Kevin Shee

P-REX1 creates a positive feedback loop to activate growth factor receptor, PI3K/AKT and MEK/ERK signaling in breast cancer

Phosphatidylinositol 3-kinase (PI3K) promotes cancer cell survival, migration, growth and proliferation by generating phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the inner leaflet of the plasma membrane. PIP3 recruits pleckstrin homology domain-containing proteins to the membrane to activate oncogenic signaling cascades. Anticancer therapeutics targeting the PI3K/AKT/mTOR (mammalian target of rapamycin) pathway are in clinical development. In a mass spectrometric screen to identify PIP3-regulated proteins in breast cancer cells, levels of the Rac activator PIP3-dependent Rac exchange factor-1 (P-REX1) increased in response to PI3K inhibition, and decreased upon loss of the PI3K antagonist phosphatase and tensin homolog (PTEN). P-REX1 mRNA and protein levels were positively correlated with ER expression, and inversely correlated with PI3K pathway activation in breast tumors as assessed by gene expression and phosphoproteomic analyses. P-REX1 increased activation of Rac1, PI3K/AKT and MEK/ERK signaling in a PTEN-independent manner, and promoted cell and tumor viability. Loss of P-REX1 or inhibition of Rac suppressed PI3K/AKT and MEK/ERK, and decreased viability. P-REX1 also promoted insulin-like growth factor-1 receptor activation, suggesting that P-REX1 provides positive feedback to activators upstream of PI3K. In support of a model where PIP3-driven P-REX1 promotes both PI3K/AKT and MEK/ERK signaling, high levels of P-REX1 mRNA (but not phospho-AKT or a transcriptomic signature of PI3K activation) were predictive of sensitivity to PI3K inhibitors among breast cancer cell lines. P-REX1 expression was highest in estrogen receptor-positive breast tumors compared with many other cancer subtypes, suggesting that neutralizing the P-REX1/Rac axis may provide a novel therapeutic approach to selectively abrogate oncogenic signaling in breast cancer cells.

Strategically Timing Inhibition of Phosphatidylinositol 3-Kinase to Maximize Therapeutic Index in Estrogen Receptor Alpha–Positive, PIK3CA-Mutant Breast Cancer

Purpose: Phosphatidylinositol 3-kinase (PI3K) inhibitors are being developed for the treatment of estrogen receptor α (ER)–positive breast cancer in combination with antiestrogens. Understanding the temporal response and pharmacodynamic effects of PI3K inhibition in ER+ breast cancer will provide a rationale for treatment scheduling to maximize therapeutic index. Experimental Design: Antiestrogen-sensitive and antiestrogen-resistant ER+ human breast cancer cell lines and mice bearing PIK3CA-mutant xenografts were treated with the antiestrogen fulvestrant, the PI3K inhibitor GDC-0941 (pictilisib; varied doses/schedules that provided similar amounts of drug each week), or combinations. Cell viability, signaling pathway inhibition, proliferation, apoptosis, tumor volume, and GDC-0941 concentrations in plasma and tumors were temporally measured. Results: Treatment with the combination of fulvestrant and GDC-0941, regardless of dose/schedule, was significantly more effective than that with single-agent treatments in fulvestrant-resistant tumors. Short-term, complete PI3K inhibition blocked cell growth in vitro more effectively than chronic, incomplete inhibition. Longer-term PI3K inhibition hypersensitized cells to growth factor signaling upon drug withdrawal. Different schedules of GDC-0941 elicited similar tumor responses. While weekly high-dose GDC-0941 with fulvestrant continuously suppressed PI3K signaling for 72 hours, inducing a bolus of apoptosis and inhibiting proliferation, PI3K reactivation upon GDC-0941 washout induced a proliferative burst. Fulvestrant with daily low-dose GDC-0941 metronomically suppressed PI3K for 6 to 9 hours/day, repeatedly inducing small amounts of apoptosis and temporarily inhibiting proliferation, followed by proliferative rebound compared with fulvestrant alone. Conclusions: Continuous and metronomic PI3K inhibition elicits robust anticancer effects in ER+, PIK3CA-mutant breast cancer. Clinical exploration of alternate treatment schedules of PI3K inhibitors with antiestrogens is warranted. Clin Cancer Res; 22(9); 2250–60. ©2016 AACR. See related commentary by Toska and Baselga, p. 2099

Cell Competition During Growth and Regeneration.

Tissue growth and regeneration are autonomous, stem-cell-mediated processes in which stem cells within the organ self-renew and differentiate to create new cells, leading to new tissue. The processes of growth and regeneration require communication and interplay between neighboring cells. In particular, cell competition, which is a process in which viable cells are actively eliminated by more competitive cells, has been increasingly implicated to play an important role. Here, we discuss the existing literature regarding the current landscape of cell competition, including classical pathways and models, fitness fingerprint mechanisms, and immune system mechanisms of cell competition. We further discuss the clinical relevance of cell competition in the physiological processes of tissue growth and regeneration, highlighting studies in clinically important disease models, including oncological, neurological, and cardiovascular diseases.

Autophagy promotes escape from phosphatidylinositol 3-kinase inhibition in estrogen receptor–positive breast cancer

Hyperactivation of the PI3K pathway has been implicated in resistance to antiestrogen therapies in estrogen receptor α (ER)‐positive breast cancer, prompting the development of therapeutic strategies to inhibit this pathway. Autophagy has tumor‐promoting and ‐suppressing roles and has been broadly implicated in resistance to anticancer therapies, including antiestrogens. Chloroquine (CQ) is an antimalarial and amebicidal drug that inhibits autophagy in mammalian cells and human tumors. Herein, we observed that CQ inhibited proliferation and autophagy in ER+ breast cancer cells. PI3K inhibition with GDC‐0941 (pictilisib) induced autophagy. Inhibition of autophagy using CQ or RNA interference potentiated PI3K inhibitor‐induced apoptosis. Combined inhibition of PI3K and autophagy effectively induced mitochondrial membrane depolarization, which required the BH3‐only proapoptotic proteins Bim and PUMA. Treatment with GDC‐0941, CQ, or the combination, significantly suppressed the growth of ER+ breast cancer xenografts in mice. In an antiestrogen‐resistant xenograft model, GDC‐0941 syn‐ ergized with CQ to provide partial, but durable, tumor regression. These findings warrant clinical evaluation of therapeutic strategies to target ER, PI3K, and autophagy for the treatment of ER+ breast cancer.—Yang, W., Hosford, S. R., Traphagen, N. A., Shee, K., Demidenko, E., Liu, S., Miller, T. W. Autophagy promotes escape from phos‐ phatidylinositol 3‐kinase inhibition in estrogen receptor‐positive breast cancer. FASEB J. 32, 1222‐1235 (2018). www.fasebj.org

Practice Makes Perfect: Correlations Between Prior Experience in High-level Athletics and Robotic Surgical Performance Do Not Persist After Task Repetition

OBJECTIVE Robotic surgical skill development is central to training in urology as well as in other surgical disciplines. Here, we describe a pilot study assessing the relationships between robotic surgery simulator performance and 3 categories of activities, namely, videogames, musical instruments, and athletics. DESIGN A questionnaire was administered to preclinical medical students for general demographic information and prior experiences in surgery, videogames, musical instruments, and athletics. For follow-up performance studies, we used the Matchboard Level 1 and 2 modules on the da Vinci Skills Simulator, and recorded overall score, time to complete, economy of motion, workspace range, instrument collisions, instruments out of view, and drops. Task 1 was run once, whereas task 2 was run 3 times. SETTING All performance studies on the da Vinci Surgical Skills Simulator took place in the Simulation Center at Dartmouth-Hitchcock Medical Center. PARTICIPANTS All participants were medical students at the Geisel School of Medicine. After excluding students with prior hands-on experience in surgery, a total of 30 students completed the study. RESULTS We found a significant correlation between athletic skill level and performance for both task 1 (p = 0.0002) and task 2 (p = 0.0009). No significant correlations were found for videogame or musical instrument skill level. Students with experience in certain athletics (e.g., volleyball, tennis, and baseball) tended to perform better than students with experience in other athletics (e.g., track and field). For task 2, which was run 3 times, this association did not persist after the third repetition due to significant improvements in students with low-level athletic skill (levels 0-2). CONCLUSIONS Our study suggests that prior experience in high-level athletics, but not videogames or musical instruments, significantly influences surgical proficiency in robot-naive students. Furthermore, our study suggests that practice through task repetition can overcome initial differences that may be related to a background in athletics. These novel relationships may have broader implications for the future recruitment and training of robotic surgeons and may warrant further investigation.

Therapeutically targeting tumor microenvironment–mediated drug resistance in estrogen receptor–positive breast cancer

Drug resistance to approved systemic therapies in estrogen receptor–positive (ER+) breast cancer remains common. We hypothesized that factors present in the human tumor microenvironment (TME) drive drug resistance. Screening of a library of recombinant secreted microenvironmental proteins revealed fibroblast growth factor 2 (FGF2) as a potent mediator of resistance to anti-estrogens, mTORC1 inhibition, and phosphatidylinositol 3-kinase inhibition in ER+ breast cancer. Phosphoproteomic analyses identified ERK1/2 as a major output of FGF2 signaling via FGF receptors (FGFRs), with consequent up-regulation of Cyclin D1 and down-regulation of Bim as mediators of drug resistance. FGF2-driven drug resistance in anti-estrogen–sensitive and –resistant models, including patient-derived xenografts, was reverted by neutralizing FGF2 or FGFRs. A transcriptomic signature of FGF2 signaling in primary tumors predicted shorter recurrence-free survival independently of age, grade, stage, and FGFR amplification status. These findings delineate FGF2 signaling as a ligand-based drug resistance mechanism and highlights an underdeveloped aspect of precision oncology: characterizing and treating patients according to their TME constitution.

The curcumin analog HO-3867 selectively kills cancer cells by converting mutant p53 protein to transcriptionally active wildtype p53

p53 is an important tumor-suppressor protein that is mutated in more than 50% of cancers. Strategies for restoring normal p53 function are complicated by the oncogenic properties of mutant p53 and have not met with clinical success. To counteract mutant p53 activity, a variety of drugs with the potential to reconvert mutant p53 to an active wildtype form have been developed. However, these drugs are associated with various negative effects such as cellular toxicity, nonspecific binding to other proteins, and inability to induce a wildtype p53 response in cancer tissue. Here, we report on the effects of a curcumin analog, HO-3867, on p53 activity in cancer cells from different origins. We found that HO-3867 covalently binds to mutant p53, initiates a wildtype p53-like anticancer genetic response, is exclusively cytotoxic toward cancer cells, and exhibits high anticancer efficacy in tumor models. In conclusion, HO-3867 is a p53 mutant–reactivating drug with high clinical anticancer potential.

Therapeutic sensitivity to Rac GTPase inhibition requires consequential suppression of mTORC1, AKT, and MEK signaling in breast cancer

Rac GTPases have oncogenic roles in cell growth, survival, and migration. We tested response to the Rac inhibitor EHT1864 in a panel of breast cancer cell lines. EHT1864-induced growth inhibition was associated with dual inhibition of the PI3K/AKT/mTORC1 and MEK/ERK pathways. Breast cancer cells harboring PIK3CA mutations or HER2 overexpression were most sensitive to Rac inhibition, suggesting that such oncogenic alterations link Rac activation with PI3K/AKT/mTORC1 and MEK/ERK signaling. Interestingly, EHT1864 decreased activation of the mTORC1 substrate p70S6K earlier than AKT inhibition, suggesting that Rac may activate mTORC1/p70S6K independently of AKT. Comparison of the growth-inhibitory profile of EHT1864 to 137 other anti-cancer drugs across 656 cancer cell lines revealed significant correlation with the p70S6K inhibitor PF-4708671. We confirmed that Rac complexes contain MEK1/2 and ERK1/2, but also contain p70S6K; these interactions were disrupted by EHT1864. Pharmacokinetic profiles revealed that EHT1864 was present in mouse plasma at concentrations effective in vitro for approximately 1 h after intraperitoneal administration. EHT1864 suppressed growth of HER2+ tumors, and enhanced response to anti-estrogen treatment in ER+ tumors. Further therapeutic development of Rac inhibitors for HER2+ and PIK3CA-mutant cancers is warranted.

Pain Management in Penile Prosthetic Surgery: A Review of the Literature

INTRODUCTION The literature on perioperative pain control and management in inflatable penile prosthesis placement is not firmly established. Because inflatable penile prosthesis placement is an elective procedure, pain can be one of the many issues that influence patient decision making. Pain control also presents a unique challenge to providers in an era of widespread opiate abuse. AIM To review published data on pain management before, during, and after penile prosthetic surgery. METHODS Peer-reviewed literature and conference abstracts were analyzed for all relevant publications related to this issue. RESULTS The past several decades have seen a shift from general to local anesthesia for penile prosthetic surgery. This has been well characterized and is seen as successful with different local anesthetic options and techniques. To date, only one study has provided follow-up for longer than 1 week regarding postoperative pain management for prosthetic surgery. CONCLUSION Perioperative pain management for the patient receiving a penile prosthetic is well characterized; postoperative pain management is not. Although periprocedural local anesthesia has been well described for penile prosthesis surgery, a standardized postoperative pain management plan does not exist. This review highlights the need for further characterization of postoperative pain and the subsequent development of an algorithmic approach for management. Reinstatler L, Shee K, Gross MS. Pain Management in Penile Prosthetic Surgery: A Review of the Literature. Sex Med Rev 2018;6:162-169.

Readability analysis of online health information about overactive bladder

Despite the prevalence of overactive bladder (OAB) and the widespread accessibility of patient education information on the Internet, the readability of this information and its potential impact on patient decision‐making are not known. This study evaluates the readability of OAB material online in the context of website ownership and the Health on the Net standard for information reliability.