Strategies for the modulation of phase II metabolism in a series of PKCε inhibitors.

Extensive phase II metabolism of an advanced PKCε inhibitor resulted in sub-optimal pharmacokinetics in rat marked by elevated clearance. Synthesis of the O-glucuronide metabolite as a standard was followed by three distinct strategies to specifically temper phase II metabolic degradation of the parent molecule. In this study, it was determined that the introduction of proximal polarity to the primary alcohol generally curbed O-glucuronidation and improved PK and physical chemical properties while maintaining potency against the target. Utilization of a Jacobsen hydrolytic kinetic resolution to obtain optically enriched final compounds is also discussed.

First detection and genetic characterization of chicken infectious anemia virus in the Mekong Delta, Vietnam.

Background: Chicken infectious anemia (CIA) caused by the CIA virus (CIAV) is considered one of the most important immunosuppressive diseases affecting chickens and recently poses a great economic burden to the poultry industry worldwide. Aim: This study aims to identify the presence of CIAV in the Mekong Delta (MD), Vietnam, and to determine genotypes of CIAVs that are currently circulating in this area. Methods: Organ samples (spleen, liver, and thymus) of 144 chickens suspected with CIA from 47 poultry farms were collected. A total of 47 pooled samples, each containing 2-4 chickens from each farm, were tested for the presence of CIAV. Results: Twenty out of 47 pooled organ samples (pool of 2-4 chickens per farm) were positive for CIAV using polymerase chain reaction targeting the viral VP1 gene. The VP1 amplicons of eight representative CIAVs were subjected to sequencing and genetic characterization. Phylogenetic analysis based on partial VP1 gene sequence revealed that the CIAVs detected in the MD grouped into different genotypes of II, IIIa, and IIIc together with CIAVs previously detected in the northern Vietnam and other Asian countries. The phylogenetic analysis also confirmed that detected CIAVs genetically differed from vaccine strains. In addition, deduced amino acids of the VP1 identified several critical amino acid substitutions in the VP1 protein that are likely associated with the virulence of CIAV. Conclusion: This is the first report to detect and determine the genetic characterization of the circulating CIAVs in the MD. Therefore, this study provides an important understanding of the evolution of CIAVs and highlights the importance of implementing prompt control measures against CIAVs in the MD and Vietnam.

Application of a high-resolution in vitro human MDR1-MDCK assay and in vivo studies in preclinical species to improve prediction of CNS drug penetration.

P-glycoprotein (P-gp, MDR1) is expressed at the blood-brain barrier (BBB) and restricts penetration of its substrates into the central nervous system (CNS). In vitro MDR1 assays are frequently used to predict the in vivo relevance of MDR1-mediated efflux at the BBB. It has been well established that drug candidates with high MDR1 efflux ratios (ERs) display poor CNS penetration. Following a comparison of MDR1 transporter function between the MDR1-MDCKI cell line from National Institutes of Health (NIH) and our internal MDR1-MDCKII cell line, the former was found to provide better predictions of in vivo brain penetration than our in-house MDR1-MDCKII cell line. In particular, the NIH MDR1 assay has an improved sensitivity to differentiate the compounds with ERs of <3 in our internal cell line and is able to reduce the risk of false negatives. A better correlation between NIH MDR1 ERs and brain penetration in rat and non-human primate (NHP) was demonstrated. Additionally, a comparison of brain penetration time course of MDR1 substrates and an MDR1 non-substrate in NHP demonstrated that MDR1 interaction can delay the time to equilibrium of drug concentration in the brain with plasma. It is recommended to select highly permeable compounds without MDR1 interaction for rapid brain penetration to produce the maximal pharmacological effect in the CNS with a quicker onset.

Real-world survival analysis by tumor mutational burden in non-small cell lung cancer: a multisite U.S. study.

BACKGROUND: Tumor mutational burden (TMB) is a potential biomarker to predict tumor response to immuno-oncology agents in patients with metastatic non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: A multi-site cohort study evaluated patients diagnosed with stage IV NSCLC between 2012 and 2019 who had received comprehensive genomic profiling (CGP) and any NSCLC-related treatment at 9 U.S. cancer centers. Baseline characteristics and clinical outcomes were compared between patients with TMB <10 and TMB ≥10. RESULTS: Among the 667 patients with CGP results, most patients received CGP from Foundation Medicine (64%) or Caris (20%). Patients with TMB ≥10 (vs. TMB <10) were associated with a positive smoking history. TMB was associated with ALK (p = 0.01), EGFR (p < 0.01), and TP53 (p < 0.05) alterations. TMB >10 showed a significant association towards longer overall survival (OS) (HR: 0.43, 95% CI: 0.21-0.88, p = 0.02) and progression-free survival (PFS) (HR: 0.43, 95% CI: 0.21-0.85, p = 0.02) in patients treated with first-line immunotherapy and tested by Foundation Medicine or Caris at treatment initiation. CONCLUSIONS: TMB levels greater than or equal to 10 mut/Mb, when tested by Foundation Medicine or Caris at treatment initiation, were significantly associated with improved OS and PFS among patients treated with first-line immunotherapy-containing regimens. Additional prospective research is warranted to validate this biomarker along with PD-L1 expression.

Third-Generation Anti-CD47-Specific CAR-T Cells Effectively Kill Cancer Cells and Reduce the Genes Expression in Lung Cancer Cell Metastasis.

CD47 is a cell surface glycoprotein molecule, belonging to the immunoglobulin superfamily, binding to various proteins including integrins, thrombospondin-1, and signal regulatory protein α (SIRPα). CD47 is an important tumor antigen for the development and progression of various cancers. This study designed the chimeric antigen receptor T-cell (CAR-T) to bind to the CD47 to inhibit the expression of CD47. We used the complementarity-determining regions (CDRs) of the B6H12 mouse antibody grafted onto the IgG1 framework to create the humanized single-chain variable fragment (scFv) with linker (G4S)x3. scFv was used to design the chimeric antigen receptor with the structure CD8signal-CD47scFv-CD8a hinge-CD4TM-CD28-41BB-CD3ζ, which was then transformed into T lymphocytes by the lentivirus to create third generation of CAR-T. Results revealed that the new CAR-T cells efficiently killed A549 cancer cells. CAR-T inhibited the expression of genes involved in metastasis and invasion of cells A549 including beta actin, calreticulin, and cyclooxygenase 2 at mRNA levels.

Digital Health Integration Assessment and Maturity of the United States Biopharmaceutical Industry: Forces Driving the Next Generation of Connected Autoinjectable Devices.

Autoinjectable devices continue to provide real-life benefits for patients with chronic conditions since their widespread adoption 30 years ago with the rise of macromolecules. Nonetheless, issues surrounding adherence, patient administration techniques, disease self-management, and data outcomes at scale persist despite product design innovation. The interface of drug device combination products and digital health technologies formulates a value proposition for next-generation autoinjectable devices to power the delivery of precision care at home and achieve the full potential of biologics. Success will largely be dependent on biopharma's digital health maturity to implement this framework. This viewpoint measures the digital health maturity of the top 15 biopharmaceutical companies in the US biologics autoinjector market and establishes the framework for next-generation autoinjectable devices powering home-based precision care and the need for formal digital health training.

De-novo Assembly of Limnospira fusiformis Using Ultra-Long Reads.

The Limnospira genus is a recently established clade that is economically important due to its worldwide use in biotechnology and agriculture. This genus includes organisms that were reclassified from Arthrospira, which are commercially marketed as "Spirulina." Limnospira are photoautotrophic organisms that are widely used for research in nutrition, medicine, bioremediation, and biomanufacturing. Despite its widespread use, there is no closed genome for the Limnospira genus, and no reference genome for the type strain, Limnospira fusiformis. In this work, the L. fusiformis genome was sequenced using Oxford Nanopore Technologies MinION and assembled using only ultra-long reads (>35 kb). This assembly was polished with Illumina MiSeq reads sourced from an axenic L. fusiformis culture; axenicity was verified via microscopy and rDNA analysis. Ultra-long read sequencing resulted in a 6.42 Mb closed genome assembled as a single contig with no plasmid. Phylogenetic analysis placed L. fusiformis in the Limnospira clade; some Arthrospira were also placed in this clade, suggesting a misclassification of these strains. This work provides a fully closed and accurate reference genome for the economically important type strain, L. fusiformis. We also present a rapid axenicity method to isolate L. fusiformis. These contributions enable future biotechnological development of L. fusiformis by way of genetic engineering.

Drosophila melanogaster Cad99C, the orthologue of human Usher cadherin PCDH15, regulates the length of microvilli.

Actin-based protrusions can form prominent structures on the apical surface of epithelial cells, such as microvilli. Several cytoplasmic factors have been identified that control the dynamics of actin filaments in microvilli. However, it remains unclear whether the plasma membrane participates actively in microvillus formation. In this paper, we analyze the function of Drosophila melanogaster cadherin Cad99C in the microvilli of ovarian follicle cells. Cad99C contributes to eggshell formation and female fertility and is expressed in follicle cells, which produce the eggshells. Cad99C specifically localizes to apical microvilli. Loss of Cad99C function results in shortened and disorganized microvilli, whereas overexpression of Cad99C leads to a dramatic increase of microvillus length. Cad99C that lacks most of the cytoplasmic domain, including potential PDZ domain-binding sites, still promotes excessive microvillus outgrowth, suggesting that the amount of the extracellular domain determines microvillus length. This study reveals Cad99C as a critical regulator of microvillus length, the first example of a transmembrane protein that is involved in this process.

Environmental Drivers for Persistence of Escherichia coli and Salmonella in Manure-Amended Soils: A Meta-Analysis.

ABSTRACT: Application of organic amendments to agricultural land improves soil quality and provides nutrients essential for plant growth; however, they are also a reservoir for zoonotic pathogens whose presence poses a significant risk to public health. The persistence of bacteria in manure-amended soil, and differences in manure handling practices, are important issues from a food safety perspective. The primary objective of this study was to quantitatively summarize the variations in the rate of decline of Escherichia coli and Salmonella spp. in manure-amended soil under laboratory and field conditions, and to assess the impact of environmental factors. Available literature data on persistence of E. coli and Salmonella spp. in manure-amended soil from 42 primary research studies were extracted and statistically analyzed using a mixed-effect regression model. The results indicated that temperature (soil and air combined) was the most prominent factor affecting persistence of both E. coli and Salmonella spp. under laboratory conditions (P < 0.001), and of E. coli under field conditions (P < 0.05). The time required for a log reduction of E. coli under field conditions was significantly higher at low temperature (0 to 10°C) than at high temperature (greater than 20°C) (P < 0.05). In addition, application method was identified as a significant factor, with manure incorporation to soil inducing longer survival compared with surface application by approximately 1.2 times. The significant variation observed among primary research studies of bacterial persistence has highlighted that mitigation strategies associated with the use of manures in fresh produce production need to be improved by addressing factors such as climate, soil management, application method, and initial microbial levels. These findings may be used to support guidelines establishing exclusion periods between manure fertilization and the grazing or harvesting of crops, and may be useful for the generation of quantitative microbial risk models for fresh produce.

Tumor mutational burden in lung cancer: a systematic literature review.

Purpose: To assess the association of tumor mutational burden (TMB) with clinical outcomes, other biomarkers and patient/disease characteristics in patients receiving therapy for lung cancer. Results: In total, 4,303 publications were identified; 81 publications were included. The majority of publications assessing clinical efficacy of immunotherapy reported an association with high TMB, particularly when assessing progression-free survival and objective response rate. High TMB was consistently associated with TP53 alterations, and negatively associated with EGFR mutations. High TMB was also associated with smoking, squamous cell non-small cell lung carcinoma, and being male. Methods: A systematic literature review based upon an a priori protocol was conducted following Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and Cochrane methodologies. Searches were conducted in EMBASE, SCOPUS, Ovid MEDLINE®, and Emcare (from January 2012 until April 2018) and in two clinical trial registries. Conference abstracts were identified in EMBASE, and in targeted searches of recent major conference proceedings (from January 2016 until April 2018). Publications reporting data in patients receiving therapy for lung cancer that reported TMB and its association with clinical efficacy, or with other biomarkers or patient/disease characteristics, were included. Results are presented descriptively. Conclusion: This systematic literature review identified several clinical outcomes, biomarkers, and patient/disease characteristics associated with high TMB, and highlights the need for standardized definitions and testing practices. Further studies using standardized methodology are required to inform treatment decisions.

In utero and postnatal VX-770 administration rescues multiorgan disease in a ferret model of cystic fibrosis.

Cystic fibrosis (CF) is a multiorgan disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR). In patients with CF, abnormalities initiate in several organs before birth. However, the long-term impact of these in utero pathologies on disease pathophysiology is unclear. To address this issue, we generated ferrets harboring a VX-770 (ivacaftor)-responsive CFTR G551D mutation. In utero VX-770 administration provided partial protection from developmental pathologies in the pancreas, intestine, and male reproductive tract. Homozygous CFTR G551D/G551D animals showed the greatest VX-770-mediated protection from these pathologies. Sustained postnatal VX-770 administration led to improved pancreatic exocrine function, glucose tolerance, growth and survival, and to reduced mucus accumulation and bacterial infections in the lung. VX-770 withdrawal at any age reestablished disease, with the most rapid onset of morbidity occurring when withdrawal was initiated during the first 2 weeks after birth. The results suggest that CFTR is important for establishing organ function early in life. Moreover, this ferret model provides proof of concept for in utero pharmacologic correction of genetic disease and offers opportunities for understanding CF pathogenesis and improving treatment.

PD1/PDL1 inhibitors for the treatment of advanced urothelial bladder cancer.

INTRODUCTION: Until recently, systemic chemotherapy was the only option for treating bladder cancer and outcomes remained dismal. After a long gap of no progress for 40 years, immuno-therapy with checkpoint inhibitors (PDL1 and PD1) has revolutionized the treatment paradigm of bladder cancer, with five approved agents to treat platinum-refractory bladder cancer since the first approval of atezolizumab in May 2016. METHODS: This review summarizes the most recent data on approved checkpoint inhibitors currently used in management of advanced bladder cancer. Early- and late-phase trials of the five checkpoint inhibitors (pembrolizumab, nivolumab, atezolizumab, durvalumab, and avelumab) in advanced bladder cancer are reviewed in detail. This review also describes the potential application of PD1/PDL1 inhibitors in adjuvant and neoadjuvant settings and non-muscle-invasive bladder cancer, as well as with radiation in muscle-invasive bladder cancer treatment. The role of PDL1 and tumor-mutation burden and clinical considerations in choosing a particular immunotherapy are also discussed. RESULTS: The approved checkpoint inhibitors (PD1 and PDL1 inhibitors) have similar efficacy and safety profiles in metastatic platinum-refractory bladder cancer, but they vary in dose and frequency and cost burden. However, only pembrolizumab has shown superiority over standard chemotherapy in a randomized Phase III setting so far. In addition, in the first-line setting for cisplatin-ineligible patients, both pembrolizumab and atezolizumab are US Food and Drug Administration-approved and well tolerated. There is a lack of consensus on the utility of testing for PDL1 as a predictive biomarker, as patients with no PDL1 expression also derive some clinical benefit. Tumor-mutation burden is another predictive biomarker, but needs further validation. CONCLUSION: Immunotherapy has offered a glimmer of hope to patients with bladder cancer. The current landscape is rapidly evolving, with novel immunotherapy-combination trials to improve outcomes further and evaluate predictive biomarkers to help identify patients most likely to benefit from such therapies.

Association between in-hospital guideline adherence and postdischarge major adverse outcomes of patients with acute coronary syndrome in Vietnam: a prospective cohort study.

OBJECTIVE: We aimed to determine the association between physician adherence to prescribing guideline-recommended medications during hospitalisation and 6-month major adverse outcomes of patients with acute coronary syndrome in Vietnam. DESIGN: Prospective cohort study. SETTING: The study was carried out in two public hospitals in Vietnam between January and October 2015. Patients were followed for 6 months after discharge. PARTICIPANTS: Patients who survived during hospitalisation with a discharge diagnosis of acute coronary syndrome and who were eligible for receiving at least one of the four guideline-recommended medications. EXPOSURES: Guideline adherence was defined as prescribing all guideline-recommended medications at both hospital admission and discharge for eligible patients. Medications were antiplatelet agents, beta-blockers, ACE inhibitors or angiotensin II receptor blockers and statins. MAIN OUTCOME MEASURE: Six-month major adverse outcomes were defined as all-cause mortality or hospital readmission due to cardiovascular causes occurring during 6 months after discharge. Cox regression models were used to estimate the association between guideline adherence and 6-month major adverse outcomes. RESULTS: Overall, 512 patients were included. Of those, there were 242 patients (47.3%) in the guideline adherence group and 270 patients (52.3%) in the non-adherence group. The rate of 6-month major adverse outcomes was 30.5%. A 29% reduction in major adverse outcomes at 6 months after discharge was found for patients of the guideline adherence group compared with the non-adherence group (adjusted HR, 0.71; 95% CI, 0.51 to 0.98; p=0.039). Covariates significantly associated with the major adverse outcomes were percutaneous coronary intervention, prior heart failure and renal insufficiency. CONCLUSIONS: In-hospital guideline adherence was associated with a significant decrease in major adverse outcomes up to 6 months after discharge. It supports the need for improving adherence to guidelines in hospital practice in low-income and middle-income countries like Vietnam.

Five new, microendemic Asian Leaf-litter Frogs (Leptolalax) from the southern Annamite mountains, Vietnam.

The Leptolalax applebyi group of Asian leaf-litter frogs currently comprises four species of particularly small-bodied (<40 mm SVL) species distributed in the Central Highlands of Vietnam and northeastern Cambodia. In addition to their small size, the group is characterized by their morphological and genetic similarities, as well as their breeding habitat at headwaters of small mountain streams and seeps. A recent study suggested that at least two-thirds of the diversity of the group remained hidden within morphologically cryptic lineages. We expand upon the molecular, morphometric, and acoustic data and formally delineate and describe five of these lineages as distinct species: Leptolalax ardens sp. nov., Leptolalax kalonensis sp. nov., Leptolalax pallidus sp. nov., Leptolalax maculosus sp. nov., and Leptolalax tadungensis sp. nov. Due to habitat loss, the current ranges of these species are likely to be a fraction of their historical extent and under continued threat from deforestation.

Undiagnosed cryptic diversity in small, microendemic frogs (Leptolalax) from the Central Highlands of Vietnam.

A major obstacle in prioritizing species or habitats for conservation is the degree of unrecognized diversity hidden within complexes of morphologically similar, "cryptic" species. Given that amphibians are one of the most threatened groups of organisms on the planet, our inability to diagnose their true diversity is likely to have significant conservation consequences. This is particularly true in areas undergoing rapid deforestation, such as Southeast Asia. The Southeast Asian genus Leptolalax is a group of small-bodied, morphologically conserved frogs that inhabit the forest-floor. We examined a particularly small-bodied and morphologically conserved subset, the Leptolalax applebyi group, using a combination of molecular, morphometric, and acoustic data to identify previously unknown diversity within. In order to predict the geographic distribution of the group, estimate the effects of habitat loss and assess the degree of habitat protection, we used our locality data to perform ecological niche modelling using MaxEnt. Molecular (mtDNA and nuDNA), acoustic and subtle morphometric differences revealed a significant underestimation of diversity in the L. applebyi group; at least two-thirds of the diversity may be unrecognised. Patterns of diversification and microendemism in the group appear driven by limited dispersal, likely due to their small body size, with several lineages restricted to watershed basins. The L. applebyi group is predicted to have historically occurred over a large area of the Central Highlands of Vietnam, a considerable portion of which has already been deforested. Less than a quarter of the remaining forest predicted to be suitable for the group falls within current protected areas. The predicted distribution of the L. applebyi group extends into unsurveyed watershed basins, each potentially containing unsampled diversity, some of which may have already been lost due to deforestation. Current estimates of amphibian diversity based on morphology alone are misleading, and accurate alpha taxonomy is essential to accurately prioritize conservation efforts.

Identification of novel HSP90α/ß isoform selective inhibitors using structure-based drug design. demonstration of potential utility in treating CNS disorders such as Huntington's disease.

A structure-based drug design strategy was used to optimize a novel benzolactam series of HSP90α/ß inhibitors to achieve >1000-fold selectivity versus the HSP90 endoplasmic reticulum and mitochondrial isoforms (GRP94 and TRAP1, respectively). Selective HSP90α/ß inhibitors were found to be equipotent to pan-HSP90 inhibitors in promoting the clearance of mutant huntingtin protein (mHtt) in vitro, however with less cellular toxicity. Improved tolerability profiles may enable the use of HSP90α/ß selective inhibitors in treating chronic neurodegenerative indications such as Huntington's disease (HD). A potent, selective, orally available HSP90α/ß inhibitor was identified (compound 31) that crosses the blood-brain barrier. Compound 31 demonstrated proof of concept by successfully reducing brain Htt levels following oral dosing in rats.

Dimethyl sulfoxide-sodium bicarbonate infusion for palliative care and pain relief in patients with metastatic prostate cancer.

Prostate cancer (adenocarcinoma of the prostate) is the most widespread cancer in men. It causes significant suffering and mortality due to metastatic disease. The main therapy for metastatic prostate cancer (MPC) includes androgen manipulation, chemotherapy, and radiotherapy and/or radioisotopes. However, these therapeutic approaches are considered palliative at this stage, and their significant side effects can cause further decline in patients' quality of life and increase non-cancer-related morbidity/mortality. In this study, the authors have used the infusion of dimethyl sulfoxide-sodium bicarbonate (DMSO-SB) to treat 18 patients with MPC. The 90-day follow-up of the patients having undergone the proposed therapeutic regimen showed significant improvement in clinical symptoms, blood and biochemistry tests, and quality of life. There were no major side effects from the treatment. In searching for new and better methods for palliative treatment and pain relief, this study strongly suggested therapy with DMSO-SB infusions could provide a rational alternative to conventional treatment for patients with MPC.

Dimethyl sulfoxide and sodium bicarbonate in the treatment of refractory cancer pain.

Pain is a major concern of cancer patients and a significant problem for therapy. Pain can become a predominant symptom in advanced cancers. In this open-label clinical study, the authors have treated 26 cancer patients who have been declared as terminal without the option of conventional treatment. These patients suffered from high levels of pain that was poorly managed by all available interventional approaches recommended by World Health Organization (WHO) guideline. The results indicate that intravenous infusion of dimethyl sulfoxide (DMSO) and sodium bicarbonate (SB) solution can be a viable, effective, and safe treatment for refractory pain in cancer patients. These patients had pain due to the disease progression and complication of chemotherapy and radiation. Moreover, the preliminary clinical outcome of 96-day follow-up suggests that the application of DMSO and SB solution intravenously could lead to better quality of life for patients with nontreatable terminal cancers. The data of this clinical observation indicates that further research and application of the DMSO and SB combination may help the development of an effective, safe, and inexpensive therapy to manage cancer pain.