含有磺胺的多金属氧酸盐的合成及抑制前列腺癌细胞PC-3M作用的研究

合成了(C₆H₉N₂O₂S)₅HP₂Mo₁₈O₆₂•15H₂O (SPOM-1)和(C₆H₉N₂O₂S)H₈P₂Mo₁₅V₃O₆₂•8H₂O (SPOM-2)两种新的含有磺胺的多金属氧酸盐, 通过元素分析、IR光谱对其结构进行了表征. 在人雄激素非依赖性前列腺癌细胞系PC-3M内, 对合成的多金属氧酸盐进行了抗肿瘤活性的研究. 研究发现, SPOM-1, SPOM-2在体外能明显抑制前列腺癌PC-3M细胞, 并呈一定的量效关系, EC₅₀分别为38, 11 g•mL^－1; 治疗指数(TI)分别为12.07, 26.82; SPOM-2的抗前列腺癌PC-3M细胞活性大于SPOM-1.     

微孔板化学发光酶免疫分析法分析人血清中游离前列腺特异性抗原

建立了一种定量分析人血清中游离前列腺特异性抗原(f-PSA)的高灵敏度微孔板化学发光酶免疫分析方法,以碱性磷酸酶(alkaline phosphatase,ALP)为标记酶,4-甲氧基-4-(3″-磷酰氧基苯)-螺旋-(1,2-二氧杂环丁烷-3,2′-金刚烷)(4-methoxy-4-(3″-phosphate-phenyl)-spiro-(1,2-dioxetane-3,2′-adamantane),AMPPD)-ALP高灵敏的化学发光反应为检测体系,通过检测发光强度对人血清中f-PSA进行定量。对几种物理化学参数如温育时间、免疫反应步骤以及检测时间等进行了优化,采用了一步双抗体夹心法,37℃恒温静置温育2h,洗涤后加入50μL AMPPD,30~80min内检测。该方法线性范围为0.15~20μg/L,相关系数大于0.998;检出限可达0.01μg/L,批内和批间相对标准偏差(C.V.)均小于7%;回收率在88%~108%之间。对人体血清中共存的6种常见肿瘤标志物CA50、CA125、CA15-3、CA242、CEA和AFP的偶联反应进行考察,特异性良好。为了验证该方法用于商业试剂盒的可行性,在4℃和37℃条件下分别进行了3d,5d,7d的稳定性考察,其线性相关系数仍均大于0.998,相对标准偏差小于6%。对98例实际血样进行测定,并与进口试剂盒(Monobind.USA)作临床比对,相关性良好。这些结果均表明,该分析体系稳定,可靠,可以用于商业化试剂盒的开发,在临床分析人血清中f-PSA的辅助诊断前列腺癌方面具有很高的应用价值。     

An Extensive Pharmacological Evaluation of New Anti-Cancer Triterpenoid (Nummularic Acid) from Ipomoea batatas through In Vitro,In Silico,and In Vivo Studies

Prostate cancer (PCa) is the most common cancer in men, accounting for approximately 10% of all new cases in the United States. Plant-derived bioactive compounds, such as pentacyclic triterpenoids (PTs), have the ability to inhibit PCa cell proliferation. We isolated and characterized nummularic acid (NA), a potent PT, as a major chemical constituent of Ipomoea batatas, a medicinal food plant used in ethnomedicine for centuries. In the current study, in vitro antiproliferative potential against PCa cells (DU145 and PC3) via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay; Western blot protein expression analysis; absorption, distribution, metabolism, excretion (ADME); pharmacokinetic prediction studies; and bisphenol A (BPA)-induced prostate inhibition in Sprague Dawley rats were conducted to gauge the anti-cancer ability of NA. Significant (p < 0.05 and p < 0.01) time- and dose-dependent reductions in proliferation of PCa cells, reduced migration, invasion, and increased apoptotic cell population were recorded after NA treatment (3–50 µM). After 72 h of treatment, NA displayed significant IC₅₀ of 21.18 ± 3.43 µM against DU145 and 24.21 ± 3.38 µM against PC3 cells in comparison to the controls cabazitaxel (9.56 ± 1.45 µM and 12.78 ± 2.67 µM) and doxorubicin (10.98 ± 2.71 µM and 15.97 ± 2.77 µM). Further deep mechanistic studies reveal that NA treatment considerably increased the cleavage of caspases and downstream PARP, upregulated BAX and P53, and downregulated BCL-2 and NF-κB, inducing apoptosis in PCa cells. Pharmacokinetic and ADME characterization indicate that NA has a favorable physicochemical nature, with high gastrointestinal absorption, low blood–brain barrier permeability, no hepatotoxicity, and cytochrome inhibition. BPA-induced perturbations of prostate glands in Sprague Dawley rats show a potential increase (0.478 ± 0.28 g) in prostate weight compared to the control (0.385 ± 0.13 g). Multi-dose treatment with NA (10 mg/kg) significantly reduced the prostate size (0.409 ± 0.21 g) in comparison to the control. NA-treated groups exhibited substantial restoration of hematological and histological parameters, reinstatement of serum hormones, and suppression of inflammatory markers. This multifaceted analysis suggests that NA, as a novel small molecule with a strong pharmacokinetic and pharmacological profile, has the potential to induce apoptosis and death in PCa cells.     

Nanoparticles of Costus speciosus Ameliorate Diabetes-Induced Structural Changes in Rat Prostate through Mediating the Pro-Inflammatory Cytokines IL 6, IL1β and TNF-α

Diabetes mellitus is a common global health problem. Among the complications that are frequently associated with DM are the alternation of sexual function and fertility, especially in young men. This study aimed to assess the efficacy of nanoparticles of Costus speciosus (C. speciosus) in preserving the prostatic structure of diabetic rats and to explore the mechanism behind this effect. A model of DM was induced in male albino rats by a single intraperitoneally injection of streptozotocin (STZ, 60 mg/kg body weight). Five groups (n = 10 each) of rats were included in this study: the control, C. speciosus gold nanoparticles-treated (150 mg/kg body weight through gastric intubation for 30 days), untreated diabetic, metformin-treated diabetic (500 mg/kg/day gastric intubation for 30 days) and the C. speciosus-treated diabetic group. The blood glucose, insulin and testosterone levels as well as oxidants/antioxidants status were assessed in the serum. Gene expression of proinflammatory cytokines TNF-α, IL1β and IL-6 were assessed in the prostate homogenate. At the end of the experiment, the rats were sacrificed and the prostate was dissected out and prepared for histopathological and immunohistochemistry study using Ki67 and Bcl-2. C. Speciosus nanoparticles significantly decreased (p = 0.03) the blood glucose level while significantly increasing insulin (p = 0.01) and testosterone (p = 0.04) levels compared to the untreated diabetic rats. Oxidants/antioxidants status was markedly improved after administration of C. speciosus. Prostatic expression of the mRNA of pro-inflammatory cytokines IL-6, IL1β and TNF-α was down-regulated in metformin- and C. speciosus-treated rats. The histological structure of the ventral prostate was preserved in metformin- and C. speciosus-treated diabetic rats with a significantly thicker epithelial cell layer and significant increase immunoexpression in Bcl-2 and Ki67. In conclusion, the protective effect induced by C. speciosus nanoparticles on the prostate of diabetic rats might be directly mediated through the down-regulation of inflammatory cytokines and the up-regulation of antioxidant activity and indirectly mediated through the anti-hyperglycemic effect through enhancing insulin secretion.     

Phytochemical Profile and Biological Activity of the Ethanolic Extract from the Aerial Part of Crocus alatavicus Regel & Semen Growing Wildly in Southern Kazakhstan

The composition of the ethanolic extract from the aerial parts of Crocus alatavicus Regel & Semen from southern Kazakhstan spontaneous flora was analyzed together with the determination of its antibacterial, antifungal, antiviral and anticancer activity. The phytochemical profile analysis by high-performance liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry (HPLC/ESI-QTOF-MS) revealed the presence of multiple kaempferol derivatives. High-performance reverse-phase liquid chromatography combined with a photodiode-array detection (RP-HPLC/PDA) found that kaempferol 3-O-dihexoside and kaempferol 3-O-acyltetrahexoside accounted for 70.5% of the kaempferol derivatives. The minimum inhibitory concentration (MIC) values of the extract for all the tested reference microorganisms were high, reaching 10 mg/mL for yeasts and 20 mg/mL for bacteria. In contrast, antiviral activity was observed at 2 mg/mL, resulting in the inhibition of the HSV-1-induced cytopathic effect and the reduction in virus infectious titer by 1.96 log, as well as the viral load by 0.85 log. Among the tested prostate cancer cell lines, significant cytotoxic activity of the extract was noted only on the LNCaP cell line, with an IC₅₀ value of 1.95 mg/mL. The LNCaP cell line treated with 2 mg/mL of the extract showed a noticeably reduced number of spindle-shaped cells with longer cellular projections, a significant increase in the peak corresponding to the population of apoptotic cells in the sub-G1 phase and a decreased intracellular glutathione (GSH) level, suggesting the prooxidative properties of the extract. The obtained data provide novel information about the flavonoids present in the aerial part of C. alatavicus and suggest its potential application as a source of the compounds active against HSV-1 and metastatic, androgen-sensitive prostate cancer.     

Pharmacokinetics,Prostate Distribution and Metabolic Characteristics of Four Representative Flavones after Oral Administration of the Aerial Part of Glycyrrhiza uralensis in Rats

(1) Background: The aerial part of G. uralensis had pharmacological effects against chronic non-bacterial prostatitis (CNP), and flavonoids are the main efficacy components. The purpose of this study was to obtain the pharmacokinetics, prostate distribution and metabolic characteristics of some flavonoids in rats. (2) Methods: The prototype flavones and the metabolites of four representative flavonoids, namely puerarin, luteolin, kaempferol and pinocembrin in plasma, prostate, urine and feces of rats were analyzed by UPLC-Q-Exactive Orbitrap-MS. In addition, the pharmacokinetic parameters in plasma and distribution of prostate of four components were analyzed by HPLC-MS/MS. (3) Results: In total, 22, 17, 22 and 11 prototype flavones were detected in the prostate, plasma, urine and feces, respectively. The metabolites of puerarin in the prostate are hydrolysis and glucose-conjugated products, the metabolites of kaempferol and luteolin in the prostate are methylation and glucuronidation, and the metabolites of pinocembrin in the prostate are naringenin, oxidation, sulfation, methylation and glucuronidation products. The t_1/2 of puerarin, luteolin, kaempferol and pinocembrin was 6.43 ± 0.20, 31.08 ± 1.17, 18.98 ± 1.46 and 13.18 ± 0.72 h, respectively. The concentrations of the four flavonoids in prostate were ranked as kaempferol > pinocembrin > luteolin > puerarin. (4) Conclusions: Methylation and glucuronidation metabolites were the main metabolites detected in the prostate. A sensitive and validated HPLC–MS/MS method for simultaneous determination of puerarin, luteolin, kaempferol and pinocembrin in rat plasma and prostate was described, and it was successfully applied to the pharmacokinetic and prostate distribution studies.     

In Vitro Study of Calcium Microsecond Electroporation of Prostate Adenocarcinoma Cells

Electroporation, applied as a non-thermal ablation method has proven to be effective for focal prostate treatment. In this study, we performed pre-clinical research, which aims at exploring the specific impact of this so-called calcium electroporation on prostate cancer. First, in an in-vitro study of DU 145 cell lines, microsecond electroporation (μsEP) parameters were optimized. We determined hence the voltage that provides both high permeability and viability of these prostate cancer cells. Subsequently, we compared the effect of μsEP on cells’ viability with and without calcium administration. For high-voltage pulses, the cell death’s mechanism was evaluated using flow-cytometry and confocal laser microscopy. For lower-voltage pulses, the influence of electroporation on prostate cancer cell mobility was studied using scratch assays. Additionally, we applied calcium-binding fluorescence dye (Fluo-8) to observe the calcium uptake dynamic with the fluorescence microscopy. Moreover, the molecular dynamics simulation visualized the process of calcium ions inflow during μsEP. According to our results calcium electroporation significantly decreases the cells viability by promoting apoptosis. Furthermore, our data shows that the application of pulsed electric fields disassembles the actin cytoskeleton and influences the prostate cancer cells’ mobility.     

Developments in Vascular-Targeted Photodynamic Therapy for Urologic Malignancies

With improved understanding of cancer biology and technical advancements in non-invasive management of urological malignancies, there is renewed interest in photodynamic therapy (PDT) as a means of focal cancer treatment. The application of PDT has also broadened as a result of development of better-tolerated and more effective photosensitizers. Vascular-targeted PDT (VTP) using padeliporfin, which is a water-soluble chlorophyll derivative, allows for tumor-specific cytotoxicity and has demonstrated efficacy in the management of urologic malignancies. Herein, we describe the evolution of photodynamic therapy in urologic oncology and the role of VTP in emerging treatment paradigms.     

Quo Vadis Advanced Prostate Cancer Therapy? Novel Treatment Perspectives and Possible Future Directions

Prostate cancer is a very common disease, which is, unfortunately, often the cause of many male deaths. This is underlined by the fact that the early stages of prostate cancer are often asymptomatic. Therefore, the disease is usually detected and diagnosed at late advanced or even metastasized stages, which are already difficult to treat. Hence, it is important to pursue research and development not only in terms of novel diagnostic methods but also of therapeutic ones, as well as to increase the effectiveness of the treatment by combinational medicinal approach. Therefore, in this review article, we focus on recent approaches and novel potential tools for the treatment of advanced prostate cancer; these include not only androgen deprivation therapy, antiandrogen therapy, photodynamic therapy, photothermal therapy, immunotherapy, multimodal therapy, but also poly(ADP-ribose) polymerase, Akt and cyclin-dependent kinase inhibitors.     

Suitable reference genes for relative quantification of miRNA expression in prostate cancer

Real time quantitative PCR (qPCR) is the method of choice for miRNA expression studies. For relative quantification of miRNAs, normalization to proper reference genes is mandatory. Currently, no validated reference genes for miRNA qPCR in prostate cancer are available. In this study, the expression of four putative reference genes (hsa-miR-16, hsa-miR-130b, RNU6-2, SNORD7) was examined with regard to their use as normalizer. After SNORD7 was already shown an inappropriate reference gene in preliminary experiments using total RNA pools, we studied the expression of the putative reference genes in tissue and normal adjacent tissue sample pairs from 76 men with untreated prostate carcinoma collected after radical prostatectomy. hsa-miR-130b and RNU6-2 showed no significantly different expression between the matched malignant and non-malignant tissue samples, whereas hsa-miR-16 was significantly underexpressed in malignant tissue. Softwares geNorm and Normfinder predicted hsa- miR-130b and the geometric mean of hsa-miR-130b and RNU6-2 as the most stable reference genes. Normalization of the four miRNAs hsa-miR-96, hsa- miR-125b, hsa-miR-205, and hsa-miR-375, which were previously shown to be regulated, shows that normalization to hsa-mir-16 can lead to biased results. We recommend using hsa-miR-130b or the geometric mean of hsa-miR-130b and small RNA RNU6-2 for normalization in miRNA expression studies of prostate cancer.