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Forests boast essential resources and potential to mitigate climate change, meriting the development of conservation policies on all governmental scales. Ecosystem services provided by forests, including biodiversity, air quality, and food and fuel production, make forests valuable assets for climate-vulnerable communities that often lack the means to cope with ecosystem service degradation resulting from climate change. Historically, these vulnerable communities are previously marginalized and socio-economically limited, and climate change augments already-existing injustices. Policy discussions around managing forests and carbon, therefore, must consider environmental justice as well as diversity, equity, and inclusion to better meet the needs of all constituents. Using R, we perform a review of forest, climate, and policy peer-reviewed literature published between 2018 and 2021 for prevalence of topics related to diversity, equity, inclusion, and justice (DEIJ). We select DEIJ terms a priori and a posteriori based on our understanding of DEIJ and common considerations of the literature. Out of 2891 unique articles, 15.7% of literature mentioned at least one DEIJ term in the title, keyword list, or abstract. We identify which journals have published DEIJ literature more often in the context of forest, climate, and policy, and we perform a co-occurrence analysis of additional common themes. Concepts such as ecosystem services and economics appeared often in the literature, as well as REDD+ as a specifically mentioned policy. We call for increased consideration of DEIJ in forest, climate, and policy discussions and literature, as vulnerable communities historically have been excluded from and victimized by the conversations held among large, economically motivated entities., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Ecosystem services (ESs) associated with surficial processes may change according to shifts in land use, land cover, and climate parameters. Estimating these shifts can be important for land development planning, as urbanization alters soil processes that can manifest legacy effects. We employed the InVEST suite of models for sediment retention, nutrient delivery, and carbon storage to postulate how these ESs will change in the Upstate of South Carolina under future precipitation and land use and land cover (LULC) scenarios. We used the average precipitation from 1981-2010 and WorldClim precipitation projections for 2021-2040 and 2041-2060 to embody climatic precipitation shifts. For our LULC scenarios, we used 2011 and 2016 NLCD landscapes, then projected future LULC to hypothesize four future scenarios. We found that for the ES models that included both precipitation and LULC as inputs, precipitation dictated ES delivery far more heavily than land use or land cover. LULC scenarios produced consistent changes in ES delivery for all models except sediment export. Phosphorus and sediment exports increased between 2011 and 2016 due to LULC change, while nitrogen export stayed the same and carbon storage decreased. Land development that prioritizes forest cover will cause the least change in ESs, but allowing for continued forest loss to low-density development will have the most intense implications for ESs. Prioritization of land uses that preserve ESs associated with surficial processes will be critical to the longevity of agriculture and ecosystem integrity in this rapidly developing region. Land development planners should integrate consideration of ESs associated with surficial processes into future regional planning., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Data are needed to assess the relationships between urbanization and biodiversity to establish conservation priorities. However, many of these relationships are difficult to fully assess using traditional research methods. To address this gap and evaluate new acoustic sensors and associated data, we conducted a multimethod analysis of biodiversity in a rapidly urbanizing county: Greenville, South Carolina, USA. We conducted audio recordings at 25 points along a development gradient. At the same locations, we used refugia tubes, visual assessments, and an online database. Analysis focused on species identification of both audio and visual data at each point along the trail to determine relationships between both herpetofauna and acoustic indices (as proxies for biodiversity) and environmental gradient of land use and land cover. Our analysis suggests the use of a multitude of different sampling methods to be conducive to the completion of a more comprehensive occupancy measure. Moving forward, this research protocol can potentially be useful in the establishment of more effective wildlife occupancy indices using acoustic sensors to move toward future conservation policies and efforts concerning urbanization, forest fragmentation, and biodiversity in natural, particularly forested, ecosystems.

Protected areas (PA) are one of the primary tools for conserving and protecting biodiversity, but their goals have evolved overtime beyond nature conservation to include supporting human communities within and adjacent to the PA. UNESCO World Heritage Sites (WHS) offer a unique perspective on the success of PAs as they fall under three categories, cultural, natural, and mixed heritage sites. The nature of these categories encapsulates the inclusion of human communities in the goals of the WHS. To understand the impact and relationship the WHS has with its surrounding landscape, we assessed changes in three indicators, land use and land cover (LULC), human footprint (HF), and forest landscape integrity index (FLII), across three spatial scales, 1, 10, 100 km from the WHS boundary. We found that there is a conservation spillover effect at least within 1 km of the WHS boundary. In this buffer zone, HF was low and FLII was high. FLII was lower and HF was higher at larger spatial scales. The relationship between the WHS and its surrounding landscape is one reason to support the WHS network, however, management of PAs should be more explicit about this relationship as well as relationships between individual PAs., Competing Interests: The authors declare that they have no competing interests., (© 2023 Hyland and Quinn.)

Epigenetic regulation is a critical mechanism in controlling virulence, differentiation, and survival of the human malaria parasite Plasmodium (P.) falciparum . Bromodomain proteins contribute to this process by binding to acetylated lysine residues of histones and thereby targeting the gene regulatory machinery to gene promoters. A protein complex containing the P. falciparum bromodomain proteins (PfBDP) 1 and PfBDP2 (BDP1/BDP2 core complex) was previously shown to play an essential role for the correct transcription of invasion related genes. Here, we performed a functional characterization of a third component of this complex, which we dubbed PfBDP7, because structural modelling predicted a typical bromodomain fold. We confirmed that PfBDP7 is a nuclear protein that interacts with PfBDP1 at invasion gene promoters in mature schizont stage parasites and contributes to their transcription. Although partial depletion of PfBDP7 showed no significant effect on parasite viability, conditional knock down of either PfBDP7 or PfBDP1 resulted in the de-repression of variant surface antigens (VSA), which are important pathogenicity factors. This de-repression was evident both on mRNA and protein level. To understand the underlying mechanism, we mapped the genome wide binding sites of PfBDP7 by ChIPseq and showed that in early schizonts, PfBDP7 and PfBDP1 are commonly enriched in heterochromatic regions across the gene body of all VSA families, including genes coding for PfEMP1, RIFIN, STEVOR, and PfMC-2TM. This suggests that PfBDP7 and PfBDP1 contribute to the silencing of VSAs by associating with heterochromatin. In conclusion, we identified PfBDP7 as a chromatin binding protein that is a constitutive part of the P. falciparum BDP1/BDP2 core complex and established PfBDP1 and PfBDP7 as novel players in the silencing of heterochromatin regulated virulence gene families of the malaria parasite P. falciparum ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Quinn, Jeninga, Limm, Pareek, Meißgeier, Bachmann, Duffy and Petter.)

Collisions with buildings cause up to 1 billion bird fatalities annually in the United States and Canada. However, efforts to reduce collisions would benefit from studies conducted at large spatial scales across multiple study sites with standardized methods and consideration of species- and life-history-related variation and correlates of collisions. We addressed these research needs through coordinated collection of data on bird collisions with buildings at sites in the United States (35), Canada (3), and Mexico (2). We collected all carcasses and identified species. After removing records for unidentified carcasses, species lacking distribution-wide population estimates, and species with distributions overlapping fewer than 10 sites, we retained 269 carcasses of 64 species for analysis. We estimated collision vulnerability for 40 bird species with ≥2 fatalities based on their North American population abundance, distribution overlap in study sites, and sampling effort. Of 10 species we identified as most vulnerable to collisions, some have been identified previously (e.g., Black-throated Blue Warbler [Setophaga caerulescens]), whereas others emerged for the first time (e.g., White-breasted Nuthatch [Sitta carolinensis]), possibly because we used a more standardized sampling approach than past studies. Building size and glass area were positively associated with number of collisions for 5 of 8 species with enough observations to analyze independently. Vegetation around buildings influenced collisions for only 1 of those 8 species (Swainson's Thrush [Catharus ustulatus]). Life history predicted collisions; numbers of collisions were greatest for migratory, insectivorous, and woodland-inhabiting species. Our results provide new insight into the species most vulnerable to building collisions, making them potentially in greatest need of conservation attention to reduce collisions and into species- and life-history-related variation and correlates of building collisions, information that can help refine collision management., (© 2020 Society for Conservation Biology.)

Apicomplexan parasites are protozoan organisms that are characterised by complex life cycles and they include medically important species, such as the malaria parasite Plasmodium and the causative agents of toxoplasmosis ( Toxoplasma gondii ) and cryptosporidiosis ( Cryptosporidium spp.). Apicomplexan parasites can infect one or more hosts, in which they differentiate into several morphologically and metabolically distinct life cycle stages. These developmental transitions rely on changes in gene expression. In the last few years, the important roles of different members of the ApiAP2 transcription factor family in regulating life cycle transitions and other aspects of parasite biology have become apparent. Here, we review recent progress in our understanding of the different members of the ApiAP2 transcription factor family in apicomplexan parasites.

Frequent exposure to green space has been linked to positive health and well-being in varying populations. Yet, there is still limited research exploring the restorative benefits associated with differing types of green space use among students living in the university setting. To address this gap, we explored green space use amongst a population of undergraduate students (n = 207) attending a university with abundant opportunities to access the restorative properties of nature. The purpose of this study was to examine the type and frequency of green space interactions that are most strongly associated with indicators of health and well-being, and investigate student characteristics associated with frequent use of green space. Results revealed that students who frequently engage with green spaces in active ways report higher quality of life, better overall mood, and lower perceived stress. Passive green space interactions were not strongly associated with indicators of health and well-being. Having had daily interactions with green space in childhood was associated with frequent green space use as a university student, and identified barriers to green space use included "not enough time," and "not aware of opportunities" These results could assist in the tailoring of "green exercise" interventions conducted in the university setting.

Objectives: The aim of this study was to investigate the relationship between BRCA1 and mitotic arrest deficiency protein 2 (MAD2) protein expression, as determined by immunohistochemistry, and clinical outcomes in epithelial ovarian carcinoma (EOC)., Methods: A tissue microarray consisting of 94 formalin-fixed paraffin-embedded EOC with fully matched clinicopathological data were immunohistochemically stained with anti-BRCA1 and anti-MAD2 antibodies. The cores were scored in a semiquantitative manner evaluating nuclear staining intensity and extent. Coexpression of BRCA1 and MAD2 was evaluated, and patient survival analyses were undertaken., Results: Coexpression of BRCA1 and MAD2 was assessed in 94 EOC samples, and survival analysis was performed on 65 high-grade serous carcinomas (HGSCs). There was a significant positive correlation between BRCA1 and MAD2 expression in this patient cohort (P < 0.0001). Both low BRCA1 and low MAD2 are independently associated with overall survival because of HGSC. Low coexpression of BRCA1 and MAD2 was also significantly associated with overall survival and was driven by BRCA1 expression., Conclusion: BRCA1 and MAD2 expressions are strongly correlated in EOC, but BRCA1 expression remains the stronger prognostic factor in HGSC.

The interest in improved environmental sustainability of agriculture via biodiversity provides an opportunity for placed-based research on the conceptualization and articulation of ecosystem services. Yet, few studies have explored how farmers conceptualize the relationship between their farm and nature and by extension ecosystem services. Examining how farmers in the Southern Piedmont of South Carolina discuss and explain the role of nature on their farm, we create a detail-rich picture of how they perceive ecosystem services and their contributions to the agroeconomy. Using 34 semi-structured interviews, we developed a detail-rich qualitative portrait of these farmers' conceptualizations of ecosystem services. Farmers' conceptualization of four ecosystem services: provisioning, supporting, regulating, and cultural are discussed, as well as articulation of disservices. Results of interviews show that most interviewees expressed a basic understanding of the relationship between nature and agriculture and many articulated benefits provided by nature to their farm. Farmers referred indirectly to most services, though they did not attribute services to biodiversity or ecological function. While farmers have a general understanding and appreciation of nature, they lack knowledge on specific ways biodiversity benefits their farm. This lack of knowledge may ultimately limit farmer decision-making and land management to utilize ecosystem services for environmental and economic benefits. These results suggest that additional communication with farmers about ecosystem services is needed as our understanding of these benefits increases. This change may require collaboration between conservation biology professionals and extension and agriculture professionals to extended successful biomass provisioning services to other ecosystem services.

Background: Evidence-based approaches to cognitive rehabilitation are limited; however, new technologies such as brain-training computer programs provide opportunities for novel interventions., Objective: This paper describes a randomized controlled training study in a military treatment facility with service members who had combat-related cognitive symptoms. It examines challenges in study design and implementation, and provides "lessons learned" with proposed solutions., Methods: Participants were randomly assigned to one of two 6-week computer-based cognitive training (CBCT) programs or a treatment-as-usual (TAU) control group. Feasibility assessments included reasons for consent refusal, compliance, and drop-out rates., Results: The intended sample size for the study was 114 participants before attrition. Of 291 patients referred over 2.5 years, 120 were eligible, 38 consented to participate, and 18 completed the study. Forty-two percent of the participants assigned to CBCT groups completed the required 30 sessions in 6.5 to 32 weeks. Study-design factors that affected enrollment and compliance included eligibility restrictions, lack of a computer-based control condition, and inflexible scheduling., Conclusions: Successful implementation of a high-dose computer-based clinical trial will require design changes such as expanded inclusion criteria, control by sham computer program or wait-list, dosing flexibility, and web-based options.

Background: There is no method routinely used to predict response to anthracycline and cyclophosphamide-based chemotherapy in the clinic; therefore patients often receive treatment for breast cancer with no benefit. Loss of the Fanconi anemia/BRCA (FA/BRCA) DNA damage response (DDR) pathway occurs in approximately 25% of breast cancer patients through several mechanisms and results in sensitization to DNA-damaging agents. The aim of this study was to develop an assay to detect DDR-deficient tumors associated with loss of the FA/BRCA pathway, for the purpose of treatment selection., Methods: DNA microarray data from 21 FA patients and 11 control subjects were analyzed to identify genetic processes associated with a deficiency in DDR. Unsupervised hierarchical clustering was then performed using 60 BRCA1/2 mutant and 47 sporadic tumor samples, and a molecular subgroup was identified that was defined by the molecular processes represented within FA patients. A 44-gene microarray-based assay (the DDR deficiency assay) was developed to prospectively identify this subgroup from formalin-fixed, paraffin-embedded samples. All statistical tests were two-sided., Results: In a publicly available independent cohort of 203 patients, the assay predicted complete pathologic response vs residual disease after neoadjuvant DNA-damaging chemotherapy (5-fluorouracil, anthracycline, and cyclophosphamide) with an odds ratio of 3.96 (95% confidence interval [Cl] =1.67 to 9.41; P = .002). In a new independent cohort of 191 breast cancer patients treated with adjuvant 5-fluorouracil, epirubicin, and cyclophosphamide, a positive assay result predicted 5-year relapse-free survival with a hazard ratio of 0.37 (95% Cl = 0.15 to 0.88; P = .03) compared with the assay negative population., Conclusions: A formalin-fixed, paraffin-embedded tissue-based assay has been developed and independently validated as a predictor of response and prognosis after anthracycline/cyclophosphamide-based chemotherapy in the neoadjuvant and adjuvant settings. These findings warrant further validation in a prospective clinical study.

Here, we show for the first time, that the familial breast/ovarian cancer susceptibility gene BRCA1 activates the Notch pathway in breast cells by transcriptional upregulation of Notch ligands and receptors in both normal and cancer cells. We demonstrate through chromatin immunoprecipitation assays that BRCA1 is localized to a conserved intronic enhancer region within the Notch ligand Jagged-1 (JAG1) gene, an event requiring ΔNp63. We propose that this BRCA1/ΔNp63-mediated induction of JAG1 may be important the regulation of breast stem/precursor cells, as knockdown of all three proteins resulted in increased tumoursphere growth and increased activity of stem cell markers such as Aldehyde Dehydrogenase 1 (ALDH1). Knockdown of Notch1 and JAG1 phenocopied BRCA1 knockdown resulting in the loss of Estrogen Receptor-α (ER-α) expression and other luminal markers. A Notch mimetic peptide could activate an ER-α promoter reporter in a BRCA1-dependent manner, whereas Notch inhibition using a γ-secretase inhibitor reversed this process. We demonstrate that inhibition of Notch signalling resulted in decreased sensitivity to the anti-estrogen drug Tamoxifen but increased expression of markers associated with basal-like breast cancer. Together, these findings suggest that BRCA1 transcriptional upregulation of Notch signalling is a key event in the normal differentiation process in breast tissue.

Objectives: The objective of this study was to investigate the relationship between BRCA1 protein expression, as determined by immunohistochemistry, and clinical outcome in uterine serous carcinoma (USC)., Methods: A tissue microarray containing duplicate cores of 73 cases of USC was immunohistochemically stained with mouse anti-BRCA1 (Ab-1) mouse monoclonal (MS110) antibody. The cores were scored in a semiquantitative manner evaluating both the distribution and intensity of nuclear staining. BRCA1 protein expression was correlated with progression-free survival., Results: Seventy-two of 73 cases were assessable, and there was a statistically significant decreased progression-free survival for those cases exhibiting tumor cell nuclei staining of 76% or greater (P = 0.0023)., Conclusions: Our study illustrates that a low level of BRCA1 protein expression is a favorable prognostic indicator in USC, similar to what is observed in high-grade serous ovarian carcinoma. Further studies should focus on the BRCA1 status of USCs at a molecular level and also investigate whether BRCA1 protein expression is associated with response to chemotherapy in USC.

Up to 50% of epithelial ovarian cancers (EOC) display defects in the homologous recombination (HR) pathway. We sought to determine the ramifications of the homologous recombination-deficient (HRD) status on the clinicopathologic features, chemotherapy response, and survival outcomes of patients with EOCs. HR status was determined in primary cultures from ascitic fluid in 50 chemotherapy-naïve patients by a functional RAD51 immunofluorescence assay and correlated with in vitro sensitivity to the PARP inhibitor (PARPi), rucaparib. All patients went on to receive platinum-based chemotherapy; platinum sensitivity, tumor progression, and overall survival were compared prospectively in HR-competent versus HRD patients. Compared with HR-competent patients, the HRD group was predominantly serous with a higher median CA125 at presentation. HRD was associated with higher ex vivo PARPi sensitivity and clinical platinum sensitivity. Median follow-up duration was 14 months; patients in the HRD group had lower tumor progression rates at 6 months, lower overall/disease-specific death rates at 12 months, and higher median survival. We therefore suggest that HRD as predicted by a functional RAD51 assay correlates with in vitro PARPi sensitivity, clinical platinum sensitivity, and improved survival outcome., (©2012 AACR.)

Therapeutic options for malignant pleural mesothelioma (MPM) are limited despite the increasing incidence globally. The vinca alkaloid vinorelbine exhibits clinical activity; however, to date, treatment optimization has not been achieved using biomarkers. BRCA1 regulates sensitivity to microtubule poisons; however, its role in regulating vinorelbine-induced apoptosis in mesothelioma is unknown. Here we demonstrate that BRCA1 plays an essential role in mediating vinorelbine-induced apoptosis, as evidenced by (1) the strong correlation between vinorelbine sensitivity and BRCA1 expression level; (2) induction of resistance to vinorelbine by BRCA1 using siRNA oligonucleotides; (3) dramatic down-regulation of BRCA1 following selection for vinorelbine resistance; and (4) the re-activation of vinorelbine-induced apoptosis following re-expression of BRCA1 in resistant cells. To determine whether loss of BRCA1 expression in mesothelioma was potentially relevant in vivo, BRCA1 immunohistochemistry was subsequently performed on 144 primary mesothelioma specimens. Loss of BRCA1 protein expression was identified in 38.9% of samples. Together, these data suggest that BRCA1 plays a critical role in mediating apoptosis by vinorelbine in mesothelioma, warranting its clinical evaluation as a predictive biomarker., (Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Novel therapeutic approaches and outcome data are needed for cognitive rehabilitation for patients with a traumatic brain injury; computer-based programs may play a critical role in filling existing knowledge gaps. Brain-fitness computer programs can complement existing therapies, maximize neuroplasticity, provide treatment beyond the clinic, and deliver objective efficacy data. However, these approaches have not been extensively studied in the military and traumatic brain injury population. Walter Reed National Military Medical Center established its Brain Fitness Center (BFC) in 2008 as an adjunct to traditional cognitive therapies for wounded warriors. The BFC offers commercially available "brain-training" products for military Service Members to use in a supportive, structured environment. Over 250 Service Members have utilized this therapeutic intervention. Each patient receives subjective assessments pre and post BFC participation including the Mayo-Portland Adaptability Inventory-4 (MPAI-4), the Neurobehavioral Symptom Inventory (NBSI), and the Satisfaction with Life Scale (SWLS). A review of the first 29 BFC participants, who finished initial and repeat measures, was completed to determine the effectiveness of the BFC program. Two of the three questionnaires of self-reported symptom change completed before and after participation in the BFC revealed a statistically significant reduction in symptom severity based on MPAI and NBSI total scores (p &lt; .05). There were no significant differences in the SWLS score. Despite the typical limitations of a retrospective chart review, such as variation in treatment procedures, preliminary results reveal a trend towards improved self-reported cognitive and functional symptoms.

Objectives: We investigated the relationship between BRCA1 protein expression by immunohistochemistry (IHC) and clinical outcome following platinum and platinum/taxane chemotherapy in sporadic epithelial ovarian cancer (EOC)., Methods: BRCA1 IHC was performed on a cohort of 292 ovarian tumours from two UK oncology centres. BRCA1 protein expression levels were correlated with overall survival (OS), progression free survival (PFS) and clinical response to chemotherapy by multivariate analysis., Results: EOC patients with absent/low BRCA1 protein expression (41%) had a better chance of clinical response following chemotherapy as compared to patients with high BRCA1 expression (odds ratio 2.47: 95%CI 1.10-5.55, p=0.029). Patients with absent/low BRCA1 had a higher probability of clinical response following single agent platinum compared to high BRCA1 expressing patients (68.5% vs. 46.8%), while addition of a taxane increased response rates independent of BRCA1. Overall, patients with absent/low BRCA1 had a better clinical outcome compared to patients with high BRCA1 protein expression in terms of both OS (HR=0.65: 95%CI 0.48-0.88, p=0.006) and PFS (HR=0.74, 95%CI 0.55-0.98, p=0.040)., Conclusions: We confirm that absent/low BRCA1 protein expression is a favourable prognostic marker. However, we also provide the first evidence that absent/low BRCA1 protein expression in sporadic EOC patients predicts for an improved clinical response to chemotherapy., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Evasion of apoptosis contributes to both tumourigenesis and drug resistance in non-small cell lung carcinoma (NSCLC). The pro-apoptotic BCL-2 family proteins BAX and BAK are critical regulators of mitochondrial apoptosis. New strategies for targeting NSCLC in a mitochondria-independent manner should bypass this common mechanism of apoptosis block. BRCA1 mutation frequency in lung cancer is low; however, decreased BRCA1 mRNA and protein expression levels have been reported in a significant proportion of lung adenocarcinomas. BRCA1 mutation/deficiency confers a defect in homologous recombination DNA repair that has been exploited by synthetic lethality through inhibition of PARP (PARPi) in breast and ovarian cells; however, it is not known whether this same synthetic lethal mechanism exists in NSCLC cells. Additionally, it is unknown whether the mitochondrial apoptotic pathway is required for BRCA1/PARPi-mediated synthetic lethality. Here we demonstrate that silencing of BRCA1 expression by RNA interference sensitizes NSCLC cells to PARP inhibition. Importantly, this sensitivity was not attenuated in cells harbouring mitochondrial apoptosis block induced by co-depletion of BAX and BAK. Furthermore, we demonstrate that BRCA1 inhibition cannot override platinum resistance, which is often mediated by loss of mitochondrial apoptosis signalling, but can still sensitize to PARP inhibition. Finally we demonstrate the existence of a BRCA1-deficient subgroup (11-19%) of NSCLC patients by analysing BRCA1 protein levels using immunohistochemistry in two independent primary NSCLC cohorts. Taken together, the existence of BRCA1-immunodeficient NSCLC suggests that this molecular subgroup could be effectively targeted by PARP inhibitors in the clinic and that PARP inhibitors could be used for the treatment of BRCA1-immunodeficient, platinum-resistant tumours., (Copyright © 2011 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Expression profiling of BRCA1-deficient tumours has identified a pattern of gene expression similar to basal-like breast tumours. In this study, we examine whether a BRCA1-dependent transcriptional mechanism may underpin the link between BRCA1 and basal-like phenotype. In methods section, the mRNA and protein were harvested from a number of BRCA1 mutant and wild-type breast cancer cell lines and from matched isogenic controls. Microarray-based expression profiling was used to identify potential BRCA1-regulated transcripts. These gene targets were then validated (by in silico analysis of tumour samples) by real-time PCR and Western blot analysis. Chromatin immunoprecipitation (ChIP) assays were used to confirm recruitment of BRCA1 to specific promoters. In results, we demonstrate that functional BRCA1 represses the expression of cytokeratins 5(KRT5) and 17(KRT17) and p-Cadherin (CDH3) in HCC1937 and T47D breast cancer cell lines at both mRNA and protein level. ChIP assays demonstrate that BRCA1 is recruited to the promoters of KRT5, KRT17 and CDH3, and re-ChIP assays confirm that BRCA1 is recruited independently to form c-Myc and Sp1 complexes on the CDH3 promoter. We show that siRNA-mediated inhibition of endogenous c-Myc (and not Sp1) results in a marked increase in CDH3 expression analogous to that observed following the inhibition of endogenous BRCA1. The data provided suggest a model whereby BRCA1 and c-Myc form a repressor complex on the promoters of specific basal genes and represent a potential mechanism to explain the observed overexpression of key basal markers in BRCA1-deficient tumours.

Objectives: Treatment of epithelial ovarian cancer (EOC) remains a challenge, despite advances in surgery and chemotherapy. Hereditary ovarian cancer is primarily due to germline mutations in the BRCA1 tumour suppressor gene. In addition, sporadic EOC tumours display significant of loss of BRCA1 function due to epigenetic inactivation of the BRCA1 gene. This article reviews the preclinical and clinical evidence to support a role for BRCA1 as a potential predictive biomarker of response to both platinum and taxane based chemotherapy in EOC., Methods: We conducted a Medline and Pubmed search for reports between 1990 and 2008 using the search terms: BRCA1 and hereditary ovarian cancer, BRCA1 and sporadic ovarian cancer, ovarian cancer and chemotherapy, ovarian cancer and taxanes, ovarian cancer and platinums, ovarian cancer and clinical response, BRCA1 and DNA damage, BRCA1 and DNA repair, BRCA1 and mitotic checkpoint. If reports identified by these criteria referred to other papers not in the initial search, then these were also reviewed if relevant to BRCA1 and ovarian cancer., Results: The BRCA1 pathway plays a significant role in the development of both hereditary and sporadic EOC. Evidence suggests that BRCA1 is a potential biomarker of response to platinum chemotherapy in EOC with BRCA1 deficiency predicting for enhanced response. In contrast, initial evidence suggests that loss of BRCA1 function results in reduced response to antimicrotubule-based chemotherapy. The ability of BRCA1 to differentially modulate response to these agents involves loss of BRCA1 mediated DNA repair and mitotic checkpoint control, respectively., Conclusions: Standard first line treatment of EOC consists of a combination of platinum and taxane chemotherapy, however clinically useful biomarkers for predicting response to these agents have yet to be established. BRCA1 may prove useful as a biomarker in EOC for assigning chemotherapy treatments based on the presence or absence of BRCA1 function.

Purpose: We investigated whether BRCA1 mRNA expression levels may represent a biomarker of survival in sporadic epithelial ovarian cancer following chemotherapy treatment., Experimental Design: The effect of loss of BRCA1 expression on chemotherapy response in ovarian cancer was measured in vitro using dose inhibition assays and Annexin V flow cytometry. Univariate and multivariate analyses were done to evaluate the relationship between BRCA1 mRNA expression levels and survival after chemotherapy treatment in 70 fresh frozen ovarian tumors., Results: We show that inhibition of endogenous BRCA1 expression in ovarian cancer cell lines results in increased sensitivity to platinum therapy and decreased sensitivity to antimicrotubule agents. In addition, we show that patients with low/intermediate levels of BRCA1 mRNA have a significantly improved overall survival following treatment with platinum-based chemotherapy in comparison with patients with high levels of BRCA1 mRNA (57.2 versus 18.2 months; P = 0.0017; hazard ratio, 2.9). Furthermore, overall median survival for higher-BRCA1-expressing patients was found to increase following taxane-containing chemotherapy (23.0 versus 18.2 months; P = 0.12; hazard ratio, 0.53)., Conclusions: We provide evidence to support a role for BRCA1 mRNA expression as a predictive marker of survival in sporadic epithelial ovarian cancer.

Background: BRCA1-mutant breast tumors are typically estrogen receptor alpha (ER alpha) negative, whereas most sporadic tumors express wild-type BRCA1 and are ER alpha positive. We examined a possible mechanism for the observed ER alpha-negative phenotype of BRCA1-mutant tumors., Methods: We used a breast cancer disease-specific microarray to identify transcripts that were differentially expressed between paraffin-embedded samples of 17 BRCA1-mutant and 14 sporadic breast tumors. We measured the mRNA levels of estrogen receptor 1 (ESR1) (the gene encoding ER alpha), which was differentially expressed in the tumor samples, by quantitative polymerase chain reaction. Regulation of ESR1 mRNA and ER alpha protein expression was assessed in human breast cancer HCC1937 cells that were stably reconstituted with wild-type BRCA1 expression construct and in human breast cancer T47D and MCF-7 cells transiently transfected with BRCA1-specific short-interfering RNA (siRNA). Chromatin immunoprecipitation assays were performed to determine if BRCA1 binds the ESR1 promoter and to identify other interacting proteins. Sensitivity to the antiestrogen drug fulvestrant was examined in T47D and MCF-7 cells transfected with BRCA1-specific siRNA. All statistical tests were two-sided., Results: Mean ESR1 gene expression was 5.4-fold lower in BRCA1-mutant tumors than in sporadic tumors (95% confidence interval [CI] = 2.6-fold to 40.1-fold, P = .0019). The transcription factor Oct-1 recruited BRCA1 to the ESR1 promoter, and both BRCA1 and Oct-1 were required for ER alpha expression. BRCA1-depleted breast cancer cells expressing exogenous ER alpha were more sensitive to fulvestrant than BRCA1-depleted cells transfected with empty vector (T47D cells, the mean concentration of fulvestrant that inhibited the growth of 40% of the cells [IC40] for empty vector versus ER alpha: >10(-5) versus 8.0 x 10(-9) M [95% CI = 3.1 x 10(-10) to 3.2 x 10(-6) M]; MCF-7 cells, mean IC40 for empty vector versus ER alpha: >10(-5) versus 4.9 x 10(-8) M [95% CI = 2.0 x 10(-9) to 3.9 x 10(-6) M])., Conclusions: BRCA1 alters the response of breast cancer cells to antiestrogen therapy by directly modulating ER alpha expression.

To date, estrogen receptor, progestogen receptor, and HER2/neu represent molecular biomarkers currently used in routine clinical practice to aid treatment decisions. Over the last few years, a large body of preclinical and retrospective clinical data has accumulated that suggests that BRCA1 mutation functions as a novel predictive marker of response to chemotherapy. This article reviews the role of BRCA1 as a predictive marker of chemotherapy response in breast cancer and examines the link between BRCA1 deficiency and the basal-like phenotype. Search strategy. Data for this article were identified through MEDLINE and PubMed searches for published reports using the terms BRCA1, breast cancer, basal-like, chemotherapy, prognosis, and predictive markers. In some cases, due to the restriction of space, readers are referred to review articles to allow further reading. Only articles published in English were included.

The exact functions of BRCA1 have not been fully described but it now seems apparent that it has roles in DNA damage repair, transcriptional regulation, cell cycle control and most recently in ubiquitylation. These functions of BRCA1 are most likely interdependent but this review will focus on the role of BRCA1 in relation to transcriptional regulation and in particular how this impacts upon cell cycle control. We will (i) describe the structure of BRCA1 and how it may contribute to its transcription function; (ii) describe the interaction of BRCA1 with the core transcriptional machinery (RNA polII); (iii) describe how BRCA1 may regulate transcription at an epigenetic level through chromatin modification; (iv) discuss the role of BRCA1 in modulating transcription through its association with sequence-specific transcription factors. Finally, we will discuss the possible effects of BRCA1 transcriptional regulation on downstream targets with known roles in cell cycle control.

Evidence is accumulating to suggest that some of the diverse functions associated with BRCA1 may relate to its ability to transcriptionally regulate key downstream target genes. Here, we identify S100A7 (psoriasin), S100A8, and S100A9, members of the S100A family of calcium-binding proteins, as novel BRCA1-repressed targets. We show that functional BRCA1 is required for repression of these family members and that a BRCA1 disease-associated mutation abrogates BRCA1-mediated repression of psoriasin. Furthermore, we show that BRCA1 and c-Myc form a complex on the psoriasin promoter and that BRCA1-mediated repression of psoriasin is dependent on functional c-Myc. Finally, we show that psoriasin expression is induced by the topoisomerase IIalpha poison, etoposide, in the absence of functional BRCA1 and increased psoriasin expression enhances cellular sensitivity to this chemotherapeutic agent. Therefore, we identified a novel transcriptional mechanism that is likely to contribute to BRCA1-mediated resistance to etoposide.

BRCA1 has been reported to have roles in DNA damage repair, cell cycle checkpoint control, transcriptional regulation and ubiquitination. We have previously demonstrated that BRCA1 is a potent activator of a subset of interferon (IFN)-regulated genes and that BRCA1 synergistically activated a number of these genes in the presence of IFN-gamma, but not type I IFNs. Here we report that one of these targets, 2,5 oligoadenylate synthetase (2,5 OAS), is a mediator of BRCA1/IFN-gamma-induced apoptosis. We show that the induction of 2,5 OAS in response to IFN-gamma is BRCA1 and STAT1 dependent. Consistent with a role as a negative regulator of proliferation, transient transfection of 2,5 OAS into breast cancer cell lines results in decreased colony growth and apoptosis. Furthermore we show that IFN-gamma-induced apoptosis is dependent on functional BRCA1 and STAT1 and we demonstrate that IFN-gamma-induced apoptosis is dependent on 2,5 OAS induction. 2,5 OAS is the only known upstream regulator of RNaseL, a recently identified hereditary prostate tumour suppressor gene implicated in apoptosis. We propose that BRCA1 may be an upstream regulator of RNaseL, acting in concert with IFN-gamma to transcriptionally activate 2,5 OAS, leading to the downstream activation of RNaseL and apoptosis.

Objective: Archipelago (AGO, also known as hCdc4, Fbw7, or Sel-10) is an F-box containing component of the SCF complex implicated in the ubiquitination and proteolysis of cyclin E and c-Myc, and found to be mutated in 16% of endometrial carcinomas. We have previously reported somatic mutations in AGO in 3/10 ovarian cancer cell lines, but the frequency of such mutations in primary ovarian cancer is unknown., Methods: The coding sequence of AGO was analyzed in 95 primary sporadic ovarian tumors and 16 cases of familial ovarian cancer, and correlated with levels of cyclin E and c-Myc protein expression. Constructs encoding mutations in AGO were transfected into an AGO-null cell line to directly test their ability to regulate cyclin E and c-Myc levels., Results: Mutations were present in only 2 of 95 sporadic cases: a premature stop within the WD domain (471 Ter) and a missense change near the F-box (S245T). Both primary tumor specimens containing these mutations showed high levels of cyclin E and c-Myc, but reconstitution of an AGO-null cell line with constructs encoding these mutations showed 471 Ter to be inactive in regulating endogenous cyclin E and c-Myc levels, while the S245T mutant was indistinguishable from wild-type. No germ-line mutations were found in familial cases of ovarian cancer., Conclusion: Somatic AGO mutations are infrequent in primary ovarian cancers and are unlikely to contribute to familial ovarian cancer. Reconstitution experiments, rather than measuring tumor levels of cyclin E and c-Myc, provide an effective approach to determine the functional significance of AGO mutations identified in human cancers.

Osteoprotegerin (OPG), a critical regulator of osteoclastogenesis, is expressed by prostate cancer cells, and OPG levels are increased in patients with prostate cancer bone metastases. The objective of this study was to investigate the effects of OPG overexpression on prostate cancer cells and prostate cancer/bone cell interactions in vitro and in vivo. OPG-transfected C4-2 cells expressed 8.0 ng OPG per mL per 10(6) cells, whereas no OPG was detected in the media of C4-2 cells transfected with a control plasmid. OPG overexpressed by C4-2 cells protected these cells from tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis and decreased osteoclast formation. Subcutaneous OPG-C4-2 and pcDNA-C4-2 tumors exhibited similar growth and take-rate characteristics. However, when grown in bone, tumor volume was decreased in OPG-C4-2 versus pcDNA-C4-2 (P=0.0017). OPG expressed by C4-2 cells caused increases in bone mineral density (P=0.0074) and percentage of trabecular bone volume (P=0.007), and decreases in numbers of osteoblasts and osteoclasts when compared with intratibial pcDNA-C4-2 tumors (P=0.003 and P=0.019, respectively). In summary, our data show that increased expression of OPG in C4-2 cells does not directly affect proliferation of prostate cancer cells but indirectly decreases growth of C4-2 tumors in the bone environment. Our data also show that OPG expressed by C4-2 cells inhibits bone lysis associated with C4-2 bone metastasis, which results in net increases in bone volume. We therefore hypothesize that OPG expressed in prostate cancer patient bone metastases may be at least partially responsible for the osteoblastic character of most prostate cancer bone lesions.

Zoledronic acid (ZA) has been shown to inhibit prostate tumor growth in vitro and have beneficial effects in patients with advanced prostate cancer (CaP). The aim of this study was to determine whether ZA exhibits direct anti-tumor effects on CaP cells in vivo. To distinguish the effects of inhibition of osteolysis and direct anti-tumor activity of ZA in vivo, we compared the results of treatment with ZA and osteoprotegerin (Fc-OPG), which inhibits osteolysis, but without significant direct anti-tumor effects. In vitro Fc-OPG had no significant effects on C4-2 proliferation, whereas ZA decreased proliferation. However, both agents decreased tumor growth in bone. Moreover, both increased bone volume and prevented the overall decreases in BMD associated with growth of C4-2 cells in bone. Our study provides novel and significant observations that the in vivo effects of ZA are consistent with indirect effects mediated by osteoclasts.

Germline mutations of the BRCA1 gene account for approximately 5% of breast and ovarian cancer cases, and lower than normal BRCA1 expression or function may be an important contributing factor in sporadic cancers. The major role of BRCA1 is to respond to DNA damage by participating in cellular pathways for DNA repair, mRNA transcription, cell cycle regulation, and protein ubiquitination. Because most chemotherapeutic agents function by directly or indirectly damaging DNA, the role of BRCA1 as a regulator of chemotherapy-induced DNA damage has been the subject of an increasing number of investigations. We review published preclinical and clinical evidence that the level of BRCA1 function in an individual patient's tumor can guide the choice of chemotherapeutic agents for breast and ovarian cancer. We conclude that a loss of BRCA1 function is associated with sensitivity to DNA-damaging chemotherapy and may also be associated with resistance to spindle poisons. We recommend that prospective clinical studies investigating the role of BRCA1 in the response to chemotherapy be conducted.

Background: LNCaP and its derivative cell lines, which include C4-2 (and the related C4-2B) and CL1, are used as models of prostate cancer. Unlike LNCaP, the other cell lines show features of progressed disease such as metastatic capability and hormone independence. Analyses were done to determine if C4-2 or CL1 cells were selected from pre-existent subpopulations in LNCaP., Methods: Prostate cancer cells were characterized by cluster designation (CD) phenotyping. Specific cell populations were sorted by flow cytometry. DNA array analysis was used to probe differential gene expression., Results: CD phenotyping showed that CL1 and C4-2 (and C4-2B) were very dissimilar, and C4-2 was more similar to LNCaP. One common difference between LNCaP and its derivatives was CD26, in which virtually all C4-2 or CL1 cells were CD26(+) but only approximately 10% of LNCaP cells were CD26(+). The CD26(+) subpopulation of LNCaP was isolated and cultured in vitro. After culture, a high percentage of the cells (descended from the sorted cells) were CD26(+), in contrast to those sorted by CD13 or CD44. The cultured CD13 and CD44 populations did not show a high percentage of CD13(+) and CD44(+) cells, respectively. CD13 and CD44 are markers, in addition to CD26, for CL1 but not for C4-2., Conclusions: C4-2 arose probably from CD26(+) LNCaP cells, while CL1 arose de novo., (Copyright 2004 Wiley-Liss, Inc.)

BRCA1 has been implicated in a number of cellular processes, including transcriptional regulation, DNA damage repair, cell cycle arrest, and apoptosis. We identified mitogen-activated protein kinase (MAPK) kinase kinase 3 (MEKK3), an upstream regulator of the c-Jun NH(2)-terminal kinase/stress-activated protein kinase and p38/MAPK pathways, as a novel BRCA1-interacting protein in a yeast two-hybrid screen and confirmed the interaction by coimmunoprecipitation in mammalian cells. Deletion mapping demonstrated that amino acids 1611-1863 are required to mediate the interaction with MEKK3 in yeast. BRCA1 disease-associated mutations abrogated the interaction in yeast, and BRCA1 failed to interact with MEKK3 in BRCA1 mutant HCC1937 breast cancer cells. We demonstrate that small interfering RNA-based inhibition of endogenous BRCA1 reduces MEKK3 kinase activity and conversely that inducible expression of BRCA1 activates MEKK3 and p38/MAPK. Finally, we demonstrate using complementary approaches that BRCA1 is required for paclitaxel-induced activation of MEKK3. These data indicate that BRCA1 is a key regulator of the paclitaxel-induced stress response pathway and suggest that the ability of BRCA1 to associate with, and mediate the activation of, MEKK3 represents a potential mechanism through which this pathway is regulated.

Osteoprotegerin (OPG) plays a central role in controlling bone resorption. Exogenous administration of OPG has been shown to be effective in preventing osteolysis and limiting the growth of osteolytic metastasis. The objective of this study was to investigate the effects of OPG on osteoblastic prostate cancer (CaP) metastases in an animal model. LuCaP 23.1 cells were injected intra-tibially and Fc-OPG (6.0 mg/kg) was administered subcutaneously three times a week starting either 24 hours prior to cell injection (prevention regimen) or at 4 weeks post-injection (treatment regimen). Changes in bone mineral density at the tumor site were determined by dual x-ray absorptiometry. Tumor growth was monitored by evaluating serum prostate specific antigen (PSA). Fc-OPG did not inhibit establishment of osteoblastic bone lesions of LuCaP 23.1, but it decreased growth of the tumor cells, as determined by decreases in serum PSA levels of 73.0 +/- 44.3% (P < 0.001) and 78.3 +/- 25.3% (P < 0.001) under the treatment and prevention regimens, respectively, compared to the untreated tumor-bearing animals. Administration of Fc-OPG decreased the proliferative index by 35.0% (P = 0.1838) in the treatment group, and 75.2% (P = 0.0358) in the prevention group. The results of this study suggest a potential role for OPG in the treatment of established osteoblastic CaP bone metastases.

Unlabelled: New well-characterized preclinical models of prostate cancer (CaP) bone metastases are needed to improve our understanding of the development of CaP-related bone disease in patients. Here we describe characterization of a model consisting of direct injection of C4-2 cells into tibias., Introduction: Prostate cancer (CaP) has a high proclivity to metastasize to bone. Development and characterization of preclinical models of CaP bone metastases are of high interest. The objective of this study was to characterize C4-2 bone metastases and their response to castration., Materials and Methods: Cell suspensions of C4-2, a subline of LNCaP, were injected directly into the tibias of intact male mice. In groups A (n = 7) and B (n = 5), animals were killed 3 and 8 weeks after injection of C4-2 cells, respectively. In group C (n = 7), animals were castrated 3 weeks after injection and killed 5 weeks after castration. Serum prostate-specific antigen (PSA) levels and bone mineral density (BMD) were measured, and bone histomorphometric analysis was performed., Results: C4-2 cells decreased BMD of the injected tibias by 36.1% and bone volume by 74.1% versus normal tibias. Castration caused a 32.3% drop in serum PSA (p = 0.0438), with a nadir at day 14, after which it began to rise again. Bone destruction in the tumorous tibias of castrated animals was decreased by 15.9% versus tumorous tibias of intact animals (p = 0.0392). However, BMD in the tumorous tibias of castrated mice was still lower than in normal tibias of intact animals. Castration also decreased BMD and bone volume in nontumorous tibias (p = 0.0406 and 0.0232, respectively)., Conclusions: The C4-2 model of bone metastasis recapitulates the response to androgen deprivation observed in CaP patients with bone metastases and is suitable for study of interactions between tumor and bone cells and evaluation of new therapeutic modalities.

We have evaluated the role played by BRCA1 in mediating the phenotypic response to a range of chemotherapeutic agents commonly used in cancer treatment. Here we provide evidence that BRCA1 functions as a differential mediator of chemotherapy-induced apoptosis. Specifically, we demonstrate that BRCA1 mediates sensitivity to apoptosis induced by antimicrotubule agents but conversely induces resistance to DNA-damaging agents. These data are supported by a variety of experimental models including cells with inducible expression of BRCA1, siRNA-mediated inactivation of endogenous BRCA1, and reconstitution of BRCA1-deficient cells with wild-type BRCA1. Most notably we demonstrate that BRCA1 induces a 10-1000-fold increase in resistance to a range of DNA-damaging agents, in particular those that give rise to double-strand breaks such as etoposide or bleomycin. In contrast, BRCA1 induces a >1000-fold increase in sensitivity to the spindle poisons, paclitaxel and vinorelbine. Fluorescence-activated cell sorter analysis demonstrated that BRCA1 mediates G(2)/M arrest in response to both antimicrotubule and DNA-damaging agents. However, poly(ADP-ribose) polymerase and caspase-3 cleavage assays indicate that the differential effect mediated by BRCA1 in response to these agents occurs through the inhibition or induction of apoptosis. Therefore, our data suggest that BRCA1 acts as a differential modulator of apoptosis depending on the nature of the cellular insult.

Objectives: Many emergency departments (EDs) receive patients from concert or other mass gathering events. The study objective was to determine whether routine emergency medical services (EMS) transport to a hospital from an indoor arena facility is warranted., Methods: Retrospective review of transport medical records from an approximately 20,000-seat arena in a major metropolitan area from January 1, 1998, to June 30, 1999. Outcomes studied included inpatient admission rate, diagnoses, treatments, and length of ED stay., Results: The authors reviewed 96 patients transported from 29 separate concert or professional wrestling events. The mean age was 23.2 years (SD, 10.4 yr). Only three patients (3.1%, 95% confidence interval [CI], 0.65%-8.9%) required inpatient admission. The mean length of stay in the ED was 158 minutes (95% CI, 132-185). Thirty-one percent of the patients had diagnoses of alcohol or drug use; trauma accounted for 33%; and medical reasons represented 35% of transports. Alcohol and/or drug cases stayed in the ED 207 minutes (mean), which was 59% longer than trauma cases (130 min) and 47% longer than the 141 minutes for medical cases (p = 0.007). Rock concerts had 45% alcohol/drug-related cases versus 22% for pop concerts and 0% for professional wrestling events (p = 0.001)., Conclusions: Patients transported from indoor arena events rarely result in inpatient admissions. Alcohol- and drug-related problems were the primary diagnoses in 31% of these patients and required the most time in the ED. Rock concerts had more alcohol and drug cases than other events.

Background: Spontaneous metastasis following implantation at the orthotopic site is a highly desired feature in prostate cancer (CaP) models, since it would enable studies of mechanisms associated with tumor cell dissemination., Methods: LuCaP 23.8 and LuCaP 35, hormone-sensitive CaP xenografts that express PSA and the wild-type androgen receptor, were grown orthotopically in SCID mice. When tumor volumes reached approximately 250-500 mg, primary tumors were removed to allow micrometastases to grow., Results: Using this procedure we generated macroscopic metastases (>20 mg) in 71% (LuCaP 23.8) and 100% (LuCaP 35) of animals, respectively., Conclusions: These models may be used to evaluate new treatment modalities and study mechanisms associated with development of metastases., (Copyright 2003 Wiley-Liss, Inc.)

Cytogenetic and molecular studies have suggested that deletion or rearrangement of sequences that map to the short arm of chromosome 8 may be permissive for tumorigenesis in several organ systems, and in human prostate tumors in particular. In this study, we hypothesized that genes deleted for one copy and localized to the 8p chromosomal region may be transcriptionally down-regulated or ablated in affected human prostate tumor tissues. To test this hypothesis, we used cDNA microarray analysis to determine the transcriptional profiles for 259 transcribed sequences mapping to the 8p chromosomal region for seven human prostate tumor xenografts, completely characterized for numerical and structural alterations on chromosome 8, and five normal human prostate tissues. These experiments identified 33 genes differentially expressed between normal and malignant prostate tissues, the majority of which (28/33, 85%) were transcriptionally down-regulated in malignant compared to normal human prostate tissues. These findings, that haploinsufficiency and transcriptional down-regulation for genes mapping to 8p are largely coincident in human prostate tumors, should provide a powerful tool for the identification of tumor-suppressor genes associated with human prostate cancer initiation and progression., (Copyright 2003 Wiley-Liss, Inc.)

Background: Generation of suitable in vivo models is critical for understanding of processes associated with development and progression of prostate cancer (CaP)., Methods: Lymph nodes containing metastatic androgen-independent CaP were implanted into athymic mice. A xenograft designated LuCaP 35 and its hormone-independent variant LuCaP 35V were established and characterized., Results: LuCaP 35 is an androgen-sensitive, prostate-specific antigen (PSA)-producing xenograft. It expresses the wild-type androgen receptor and exhibits deletions in chromosome 8p, but not in chromosome 10. The response of LuCaP 35 to androgen ablation is similar to that observed in man. Using recurring LuCaP 35 tumors we have also established an androgen-insensitive variant of LuCaP 35., Conclusions: The availability of hormone-dependent and -independent variants of LuCaP 35, which exhibit many properties analogous to those of CaP in man, provides an excellent model system to study the processes associated with development of androgen independence and to evaluate new treatment modalities., (Copyright 2003 Wiley-Liss, Inc.)

Objectives: A subset of prostate carcinomas is composed predominantly, even exclusively, of neuroendocrine (NE) cells. In this report, we sought to characterize the gene expression profile of a prostate small cell NE carcinoma by assessing the diversity and abundance of transcripts in the LuCaP 49 prostate small cell carcinoma xenograft., Methods: We constructed a cDNA library (PRCA3) from the LuCap 49 prostate small cell xenograft. Single pass DNA sequencing of randomly selected cDNA clones followed by sequence assembly and annotation produced a library of Expressed Sequence Tags (ESTs) representing the LuCaP 49 transcriptome. Comparative sequence analysis with ESTs derived from prostate adenocarcinoma libraries was performed using statistical algorithms designed to identify differentially expressed sequences. Putative NE cell-specific genes were further examined by Northern analysis., Results: Sequence assembly and analysis identified 1,447 distinct genes expressed in the LuCaP 49 cDNA library. These include cDNAs encoding the NE markers secretogranin (SCG2), CD24, and ENO2. Northern analysis revealed that three additional genes, ASCL1, INA, and SV2B are expressed in LuCaP 49 but not in various prostate cancer cell lines or xenografts. Fifteen genes were identified with a statistical probability (P > 0.9) of being up-regulated in LuCaP 49 small cell carcinoma relative to prostate adenocarcinoma (two primary prostate adenocarcinomas and the LNCaP prostate adenocarcinoma cell line)., Conclusions: Prostate small cell carcinoma expresses a diverse repertoire of genes that reflect characteristics of their NE cell of origin. ASCL1, INA, and SV2B are potential molecular markers for small cell NE tumors and NE cells of the prostate. This small cell NE carcinoma gene expression profile may yield insights into the development, progression, and treatment of subtypes of prostate cancer., (Copyright 2003 Wiley-Liss, Inc.)

BRCA1 (breast-cancer susceptibility gene 1) is a tumour suppressor gene that is mutated in the germline of women with a genetic predisposition to breast and ovarian cancer. In this review, we examine the role played by BRCA1 in mediating the cellular response to stress. We review the role played by BRCA1 in detecting and signalling the presence of DNA damage, particularly double-strand DNA breaks, and look at the evidence to support a role for BRCA1 in regulating stress response pathways such as the c-Jun N-terminal kinase/stress-activated protein kinase pathway. In addition, we examine the role played by BRCA1 in mediating both cell-cycle arrest and apoptosis following different types of cellular insult, and how this may be modulated by the presence or absence of associated proteins such as p53. Finally, we explore the possibility that many of the functions associated with BRCA1 may be based on transcriptional regulation of key downstream genes that have been implicated in the regulation of these specific cellular pathways.

Purpose: In this study we have examined the effects of zoledronic acid (ZA), a new-generation bisphosphonate, on prostate cancer (CaP) cells in vitro, and on both osteoblastic and osteolytic CaP metastases in animal models., Experimental Design: In vitro, CaP cells were treated with ZA, and the effects on proliferation, cell cycle, and apoptosis were determined. In vivo, PC-3, and LuCaP 23.1 s.c. and tibial tumors were treated with ZA. Effects on bone and tumor were determined by histomorphometry and immunohistochemistry., Results: ZA decreased proliferation of CaP cells, and caused G(1) arrest and apoptosis of CaP cells in vitro. In vivo, s.c. CaP tumor growth was not affected by ZA. However, growth of osteoblastic and osteolytic metastases of CaP was inhibited significantly in vivo. Matrix metalloproteinase-2, matrix metalloproteinase-9, and Cathepsin K levels were decreased in osteolytic bone metastases after ZA administration., Conclusions: In conclusion, we have shown that ZA has significant antitumor effects on CaP cells in vitro and in vivo. Antiosteolytic activity and the antitumor effects of this compound could benefit CaP patients with bone metastases.

The BRCA1 gene was cloned in 1994 as one of the genes that conferred genetic predisposition to early-onset breast and ovarian cancer. Since then, a genetic test for identification of high-risk individuals has been developed. Despite being implicated in many important cellular pathways, including DNA repair and regulation of transcription, the exact mechanism by which inactivation of BRCA1 might lead to malignant transformation of cells remains unknown. We examine the mechanisms that underlie inactivation of BRCA1 and assess how they affect management of patients, in terms of both primary and secondary cancer prevention strategies. Furthermore, we look at the potential usefulness of BRCA1 as a prognostic tool and as a predictive marker of response to different classes of drugs. Finally, throughout this review, we draw links between the functional consequences of BRCA1 inactivation, in terms of key cellular signalling pathways, and how they might explain specific clinical observations in individuals who carry mutations in the gene.

BRCA1 is a tumor suppressor gene implicated in transcriptional regulation. We have generated cell lines with inducible expression of BRCA1 as a tool to identify downstream targets that may be important mediators of BRCA1 function. Oligonucleotide array-based expression profiling identified 11 previously described interferon regulated genes that were up-regulated following inducible expression of BRCA1. Northern blot analysis revealed that a subset of the identified targets including IRF-7, MxA, and ISG-54 were synergistically up-regulated by BRCA1 in the presence of interferon gamma (IFN-gamma) but not interferons alpha or beta. Importantly, IFN-gamma-mediated induction of IRF-7 and MxA was attenuated in the BRCA1 mutant cell line HCC1937, an effect that was rescued following reconstitution of exogenous wild type BRCA1 in these cells. Furthermore, reconstituted BRCA1 sensitized HCC1937 cells to IFN-gamma-induced apoptotic cell death. This study identifies BRCA1 as a component of the IFN-gamma-regulated signaling pathway and suggests that BRCA1 may play a role in the regulation of IFN-gamma-mediated apoptosis.

Background: To improve the therapy of advanced prostate cancer (CaP), it is critical to develop animal models that mimic CaP bone metastases. Unlike the human disease, CaP xenograft models rarely metastasize spontaneously to bone from the orthotopic site of primary tumor growth., Methods: Single-cell suspensions of LNCaP, PC-3, LuCaP 35, and LuCaP 23.1 CaP cells were injected directly into tibia of SCID mice. Immunohistochemistry and bone histomorphometrical analyses were performed to characterize these osseous-CaP models., Results: LuCaP 23.1 yields an osteoblastic response, LNCaP yields mixed lesions, and LuCaP 35 and PC-3 result in osteolytic responses. We have detected osteoprotegerin, RANK ligand, parathyroid hormone-related protein, and endothelin-1, proteins associated with bone growth and remodeling, in the CaP cells grown in the bone., Conclusions: These animal models can be used to study biological interactions, pathways, and potential therapeutic targets, and also to evaluate new agents for treatment and prevention of CaP bone metastasis., (Copyright 2002 Wiley-Liss, Inc.)

Purpose: Estrogen treatment has long been known to be of benefit in prostate cancer (CaP), but its mechanism was thought to involve merely a reduction in androgen levels. However, new evidence indicates that estrogen may exert effects on CaP cells in the absence of androgens., Experimental Design: Implantation of CaP xenografts (LuCaP 35, LuCaP 49, LuCaP 58, LuCaP 73, PC-3, and LNCaP) into intact and ovariectomized female mice was done to characterize growth and take rates in the absence of androgens. Ovariectomized female mice were supplemented with 17beta-estradiol, and LuCaP 35 CaP xenograft take and growth rates were determined. Reverse transcription-PCR was used to evaluate the presence of the estrogen receptor messages in CaP xenografts., Results: We have observed significant inhibition of CaP growth in intact versus ovariectomized female animals in five of six CaP xenograft lines. 17beta-Estradiol supplements given to ovariectomized female mice led to inhibition of tumor establishment and diminished growth of LuCaP 35 similar to that observed in intact female mice. Using reverse transcription-PCR, we have shown that these xenografts express the estrogen receptor beta message., Conclusions: We have determined that 17beta-estradiol supplementation causes inhibition of CaP growth in an animal model by mechanisms that are independent of androgen action. This gives rise to the possibility that estrogen therapy may be of potential use with hormone-refractory cancers. The xenograft models we describe herein may be useful as well in elucidating the pathways mediating the androgen-independent effects of estrogen on CaP.

Mutations in the CWH8 gene, which encodes an ER transmembrane protein with a phosphate binding pocket in Saccharomyces cerevisiae, result in a deficiency in dolichyl pyrophosphate (Dol-P-P)-linked oligosaccharide intermediate synthesis and protein N-glycosylation (van Berkel, M. A., Rieger, M., te Heesen, S., Ram, A. F., van den Ende, H., Aebi, M., and Klis, F. M. (1999) Glycobiology 9, 243-253). Genetic, enzymological, and topological approaches were taken to investigate the potential role of Cwh8p in Dol-P-P/Dol-P metabolism. Overexpression of Cwh8p in the yeast double mutant strain, lacking LPP1/DPP1, resulted in an impressive increase in Dol-P-P phosphatase activity, a relatively small increase in Dol-P phosphatase activity, but no change in phosphatidate (PA) phosphatase activity in microsomal fractions. The Dol-P-P phosphatase encoded by CWH8 is optimally active in the presence of 0.5% octyl glucoside and relatively unstable in Triton X-100, distinguishing this activity from the lipid phosphatases encoded by LPP1 and DPP1. Stoichiometric amounts of P(i) and Dol-P are formed during the enzymatic reaction indicating that Cwh8p cleaves the anhydride linkage in Dol-P-P. Membrane fractions from Sf-9 cells expressing Cwh8p contained a 30-fold higher level of Dol-P-P phosphatase activity, a slight increase in Dol-P phosphatase activity, but no increase in PA phosphatase relative to controls. This is the first report of a lipid phosphatase that hydrolyzes Dol-P-P/Dol-P but not PA. In accord with this enzymatic function, Dol-P-P accumulated in cells lacking the Dol-P-P phosphatase. Topological studies using different approaches indicate that Cwh8p is a transmembrane protein with a luminally oriented active site. The specificity, subcellular location, and topological orientation of this novel enzyme are consistent with a role in the re-utilization of the glycosyl carrier lipid for additional rounds of lipid intermediate biosynthesis after its release during protein N-glycosylation reactions.

BRCA1 is a tumour suppressor gene implicated in the predisposition to early onset breast and ovarian cancer. We have generated cell lines with inducible expression of BRCA1 to evaluate its role in mediating the cellular response to various chemotherapeutic drugs commonly used in the treatment of breast and ovarian cancer. Induction of BRCA1 in the presence of Taxol and Vincristine resulted in a dramatic increase in cell death; an effect that was preceded by an acute arrest at the G2/M phase of the cell cycle and which correlated with BRCA1 mediated induction of GADD45. A proportion of the arrested cells were blocked in mitosis suggesting activation of both a G2 and a mitotic spindle checkpoint. In contrast, no specific interaction was observed between BRCA1 induction and treatment of cells with a range of DNA damaging agents including Cisplatin and Adriamycin. Inducible expression of GADD45 in the presence of Taxol induced both G2 and mitotic arrest in these cells consistent with a role for GADD45 in contributing to these effects. Our results support a role for both BRCA1 and GADD45 in selectively regulating a G2/M checkpoint in response to antimicrotubule agents and raise the possibility that their expression levels in cells may contribute to the toxicity observed with these compounds.