The Prolyl Isomerase Pin1 in Breast Development and Cancer

The prolyl isomerase Pin1 specifically isomerizes certain phosphorylated Ser/Thr-Pro bonds and thereby regulates various cellular processes. Pin1 is a target of several oncogenic pathways and is overexpressed in human breast cancer. Its overexpression can lead to upregulation of cyclin D1 and transformation of breast epithelial cells in collaboration with the oncogenic pathways. In contrast, inhibition of Pin1 can suppress the transformation of breast epithelial cells. In addition, Pin1 knockout in mice prevents massive proliferation of breast epithelial cells during pregnancy. Pin1 plays a pivotal role in breast development and may be a promising new anticancer target

Prolyl Isomerase Pin1 is Highly Expressed in Her2-Positive Breast Cancer and Regulates erbB2 Protein Stability

Overexpression of HER-2/Neu occurs in about 25–30% of breast cancer patients and is indicative of poor prognosis. While Her2/Neu overexpression is primarily a result of erbB2 amplification, it has recently been recognized that erbB2 levels are also regulated on the protein level. However, factors that regulate Her2/Neu protein stability are less well understood. The prolyl isomerase Pin1 catalyzes the isomerization of specific pSer/Thr-Pro motifs that have been phosphorylated in response to mitogenic signaling. We have previously reported that Pin1-catalyzed postphosphorylational modification of signal transduction modulates the oncogenic pathways downstream from c-neu. The goal of this study was to examine the expression of prolyl isomerase Pin1 in human Her2+ breast cancer, and to study if Pin1 affects the expression of Her2/Neu itself. Methods: Immunohistochemistry for Her2 and Pin1 were performed on two hundred twentythree human breast cancers, with 59% of the specimen from primary cancers and 41% from metastatic sites. Pin1 inhibition was achieved using siRNA in Her2+ breast cancer cell lines, and its effects were studied using cell viability assays, immunoblotting and immunofluorescence. Results: Sixty-four samples (28.7%) stained positive for Her2 (IHC 3+), and 54% (122/223) of all breast cancers stained positive for Pin1. Of the Her2-positive cancers 40 (62.5%) were also Pin1-positive, based on strong nuclear or nuclear and cytoplasmic staining. Inhibition of Pin1 via RNAi resulted in significant suppression of Her2-positive tumor cell growth in BT474, SKBR3 and AU565 cells. Pin1 inhibition greatly increased the sensitivity of Her2-positive breast cancer cells to the mTOR inhibitor Rapamycin, while it did not increase their sensitivity to Trastuzumab, suggesting that Pin1 might act on Her2 signaling. We found that Pin1 interacted with the protein complex that contains ubiquitinated erbB2 and that Pin1 inhibition accelerated erbB2 degradation, which could be prevented by treatments with the proteasome inhibitor ALLnL. Conclusion: Pin1 is a novel regulator of erbB2 that modulates the ubiquitin-mediated degradation of erbB2. The overexpression of Pin1 in a majority of Her2-overexpressing breast cancer may contribute to maintain erbB2 levels. Pin1 inhibition alone and in conjunction with mTOR inhibition suppresses the growth of Her2+ breast cancer cells

Supplementary Figures S1 to S11 from Allosteric PI3Kα Inhibition Overcomes On-target Resistance to Orthosteric Inhibitors Mediated by Secondary <i>PIK3CA</i> Mutations

Supplementary Figure S1: PI3K pathway activity in selected cases with acquired PTEN alteration. Supplementary Figure S2. Validation of AKT constructs expression in T47D cells. Supplementary Figure S3. AKT activating mutations confer resistance to PI3Ka inhibitors. Supplementary Figure S4: Free energy calculations predict resistance to orthosteric PI3K inhibitors due to specific double PIK3CA mutants. Supplementary Figure S5: Free energy perturbation predicts reduced binding of orthosteric PI3K inhibitors to double mutants. Supplementary Figure S6. Expression of PIK3CA mutations in T47D cells. Supplementary Figure S7. MCF7 cells expressing W780R or Q859H double mutants show differential response to PIK3CA orthosteric inhibitors. Supplementary Figure S8: Chemical structure of RLY-2608. Supplementary Figure S9: Surface plasmon resonance (SPR) binding assay. Supplementary Figure S10. Alpelisib shows reduced potency of downstream signaling inhibition in the presence of W780R or Q859H/K. Supplementary Figure S11. T47D cells expressing I817F or E726K double mutants do not show a differential response to inavolisib (A) or RLY-2608 (B).</p

Supplementary Tables S1 to S5 from Allosteric PI3Kα Inhibition Overcomes On-target Resistance to Orthosteric Inhibitors Mediated by Secondary <i>PIK3CA</i> Mutations

Table S1: Eligibility Criteria. Table S2: Genomic alterations within the PIK3CA pathway and other documented alterations for this study. Table S3: VAF of EOT alterations. Table S4: Comparison of Acquired PIK3CA mutations based on baseline activating mutation. Table S5: Comparison of Acquired PIK3CA mutations based on number of baseline activating PIK3CA mutations.</p

Supplementary Figures S1 to S11 from Allosteric PI3Kα Inhibition Overcomes On-target Resistance to Orthosteric Inhibitors Mediated by Secondary <i>PIK3CA</i> Mutations

Supplementary Figure S1: PI3K pathway activity in selected cases with acquired PTEN alteration. Supplementary Figure S2. Validation of AKT constructs expression in T47D cells. Supplementary Figure S3. AKT activating mutations confer resistance to PI3Ka inhibitors. Supplementary Figure S4: Free energy calculations predict resistance to orthosteric PI3K inhibitors due to specific double PIK3CA mutants. Supplementary Figure S5: Free energy perturbation predicts reduced binding of orthosteric PI3K inhibitors to double mutants. Supplementary Figure S6. Expression of PIK3CA mutations in T47D cells. Supplementary Figure S7. MCF7 cells expressing W780R or Q859H double mutants show differential response to PIK3CA orthosteric inhibitors. Supplementary Figure S8: Chemical structure of RLY-2608. Supplementary Figure S9: Surface plasmon resonance (SPR) binding assay. Supplementary Figure S10. Alpelisib shows reduced potency of downstream signaling inhibition in the presence of W780R or Q859H/K. Supplementary Figure S11. T47D cells expressing I817F or E726K double mutants do not show a differential response to inavolisib (A) or RLY-2608 (B).</p

Supplementary Tables S1 to S5 from Allosteric PI3Kα Inhibition Overcomes On-target Resistance to Orthosteric Inhibitors Mediated by Secondary <i>PIK3CA</i> Mutations

Table S1: Eligibility Criteria. Table S2: Genomic alterations within the PIK3CA pathway and other documented alterations for this study. Table S3: VAF of EOT alterations. Table S4: Comparison of Acquired PIK3CA mutations based on baseline activating mutation. Table S5: Comparison of Acquired PIK3CA mutations based on number of baseline activating PIK3CA mutations.</p