Development and Validation of a Novel LC–MS/MS Method for Simultaneous Determination of Abiraterone and its Seven Steroidal Metabolites in Human Serum: Innovation in Separation of Diastereoisomers Without Use of a Chiral Column

Abiraterone acetate (AA), the prodrug of abiraterone, is FDA-approved for the treatment of castration-resistant prostate cancer. Abiraterone is metabolized in patients to a more potent analogue, D4A. However, we have recently reported that this analogue is further metabolized to additional metabolites in patients treated with AA. Here, we present a liquid chromatography-tandem mass spectrometry method developed to resolve and detect abiraterone and its seven metabolites in human serum using an AB Sciex Qtrap 5500 mass analyzer coupled with a Shimadzu Nexera UPLC station. Analytes and the internal standard (abiraterone-d4) were extracted from human serum using the liquid–liquid extraction procedure. The analytes were separated using a Zorbax Eclipse Plus C18 150 × 2.1 mm, 3.5 μm column at 40 °C and an isocratic mobile phase 35% A (0.1% formic acid in water), 65% B (0.1% formic acid in methanol:acetonitrile; 60:40). Electrospray ionization in positive mode was applied with multiple reaction monitoring in a total run time of 13 min. Abiraterone detection was linear in the range 2–400 ng/mL and all metabolites from 0.1–20 ng/mL. The method was validated following US FDA guidelines for bioanalytical method validation, and all the metabolite results were within the acceptance limits. Despite the similarity in structure and mass transition between the metabolites, the validated method separated all the metabolites, including diastereomers, to allow accurate identification and quantitation of each compound

FORMULATION AND EVALUATION OF BILAYERED FLOATING TABLET OF DILTIAZEM DRUG

Aim of study was to develop bilayered floating drug delivery for treatment of hypertension by delivering loading and maintenance dose for fast achievement of peak plasma concentration and maintaining the same respectively. The prepared drug loaded bilayered floating tablets were evaluated for pre and post compression parameters. Stability study of the promising formulation was also performed. The tablets were prepared by direct compression method. The loading dose was delivered in the form of immediate release layer prepared by different super-disintegrations and maintenance dose was delivered through sustained release layer prepared by using polymers like HPMC K15M and Carbopol 934P. Both the immediate release layer and sustained release layers were separately optimized and then combined to optimize the bilayered floating tablets. No interactions were found between drug and excipients. Formulation containing crosscarmellose sodium shows immediate drug release. Formulation Containing HPMC K15M shows sustained release action and bilayered formulations FB7 shows releases up to 12 hours with good buoyancy and total floating time. All the Bilayered floating formulations buoyant up to 12 hrs. Bilayered floating tablets with release characteristics offer critical advantages such as, site specificity with improved absorption and efficacy. This technology can be inculcated to various medicaments which have stomach as the major site of absorption. Key words: Diltiazem, Bilayered floating tablet, sustain release table

Redirecting abiraterone metabolism to fine tune prostate cancer anti-androgen therapy

Abiraterone blocks androgen synthesis and prolongs survival in castration-resistant prostate cancer, which is otherwise driven by intratumoral androgen synthesis1,2. Abiraterone is metabolized in patients to D4A, which has even greater anti-tumor activity and structural similarities to endogenous steroidal 5α-reductase substrates, such as testosterone3. Here, we show that D4A is converted to at least 3 5α-reduced and 3 5β-reduced metabolites. The initial 5α-reduced metabolite, 3-keto-5α-abi, is more abundant than D4A in patients with prostate cancer taking abiraterone, and is an androgen receptor (AR) agonist, which promotes prostate cancer progression. In a clinical trial of abiraterone alone, followed by abiraterone plus dutasteride (a 5α-reductase inhibitor), 3-keto-5α-abi and downstream metabolites are depleted, while D4A concentrations rise, effectively blocking production of a tumor-promoting metabolite and permitting D4A accumulation. Furthermore, dutasteride does not deplete three 5β-reduced metabolites, which were also clinically detectable, demonstrating the specific biochemical effects of pharmacologic 5α-reductase inhibition on abiraterone metabolism. Our findings suggest a previously unappreciated and biochemically specific method of clinically fine-tuning abiraterone metabolism to optimize therapy

Global burden of 288 causes of death and life expectancy decomposition in 204 countries and territories and 811 subnational locations, 1990–2021: a systematic analysis for the Global Burden of Disease Study 2021

BACKGROUND Regular, detailed reporting on population health by underlying cause of death is fundamental for public health decision making. Cause-specific estimates of mortality and the subsequent effects on life expectancy worldwide are valuable metrics to gauge progress in reducing mortality rates. These estimates are particularly important following large-scale mortality spikes, such as the COVID-19 pandemic. When systematically analysed, mortality rates and life expectancy allow comparisons of the consequences of causes of death globally and over time, providing a nuanced understanding of the effect of these causes on global populations. METHODS The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 cause-of-death analysis estimated mortality and years of life lost (YLLs) from 288 causes of death by age-sex-location-year in 204 countries and territories and 811 subnational locations for each year from 1990 until 2021. The analysis used 56 604 data sources, including data from vital registration and verbal autopsy as well as surveys, censuses, surveillance systems, and cancer registries, among others. As with previous GBD rounds, cause-specific death rates for most causes were estimated using the Cause of Death Ensemble model-a modelling tool developed for GBD to assess the out-of-sample predictive validity of different statistical models and covariate permutations and combine those results to produce cause-specific mortality estimates-with alternative strategies adapted to model causes with insufficient data, substantial changes in reporting over the study period, or unusual epidemiology. YLLs were computed as the product of the number of deaths for each cause-age-sex-location-year and the standard life expectancy at each age. As part of the modelling process, uncertainty intervals (UIs) were generated using the 2·5th and 97·5th percentiles from a 1000-draw distribution for each metric. We decomposed life expectancy by cause of death, location, and year to show cause-specific effects on life expectancy from 1990 to 2021. We also used the coefficient of variation and the fraction of population affected by 90% of deaths to highlight concentrations of mortality. Findings are reported in counts and age-standardised rates. Methodological improvements for cause-of-death estimates in GBD 2021 include the expansion of under-5-years age group to include four new age groups, enhanced methods to account for stochastic variation of sparse data, and the inclusion of COVID-19 and other pandemic-related mortality-which includes excess mortality associated with the pandemic, excluding COVID-19, lower respiratory infections, measles, malaria, and pertussis. For this analysis, 199 new country-years of vital registration cause-of-death data, 5 country-years of surveillance data, 21 country-years of verbal autopsy data, and 94 country-years of other data types were added to those used in previous GBD rounds. FINDINGS The leading causes of age-standardised deaths globally were the same in 2019 as they were in 1990; in descending order, these were, ischaemic heart disease, stroke, chronic obstructive pulmonary disease, and lower respiratory infections. In 2021, however, COVID-19 replaced stroke as the second-leading age-standardised cause of death, with 94·0 deaths (95% UI 89·2-100·0) per 100 000 population. The COVID-19 pandemic shifted the rankings of the leading five causes, lowering stroke to the third-leading and chronic obstructive pulmonary disease to the fourth-leading position. In 2021, the highest age-standardised death rates from COVID-19 occurred in sub-Saharan Africa (271·0 deaths [250·1-290·7] per 100 000 population) and Latin America and the Caribbean (195·4 deaths [182·1-211·4] per 100 000 population). The lowest age-standardised death rates from COVID-19 were in the high-income super-region (48·1 deaths [47·4-48·8] per 100 000 population) and southeast Asia, east Asia, and Oceania (23·2 deaths [16·3-37·2] per 100 000 population). Globally, life expectancy steadily improved between 1990 and 2019 for 18 of the 22 investigated causes. Decomposition of global and regional life expectancy showed the positive effect that reductions in deaths from enteric infections, lower respiratory infections, stroke, and neonatal deaths, among others have contributed to improved survival over the study period. However, a net reduction of 1·6 years occurred in global life expectancy between 2019 and 2021, primarily due to increased death rates from COVID-19 and other pandemic-related mortality. Life expectancy was highly variable between super-regions over the study period, with southeast Asia, east Asia, and Oceania gaining 8·3 years (6·7-9·9) overall, while having the smallest reduction in life expectancy due to COVID-19 (0·4 years). The largest reduction in life expectancy due to COVID-19 occurred in Latin America and the Caribbean (3·6 years). Additionally, 53 of the 288 causes of death were highly concentrated in locations with less than 50% of the global population as of 2021, and these causes of death became progressively more concentrated since 1990, when only 44 causes showed this pattern. The concentration phenomenon is discussed heuristically with respect to enteric and lower respiratory infections, malaria, HIV/AIDS, neonatal disorders, tuberculosis, and measles. INTERPRETATION Long-standing gains in life expectancy and reductions in many of the leading causes of death have been disrupted by the COVID-19 pandemic, the adverse effects of which were spread unevenly among populations. Despite the pandemic, there has been continued progress in combatting several notable causes of death, leading to improved global life expectancy over the study period. Each of the seven GBD super-regions showed an overall improvement from 1990 and 2021, obscuring the negative effect in the years of the pandemic. Additionally, our findings regarding regional variation in causes of death driving increases in life expectancy hold clear policy utility. Analyses of shifting mortality trends reveal that several causes, once widespread globally, are now increasingly concentrated geographically. These changes in mortality concentration, alongside further investigation of changing risks, interventions, and relevant policy, present an important opportunity to deepen our understanding of mortality-reduction strategies. Examining patterns in mortality concentration might reveal areas where successful public health interventions have been implemented. Translating these successes to locations where certain causes of death remain entrenched can inform policies that work to improve life expectancy for people everywhere. FUNDING Bill & Melinda Gates Foundation

Heterogeneity in glycan composition on the surface of HIV-1 envelope determines virus sensitivity to lectins.

Lectins that target N-glycans on the surface of HIV-1 envelope (Env) glycoprotein have the potential for use as antiviral agents. Although progress has been made in deciphering the molecular details of lectin and Env glycan interaction, further studies are needed to better understand Env glycan heterogeneity among HIV-1 isolates and its influence on virus-neutralization sensitivity to lectins. This study evaluated a panel of lectins with fine specificity for distinct oligosaccharides and assessed their ability to inhibit infection of HIV-1 viruses known to have differing sensitivity to anti-HIV Env antibodies. The results showed that HIV-1 isolates have different sensitivity to lectins specific for α1-3Man, α1-6Man, and α1-2Man binding lectins. Considering that lectins exclusively recognize the oligosaccharide components of virus Env, these data suggest that glycan heterogeneity among HIV-1 isolates may explain this differential sensitivity. To evaluate this further, chronic and acute viruses were produced in the presence of different glycosidase inhibitors to express more homogenous glycans. Viruses enriched for α1-2Man terminating Man5-9GlcNAc2 glycans became similarly sensitive to α1-2Man-binding lectins. The α1-3Man- and α1-6Man-binding lectins also were more potent against viruses expressing predominantly Man5GlcNAc2 and hybrid type glycans with terminal α1-3Man and α1-6Man. Furthermore, lectin-mediated inhibition was competitively alleviated by mannan and this effect was augmented by enrichment of mannose-type glycans on the virus. In addition, while Env of viruses enriched with mannose-type glycans were sensitive to Endo-H deglycosylation, Env of untreated viruses were partially resistant, indicating that HIV-1 Env glycans are heterogeneously comprised of complex, hybrid, and mannose types. Overall, our data demonstrate that HIV-1 isolates display differential sensitivity to lectins, in part due to the microheterogeneity of N-linked glycans expressed on the surface of the virus Env glycoprotein

Development and Validation of a Novel LC–MS/MS Method for Simultaneous Determination of Abiraterone and its Seven Steroidal Metabolites in Human Serum: Innovation in Separation of Diastereoisomers Without Use of a Chiral Column

Abiraterone acetate (AA), the prodrug of abiraterone, is FDA-approved for the treatment of castration-resistant prostate cancer. Abiraterone is metabolized in patients to a more potent analogue, D4A. However, we have recently reported that this analogue is further metabolized to additional metabolites in patients treated with AA. Here, we present a liquid chromatography-tandem mass spectrometry method developed to resolve and detect abiraterone and its seven metabolites in human serum using an AB Sciex Qtrap 5500 mass analyzer coupled with a Shimadzu Nexera UPLC station. Analytes and the internal standard (abiraterone-d4) were extracted from human serum using the liquid–liquid extraction procedure. The analytes were separated using a Zorbax Eclipse Plus C18 150 × 2.1 mm, 3.5 μm column at 40 °C and an isocratic mobile phase 35% A (0.1% formic acid in water), 65% B (0.1% formic acid in methanol:acetonitrile; 60:40). Electrospray ionization in positive mode was applied with multiple reaction monitoring in a total run time of 13 min. Abiraterone detection was linear in the range 2–400 ng/mL and all metabolites from 0.1–20 ng/mL. The method was validated following US FDA guidelines for bioanalytical method validation, and all the metabolite results were within the acceptance limits. Despite the similarity in structure and mass transition between the metabolites, the validated method separated all the metabolites, including diastereomers, to allow accurate identification and quantitation of each compound

Postpartum depression in India: a systematic review and meta-analysis

To provide an estimate of the burden of postpartum depression in Indian mothers and investigate some risk factors for the condition. Methods We searched PubMed®, Google Scholar and Embase® databases for articles published from year 2000 up to 31 March 2016 on the prevalence of postpartum depression in Indian mothers. The search used subject headings and keywords with no language restrictions. Quality was assessed via the Newcastle-Ottawa quality assessment scale. We performed the meta-analysis using a random effects model. Subgroup analysis and meta-regression was done for heterogeneity and the Egger test was used to assess publication bias. Thirty-eight studies involving 20 043 women were analysed. Studies had a high degree of heterogeneity (I2 = 96.8%) and there was evidence of publication bias (Egger bias = 2.58; 95% confidence interval, CI: 0.83-4.33). The overall pooled estimate of the prevalence of postpartum depression was 22% (95% CI: 19-25). The pooled prevalence was 19% (95% CI: 17-22) when excluding 8 studies reporting postpartum depression within 2 weeks of delivery. Small, but non-significant differences in pooled prevalence were found by mother\u27s age, geographical location and study setting. Reported risk factors for postpartum depression included financial difficulties, presence of domestic violence, past history of psychiatric illness in mother, marital conflict, lack of support from husband and birth of a female baby. The review shows a high prevalence of postpartum depression in Indian mothers. More resources need to be allocated for capacity-building in maternal mental health care in India

Effect of <i>N</i>-glycan modification on virus sensitivity to lectins.

<p>JRFL.JB and REJO4541.67 viruses were produced in 293T cells in the presence of glycosylation-pathway inhibitors kifunensine or swainsonine, or in 293S GnTI^-/- cells lacking GlcNAc transferase I. Lectins were serially diluted; viruses were then added at 200 TCID₅₀ and incubated for 1 h at 37°C before addition of TZM-bl cells. After 48 h, virus infection was measured based on β-galactosidase activity. Cells infected with viruses only, in the absence of lectins, represent 100% infection. Percentage of virus inhibition by lectin was calculated using virus control (virus and cells only, 0% inhibition) and cell control (cell and no virus, 100% inhibition). All experiments were performed in duplicate and repeated three times. Data are shown as the averages and standard deviations from all 3 experiments.</p

Effect of <i>N</i>-glycan modification on Env incorporation to virions.

<p>Viruses were produced in 293T HEK cells in the presence or absence of glycosidase inhibitors kifunensine and swainsonine or in 293S HEK GnTI-/- cells. A) JRFL virus preparations were loaded to SDS-PAGE after normalization for CA-p24 content. Env was detected by Western blot with anti-gp120 mAbs and quantified using Image Lab (BioRad) software. B) The relative contents of Env associated with N-glycan-modified JRFL viruses were calculated in comparison with Env of wild type control (set at 100%). C) The relative Env incorporation to N-glycan modified vs wild type REJO viruses. Data represent averages and standard errors from 2 experiments (each in duplicate) and were analyzed using unpaired t-test. ***p <0.0005, ** p <0.005. D) Correlation of Env incorporation to virions and virus infectivity by spearman’s rank test.</p