Increasing carotenoid production in Xanthophyllomyces dendrorhous/Phaffia rhodozyma: SREBP pathway activation and promoter engineering.

The yeast Xanthophyllomyces dendrorhous synthesizes astaxanthin, a high-value carotenoid with biotechnological relevance in the nutraceutical and aquaculture industries. However, enhancing carotenoid production through strain engineering remains an ongoing challenge. Recent studies have demonstrated that carotenogenesis in X. dendrorhous is regulated by the SREBP pathway, which includes the transcription factor Sre1, particularly in the mevalonate pathway that also produces precursors used for ergosterol synthesis. In this study, we explored a novel approach to enhance carotenoid synthesis by replacing the native crtE promoter, which drives geranylgeranyl pyrophosphate synthesis (the step where carotenogenesis diverges from ergosterol biosynthesis), with the promoter of the HMGS gene, which encodes 3-hydroxy-3-methylglutaryl-CoA synthase from the mevalonate pathway. The impact of this substitution was evaluated in two mutant strains that already overproduce carotenoids due to the presence of an active Sre1 transcription factor: CBS.cyp61-, which does not produce ergosterol and strain CBS.SRE1N.FLAG, which constitutively expresses the active form of Sre1. Wild-type strain CBS6938 was used as a control. Our results showed that this modification increased the crtE transcript levels more than threefold and fourfold in CBS.cyp61-.pHMGS/crtE and CBS.SRE1N.FLAG.pHMGS/crtE, respectively, resulting in 1.43-fold and 1.22-fold increases in carotenoid production. In contrast, this modification did not produce significant changes in the wild-type strain, which lacks the active Sre1 transcription factor under the same culture conditions. This study highlights the potential of promoter substitution strategies involving genes regulated by Sre1 to enhance carotenoid production, specifically in strains where the SREBP pathway is activated, offering a promising avenue for strain improvement in industrial applications.

From the Individual to the Group: Using Idiographic Analyses and Two-Stage Random Effects Meta-Analysis to Obtain Population Level Inferences for within-Person Processes.

In psychology, the use of portable technology and wearable devices to ease participant burden in data collection is on the rise. This creates increased interest in collecting real-time or near real-time data from individuals within their natural environments. As a result, vast amounts of observational time series data are generated. Often, motivation for collecting this data hinges on understanding within-person processes that underlie psychological phenomena. Motivated by the body of Dr. Peter Molenaar's life work calling for analytical approaches that consider potential heterogeneity and non-ergodicity, the focus of this paper is on using idiographic analyses to generate population inferences for within-person processes. Meta-analysis techniques using one-stage and two-stage random effects meta-analysis as implemented in single-case experimental designs are presented. The case for preferring a two-stage approach for meta-analysis of single-subject observational time series data is made and demonstrated using an empirical example. This provides a novel implementation of the methodology as prior implementations focus on applications to short time series with experimental designs. Inspired by Dr. Molenaar's work, we describe how an approach, two-stage random effects meta-analysis (2SRE-MA), aligns with recent calls to consider idiographic approaches when making population-level inferences regarding within-person processes.

Impact of Somatic DNA Repair Mutations on the Clinical Outcomes of Bone Metastases from Castration-Resistant Prostate Cancer.

Up to 80% of castration-resistant prostate cancer (CRPC) patients develop bone metastases during the natural history of disease and about 25% harbor mutations in DNA damage repair (DDR) genes. This retrospective observational study evaluated the prevalence of DDR alterations in CRPC patients and their effect on the clinical outcomes associated with bone metastases. The mutational status of CRPC patients was analyzed per FoundationOne® analysis in tissue biopsy or, when it was not possible, in liquid biopsy performed at the onset of metastatic CRPC (mCRPC). The impact of DDR gene mutations on bone-related efficacy endpoints was evaluated at the time of mCRPC diagnoses. In total, 121 mCRPC patients with bone metastases were included: 38 patients had mutations in at least one DDR gene, the remaining 83 ones had a non-mutated DDR status. DDR mutated status was associated with bone metastases volume (p = 0.006), but did not affect SRE (skeletal-related events) incidence and time to SRE onset. Liquid and tissue biopsies were both available for 61 patients with no statistically significant difference in terms of incidence and type of molecular DDR alterations. Mutated DDR status was associated with higher bone metastasic volume, although a not detrimental effect on the other bone-related efficacy endpoints was observed.

3-T magnetic resonance-guided high-intensity focused ultrasound (3 T-MR-HIFU) for the treatment of pain from bone metastases of solid tumors.

INTRODUCTION: Bone metastases (BM) are still the main cause of morbidity and mortality in cancer patients, not only because of their complications, defined as skeletal-related events (SREs), but also because of the negative impact bone pain has on quality of life (QoL) and survival, especially when opioid analgesics and locoregional treatments fail. MATERIALS AND METHODS: A single-center prospective study was carried out on 12 patients with symptomatic BM treated with MRI-guided focused ultrasound (MR-HIFU). The primary endpoint was the effectiveness of MR-HIFU in reducing current and breakthrough cancer pain (BTCP) scores. The main secondary aims were the evaluation of circulating markers at different time-points and their relation to pain and procedure efficacy. Other secondary objectives included temporal evolution of pain response, evaluation of QoL, and side effects of the treatment. Descriptive statistics were used to evaluate primary and secondary endpoints. Questionnaires on pain and QoL completed at baseline and at 30 days were compared using appropriate statistical tests with exploratory intent. RESULTS: MR-HIFU was successfully completed in all 12 patients enrolled between September 2015 and December 2018. On day 30, 6 (50.0%) patients showed a complete response of current pain and 6 a partial response, while 5 (41.7%) obtained a complete BTCP response. A partial response of BM evaluated by MD Anderson criteria was obtained in 9 (81.8%) patients. Only one patient progressed in the target lesion after MR-HIFU. No treatment-related adverse events were recorded. Bone turnover markers CTX/RANK-L (P) do not demonstrate any significant change with the pain or BM response. CONCLUSION: In our patients, targeted therapy of painful BM with MRI-guided focused ultrasound ablation was safe and showed encouraging early-onset and functional results.

Life-threatening sustained hypocalcemia following Denosumab use in metastatic prostate cancer.

INTRODUCTION: Prostate cancer is the second most frequently diagnosed cancer among men worldwide in 2020. Skeletal-related events (SRE) like pathologic fracture or spinal cord compression are commonly seen in metastatic prostate cancer. Denosumab, a monoclonal antibody, acts by inhibiting osteoclast-mediated bone resorption in bone metastasis from solid tumors and reduces bone turnover and destruction. However, there is an increased risk of life-threatening denosumab-induced hypocalcemia with an incidence of 0.1 to 12.8%. CASE REPORT: Our patient is a 69-year-old man with widespread skeletal metastatic disease from primary prostate cancer who presented to the hospital complaining of generalized fatigue and joint pain. Due to severe debilitating low back pain secondary to osteochondral lesions, the patient was started on Denosumab 120 mg. On presentation, serum calcium was found to be severely low at 5.9 mg/dl (serum calcium level prior to Denosumab was 9.1 mg/dl). MANAGEMENT AND OUTCOME: Denosumab was discontinued immediately, and the patient was started on IV calcium gluconate. Repeat serum calcium level continued to be low at 6.7 likely due to the long elimination half-life of Denosumab (25-30 days). He was transferred to a long-term acute care facility for long-term IV calcium replacement, where he succumbed to illness six weeks later. DISCUSSION: Denosumab, an anti-resorptive treatment for skeletal metastasis from solid tumors, is shown to cause severe life-threatening hypocalcemia. The maximum serum drug level of Denosumab reaches 7-21 days after administration. Sustained hypocalcemia is rare and life-threatening. Clinicians should use this medication with caution due to its unpredictable side effect profile.

Characterization, Behavior, and Risk Assessment of Polycyclic Aromatic Hydrocarbons (PAHs) in the Estuary Sediments.

Polycyclic aromatic hydrocarbons (PAHs) are persistent toxic substances that have ubiquitous presence in water, air, soil, and sediment environments. The growth of PAH toxicities and related ecotoxicology risk in estuary sediment has a serious concern. Present study examined the PAHs concentration, sources, and ecological risk from selected sites in Subarnarekha River estuary (SRE) sediment deposits. The sum of toxic 16 PAHs was ranged from 36.8 to 670.8 ng/g (mean = 223.46 ± 196.35 ng/g). The total PAH concentration varied significantly among the sampling sites (range 511.3 ng/g to 233.8 ng/g) based on allochthonous contaminant loads. Among the 16 compounds, Phen had the highest concentration (40.18 ng/g), followed by Pye (31.86 ng/g), Flur (29.36 ng/g), and NA (19.33 ng/g). Most of the sampling sites contained abundant 3-ring and 4-5-ring PAHs. Based on diagnostic ratios and PCA analysis petroleum combustion, biomass, and coal-burning have been identified as the major sources. The PAHs had high mutagenic equivalent factor and toxic equivalent factor values posing great ecological threats and health risks.

A phase II study of palliative radiotherapy combined with zoledronic acid hydrate for metastatic bone tumour from renal cell carcinoma.

PURPOSE: Palliative radiotherapy is the standard of care for bone metastases. However, skeletal-related events, defined as a pathologic fracture, paraplegia, surgery or radiotherapy for local recurrence, or severe pain in previously irradiated bone with radio-resistant histology type still present high incidence. The primary objective of this study was to determine whether zoledronic acid hydrate and palliative radiotherapy could prevent local skeletal-related events. METHODS: Eligible patients with bone metastases from renal cell carcinoma were treated with zoledronic acid hydrate every 3 or 4 weeks and concurrent palliative radiotherapy of 30 Gy in 3 Gy fractions. The criteria for radiotherapy were established by the treating physician, but patients with complicated bone metastases (impending pathological fracture or spinal cord compression) which needed immediate surgery were excluded. The primary endpoint was the local skeletal-related event-free survival rate at 1 year. RESULTS: Twenty-seven patients were included in the study. The median age was 65 (range, 50-84) years. Radiotherapy dose was 30 Gy for all patients except 1 whose radiotherapy was terminated due to brain metastasis progression at 18 Gy. Zoledronic acid hydrate was administered in a median of 12 (range, 0-34) times. The median follow-up period was 12 months and 19 months in patients who were still alive. Of 27 patients in the efficacy analysis, the 1-year local skeletal-related event-free rate was 77.6% (80% confidence interval, 66.2-89.0). Common grade 3 toxicities were hypocalcemia (1 [4%]), sGPT level increase (1 [4%]) and sGOT level increase (1 [4%]). There was no grade 4 or 5 toxicity. CONCLUSION: Zoledronic acid hydrate administration and palliative radiotherapy were a well-tolerated and promising treatment reducing skeletal-related events for bone metastases from renal cell carcinoma.

The SREBP (Sterol Regulatory Element-Binding Protein) pathway: a regulatory bridge between carotenogenesis and sterol biosynthesis in the carotenogenic yeast Xanthophyllomyces dendrorhous.

Xanthophyllomyces dendrorhous is a basidiomycete yeast that naturally produces the red-orange carotenoid astaxanthin, which has remarkable antioxidant properties. The biosynthesis of carotenoids and sterols share some common elements that have been studied in X. dendrorhous. For example, their synthesis requires metabolites derived from the mevalonate pathway and in both specific pathways, cytochrome P450 enzymes are involved that share a single cytochrome P450 reductase, CrtR, which is essential for astaxanthin biosynthesis, but is replaceable for ergosterol biosynthesis. Research on the regulation of carotenoid biosynthesis is still limited in X. dendrorhous; however, it is known that the Sterol Regulatory Element-Binding Protein (SREBP) pathway, which is a conserved regulatory pathway involved in the control of lipid metabolism, also regulates carotenoid production in X. dendrorhous. This review addresses the similarities and differences that have been observed between mammal and fungal SREBP pathways and what it is known about this pathway regarding the regulation of the production of carotenoids and sterols in X. dendrorhous.

Clinicopathologic characteristics, metastasis-free survival, and skeletal-related events in 628 patients with skeletal metastases in a tertiary orthopedic and trauma center.

BACKGROUND: Skeletal-related events (SREs) due to bone metastases (BM) significantly impact the morbidity and mortality of cancer patients. The present study sought to investigate clinicopathological characteristics, metastasis-free survival (MFS), and SREs in patients referred to a tertiary orthopedic and trauma center. METHODS: Data were retrieved from electronic health records (n=628). Survival curves were estimated utilizing the Kaplan-Meier method. The Cox regression model was used to determine factors influencing MFS based on estimated hazard ratios (HRs). RESULTS: Breast (55.8%) and lung (18.2%), and lung (32.9%) and prostate (16.8%) cancer were the most common cancer types in our cohort in women and men, respectively. Fifteen percent of patients presented with BM as the first manifestation of tumor disease, 23% had metastasis diagnosis on the same day of primary tumor diagnosis or within 3 months, and 62% developed BM at least 3 months after primary tumor diagnosis. Osteolytic BM were predominant (72.3%) and most commonly affecting the spine (23%). Overall median MFS was 45 months (32 (men) vs. 53 (women) months). MFS was shortest in the lung (median 15 months, 95% CI 8.05-19) and longest in breast cancer (median 82 months, 95% CI 65.29-94). Age (≥ 60 vs. < 60 years) and primary cancer grading of ≥2 vs. 1 revealed prognostic relevance. CONCLUSION: Women with breast or lung cancer, men with lung or prostate cancer, age ≥60 years, male sex, and primary cancer grading ≥2 are associated with increased risk for MBD. Intensified follow-up programs may reduce the risk of SREs and associated morbidity and mortality.

Metallopeptidase Stp1 activates the transcription factor Sre1 in the carotenogenic yeast Xanthophyllomyces dendrorhous.

Xanthophyllomyces dendrorhous is a basidiomycete yeast known as a natural producer of astaxanthin, a carotenoid of commercial interest because of its antioxidant properties. Recent studies indicated that X. dendrorhous has a functional SREBP pathway involved in the regulation of isoprenoid compound biosynthesis, which includes ergosterol and carotenoids. SREBP is a major regulator of sterol metabolism and homeostasis in mammals; characterization in fungi also provides information about its role in the hypoxia adaptation response and virulence. SREBP protease processing is required to activate SREBP pathway functions in fungi. Here, we identified and described the STP1 gene, which encodes a metallopeptidase of the M50 family involved in the proteolytic activation of the transcription factor Sre1 of the SREBP pathway, in X. dendrorhous We assessed STP1 function in Δstp1 strains derived from the wild-type and a mutant of ergosterol biosynthesis that overproduces carotenoids and sterols. Bioinformatic analysis of the deduced protein predicted the presence of characteristic features identified in homologs from mammals and fungi. The Δstp1 mutation decreased yeast growth in the presence of azole drugs and reduced transcript levels of Sre1-dependent genes. This mutation also negatively affected the carotenoid- and sterol-overproducing phenotype. Western blot analysis demonstrated that Sre1 was activated in the yeast ergosterol biosynthesis mutant and that the Δstp1 mutation introduced in this strain prevented Sre1 proteolytic activation. Overall, our results demonstrate that STP1 encodes a metallopeptidase involved in proteolytic activation of Sre1 in X. dendrorhous, contributing to our understanding of fungal SREBP pathways.

Early palliative radiation versus observation for high-risk asymptomatic or minimally symptomatic bone metastases: study protocol for a randomized controlled trial.

BACKGROUND: In patients with metastatic cancer, the bone is the third-most common site of involvement. Radiation to painful bone metastases results in high rates of pain control and is an integral part of bone metastases management. Up to one-third of inpatient consults are requested for painful bone metastases, and up to 60% of these patients had evidence of these lesions visible on prior imaging. Meanwhile recent advances have reduced potential side effects of radiation. Therefore, there is an opportunity to further improve outcomes for patients using prophylactic palliative radiation to manage asymptomatic bone metastases. METHODS/STUDY DESIGN: In this trial, 74 patients with metastatic solid tumors and high-risk asymptomatic or minimally symptomatic bone metastases will be enrolled and randomized to early palliative radiation or standard of care. This will be the first trial to assess the efficacy of prophylactic palliative radiation in preventing skeletal related events (SREs), the primary endpoint. This endpoint was selected to encompass patient-centered outcomes that impact quality of life including pathologic fracture, spinal cord compression, and intervention with surgery or radiation. Secondary endpoints include hospitalizations, Bone Pain Index, pain-free survival, pain-related quality of life, and side effects of radiation therapy. DISCUSSION: In this study, we propose a novel definition of high-risk bone metastases most likely to benefit from preventive radiation and use validated questionnaires to assess pain and impact on quality of life and health resource utilization. Observations from early patient enrollment have demonstrated robustness of the primary endpoint and need for minor modifications to Bone Pain Index and data collection for opioid use and hospitalizations. With increasing indications for radiation in the oligometastatic setting, this trial aims to improve patient-centered outcomes in the polymetastatic setting. TRIAL REGISTRATION: ISRCTN Number/Clinical trials.gov, ID: NCT03523351 . Registered on 14 May 2018.

Genome-wide association studies of the self-rating of effects of ethanol (SRE).

The level of response (LR) to alcohol as measured with the Self-Report of the Effects of Alcohol Retrospective Questionnaire (SRE) evaluates the number of standard drinks usually required for up to four effects. The need for a higher number of drinks for effects is genetically influenced and predicts higher risks for heavy drinking and alcohol problems. We conducted genome-wide association study (GWAS) in the African-American (COGA-AA, N = 1527 from 309 families) and European-American (COGA-EA, N = 4723 from 956 families) subsamples of the Collaborative Studies on the Genetics of Alcoholism (COGA) for two SRE scores: SRE-T (average of first five times of drinking, the period of heaviest drinking, and the most recent 3 months of consumption) and SRE-5 (the first five times of drinking). We then meta-analyzed the two COGA subsamples (COGA-AA + EA). Both SRE-T and SRE-5 were modestly heritable (h2 : 21%-31%) and genetically correlated with alcohol dependence (AD) and DSM-IV AD criterion count (rg : 0.35-0.76). Genome-wide significant associations were observed (SRE-T: chromosomes 6, rs140154945, COGA-EA P = 3.30E-08 and 11, rs10647170, COGA-AA+EA P = 3.53E-09; SRE-5: chromosome13, rs4770359, COGA-AA P = 2.92E-08). Chromosome 11 was replicated in an EA dataset from the National Institute on Alcohol Abuse and Alcoholism intramural program. In silico functional analyses and RNA expression analyses suggest that the chromosome 6 locus is an eQTL for KIF25. Polygenic risk scores derived using the COGA SRE-T and SRE-5 GWAS predicted 0.47% to 2.48% of variances in AD and DSM-IV AD criterion count in independent datasets. This study highlights the genetic contribution of alcohol response phenotypes to the etiology of alcohol use disorders.

Coordinate Regulation of Yeast Sterol Regulatory Element-binding Protein (SREBP) and Mga2 Transcription Factors.

The Mga2 and Sre1 transcription factors regulate oxygen-responsive lipid homeostasis in the fission yeast Schizosaccharomyces pombe in a manner analogous to the mammalian sterol regulatory element-binding protein (SREBP)-1 and SREBP-2 transcription factors. Mga2 and SREBP-1 regulate triacylglycerol and glycerophospholipid synthesis, whereas Sre1 and SREBP-2 regulate sterol synthesis. In mammals, a shared activation mechanism allows for coordinate regulation of SREBP-1 and SREBP-2. In contrast, distinct pathways activate fission yeast Mga2 and Sre1. Therefore, it is unclear whether and how these two related pathways are coordinated to maintain lipid balance in fission yeast. Previously, we showed that Sre1 cleavage is defective in the absence of mga2 Here, we report that this defect is due to deficient unsaturated fatty acid synthesis, resulting in aberrant membrane transport. This defect is recapitulated by treatment with the fatty acid synthase inhibitor cerulenin and is rescued by addition of exogenous unsaturated fatty acids. Furthermore, sterol synthesis inhibition blocks Mga2 pathway activation. Together, these data demonstrate that Sre1 and Mga2 are each regulated by the lipid product of the other transcription factor pathway, providing a source of coordination for these two branches of lipid synthesis.

FHOD1 formin is upregulated in melanomas and modifies proliferation and tumor growth.

The functional properties of actin-regulating formin proteins are diverse and in many cases cell-type specific. FHOD1, a formin expressed predominantly in cells of mesenchymal lineage, bundles actin filaments and participates in maintenance of cell shape, migration and cellular protrusions. FHOD1 participates in cancer-associated epithelial to mesenchymal transition (EMT) in oral squamous cell carcinoma and breast cancer. The role of FHOD1 in melanomas has not been characterized. Here, we show that FHOD1 expression is typically strong in cutaneous melanomas and cultured melanoma cells while the expression is low or absent in benign nevi. By using shRNA to knockdown FHOD1 in melanoma cells, we discovered that FHOD1 depleted cells are larger, rounder and have smaller focal adhesions and inferior migratory capacity as compared to control cells. Importantly, we found FHOD1 depleted cells to have reduced colony-forming capacity and attenuated tumor growth in vivo, a finding best explained by the reduced proliferation rate caused by cell cycle arrest. Unexpectedly, FHOD1 depletion did not prevent invasive growth at the tumor margins. These results suggest that FHOD1 participates in key cellular processes that are dysregulated in malignancy, but may not be essential for melanoma cell invasion.

Metals Differentially Activate Ovarian Cancer G Protein-Coupled Receptor 1 in Various Species.

Human, mouse, and zebrafish ovarian cancer G protein-coupled receptors (OGR1s) are activated by both metals and extracellular protons. In the present study, we examined whether pig, rat, chicken, and Xenopus OGR1 homologs could sense and be activated by protons and metals. We found that all homologs stimulated serum response element (SRE)-driven promoter activities when they are stimulated by protons. On the other hand, metals differentially activated the homologs. The results using chimeric receptors of human and zebrafish OGR1s indicate that the specificity of the metal-induced activation lies in the extracellular region. These results suggest that protons are an evolutionally conserved agonist of OGR1. However, the types of metals that activated the receptor differed among the homologs.

Sterol regulatory element-binding proteins are transcriptional regulators of the thyroglobulin gene in thyroid cells.

The genes encoding sodium/iodide symporter (NIS) and thyroid peroxidase (TPO), both of which are essential for thyroid hormone (TH) synthesis, were shown to be regulated by sterol regulatory element-binding proteins (SREBP)-1c and -2. In the present study we tested the hypothesis that transcription of a further gene essential for TH synthesis, the thyroglobulin (TG) gene, is under the control of SREBP. To test this hypothesis, we studied the influence of inhibition of SREBP maturation and SREBP knockdown on TG expression in FRTL-5 thyrocytes and explored transcriptional regulation of the TG promoter by reporter gene experiments in FRTL-5 and HepG2 cells, gel shift assays and chromatin immunoprecipitation. Inhibition of SREBP maturation by 25-hydroxycholesterol and siRNA-mediated knockdown of either SREBP-1c or SREBP-2 decreased mRNA and protein levels of TG in FRTL-5 thyrocytes. Reporter gene assays with wild-type and mutated TG promoter reporter truncation constructs revealed that the rat TG promoter is transcriptionally activated by nSREBP-1c and nSREBP-2. DNA-binding assays and chromatin immunoprecipitation assays showed that both nSREBP-1c and nSREBP-2 bind to a SREBP binding motif with characteristics of an E-box SRE at position -63 in the rat TG promoter. In connection with recent findings that NIS and TPO are regulated by SREBP in thyrocytes the present findings support the view that SREBP are regulators of essential steps of TH synthesis in the thyroid gland such as iodide uptake, iodide oxidation and iodination of tyrosyl residues of TG. This moreover suggests that SREBP may be molecular targets for pharmacological modulation of TH synthesis.

Factors predicting skeletal-related events in patients with bone metastatic castration-resistant prostate cancer.

BACKGROUND: Skeletal-related events (SREs) are common complications of bone metastatic castration-resistant prostate cancer (mCRPC). To the authors' knowledge, there are limited data regarding which factors predict SREs. The authors identified risk factors for SREs in men with bone mCRPC using characteristics commonly available in the medical record. METHODS: Data from 454 patients with nonmetastatic CRPC were identified from 2 Veteran Affairs Medical Centers from 2000 through 2013. Among these men, 233 (51%) developed bone metastases during follow-up and represented the study cohort. First occurrence of an SRE was abstracted from the medical records. A stepwise multivariable Cox model was used to select the strongest predictors of time to SRE. RESULTS: The median age of the patients at the time of diagnosis of bone mCRPC was 75 years (interquartile range, 68-81 years), and there were 153 nonblack patients (66%). During follow-up (median, 7.8 months [interquartile range, 2.9-18.3 months]), 88 patients (38%) had an SRE. On univariable analysis, more recent year of metastasis (hazard ratio [HR], 0.91), prostate-specific antigen doubling time of ≥9 months versus <9 months (HR, 0.50), and bone pain (HR, 3.34) were all found to be associated with SRE risk. On multivariable analysis, year of metastasis (HR, 0.93), biopsy Gleason score of 7 versus ≤6 (HR, 1.74), radiotherapy as the primary localized treatment versus none (HR, 2.33), and bone pain (HR, 3.64) were associated with SRE risk. The area under the curve for a multivariable model based upon these risk factors was 0.744. CONCLUSIONS: The authors identified several significant predictors of SREs among men with mCRPC. In particular, men with bone pain are at high risk of an SRE. If confirmed, future trials should focus on prolonging life and reducing SRE risk in patients with mCRPC with bone pain. Cancer 2017;123:1528-1535. © 2017 American Cancer Society.

New insights into cellular cholesterol acquisition: promoter analysis of human HMGCR and SQLE, two key control enzymes in cholesterol synthesis.

BACKGROUND: The two control points of cholesterol synthesis, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) and squalene monooxygenase (SQLE) are known targets of the transcription factor sterol-regulatory element binding protein-2 (SREBP-2). Yet the location of the sterol-regulatory elements (SREs) and cofactor binding sites, nuclear factor-Y (NF-Y) and specificity protein 1 (Sp1), have not been satisfactorily mapped in the human SQLE promoter, or at all in the human HMGCR promoter. METHODS: We used luciferase reporter assays to screen the sterol-responsiveness of a library of predicted SRE, Sp1 and NF-Y site mutants and hence identify bone fide binding sites. We confirmed SREs via an electrophoretic mobility shift assay (EMSA) and ChIP-PCR. RESULTS: We identified two SREs in close proximity in both the human HMGCR and SQLE promoters, as well as one NF-Y site in HMGCR and two in SQLE. In addition, we found that HMGCR expression is highly activated only when SREBP-2 levels are very high, in contrast to the low density lipoprotein receptor (LDLR), a result reflected in mouse models used in other studies. CONCLUSIONS: Both HMGCR and SQLE promoters have two SREs that may act as a homing region to attract a single SREBP-2 homodimer, with HMGCR being activated only when there is absolute need for cholesterol synthesis. This ensures preferential uptake of exogenous cholesterol via LDLR, thereby conserving energy. GENERAL SIGNIFICANCE: We provide the first comprehensive investigation of SREs and NF-Ys in the human HMGCR and SQLE promoters, increasing our fundamental understanding of the transcriptional regulation of cholesterol synthesis.

Bisphosphonate guidelines for treatment and prevention of myeloma bone disease.

Multiple myeloma (MM) is a haematological malignancy characterised by the clonal proliferation of plasma cells in the bone marrow. More than 80% of patients with MM display evidence of myeloma bone disease (MBD), characterised by the formation of osteolytic lesions throughout the axial and appendicular skeleton. MBD significantly increases the risk of skeletal-related events such as pathologic fracture, spinal cord compression and hypercalcaemia. MBD is the result of MM plasma cells-mediated activation of osteoclast activity and suppression of osteoblast activity. Bisphosphonates (BP), pyrophosphate analogues with high bone affinity, are the only pharmacological agents currently recommended for the treatment and prevention of MBD and remain the standard of care. Pamidronate and zoledronic acid are the most commonly used BP to treat MBD. Although generally safe, frequent high doses of BP are associated with adverse events such as renal toxicity and osteonecrosis of the jaw. As such, optimal duration and dosing of BP therapy is required in order to minimise BP-associated adverse events. The following guidelines provide currently available evidence for the adoption of a tailored approach when using BP for the management of MBD.

Risk factors for skeletal-related events (SREs) and factors affecting SRE-free survival for nonsmall cell lung cancer patients with bone metastases.

Skeletal-related events (SREs) for nonsmall cell lung cancer (NSCLC) patients with bone metastasis lead to serious morbidity. The aim of this study was to determine risk factors for SREs in NSCLC patients with bone metastasis and the factors influencing SRE-free survival and overall survival (OS). From 2000 to 2012, we evaluated retrospectively 835 NSCLC patients. Three hundred and thirty-five of them with bone metastasis were included in the study. SREs and the other prognostic factors were evaluated by univariate and multivariate analysis for SRE-free survival and OS. SREs were detected in 244 patients (72.8 %). The most common SREs were the need for radiotherapy (43.2 %) and malignant hypercalcemia (17.6 %). The median time to first SRE was 3.5 months at the median follow-up of 17 months. A multivariate analysis showed that the presence of bone metastasis at diagnosis (p < 0.001), the number of bone metastasis (p = 0.001), baseline hypercalcemia (p = 0.004), and the presence of palliative radiotherapy (p = 0.04) were independent prognostic factors for SRE-free survival. A logistic regression analysis identified that the presence of bone metastasis at diagnosis [odds ratio (OR), 12.6], number of bone metastasis (OR, 3.05), and baseline hypercalcemia (OR, 0.33) were found to be predictive factors in the developing of SRE. The median OS time for patients with SRE was worse than that for patients without SRE (7 vs 12 months, respectively). For OS, male gender, ECOG performance status (PS), high lactate dehydrogenase (LDH) level, hypoalbuminemia, the presence of bone metastasis at diagnosis, the number of bone metastasis, the presence of SREs, the presence of bisphosphonate therapy, and palliative radiotherapy were independent prognostic indicators for OS by the multivariate analysis. Our results indicated that the frequency of SREs was high and the presence of bone metastasis at the time of diagnosis, baseline hypercalcemia, and multiple bone metastases were significant factors predicting the occurrence of SREs. If bone metastases diagnose earlier, treatments for the prevention of SREs may be initiated earlier; thus, the deterioration of quality of life may be preserved.