a strategy to abrogate antiestrogen resistance in breast cancer

Specific Aims

Antiestrogen therapies have changed the natural history of hormone-dependent, estrogen receptor-positive (ER+) human breast cancer. However, many ER+ tumors develop drug resistance and progress, with more patients still dying from ER+ breast cancer than all other breast cancer types combined. For the majority of these cancers, mechanisms of escape from antiestrogens remain to be discovered. During the current award period, we have shown that activation of the phosphatidylinositol-3 kinase (PI3K) pathway can promote resistance to endocrine therapy, though demonstration of this mechanism awaits further confirmation in the clinic. The PI3K pathway is overall the most frequently altered oncogenic pathway in cancer. Mutations in PIK3CA, the gene encoding the p110α catalytic subunit of PI3K, are the most common somatic alterations of this pathway in breast cancer, where ≥80% occur within the helical (E542K, E545K) and kinase (H1047R) domains of p110α. These mutations confer increased PIP3-forming catalytic activity and induce a transformed phenotype including growth factor- and anchorage-independent growth, resistance to anoikis, and drug resistance. Small molecule pan-PI3K inhibitors that bind reversibly to the ATP pocket of p110 have completed phase I trials. Preclinical and few clinical studies have already suggested that ER+/PI3K mutant tumors exhibit a lower response to antiestrogens compared to ER+/PI3K wild-type tumors. Thus, we hypothesize that antiestrogens in combination with a PI3K inhibitor will be more effective against ER+/PI3K mutant breast cancers compared to the antiestrogen alone. In addition, breast cancers that do not respond to the combination will contain somatic alterations causally associated with drug resistance. To test these hypotheses, we propose the following specific aims:

Aim 1: To determine the rate of pathological complete response (path CR) in patients with ER+/HER2– breast cancer treated with the aromatase inhibitor letrozole and a PI3K inhibitor. Patients with stage II-III ER+/HER2– (with wild-type or mutant PIK3CA) breast cancer will be randomized in a 2:1 ratio to letrozole + placebo vs. letrozole + the pan-PI3K inhibitor BKM120 for 24 weeks. An ultrasound-guided biopsy and FDG-PET/CT will be performed at two weeks to determine if BKM120 increases letrozole action as measured by cellular, molecular and metabolic markers of tumor growth. The primary objective of this trial is the rate of path CR, defined as absence of cancer in the breast and lymph nodes, at the time of definitive surgery at 24 weeks.

Aim 2: To identify molecular alterations potentially associated with drug resistance in breast cancers after neoadjuvant therapy with letrozole plus BKM120. DNA will be harvested from pre-treatment and surgical (posttherapy) biopsies and subjected to whole exome sequencing. To validate copy-number variations, somatic single-nucleotide variants and insertion-deletions, and for tumors with scarce tissue, low cellularity and/or high heterogeneity, a (275) gene-targeted capture assay will be used.

Aim 3: To determine whether molecular alterations identified in post-treatment residual cancers are causally associated with resistance to the combination of letrozole and the PI3K inhibitor. Higher frequency mutations/amplifications identified in Aim 2 will be tested using conventional shRNA-based and overexpression approaches in human breast cancer cells and xenografts to determine a causal association with drug resistance. As a complementary approach and using a kinase-open reading frame (ORF) expression system, finally we will interrogate actionable mediators of resistance to antiestrogen therapy plus BKM120 in the human kinome.