Ontogeny of Human Liver Aldehyde Oxidase: Developmental Changes and Implications for Drug Metabolism.

Despite the increasing importance of aldehyde oxidase (AO) in the drug metabolism of clinical candidates, ontogeny data for AO are limited. The objective of our study was to characterize the age-dependent AO content and activity in the human liver cytosolic fraction (HLC) and human hepatocytes (HH). HLC (n = 121 donors) and HH (n = 50 donors) were analyzed for (1) AO protein content by quantitative proteomics and (2) enzyme activity using carbazeran as a probe substrate. AO activity showed high technical variability and poor correlation with the content in HLC samples, whereas hepatocyte samples showed a strong correlation between the content and activity. Similarly, AO content and activity showed no significant age-dependent differences in HLC samples, whereas the average AO content and activity in hepatocytes increased significantly (∼20-40-fold) from the neonatal levels (0-28 days). Based on the hepatocyte data, the age at which 50% of the adult AO content is reached (age50) was 3.15 years (0.32-13.97 years, 95% CI). Metabolite profiling of carbazeran revealed age-dependent metabolic switching and the role of non-AO mechanisms (glucuronidation and desmethylation) in carbazeran elimination. The content-activity correlation in hepatocytes improved significantly (R2 = 0.95; p < 0.0001) in samples showing <10% contribution of glucuronidation toward the overall metabolism, confirming that AO-mediated oxidation and glucuronidation are the key routes of carbazeran metabolism. Considering the confounding effect of glucuronidation on AO activity, AO content-based ontogeny data are a more direct reflection of developmental changes in protein expression. The comprehensive ontogeny data of AO in HH samples are more reliable than HLC data, which are important for developing robust physiologically based pharmacokinetic models for predicting AO-mediated metabolism in children.

Dissecting Parameters Contributing to the Underprediction of Aldehyde Oxidase-Mediated Metabolic Clearance of Drugs.

We investigated the effect of variability and instability in aldehyde oxidase (AO) content and activity on the scaling of in vitro metabolism data. AO content and activity in human liver cytosol (HLC) and five recombinant human AO preparations (rAO) were determined using targeted proteomics and carbazeran oxidation assay, respectively. AO content was highly variable as indicated by the relative expression factor (REF; i.e., HLC to rAO content) ranging from 0.001 to 1.7 across different in vitro systems. The activity of AO in HLC degrades at a 10-fold higher rate in the presence of the substrate as compared with the activity performed after preincubation without substrate. To scale the metabolic activity from rAO to HLC, a protein-normalized activity factor (pnAF) was proposed wherein the activity was corrected by AO content, which revealed up to sixfold higher AO activity in HLC versus rAO systems. A similar value of pnAF was observed for another substrate, ripasudil. Physiologically based pharmacokinetic (PBPK) modeling revealed a significant additional clearance (CL; 66%), which allowed for the successful prediction of in vivo CL of four other substrates, i.e., O-benzyl guanine, BIBX1382, zaleplon, and zoniporide. For carbazeran, the metabolite identification study showed that the direct glucuronidation may be contributing to around 12% elimination. Taken together, this study identified differential protein content, instability of in vitro activity, role of additional AO clearance, and unaccounted metabolic pathways as plausible reasons for the underprediction of AO-mediated drug metabolism. Consideration of these factors and integration of REF and pnAF in PBPK models will allow better prediction of AO metabolism. SIGNIFICANCE STATEMENT: This study elucidated the plausible reasons for the underprediction of aldehyde oxidase (AO)-mediated drug metabolism and provided recommendations to address them. It demonstrated that integrating protein content and activity differences and accounting for the loss of AO activity, as well as consideration of extrahepatic clearance and additional pathways, would improve the in vitro to in vivo extrapolation of AO-mediated drug metabolism using physiologically based pharmacokinetic modeling.

Interventions Designed to Improve HIV Continuum of Care Outcomes for Persons with HIV in Contact with the Carceral System in the USA.

PURPOSE OF REVIEW: To describe existing evidence and identify future directions for intervention research related to improving HIV care outcomes for persons with HIV involved in the carceral system in the USA, a population with high unmet HIV care needs. RECENT FINDINGS: Few recent intervention studies focus on improving HIV care outcomes for this population. Successful strategies to improve care outcomes include patient navigation, substance use treatment, and incentivizing HIV care outcomes. Technology-supported interventions are underutilized in this population. Notable gaps in the existing literature include intervention research addressing HIV care needs for cisgender and transgender women and those under carceral supervision in the community. Future research should address existing gaps in the literature and respond to emergent needs including understanding how the changing HIV care delivery environment resulting from the COVID-19 pandemic and the approval of new injectable ART formulation shape HIV care outcomes in this population.

Prediction of drug-induced kidney injury in drug discovery.

Drug induced kidney injury is one of the leading causes of failure of drug development programs in the clinic. Early prediction of renal toxicity potential of drugs is crucial to the success of drug candidates in the clinic. The dynamic nature of the functioning of the kidney and the presence of drug uptake proteins introduce additional challenges in the prediction of renal injury caused by drugs. Renal injury due to drugs can be caused by a wide variety of mechanisms and can be broadly classified as toxic or obstructive. Several biomarkers are available for in vitro and in vivo detection of renal injury. In vitro static and dynamic (microfluidic) cellular models and preclinical models can provide valuable information regarding the toxicity potential of drugs. Differences in pharmacology and subsequent disconnect in biomarker response, differences in the expression of transporter and enzyme proteins between in vitro to in vivo systems and between preclinical species and humans are some of the limitations of current experimental models. The progress in microfluidic (kidney-on-chip) platforms in combination with the ability of 3-dimensional cell culture can help in addressing some of these issues in the future. Finally, newer in silico and computational techniques like physiologically based pharmacokinetic modeling and machine learning have demonstrated potential in assisting prediction of drug induced kidney injury.

Role of miR-182/PDCD4 axis in aggressive behavior of prostate cancer in the African Americans.

BACKGROUND: Prostate cancer is one of the most commonly diagnosed cancers among men. African Americans (AA) are at an increased risk of developing prostate cancer compared to European Americans (EA). miRNAs play a critical role in these tumors, leading to tumor progression. In this study, we investigated the role of miR-182 in racial disparity in prostate cancer. RESULTS: We found significantly increased levels of miR-182 in prostate cancer tissues compared to BPH. Also, miR-182 shows increased expression in AA prostate cancer cell line and tissue samples compared to EA. We performed biochemical recurrence (BCR) - free survival time in AA and EA patients and found that high miR-182 expression had significantly shorter BCR-free survival than patients with low miR-182 expression (P = 0.031). To elucidate the role of miR-182, we knocked down miR-182 in EA (DU-145 and LNCaP) and AA (MDA-PCa-2b) cell lines and found an increase in apoptosis, arrest of the cell cycle, and inhibition of colony formation in the AA cell line to a greater extent than EA cell lines. CONCLUSIONS: Our results showed that PDCD4 is a direct miR-182 target and its inhibition is associated with aggressiveness and high Gleason grade in prostate cancer among AA. These findings show that miR-182 is highly expressed in AA patients and miR-182 may be a target for effective therapy in AA patients.

Polymorphism of interleukin-6 -174 G/C (rs1800795) & the corresponding interleukin-6 level as a prognostic marker of cervical cancer.

BACKGROUND & OBJECTIVES: Aetiology of cervical cancer (CaCx) is multifactorial. Besides human papillomavirus (HPV) infection, many immunogenetic factors are involved in this complex process. The present study was carried out to investigate one such factor, interleukin-6 (IL-6), a central pro-inflammatory cytokine and a polymorphism at its promoter region -174 G/C (rs1800795) with CaCx. METHODS: HPV-infected women with or without CaCx were enrolled in group I and II, respectively. Another group of uninfected healthy women was also included as group III for comparison. Polymorphism in IL-6-174 G/C and IL-6 levels were analyzed by sequence-specific primer PCR (PCR-SSP) and ELISA, respectively. RESULTS: Groups I (n=111) and II (n=87) had significantly higher frequency of IL-6-174 GG genotype [odds ratios (OR)=3.9; P<0.001 and OR=3.2; P<0.001, respectively] as compared to group III (n=163). Furthermore, individuals with GG or GC genotypes had high IL-6 levels than those with CC genotypes. IL-6 levels were significantly (P<0.001) elevated in group I. This was also significantly high in untreated cases as compared to treated (P<0.05) ones. IL-6 levels of treated group were comparable with groups II and III. INTERPRETATION & CONCLUSIONS: Our results suggested a possible association of IL-6-174 GG with CaCx, which was also associated with high IL-6 levels. Decreased levels of IL-6 following treatment indicate its possible prognostic use in CaCx cases.

Genistein Represses HOTAIR/Chromatin Remodeling Pathways to Suppress Kidney Cancer.

BACKGROUND/AIMS: Genistein, a soy isoflavone, has been shown to have anti-cancer effects in various cancers including renal cancer. Long non-coding RNA, HOX transcript antisense RNA (HOTAIR), is involved in cancer progression and metastasis, such as renal cancer. Our aim was to investigate the effects of genistein on HOTAIR chromatin remodeling functions. METHODS: We used MTS assays and Transwell migration assays to study the effects of genistein on cell proliferation and migration respectively in human renal cell carcinoma (RCC) cell lines. We used Western blots to analyze SNAIL and ZO-1 expression. We performed chromatin immunoprecipitation (ChIP) assays to study recruitment of the polycomb repressive complex 2 (PRC2) to the ZO-1 promoter. We performed RNA immunoprecipitation (RIP) assays to study interaction between HOTAIR and PRC2, SMARCB1 or ARID1A. We also performed transfection experiments to overexpress EED, HOTAIR and knockdown SMARCB1. RESULTS: Genistein reduced cell proliferation and migration of human renal cell carcinoma cell lines. ChIP assays indicated that genistein reduces recruitment of the PRC2 to the ZO-1 promoter and increased its expression. RIP assays showed that genistein inhibits HOTAIR interaction with PRC2, leading to tumor suppression. Immunoprecipitation also revealed that genistein reduced EED levels in PRC2, suggesting that decreased EED levels suppress HOTAIR interaction with PRC2. EED overexpression in the presence of genistein restored PRC2 interaction with HOTAIR and reduced ZO-1 transcription, suggesting genistein activates ZO-1 by inhibiting HOTAIR/PRC2 functions. RIP assays also showed that HOTAIR interacts with SMARCB1 and ARID1A, subunits of the human SWI/SNF chromatin remodeling complex and genistein reduces this interaction. Combination of HOTAIR overexpression and SMARCB1 knockdown in the presence of genistein revealed that genistein inhibits SNAIL transcription via the HOTAIR/SMARCB1 pathway. CONCLUSION: Genistein suppresses EED levels in PRC2 and inhibits HOTAIR/PRC2 interaction. Genistein suppresses HOTAIR/PRC2 recruitment to the ZO-1 promoter and enhances ZO-1 transcription. Genistein also inhibits SNAIL transcription via reducing HOTAIR/SMARCB1 interaction. We demonstrate that the reduction of HOTAIR interaction with chromatin remodeling factors by genistein represses HOTAIR/chromatin remodeling pathways to suppress RCC malignancy.

Activation of the Erk/MAPK signaling pathway is a driver for cadmium induced prostate cancer.

PURPOSE: Cadmium (Cd) is reported to be associated with carcinogenesis. The molecular mechanisms associated with Cd-induced prostate cancer (PCa) remain elusive. MATERIALS AND METHODS: RWPE1, PWR1E and DU 145 cells were used. RT2 Profiler Array, real-time-quantitative-PCR, immunofluorescence, cell cycle, apoptosis, proliferation and colony formation assays along with Gene Set Enrichment Analysis (GSEA) were performed. RESULT: Chronic Cd exposure of non-malignant RWPE1 and PWR1E cells promoted cell survival, proliferation and colony formation with inhibition of apoptosis. Even a two-week Cd exposure of PCa cell line (DU 145) significantly increased the proliferation and decreased apoptosis. RT2 profiler array of 84 genes involved in the Erk/MAPK pathway revealed induction of gene expression in Cd-RWPE1 cells compared to RWPE1. This was confirmed by individual TaqMan gene expression analysis in both Cd-RWPE1 and Cd-PWR1E cell lines. GSEA showed an enrichment of the Erk/MAPK pathway along with other pathways such as KEGG-ERBB, KEGG-Cell Cycle, KEGG-VEGF, KEGG-Pathways in cancer and KEGG-prostate cancer pathway. We randomly selected upregulated genes from Erk/MAPK pathway and performed profile analysis in a PCa data set from the TCGA/GDC data base. We observed upregulation of these genes in PCa compared to normal samples. An increase in phosphorylation of the Erk1/2 and Mek1/2 was observed in Cd-RWPE1 and Cd-PWR1E cells compared to parental cells, confirming that Cd-exposure induces activation of the Erk/MAPK pathway. CONCLUSION: This study demonstrates that Erk/MAPK signaling is a major pathway involved in Cd-induced malignant transformation of normal prostate cells. Understanding these dominant oncogenic pathways may help develop optimal therapeutic strategies for PCa.

Role of the PI3K/Akt pathway in cadmium induced malignant transformation of normal prostate epithelial cells.

This study investigated the role of the PI3K/Akt pathway in cadmium (Cd) induced malignant transformation of normal prostate epithelial (PWR1E and RWPE1) cells. Both PWR1E and RWPE1 cells were exposed to 10 µM Cd for one year and designated as Cd-PWR1E and Cd-RWPE1. Cd-RWPE1 cells robustly formed tumors in athymic nude mice. Functionally, Cd-exposure induced tumorigenic attributes indicated by increased wound healing, migration and invasion capabilities in both cell lines. RT2-array analysis revealed many oncogenes including P110α, Akt, mTOR, NFKB1 and RAF were induced whereas tumor suppressor (TS) genes were attenuated in Cd-RWPE1. This was validated by individual quantitative-real-time-PCR at transcriptional and by immunoblot at translational levels. These results were consistent in Cd-PWR1E vs parental PWR1E cells. Gene Set Enrichment Analysis revealed that five prostate cancer (PCa) related pathways were enriched in Cd-exposed cells compared to their normal controls. These pathways include the KEGG- Pathways in cancer, Prostate Cancer Pathway, ERBB, Apoptosis and MAPK pathways. We selected up- and down-regulated genes randomly from the PI3K/Akt pathway array and profiled these in the TCGA/GDC prostate-adenocarcinoma (PRAD) patient cohort. An upregulation of oncogenes and downregulation of TS genes was observed in PCa compared to their normal controls. Taken together, our study reveals that the PI3K/Akt signaling is one of the main molecular pathways involved in Cd-driven transformation of normal prostate epithelial cells to malignant form. Understanding the molecular mechanisms involved in the Cd-driven malignant transformation of normal prostate cells will provide a significant insight to develop better therapeutic strategies for Cd-induced prostate cancer.

A hybrid model to evaluate the impact of active uptake transport on hepatic distribution of atorvastatin in rats.

1. Mathematical modeling remains a useful tool to study the impact of transporters on overall and intracellular drug disposition. The impact of organic anion transporter protein mediated uptake on atorvastatin systemic and intracellular pharmacokinetics, specifically distribution volume, was studied in rats with mathematical modeling and conducting in vivo pharmacokinetic studies for atorvastatin in presence and absence of rifampicin. A previously developed 5-compartment explicit membrane model for the liver was combined with a compartmental model to develop a semi-physiological hybrid model for atorvastatin disposition. 2. Rifampicin treatment resulted in a decrease in systemic clearance and steady-state distribution volume, and an increase in half-life of atorvastatin. The hybrid model predicted higher unbound intracellular liver atorvastatin concentrations than unbound plasma concentrations in both rifampicin treated and untreated groups, indicating involvement of uptake transporters. The intracellular unbound concentrations during the distributive phase were unaffected by rifampicin. The dependence of clearance on blood flow as well as hepatic uptake for atorvastatin (a moderate-to-high extraction ratio drug) can explain this lack of change in intracellular concentrations if there is incomplete inhibition of transport at the tested rifampicin dose. 3. The hybrid model successfully allowed the evaluation of effect of active uptake on intracellular and plasma atorvastatin disposition.

Intracellular Unbound Atorvastatin Concentrations in the Presence of Metabolism and Transport.

Accurate prediction of drug target activity and rational dosing regimen design require knowledge of drug concentrations at the target. It is important to understand the impact of processes such as membrane permeability, partitioning, and active transport on intracellular drug concentrations. The present study aimed to predict intracellular unbound atorvastatin concentrations and characterize the effect of enzyme-transporter interplay on these concentrations. Single-pass liver perfusion studies were conducted in rats using atorvastatin (ATV, 1 µM) alone at 4°C and at 37°C in presence of rifampin (RIF, 20 µM) and 1-aminobenzotriazole (ABT, 1 mM), separately and in combination. The unbound intracellular ATV concentration was predicted with a five-compartment explicit membrane model using the parameterized diffusional influx clearance, active basolateral uptake clearance, and metabolic clearance. Chemical inhibition of uptake and metabolism at 37°C proved to be better controls relative to studies at 4°C. The predicted unbound intracellular concentration at the end of the 50-minute perfusion in the +ABT , +ABT+RIF, and the ATV-only groups was 6.5 µM, 0.58 µM, and 5.14 µM, respectively. The predicted total liver concentrations and amount recovered in bile were within 0.94-1.3 fold of the observed value in all groups. The fold difference in total liver concentration did not always extrapolate to the fold difference in predicted unbound concentration across groups. Together, these results support the use of compartmental modeling to predict intracellular concentrations in dynamic organ-based systems. These predictions can provide insight into the role of uptake transporters and metabolizing enzymes in determining drug tissue concentrations.

Enzymatic Routes for Chiral Amine Synthesis: Protein Engineering and Process Optimization.

Chiral amines are essential motifs in pharmaceuticals, agrochemicals, and specialty chemicals. While traditional chemical routes to chiral amines often lack stereoselectivity and require harsh conditions, biocatalytic methods using engineered enzymes can offer high efficiency and selectivity under sustainable conditions. This review discusses recent advances in protein engineering of transaminases, oxidases, and other enzymes to improve catalytic performance. Strategies such as directed evolution, immobilization, and computational redesign have expanded substrate scope and enhanced efficiency. Furthermore, process optimization guided by techno-economic assessments has been crucial for establishing viable biomanufacturing routes. Combining state-of-the-art enzyme engineering with multifaceted process development will enable scalable, economical enzymatic synthesis of diverse chiral amine targets.

Nanoparticle biodistribution coefficients: A quantitative approach for understanding the tissue distribution of nanoparticles.

The objective of this manuscript is to provide quantitative insights into the tissue distribution of nanoparticles. Published pharmacokinetics of nanoparticles in plasma, tumor and 13 different tissues of mice were collected from literature. A total of 2018 datasets were analyzed and biodistribution of graphene oxide, lipid, polymeric, silica, iron oxide and gold nanoparticles in different tissues was quantitatively characterized using Nanoparticle Biodistribution Coefficients (NBC). It was observed that typically after intravenous administration most of the nanoparticles are accumulated in the liver (NBC = 17.56 %ID/g) and spleen (NBC = 12.1 %ID/g), while other tissues received less than 5 %ID/g. NBC values for kidney, lungs, heart, bones, brain, stomach, intestine, pancreas, skin, muscle and tumor were found to be 3.1 %ID/g, 2.8 %ID/g, 1.8 %ID/g, 0.9 %ID/g, 0.3 %ID/g, 1.2 %ID/g, 1.8 %ID/g, 1.2 %ID/g, 1.0 %ID/g, 0.6 %ID/g and 3.4 %ID/g, respectively. Significant variability in nanoparticle distribution was observed in certain organs such as liver, spleen and lungs. A large fraction of this variability could be explained by accounting for the differences in nanoparticle physicochemical properties such as size and material. A critical overview of published nanoparticle physiologically-based pharmacokinetic (PBPK) models is provided, and limitations in our current knowledge about in vitro and in vivo pharmacokinetics of nanoparticles that restrict the development of robust PBPK models is also discussed. It is hypothesized that robust quantitative assessment of whole-body pharmacokinetics of nanoparticles and development of mathematical models that can predict their disposition can improve the probability of successful clinical translation of these modalities.

Streamlining Food Effect Assessment - Are Repeated Food Effect Studies Needed? An IQ Analysis.

Current regulatory guidelines on drug-food interactions recommend an early assessment of food effect to inform clinical dosing instructions, as well as a pivotal food effect study on the to-be-marketed formulation if different from that used in earlier trials. Study waivers are currently only granted for BCS class 1 drugs. Thus, repeated food effect studies are prevalent in clinical development, with the initial evaluation conducted as early as the first-in-human studies. Information on repeated food effect studies is not common in the public domain. The goal of the work presented in this manuscript from the Food Effect PBPK IQ Working Group was to compile a dataset on these studies across pharmaceutical companies and provide recommendations on their conduct. Based on 54 studies collected, we report that most of the repeat food effect studies do not result in meaningful differences in the assessment of the food effect. Seldom changes observed were more than twofold. There was no clear relationship between the change in food effect and the formulation change, indicating that in most cases, once a compound is formulated appropriately within a specific formulation technology, the food effect is primarily driven by inherent compound properties. Representative examples of PBPK models demonstrate that following appropriate validation of the model with the initial food effect study, the models can be applied to future formulations. We recommend that repeat food effect studies should be approached on a case-by-case basis taking into account the totality of the evidence including the use of PBPK modeling.

Isolation, biochemical characterization, and development of a biodegradable antimicrobial film from Cirrhinus mrigala scale collagen.

Collagen is a promising candidate for food and pharmaceutical applications due to its excellent biocompatibility, low antigenicity, and controlled biodegradability; however, its heavy price restricts its utilization. Fish scales generated during the processing are generally regarded as waste material and an environmental pollutant, though they are a promising source of collagen. In the present study, Cirrhinus mrigala scales were demineralized and extracted for acid-soluble collagen (ASC) using acetic acid, with a collagen yield of 2.7%. UV-Vis spectra, SDS-PAGE, FTIR analyses, and amino acid composition confirmed the type I nature of the collagen extracted. The denaturation temperature of the collagen was found to be 30.09 °C using differential scanning calorimetry (DSC). The collagen was highly soluble at acidic pH and lower NaCl concentrations while its solubility was lowered in alkaline conditions and NaCl concentrations above 0.5 M. The collagen exhibited good emulsifying potential with an emulsion activity index (EAI) and emulsion stability index (ESI) of 21.49 ± 0.22 m2 g-1 and 15.67 ± 0.13 min, respectively. Owing to the good physicochemical characteristics of the extracted collagen, collagen-chitosan-neem extract (CCN) films were prepared subsequently which showed good antimicrobial activity against Bacillus subtilis NCIM 2635, Staphylococcus aureus NCIM 2654, Escherichia coli NCIM 2832, and Pseudomonas aeruginosa NCIM 5032, suggesting the potential of collagen in the development of antimicrobial films. These results demonstrate that the collagen from fish waste could be valorized and used effectively along with chitosan and neem extract for the synthesis of novel biodegradable films with antimicrobial efficacy.

Commercial and Social Value of Pharmaceutical Industry-led Access Programs: Conceptual Framework and Descriptive Analysis.

Pharmaceutical industry-led access programs are growing in number globally and are increasingly adopting a hybrid approach intended to generate commercial and social value in parallel. We developed and applied a new conceptual framework in a descriptive analysis of observable indicators measuring commercial and social value for 91 programs registered in the Access Observatory. We found that most programs had features consistent with the generation of commercial value, directly through revenue generation (50.0%), or indirectly by creating competitive advantage (70.3%). We also found that most programs were implemented in countries where the company has commercial products registered (85.5%). While many programs had features consistent with the generation of social value, it was difficult to ascertain the level of that value because most did not share data (83.5%) and had not been evaluated (74.7%). Future efforts by the global health community and the pharmaceutical industry should focus on strengthening measurement and reporting on commercial and social indicators of industry-led access programs.

Prediction of Drug Clearance from Enzyme and Transporter Kinetics.

Accurate estimation of in vivo clearance in human is pivotal to determine the dose and dosing regimen for drug development. In vitro-in vivo extrapolation (IVIVE) has been performed to predict drug clearance using empirical and physiological scalars. Multiple in vitro systems and mathematical modeling techniques have been employed to estimate in vivo clearance. The models for predicting clearance have significantly improved and have evolved to become more complex by integrating multiple processes such as drug metabolism and transport as well as passive diffusion. This chapter covers the use of conventional as well as recently developed methods to predict metabolic and transporter-mediated clearance along with the advantages and disadvantages of using these methods and the associated experimental considerations. The general approaches to improve IVIVE by use of appropriate scalars, incorporation of extrahepatic metabolism and transport and application of physiologically based pharmacokinetic (PBPK) models with proteomics data are also discussed. The chapter also provides an overview of the advantages of using such dynamic mechanistic models over static models for clearance predictions to improve IVIVE.

Evaluation of drug-drug interaction potential between omecamtiv mecarbil and rosuvastatin, a BCRP substrate, with a clinical study in healthy subjects and using a physiologically-based pharmacokinetic model.

Omecamtiv mecarbil (OM) is a novel cardiac myosin activator in development for the treatment of heart failure. In vitro, OM is an inhibitor of BCRP. Rosuvastatin, a BCRP substrate, is one of the most commonly prescribed medications in patients with heart failure. The potential for a pharmacokinetic (PK) drug-drug interaction (DDI) was investigated, specifically to determine whether a single 50 mg dose of OM would impact the PKs of a single 10 mg dose of rosuvastatin in an open-label study in 14 healthy subjects. The ratios of the geometric least-square means (90% confidence intervals [CIs]) of rosuvastatin co-administered with OM compared to rosuvastatin alone were 127.1% (90% CI 113.8-141.9), 132.8% (90% CI 120.7-146.1), and 154.2% (90% CI 132.8-179.1) for area under the plasma-concentration time curve from time zero to infinity (AUCinf ), area under the plasma-concentration time curve from time zero to time of last quantifiable concentration (AUClast ), and maximum observed plasma concentration (Cmax ), respectively. Whereas the DDI study with rosuvastatin was conducted with the co-administration of a single dose of OM, in the clinical setting, patients receive OM at doses of 25, 37.5, or 50 mg twice daily (b.i.d.). Hence, to extrapolate the results of the DDI study to a clinically relevant scenario of continuous b.i.d. dosing with OM, physiologically-based pharmacokinetic (PBPK) modeling was performed to explore the potential of BCRP inhibition following continuous b.i.d. dosing of OM at the highest 50 mg dose. Modeling results indicated that following 50 mg b.i.d. dosing of OM, the predicted ratios of the geometric means (90% CIs) for rosuvastatin AUCinf and Cmax were 1.18 (90% CI 1.16-1.20) and 2.04 (90% CI 1.99-2.10), respectively. Therefore, these results suggest that OM, following multiple dose administration, is a weak inhibitor of BCRP substrates and is in accordance with that observed in the single dose OM DDI clinical study.

A lncRNA TCL6-miR-155 Interaction Regulates the Src-Akt-EMT Network to Mediate Kidney Cancer Progression and Metastasis.

Metastasis is the leading cause of mortality from kidney cancer, and understanding the underlying mechanism of this event will provide better strategies for its management. Here we investigated the biological, functional, and clinical significance of lncTCL6 and its interacting miR-155 in clear cell renal cell carcinoma (ccRCC). We employed a comprehensive approach to investigate the lncTCL6-miR-155-Src/Akt-mediated epithelial-to-mesenchymal transition (EMT) pathway as a novel regulatory mechanism in ccRCC progression. Expression analyses revealed that lncTCL6 is downregulated in ccRCC compared with normal tissues. Overexpression of lncTCL6 in ccRCC cell lines impaired their oncogenic functions, such as cell proliferation and migration/invasion, and induced cell-cycle arrest and apoptosis; conversely, depletion of lncTCL6 rescued these phenotypic effects. Furthermore, lncTCL6 directly interacted with miR-155. Unlike lncTCL6, miR-155 was overexpressed in ccRCC. Stable knockdown of miR-155 phenocopied the effects of lncTCL6 overexpression. Conversely, reconstitution of miR-155 and suppression of lncTCL6 in noncancerous renal cell HK2 induced tumorigenic characteristics. Patients with higher expression of lncTCL6 and lower expression of miR-155 had better survival probability. When overexpressed, lncTCL6 recruited STAU1 and mediated decay of Src mRNA, followed by a marked downregulation of an integrated network of Src target genes involved in migration, invasion, and EMT. However, the interaction between miR-155 and lncTCL6 attenuated the regulatory role of lncTCL6 on Src-mediated EMT. In conclusion, this study is the first report documenting the lncTCL6-miR155-Src/Akt/EMT network as a novel regulatory mechanism in aggressive ccRCC and a promising therapeutic target to inhibit renal cancer. SIGNIFICANCE: This study's investigation of noncoding RNA interactions in renal cell carcinoma identify miRNA-155-lncRNA TCL6-mediated regulation of the Src-Akt-EMT network as a novel mechanism of disease progression and metastasis.

Suppressor effect of catechol-O-methyltransferase gene in prostate cancer.

Catechol-estrogens can cause genetic mutations and to counteract their oncogenicity, the catechol-O-methyltransferase (COMT) gene is capable of neutralizing these reactive compounds. In this study, we determined the functional effects and regulation of COMT in prostate cancer. Both the Cancer Genome Atlas (TCGA) and immunohistochemical analysis of clinical specimens demonstrated a reduction of COMT expression in prostate cancer. Also, western analyses of prostate cancer cell lines show COMT levels to be minimal in DuPro and DU145 and thus, these cells were used for further analyses. Re-expression of COMT led to suppressed migration ability (wound healing assay) and enhanced apoptosis (flow cytometric analyses), and when challenged with 4-hydroxyestradiol, a marked reduction of cell proliferation (MTT assay) was observed. Xenograft growth in athymic mice also resulted in inhibition due to COMT. As a mechanism, western analyses show cleaved CASP3 and BID were increased whereas XIAP and cIAP2 were reduced due to COMT. As COMT expression is low in prostate cancer, its regulation was determined. Databases identified several miRNAs capable of binding COMT and of these, miR-195 was observed to be increased in prostate cancer according to TCGA. Real-time PCR validated upregulation of miR-195 in clinical prostate cancer specimens as well as DuPro and DU145 and interestingly, luciferase reporter showed miR-195 capable of binding COMT and overexpressing miR-195 could reduce COMT in cells. These results demonstrate COMT to play a protective role by activating the apoptosis pathway and for miR-195 to regulate its expression. COMT may thus be a potential biomarker and gene of interest for therapeutic development for prostate cancer.