GATA2 facilitates steroid receptor coactivator recruitment to the androgen receptor complex.

The androgen receptor (AR) is a key driver of prostate cancer (PC), even in the state of castration-resistant PC (CRPC) and frequently even after treatment with second-line hormonal therapies such as abiraterone and enzalutamide. The persistence of AR activity via both ligand-dependent and ligand-independent mechanisms (including constitutively active AR splice variants) highlights the unmet need for alternative approaches to block AR signaling in CRPC. We investigated the transcription factor GATA-binding protein 2 (GATA2) as a regulator of AR signaling and an actionable therapeutic target in PC. We demonstrate that GATA2 directly promotes expression of both full-length and splice-variant AR, resulting in a strong positive correlation between GATA2 and AR expression in both PC cell lines and patient specimens. Conversely, GATA2 expression is repressed by androgen and AR, suggesting a negative feedback regulatory loop that, upon androgen deprivation, derepresses GATA2 to contribute to AR overexpression in CRPC. Simultaneously, GATA2 is necessary for optimal transcriptional activity of both full-length and splice-variant AR. GATA2 colocalizes with AR and Forkhead box protein A1 on chromatin to enhance recruitment of steroid receptor coactivators and formation of the transcriptional holocomplex. In agreement with these important functions, high GATA2 expression and transcriptional activity predicted worse clinical outcome in PC patients. A GATA2 small molecule inhibitor suppressed the expression and transcriptional function of both full-length and splice-variant AR and exerted potent anticancer activity against PC cell lines. We propose pharmacological inhibition of GATA2 as a first-in-field approach to target AR expression and function and improve outcomes in CRPC.

Prostate cancer-associated mutations in speckle-type POZ protein (SPOP) regulate steroid receptor coactivator 3 protein turnover.

The p160 steroid receptor coactivators (SRCs) SRC-1, SRC-2 [nuclear receptor coactivator (NCOA)2], and SRC-3 [amplified in breast cancer 1 (AIB1)/NCOA3] are key pleiotropic "master regulators" of transcription factor activity necessary for cancer cell proliferation, survival, metabolism, and metastasis. SRC overexpression and overactivation occur in numerous human cancers and are associated with poor clinical outcomes and resistance to therapy. In prostate cancer (PC), the p160 SRCs play critical roles in androgen receptor transcriptional activity, cell proliferation, and resistance to androgen deprivation therapy. We recently demonstrated that the E3 ubiquitin ligase adaptor speckle-type poxvirus and zinc finger (POZ) domain protein (SPOP) interacts directly with SRC-3 and promotes its cullin 3-dependent ubiquitination and proteolysis in breast cancer, thus functioning as a potential tumor suppressor. Interestingly, somatic heterozygous missense mutations in the SPOP substrate-binding cleft recently were identified in up to 15% of human PCs (making SPOP the gene most commonly affected by nonsynonymous point mutations in PC), but their contribution to PC pathophysiology remains unknown. We now report that PC-associated SPOP mutants cannot interact with SRC-3 protein or promote its ubiquitination and degradation. Our data suggest that wild-type SPOP plays a critical tumor suppressor role in PC cells, promoting the turnover of SRC-3 protein and suppressing androgen receptor transcriptional activity. This tumor suppressor effect is abrogated by the PC-associated SPOP mutations. These studies provide a possible explanation for the role of SPOP mutations in PC, and highlight the potential of SRC-3 as a therapeutic target in PC.

Safety and Efficacy of Wharton's Jelly Connective Tissue Allograft for Rotator Cuff Tears: Findings from a Retrospective Observational Study.

With the increasing occurrence of rotator cuff injuries every year, there is a great need for a reliable treatment option. Wharton's Jelly contains several components that can positively impact the replacement and repair of musculoskeletal defects. The overall objective of this study is to evaluate the improvement of patient-reported pain scales after applying Wharton's Jelly (WJ) in rotator cuff defects. Eighty-seven patients with rotator cuff defects who failed at least eight weeks of conservative treatment were selected from the retrospective repository. A total of 2 cc of WJ flowable allograft was applied to the specific affected anatomy, the most common being supraspinatus tendon, biceps tendon insertion, labral tear, and subscapularis tear. No adverse reactions were reported. Statistically significant improvements were found from the initial to Day 90 in all scales. Patient satisfaction was calculated using minimal clinically important differences. No statistically significant differences were found in mean changes between gender, BMI, and age. Scanning electron microscopy images reveal the similarities between the collagen matrix in WJ and the rotator cuff. The significant improvement in patient outcomes coincides with the current literature analyzing WJ applications with other structural defects around the body. WJ is a promising alternative for musculoskeletal defects when the standard of care fails.

Retrospective Evaluation of Cryopreserved Human Umbilical Cord Tissue Allografts in the Supplementation of Cartilage Defects Associated with Hip Osteoarthritis.

Background: Osteoarthritis is a chronic disorder that affects the synovial joints by the progressive loss of articular cartilage. In the hip, the largest weight-bearing joint, the deterioration of articular cartilage and acetabular labrum can cause pain, diminishing the quality of life for patients. This study presents changes in reported pain scales from patients who received Wharton's jelly applications to cartilage deterioration in the hip from the observational retrospective repository at Regenative Labs. Methods: Sixty-nine patients were selected based on inclusion criteria with patient-reported pain scales, including the Numeric Pain Rating Scale and the Western Ontario and McMaster University Osteoarthritis Index, collected at the initial application, 30, and 90-day follow-up visits. Thirteen patients received a second allograft application and had additional follow-up visits at 120 and 180 days. Results: Five of the six scales used showed a statistically significant improvement in average scores across the cohort. The greatest improvements were observed in the NPRS with a 31.36% improvement after 90 days and a 44.64% improvement for patients with two applications after 180 days. The minimal clinically important difference (MCID) was also calculated to determine the perceived value of care for each patient with 44.9% of patients exceeding the MCID and 78.3% reporting at least one level of improvement. Conclusions: The positive outcomes for the patients in this cohort suggest WJ to be a promising alternative care option for patients with structural tissue degeneration in the hip refractory to the current standard of care.

miR-137 Targets p160 Steroid Receptor Coactivators SRC1, SRC2, and SRC3 and Inhibits Cell Proliferation.

The p160 family of steroid receptor coactivators (SRCs) are pleiotropic transcription factor coactivators and "master regulators" of gene expression that promote cancer cell proliferation, survival, metabolism, migration, invasion, and metastasis. Cancers with high p160 SRC expression exhibit poor clinical outcomes and resistance to therapy, highlighting the SRCs as critical oncogenic drivers and, thus, therapeutic targets. microRNAs are important epigenetic regulators of protein expression. To examine the regulation of p160 SRCs by microRNAs, we used and combined 4 prediction algorithms to identify microRNAs that could target SRC1, SRC2, and SRC3 expression. For validation of these predictions, we assessed p160 SRC protein expression and cell viability after transfection of corresponding microRNA mimetics in breast cancer, uveal melanoma, and prostate cancer (PC) cell lines. Transfection of selected microRNA mimetics into breast cancer, uveal melanoma, and PC cells depleted SRC protein expression levels and exerted potent antiproliferative activity in these cell types. In particular, microRNA-137 (miR-137) depleted expression of SRC1, SRC2, and very potently, SRC3. The latter effect can be attributed to the presence of 3 miR-137 recognition sequences within the SRC3 3'-untranslated region. Using reverse phase protein array analysis, we identified a network of proteins, in addition to SRC3, that were modulated by miR-137 in PC cells. We also found that miR-137 and its host gene are epigenetically silenced in human cancer specimens and cell lines. These results support the development and testing of microRNA-based therapies (in particular based on restoring miR-137 levels) for targeting the oncogenic family of p160 SRCs in cancer.

Androgen receptor is the key transcriptional mediator of the tumor suppressor SPOP in prostate cancer.

Somatic missense mutations in the substrate-binding pocket of the E3 ubiquitin ligase adaptor SPOP are present in up to 15% of human prostate adenocarcinomas, but are rare in other malignancies, suggesting a prostate-specific mechanism of action. SPOP promotes ubiquitination and degradation of several protein substrates, including the androgen receptor (AR) coactivator SRC-3. However, the relative contributions that SPOP substrates may make to the pathophysiology of SPOP-mutant (mt) prostate adenocarcinomas are unknown. Using an unbiased bioinformatics approach, we determined that the gene expression profile of prostate adenocarcinoma cells engineered to express mt-SPOP overlaps greatly with the gene signature of both SRC-3 and AR transcriptional output, with a stronger similarity to AR than SRC-3. This finding suggests that in addition to its SRC-3-mediated effects, SPOP also exerts SRC-3-independent effects that are AR-mediated. Indeed, we found that wild-type (wt) but not prostate adenocarcinoma-associated mutants of SPOP promoted AR ubiquitination and degradation, acting directly through a SPOP-binding motif in the hinge region of AR. In support of these results, tumor xenografts composed of prostate adenocarcinoma cells expressing mt-SPOP exhibited higher AR protein levels and grew faster than tumors composed of prostate adenocarcinoma cells expressing wt-SPOP. Furthermore, genetic ablation of SPOP was sufficient to increase AR protein levels in mouse prostate. Examination of public human prostate adenocarcinoma datasets confirmed a strong link between transcriptomic profiles of mt-SPOP and AR. Overall, our studies highlight the AR axis as the key transcriptional output of SPOP in prostate adenocarcinoma and provide an explanation for the prostate-specific tumor suppressor role of wt-SPOP.