Disease-related complications in patients with metastatic hormone-sensitive prostate cancer.

INTRODUCTION: Prostate-specific antigen-based screening for prostate cancer reportedly does not improve cancer-specific survival. However, there remain concerns about the increasing incidence of advanced disease at initial presentation. Here, we investigated the incidences and types of complications that occur during the course of disease in patients with metastatic hormone-sensitive prostate cancer (mHSPC). METHODS: This study included 100 consecutive patients who were diagnosed with mHSPC at five hospitals from January 2016 to August 2017. Analyses were conducted using patient data extracted from a prospectively collected database, along with information about complications and readmission obtained from electronic medical records. RESULTS: The median patient age was 74 years and the median serum prostate-specific antigen level at diagnosis was 202.5 ng/mL. Ninety-nine patients received androgen deprivation therapy; 17 of these patients also received chemotherapy. During a mean follow-up period of 32.9 months, 41 patients reported bone pain; of these patients, 21 developed pathologic fractures and eight had cord compression. Twenty-eight patients developed retention of urine; of these patients, 10 (36%) required surgery and 11 (39%) required long-term urethral catheter use. Among 15 patients who developed ureteral obstruction, four (27%) required ureteral stenting and four (27%) required long-term nephrostomy drainage. Other complications included anaemia (41%) and deep vein thrombosis (4%). Fifty-nine (59%) patients had ≥1 unplanned hospital admission during the course of disease; 16% of such patients had >5 episodes of readmission. CONCLUSION: Among patients with mHSPC, 70% experienced disease-related complications and unplanned hospital admissions, which substantially burdened both patients and the healthcare system.

Outcomes of transperineal and transrectal ultrasound-guided prostate biopsy.

OBJECTIVE: To compare the clinical outcomes and pathological findings of transperineal ultrasound-guided prostate biopsy (TPUSPB) and transrectal ultrasound-guided prostate biopsy (TRUSPB) in a secondary referral hospital. METHODS: This was a retrospective study of 100 TPUSPBs and 100 TRUSPBs performed in our centre. Pre-biopsy patient parameters (eg, patient age, clinical staging, serum prostate-specific antigen [PSA] level, prostate size, and PSA density), as well as pathological results and 30-day complication and readmission rates, were retrieved from the patients' medical records and compared between the two groups. RESULTS: One hundred TPUSPBs performed from January 2018 to May 2018 and 100 TRUSPBs performed from January 2016 to April 2016 were included for analysis. Mean age did not significantly differ between the groups. The TPUSPB group had a higher mean PSA level, smaller prostate size, and higher PSA density, compared with the TRUSPB group. The overall prostate cancer detection rate was similar between the TPUSPB and TRUSPB groups (35% vs 25%, P=0.123). There were no significant differences between the groups in prostate cancer detection rates after stratification according to PSA density and clinical staging. With respect to complications, no patients developed fever in the TPUSPB group, while 4% of patients in the TRUSPB group had fever and required at least 1-week admission for intravenous antibiotic administration. CONCLUSION: For prostate biopsy, TPUSPB is safer, with no infection complications, and has similar prostate cancer detection rate compared with TRUSPB.

Intracerebroventricular delivery of self-complementary adeno-associated virus serotype 9 to the adult rat brain.

Gene therapy for the central nervous system is poised to become a powerful treatment for numerous neurological disorders. Adeno-associated viral vectors based on serotype 9 (AAV9) have proven themselves to be strong candidates for delivering gene-based therapies throughout the brain and spinal cord when administered intravenously, intrathecally, intracisternally, and intracerebroventricularly (i.c.v.). Previous studies of i.c.v.-delivered self-complimentary AAV9 have been performed in neonatal mice with delivery of a single dose. However, before clinical trials can be considered, more information is required about the dose-response relationship for transduction efficiency in adult animals. In the current study, three doses of self-complementary AAV9 were administered to adult rats. High levels of transduction were observed in the hippocampus, cerebellum and cerebral cortex, and transduction increased with increasing dosage. Both neurons and astrocytes were transduced. There was no evidence of astrocytosis at the doses tested. Preliminary results from pigs receiving i.c.v. self-complementary AAV9 are also presented. The results of this study will serve to inform dosing studies in large animal models before clinical testing.

Identification of a natural product-like STAT3 dimerization inhibitor by structure-based virtual screening.

STAT3 regulates a variety of genes involved with cell proliferation, differentiation, apoptosis, angiogenesis, metastasis, inflammation, and immunity. The purpose of this study was to apply molecular docking techniques to identify STAT3 inhibitors from a database of over 90000 natural product and natural product-like compounds. The virtual screening campaign furnished 14 hit compounds, from which compound 1 emerged as a top candidate. Compound 1 inhibited STAT3 DNA-binding activity in vitro and attenuated STAT3-directed transcription in cellulo with selectivity over STAT1 and with comparable potency to the well-known STAT3 inhibitor S3I-201. Furthermore, compound 1 inhibited STAT3 dimerization and decreased STAT3 phosphorylation in cells without affecting STAT1 dimerization and phosphorylation. Compound 1 also exhibited selective anti-proliferative activity against cancer cells over normal cells in vitro. Molecular docking analysis suggested that compound 1 might putatively function as an inhibitor of STAT3 dimerization by binding to the SH2 domain. This study also validates the use of in silico techniques to identify inhibitors of protein-protein interactions, which are typically considered difficult to target with small molecules.

A cyclometallated iridium(III) complex as a c-myc G-quadruplex stabilizer and down-regulator of c-myc oncogene expression.

A new cyclometallated iridium(III) complex with the 2,2'-biquinoline N-donor ligand has been synthesized and characterized. The interaction and affinity of the complex towards c-myc G-quadruplex and duplex DNA have been investigated using UV/Vis spectroscopy and gel mobility shift assay. These studies revealed that complex 1 binds to c-myc G-quadruplexes (Pu22 and Pu27) with high affinity but does not interact with duplex DNA either by intercalation or groove binding. The ability of 1 to stabilize c-myc G-quadruplex DNA in vitro has also been examined through a PCR stop assay and a cell-based luciferase reporter assay. Complex 1 displays promising cytotoxic activity against the HeLa cell line with sub-micromolar potency.

PRAP1 is a novel executor of p53-dependent mechanisms in cell survival after DNA damage.

p53 has a crucial role in governing cellular mechanisms in response to a broad range of genotoxic stresses. During DNA damage, p53 can either promote cell survival by activating senescence or cell-cycle arrest and DNA repair to maintain genomic integrity for cell survival or direct cells to undergo apoptosis to eliminate extensively damaged cells. The ability of p53 to execute these two opposing cell fates depends on distinct signaling pathways downstream of p53. In this study, we showed that under DNA damage conditions induced by chemotherapeutic drugs, gamma irradiation and hydrogen peroxide, p53 upregulates a novel protein, proline-rich acidic protein 1 (PRAP1). We identified functional p53-response elements within intron 1 of PRAP1 gene and showed that these regions interact directly with p53 using ChIP assays, indicating that PRAP1 is a novel p53 target gene. The induction of PRAP1 expression by p53 may promote resistance of cancer cells to chemotherapeutic drugs such as 5-fluorouracil (5-FU), as knockdown of PRAP1 increases apoptosis in cancer cells after 5-FU treatment. PRAP1 appears to protect cells from apoptosis by inducing cell-cycle arrest, suggesting that the induction of PRAP1 expression by p53 in response to DNA-damaging agents contributes to cancer cell survival. Our findings provide a greater insight into the mechanisms underlying the pro-survival role of p53 in response to cytotoxic treatments.

Comparative in vitro cytotoxicity study on uncoated magnetic nanoparticles: effects on cell viability, cell morphology, and cellular uptake.

Magnetic iron oxide nanoparticles (MIONPs) must be biocompatible, and a thorough knowledge on their potential cytotoxicity is crucial for their biomedical applications. However, the detailed study about the effects of iron oxide nanoparticles on cell viability, cell morphology, and cellular uptake of different mammalian cells is still insufficient. In this paper, comparative cytotoxicity study of uncoated magnetite nanoparticles at different concentrations was performed on human cervical cancer cell line (HeLa) and immortalized normal human retinal pigment epithelial cell line (RPE). The size, structure, and magnetic behavior of the MIONPs were characterized using transmission electron microscopy (TEM), X-ray diffractometry (XRD), and vibrating sample magnetometry (VSM) respectively. After 24-hour incubation with the MIONPs, the cell viability was determined by live/dead assay, the cell morphology at high magnification was observed under scanning electron microscopy (SEM), and the cellular uptake of MIONPs was measured under TEM and verified by energy-dispersive X-ray spectroscopy (EDX) analysis. Our results indicate that the uncoated MIONPs at a high concentration (0.40 mg/ml) were toxic to both HeLa and RPE cells. However, the cytotoxicity of uncoated MIONPs at low concentrations was cell-type specific, and RPE cells were more susceptible to these MIONPs than HeLa cells. The effects of the MIONPs on cell morphology and the nanoparticles uptake also showed different features between these two cell lines. Hence cell type should be taken into consideration in the in vitro cytotoxicity study of uncoated MIONPs. Additionally, it should be noticed that the cell morphological changes and the uptake of nanoparticles can take place even though no toxic effect of these MIONPs at low concentrations was reflected in the traditional cell viability assay.

Reversal of multidrug resistance in cancer cells by Rhizoma Alismatis extract.

Prolonged chemotherapy may lead to the selective proliferation of multidrug resistant (MDR) cancer cells. In MDR HepG2-DR and K562-DR cells that over-expressed P-glycoprotein (Pgp), the extract of the rhizomes of Alisma orientalis (Sam) Juzep. showed a synergistic growth inhibitory effect with cancer drugs that are Pgp substrates including actinomycin D, puromycin, paclitaxel, vinblastine and doxorubicin. At the same toxicity levels the herbal extract was more effective than verapamil, a standard Pgp inhibitor, in enhancing cellular doxorubicin accumulation and preventing the efflux of rhodamin-123 from the MDR cells. The extract restored the effect of vinblastine on the induction of G(2)/M arrest in MDR cells. Our data suggest that A. orientalis may contain components that are effective inhibitors of Pgp.

A randomized controlled trial of surgery vs steroid injection for carpal tunnel syndrome.

BACKGROUND: Decompressive surgery and steroid injection are widely used forms of treatment for carpal tunnel syndrome (CTS) but there is no consensus on their effectiveness in comparison to each other. The authors evaluated the efficacy of surgery vs steroid injection in relieving symptoms in patients with CTS. METHODS: The authors conducted a randomized, single blind, controlled trial. Fifty patients with electrophysiologically confirmed idiopathic CTS were randomized and assigned to open carpal tunnel release (25 patients) or to a single injection of steroid (25 patients). Patients were followed up at 6 and 20 weeks. The primary outcome was symptom relief in terms of the Global Symptom Score (GSS), which rates symptoms on a scale of 0 (no symptoms) to 50 (most severe). Nerve conduction studies and grip strength measurements were used as secondary outcome assessments. RESULTS: At 20 weeks after randomization, patients who underwent surgery had greater symptomatic improvement than those who were injected. The mean improvement in GSS after 20 weeks was 24.2 (SD 11.0) in the surgery group vs 8.7 (SD 13.0) in the injection group (p < 0.001); surgical decompression also resulted in greater improvement in median nerve distal motor latencies and sensory nerve conduction velocity. Mean grip strength in the surgical group was reduced by 1.7 kg (SD 5.1) compared with a gain of 2.4 kg (SD 5.5) in the injection group. CONCLUSION: Compared with steroid injection, open carpal tunnel release resulted in better symptomatic and neurophysiologic outcome but not grip strength in patients with idiopathic carpal tunnel syndrome over a 20-week period.

Inhibition of histone deacetylase 2 increases apoptosis and p21Cip1/WAF1 expression, independent of histone deacetylase 1.

Histone deacetylases (HDACs) 1 and 2 share a high degree of homology and coexist within the same protein complexes. Despite their close association, each possesses unique functions. We show that the upregulation of HDAC2 in colorectal cancer occurred early at the polyp stage, was more robust and occurred more frequently than HDAC1. Similarly, while the expression of HDACs1 and 2 were increased in cervical dysplasia and invasive carcinoma, HDAC2 expression showed a clear demarcation of high-intensity staining at the transition region of dysplasia compared to HDAC1. Upon HDAC2 knockdown, cells displayed an increased number of cellular extensions reminiscent of cell differentiation. There was also an increase in apoptosis, associated with increased p21Cip1/WAF1 expression that was independent of p53. These results suggest that HDACs, especially HDAC2, are important enzymes involved in the early events of carcinogenesis, making them candidate markers for tumor progression and targets for cancer therapy.

Involvement of NF-kappaB and c-myc signaling pathways in the apoptosis of HL-60 cells induced by alkaloids of Tripterygium hypoglaucum (levl.) Hutch.

Tripterygium hypoglaucum (levl.) Hutch (Celastraceae) (THH) root is a Chinese medicinal herb commonly used for treating autoimmune diseases. In the present study, alkaloids of THH were prepared and their cytotoxicity against the HL-60 cell was investigated. THH-induced apoptosis was observed using flow cytometry, confocal fluorescence microscope, and DNA laddering and caspase assays. The molecular mechanism involved in the induction of HL-60 cell apoptosis by THH alkaloids was examined using cDNA microarrays containing 3000 human genes derived from a leukocyte cDNA library. Sixteen genes were identified to be differentially expressed in HL-60 cells upon THH treatment. Several genes related to the NF-kappaB signaling pathway and cell apoptosis (such as NFKBIB, PRG1 and B2M) were up-regulated. In addition, c-myc binding protein and apoptosis-related cysteine proteases caspase-3 and caspase-8 were also regulated. The changes in c-Myc RNA expression and c-myc protein level were further confirmed by RT-PCR and Western blot analysis. The results demonstrated that THH alkaloids induced apoptosis of HL-60 cells though c-myc and NF-kappaB signaling pathways.

LINAC radiosurgery in recurrent Cushing's disease after transsphenoidal surgery: a series of 5 cases.

INTRODUCTION: Oversecretion of ACTH from pituitary adenomas is associated with morbidity and reduced life expectancies. We hypothesized that radiosurgical treatment may provide effective tumour and hormonal control associated with minimal pituitary insufficiency. METHOD: Data of five patients who underwent LINAC radiosurgery for recurrent or residual Cushing's disease between 1999 and 2002 were prospectively collected. RESULT: Follow-up period ranged from 27 to 49 months (mean 38 months). All patients attained remission in 6-18 months (mean 8.4 months). One patient (20 %) developed biochemical recurrence 12 months after remission. One patient (20 %) developed hydrocortisone deficiency 24 months after radiosurgery. None of the patients had new visual field defect detected on follow-up. CONCLUSION: In this small case series, LINAC radiosurgery was shown to be an effective and safe treatment of persistent or recurrent Cushing's disease following transsphenoidal surgery. Long-term follow-up and larger patient series is recommended for further clarification.

PKCdelta-dependent deubiquitination and stabilization of Gadd45 in A431 cells overexposed to EGF.

Epidermal growth factor (EGF) receptor-overexpressing p53-deficient A431 cells response to toxic dose of EGF by G1 arrest and apoptosis was studied. We previously reported an increased expression of growth arrest and DNA-damage-inducible gene, Gadd45, in EGF-overexposed A431 cells. The mechanism for this induction was increased half-lives of mRNA and protein. In this study, using phorbol ester (a PKC activator) and specific inhibitors of PKC isoforms, we showed that protein kinase C-delta (PKCdelta) was involved in the increase of Gadd45 protein stability. We further demonstrated that Gadd45 is ubiquitinated and is regulated by proteolysis. While EGF induced ubiquitination of total cellular proteins, there was a decrease in Gadd45 ubiquitination, which could be inhibited by Rottlerin, a PKCdelta-specific inhibitor. These results suggest that an increase in Gadd45 stability may involve PKCdelta-dependent ubiquitin-proteasome pathway.

Spinal cord pilocytic astrocytoma with cranial meningeal metastases.

Dissemination of tumour cells along the cerebrospinal fluid (CSF) pathway has been reported mostly in medulloblastomas, germ cell tumours or high grade gliomas. Juvenile pilocytic astrocytoma (JPA) is usually a benign astrocytoma. However, drop metastases of indolent nature from intracranial tumours to the spinal cord are documented. All of the previously reported cases represent metastases of cerebellar or hypothalamic tumours spreading to the spinal cord. We document in this paper the first report of a spinal cord pilocytic astrocytoma spreading via the CSF to the cerebral meninges. A 9 year old girl had a JPA of C5 to C7 subtotally resected. Two and a half years later she presented with hydrocephalus with radiologically meningeal enhancement. The meninges were biopsied which showed metastatic JPA. The girl was relatively well 4 years after initial surgery with residual tumour. Spinal cord JPA can rarely metastasize to the cranial meninges. Similar to intracranial tumours which spread to the spinal cord, such metastatic lesions are indolent.

Epidermal growth factor induces Gadd45 (growth arrest and DNA damage inducible protein) expression in A431 cells.

Epidermal growth factor (EGF) receptor over-expressing, p53-deficient A431 cells response to toxic dose of EGF by G1 arrest and apoptosis. Applying cDNA expression array technology we demonstrated that EGF increased the levels of Gadd45 mRNA. Northern blot and Western blot analyses confirmed that both Gadd45 mRNA and protein were increased. Concurrently half-lives of Gadd45 mRNA and protein also increased. Nuclear run-on experiments did not show a large increase of Gadd45 mRNA transcription rate. Gadd45 mRNA and protein started to increase after 1 h of EGF treatment and remained high for up to 10 h. We have also confirmed previous studies which showed that in EGF-stimulated A431 cells p21(Cip1/Waf1) (cyclin-dependent kinase interacting protein 1) was up-regulated within the same time frame. Thus it appears that in addition to inducing G2 arrest by directly disrupting Cdc2/Cyclin B1 complex in genotoxic-stressed cells, Gadd45 may also participate in G1 arrest in growth factor overexposed cells.

Toxicity and DNA binding of dextran-doxorubicin conjugates in multidrug-resistant KB-V1 cells: optimization of dextran size.

We previously showed that conjugating doxorubicin to very large 70-500 kDa dextran decreased its removal rate from P-glycoprotein (P-gp) over-expressing, multidrug-resistant KB-V1 cells. Furthermore these conjugates could act synergistically with other cancer drugs. In the drug-sensitive 3-1 clone, but not in the V1 subclone which was 300-fold more resistant to free doxorubicin, conjugation led to a size-related decrease in toxicity. Here we identified the optimal size of dextran for avoiding P-gp-mediated efflux and yet preserving as much as possible doxorubicin toxicity. Chemically reduced, intracellularly stable 3.4-10 kDa conjugates were prepared. Confocal microscopy and fluorescence quenching experiments showed that these conjugates entered nuclei and interacted with DNA. In 3-1 cells, but not in V1 cells, cytotoxicity of conjugates decreased 14- to 45-fold linearly related to log size of the carrier (r=0.95). In V1 cells toxicity of the 10 kDa conjugate exceeded that of free doxorubicin. After conjugation the equilibrium binding constant of the DNA-drug complex (KA) decreased only by up to 3-fold. In 3-1 cells, but not in VI cells, DNA binding kinetics was an important factor and toxicity could be linearly correlated to 1/KA of conjugate (r=0.94). Drug accumulation decreased with an increase in dextran size but drug removal was decreased only in V1 cells. It appeared that drug uptake was also sensitive to dextran conjugation. In Vl cells drug removal was sensitive to the P-gp inhibitor verapamil or energy starvation. Ratios of V1/3-1 toxicity, drug accumulation and drug removal correlated linearly with log dextran size. When these ratios equaled 1, dextran sizes were estimated to be 32, 103 and 21 kDa, respectively.

Rapid detection of the mecA gene in methicillin resistant staphylococci using a colorimetric cycling probe technology.

A Cycling Probe Technology (CPT) assay was developed for the detection of the mecA gene from methicillin resistant staphylococcal cultures. The assay is based on a colorimetric enzyme-immuno-assay (EIA) and uses a mecA probe (DNA-RNA-DNA) labeled with fluorescein at the 5'-terminus and biotin at the 3'-terminus. The reaction occurs at a constant temperature that allows the target DNA to anneal to the probe. RNase H cuts the RNA portion, allowing the cut fragments to dissociate from the target, making it available for further cycling. CPT-EIA uses streptavidin-coated microplate wells to capture uncut probe followed by detection with horseradish-peroxidase conjugated anti-fluorescein antibody. The assay was compared to PCR and shown to accurately detect the presence or absence of the mecA gene in 159 staphylococcal clinical isolates. The CPT-EIA assay takes two hours starting from cultured cells compared with the 24-48 h required for detection of methicillin resistance by conventional susceptibility tests.

Functional evidence for divergent receptor activation mechanisms of luteotrophic and luteolytic events in the human corpus luteum.

Using a dispersed human luteal cell culture model, progesterone synthesis following treatment by incremental doses of human chorionic gonadotrophin (HCG) and the stable prostaglandin F2alpha (PGF2alpha) analogue cloprostenol, alone or in combination, was related to corpora lutea (CL) mRNA transcript abundance coding for the luteinizing hormone (LH)/HCG receptor (LH-R) and PGF2alpha-receptor (FP) by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) in 33 otherwise healthy women, scheduled for surgery due to benign conditions. CL were grouped according to age, based on the occurrence of a preovulatory LH surge where post-LH days 2-5 were designated as early luteal phase, days 6-10 as mid-luteal phase and days 11-14 as late luteal phase. When exposed to HCG, maximal progesterone output was raised 2.2-fold (P = 0.08, n = 5) compared with untreated controls in the early CL, while it increased 5.7- and 4.6-fold in the mid- and late groups respectively (P<0.05, n = 4 mid-luteal phase, n = 3 late luteal phase). This stimulation pattern was found to be concordant with the value of mRNA coding for LH-R in all groups (n = 6 early luteal phase, n = 5 mid-luteal phase, n = 6 late luteal phase). The integrated response to HCG and cloprostenol showed a dose-dependent 60% inhibition of progesterone production; but only in late luteal phase luteal cells (P<0.01, n = 3). FP mRNA values were lowest in early luteal phase, and increased with the age of the CL. Interestingly, lowest CL tissue concentrations of the natural FP agonist PGF2alpha were found during mid-luteal phase while it increased again 1.6-fold during late luteal phase (P<0.05, n = 8 versus mid-luteal phase, n = 6). Collectively, these data demonstrate that (i) the extrinsic functional control (or rescue of CL in the event of pregnancy) occurs when the sensitivity towards LH/HCG is maximal; and (ii) the demise of CL function is mediated via an acquisition of sensitivity towards the intrinsic luteolytic signal, PGF2alpha in the ageing CL.