Cell cycle-coupled expansion of AR activity promotes cancer progression.

The androgen receptor (AR) is required for prostate cancer (PCa) survival and progression, and ablation of AR activity is the first line of therapeutic intervention for disseminated disease. While initially effective, recurrent tumors ultimately arise for which there is no durable cure. Despite the dependence of PCa on AR activity throughout the course of disease, delineation of the AR-dependent transcriptional network that governs disease progression remains elusive, and the function of AR in mitotically active cells is not well understood. Analyzing AR activity as a function of cell cycle revealed an unexpected and highly expanded repertoire of AR-regulated gene networks in actively cycling cells. New AR functions segregated into two major clusters: those that are specific to cycling cells and retained throughout the mitotic cell cycle ('Cell Cycle Common'), versus those that were specifically enriched in a subset of cell cycle phases ('Phase Restricted'). Further analyses identified previously unrecognized AR functions in major pathways associated with clinical PCa progression. Illustrating the impact of these unmasked AR-driven pathways, dihydroceramide desaturase 1 was identified as an AR-regulated gene in mitotically active cells that promoted pro-metastatic phenotypes, and in advanced PCa proved to be highly associated with development of metastases, recurrence after therapeutic intervention and reduced overall survival. Taken together, these findings delineate AR function in mitotically active tumor cells, thus providing critical insight into the molecular basis by which AR promotes development of lethal PCa and nominate new avenues for therapeutic intervention.

Targeting cell cycle and hormone receptor pathways in cancer.

The cyclin/cyclin-dependent kinase (CDK)/retinoblastoma (RB)-axis is a critical modulator of cell cycle entry and is aberrant in many human cancers. New nodes of therapeutic intervention are needed that can delay or combat the onset of malignancies. The antitumor properties and mechanistic functions of PD-0332991 (PD; a potent and selective CDK4/6 inhibitor) were investigated using human prostate cancer (PCa) models and primary tumors. PD significantly impaired the capacity of PCa cells to proliferate by promoting a robust G1-arrest. Accordingly, key regulators of the G1-S cell cycle transition were modulated including G1 cyclins D, E and A. Subsequent investigation demonstrated the ability of PD to function in the presence of existing hormone-based regimens and to cooperate with ionizing radiation to further suppress cellular growth. Importantly, it was determined that PD is a critical mediator of PD action. The anti-proliferative impact of CDK4/6 inhibition was revealed through reduced proliferation and delayed growth using PCa cell xenografts. Finally, first-in-field effects of PD on proliferation were observed in primary human prostatectomy tumor tissue explants. This study shows that selective CDK4/6 inhibition, using PD either as a single-agent or in combination, hinders key proliferative pathways necessary for disease progression and that RB status is a critical prognostic determinant for therapeutic efficacy. Combined, these pre-clinical findings identify selective targeting of CDK4/6 as a bona fide therapeutic target in both early stage and advanced PCa and underscore the benefit of personalized medicine to enhance treatment response.

Cyclin D1b protein expression in breast cancer is independent of cyclin D1a and associated with poor disease outcome.

Aberrant expression of cyclin D1 protein is a common feature of breast cancer. However, the CCND1 gene encodes two gene products, cyclin D1a and cyclin D1b, which have discrete mechanisms of regulation and impact on cell behavior. A polymorphism at nucleotide 870 in the CCND1 gene, rs603965, influences the relative production of the encoded proteins and can impart increased risk for tumor development. Here, the impact of both the G/A870 polymorphism and cyclin D1b protein production on breast cancer risk, disease phenotype and patient outcome was analysed. In a large multiethnic case-control study, the G/A870 polymorphism conferred no significant risk for breast cancer overall or by stage or estrogen receptor (ER) status. However, the cyclin D1b protein was found to be upregulated in breast cancer, independent of cyclin D1a levels, and exhibited heterogeneous levels in breast cancer specimens. High cyclin D1a expression inversely correlated with the Ki67 proliferation marker and was not associated with clinical outcome. In contrast, elevated cyclin D1b expression was independently associated with adverse outcomes, including recurrence, distant metastasis and decreased survival. Interestingly, cyclin D1b was particularly associated with poor outcome in the context of ER-negative breast cancer. Thus, specific cyclin D1 isoforms are associated with discrete forms of breast cancer and high cyclin D1b protein levels hold prognostic potential.

Cyclin D3 action in androgen receptor regulation and prostate cancer.

Prostate cancer (PCa) cell proliferation is dependent on activation of the androgen receptor (AR), a ligand-dependent transcription factor. AR activation controls G1-S phase progression through fostering enhanced translation of the D-type cyclins, which promote cell cycle progression through activation of CDK4/6. However, the D-type cyclins harbor additional, CDK4/6 kinase-independent, functions through manipulation of transcription factors, including AR. It was previously established that cyclins D1 and D3 have the potential to modulate AR, and with regard to cyclin D1, disruption of this function occurs in human tumors. Therefore, it was essential to interrogate cyclin D3 function in this tumor type. Here, we show that cyclin D3 is found in association with AR in PCa cells, as mediated through a conserved motif. Cyclin D3 functions to attenuate AR activity through defined mechanisms that include modulation of ligand-dependent conformational changes and modulation of chromatin binding activity. Accumulated cyclin D3 slows cell proliferation in AR-dependent cells, thus suggesting that androgen-induced D-type cyclin production serves to temper the mitogenic response to androgen. Supporting this hypothesis, it is shown that cyclin D3 expression is reduced in primary PCas as a function of tumor grade, and inversely correlates with the proliferative index. In total, these data identify cyclin D3 as a critical modulator of the androgen response, whose deregulation may foster unchecked AR activity in PCa.

Impact of differential cyclin D1 expression and localisation in prostate cancer.

Cyclin D1 is a critical regulator of androgen-dependent transcription and cell cycle progression in prostate cancer cells. Despite the influence of D-type cyclins on prostate cancer proliferation, few studies have examined the expression of cyclin D1 in localised tumours or challenged its relevance to disease progression. Cyclin D1 status was characterised using immunohistochemistry in 38 non-neoplastic prostate samples, 138 primary human prostate carcinomas, and three lymph node metastatic specimens. Relevance of cyclin D1 to preoperative prostate-specific antigen (PSA) levels, Ki-67 index, and p21Cip1 status was also examined. Cyclin D1-positive phenotype was increased in primary carcinoma compared to non-neoplastic tissue, and was evident in all lymph node metastases cases. Interestingly, at least three distinct localisation patterns were observed in the cyclin D1-positive cohort, wherein cytoplasmic localisation was identified in a large fraction, and this pattern was predominant in lower grade tumours. Relevance of altered cyclin D1 status was observed, wherein cyclin D1-positive tumours were associated with low preoperative PSA levels, consistent with in vitro reports that cyclin D1 may alter the expression of this tumour marker. Moreover, tumours with predominantly cytoplasmic cyclin D1 showed the lowest Ki-67 index, whereas nuclear cyclin D1 was associated with higher grade, elevated Ki-67, and increased nuclear p21Cip1. These data demonstrate that differential cyclin D1 status may influence clinicopathological parameters, and reveal new insight as to the regulation and potential consequence of cyclin D1 expression in prostate cancer.

Rates and causes of disagreement in interpretation of full-field digital mammography and film-screen mammography in a diagnostic setting.

OBJECTIVE: This study was performed to determine the rates and causes of disagreements in interpretation between full-field digital mammography and film-screen mammography in a diagnostic setting. SUBJECTS AND METHODS: Patients undergoing diagnostic mammography were invited to participate in the digital mammography study. Three views, selected by the radiologist interpreting the film-screen mammography, were obtained in both film-screen mammography and digital mammography. Radiologists independently assigned a Breast Imaging Reporting and Data System (BI-RADS) category to the film-screen mammography and the digital mammography images. The BI-RADS categories were grouped into the general categories of agreement, partial agreement, or disagreement. A third and different radiologist reviewed all cases of disagreement, reached a decision as to management, and determined the primary cause of disagreement. RESULTS: Six radiologists reviewed digital mammography and film-screen mammography diagnostic images in a total of 1147 breasts in 692 patients. Agreement between digital mammography and final film-screen mammography assessment was present in 937 breasts (82%), partial agreement in 159 (14%), and disagreement in 51 (4%), for a kappa value of 0.29. The primary causes of disagreement were differences in management approach of the radiologists (52%), information derived from sonography or additional film-screen mammograms (34%), and technical differences between the two mammographic techniques (10%). CONCLUSION: Significant disagreement between film-screen mammography and digital mammography affecting follow-up management was present in only 4% of breasts. The most frequent cause of disagreement in interpretation was a difference in management approach between radiologists (interobserver variability). This source of variability was larger than that due to differences in lesion visibility between film-screen mammography and digital mammography.

Inter-observer variability in ultrasonic evaluation of abdominal aortic aneurysms.

This study was undertaken to assess the consistency in 16 independent observations of the diameters of abdominal aortic aneurysms by diagnostic ultrasound. Confidence intervals were developed for the anterior-posterior dimension and the transverse dimension. Clinical treatment decisions should be made with appreciation that measurements of diameter increases greater than 0.21 cm in the anterior-posterior dimension and 0.31 cm in the transverse dimension are significant and are not a result of inter-observer variation.

Development of turbulence in steady flow through models of abdominal aortic aneurysms.

As part of a general examination of the effects of blood flow patterns on the risk of rupture of abdominal aortic aneurysms, we have investigated steady flow through a series of polyvinyl chloride models of fusiform aneurysms. A series of steady flow rates between 460 ml/min and 2.0 l/min have been delivered through the models, and the resulting flow patterns have been evaluated by color Doppler flow imaging. At low volume flow rates, the pattern of flow through the model is smooth and laminar. At higher flow rates, however, this is replaced by an irregular, fluctuating, and apparently turbulent motion. This finding is consistent with previous clinical observations. Turbulence appears initially at the distal end of the model aneurysm and then spreads proximally to fill the whole model only if the flow rate continues to increase. The onset of irregular, turbulent flow is found to be governed by both the overall flow rate through the model and the model diameter. Critical conditions for the appearance of turbulence are presented and discussed, and the critical value of the Reynolds number for both the initial distal transition and the final full transition is shown to decrease as the size of the model aneurysm increases.

A qualitative investigation of turbulence in flow through a model abdominal aortic aneurysm.

RATIONALE AND OBJECTIVES: As part of a general examination of the effect of blood flow patterns on the growth and risk of rupture of abdominal aortic aneurysms, the authors investigated the flow through an experimental model of a fusiform aneurysm. MATERIALS AND METHODS: Aneurysms were simulated in polyvinyl chloride tubing. A steady flow of 450 mL/minute to 2 L/minute was investigated by color Doppler flow imaging. RESULTS: Two types of flow patterns existed in the model aneurysm, depending on the overall flow rate through the model. At low velocity flow, this pattern is smooth, steady, and laminar, whereas at higher flows it is irregular and turbulent. CONCLUSION: This finding corresponds to recent clinical observations. The onset of unsteadiness and irregularity in the flow through the model is related to flow velocity.