Biochemical and metabolic characterization of a G6PC2 inhibitor.

Three glucose-6-phosphatase catalytic subunits, that hydrolyze glucose-6-phosphate (G6P) to glucose and inorganic phosphate, have been identified, designated G6PC1-3, but only G6PC1 and G6PC2 have been implicated in the regulation of fasting blood glucose (FBG). Elevated FBG has been associated with multiple adverse clinical outcomes, including increased risk for type 2 diabetes and various cancers. Therefore, G6PC1 and G6PC2 inhibitors that lower FBG may be of prophylactic value for the prevention of multiple conditions. The studies described here characterize a G6PC2 inhibitor, designated VU0945627, previously identified as Compound 3. We show that VU0945627 preferentially inhibits human G6PC2 versus human G6PC1 but activates human G6PC3. VU0945627 is a mixed G6PC2 inhibitor, increasing the Km but reducing the Vmax for G6P hydrolysis. PyRx virtual docking to an AlphaFold2-derived G6PC2 structural model suggests VU0945627 binds two sites in human G6PC2. Mutation of residues in these sites reduces the inhibitory effect of VU0945627. VU0945627 does not inhibit mouse G6PC2 despite its 84% sequence identity with human G6PC2. Mutagenesis studies suggest this lack of inhibition of mouse G6PC2 is due, in part, to a change in residue 318 from histidine in human G6PC2 to proline in mouse G6PC2. Surprisingly, VU0945627 still inhibited glucose cycling in the mouse islet-derived ßTC-3 cell line. Studies using intact mouse liver microsomes and PyRx docking suggest that this observation can be explained by an ability of VU0945627 to also inhibit the G6P transporter SLC37A4. These data will inform future computational modeling studies designed to identify G6PC isoform-specific inhibitors.

Ten-Year Outcomes: Community Health Center/Academic Medicine Partnership for Rural Family Medicine Training.

BACKGROUND AND OBJECTIVES: The widening gap between urban and rural health outcomes is exacerbated by physician shortages that disproportionately affect rural communities. Rural residencies are an effective mechanism to increase physician placement in rural and medically underserved areas yet are limited in number due to funding. Community health center/academic medicine partnerships (CHAMPs) can serve as a collaborative framework for expansion of academic primary care residencies outside of traditional funding models. This report describes 10-year outcomes of a rural training pathway developed as part of a CHAMP collaboration. METHODS: Using data from internal registries and public sources, our retrospective study examined demographic and postgraduation practice characteristics for rural pathway graduates. We identified the rates of postgraduation placement in rural (Federal Office of Rural Health Policy grant-eligible) and federally designated Medically Underserved Areas/Populations (MUA/Ps). We assessed current placement for graduates >3 years from program completion. RESULTS: Over a 10-year period, 25 trainees graduated from the two residency expansion sites. Immediately postgraduation, 84% (21) were in primary care Health Professional Shortage Areas (HPSAs), 80% (20) in MUA/Ps, and 60% (15) in rural locations. Sixteen graduates were >3 years from program completion, including 69% (11) in primary care HPSAs, 69% (11) in MUA/Ps, and 50% (5) in rural locations. CONCLUSIONS: This CHAMP collaboration supported development of a rural pathway that embedded family medicine residents in community health centers and effectively increased placement in rural and MUA/Ps. This report adds to national research on rural workforce development, highlighting the role of academic-community partnerships in expanding rural residency training outside of traditional funding models.

Integrative analysis of pathogenic variants in glucose-6-phosphatase based on an AlphaFold2 model.

Mediating the terminal reaction of gluconeogenesis and glycogenolysis, the integral membrane protein glucose-6-phosphate catalytic subunit 1 (G6PC1) regulates hepatic glucose production by catalyzing hydrolysis of glucose-6-phosphate (G6P) within the lumen of the endoplasmic reticulum. Consistent with its vital contribution to glucose homeostasis, inactivating mutations in G6PC1 causes glycogen storage disease (GSD) type 1a characterized by hepatomegaly and severe hypoglycemia. Despite its physiological importance, the structural basis of G6P binding to G6PC1 and the molecular disruptions induced by missense mutations within the active site that give rise to GSD type 1a are unknown. In this study, we determine the atomic interactions governing G6P binding as well as explore the perturbations imposed by disease-linked missense variants by subjecting an AlphaFold2 G6PC1 structural model to molecular dynamics simulations and in silico predictions of thermodynamic stability validated with robust in vitro and in situ biochemical assays. We identify a collection of side chains, including conserved residues from the signature phosphatidic acid phosphatase motif, that contribute to a hydrogen bonding and van der Waals network stabilizing G6P in the active site. The introduction of GSD type 1a mutations modified the thermodynamic landscape, altered side chain packing and substrate-binding interactions, and induced trapping of catalytic intermediates. Our results, which corroborate the high quality of the AF2 model as a guide for experimental design and to interpret outcomes, not only confirm the active-site structural organization but also identify previously unobserved mechanistic contributions of catalytic and noncatalytic side chains.

Pharmacists Colocated With Primary Care Physicians: Understanding Delivery of Interprofessional Primary Care.

BACKGROUND: While evidence supports interprofessional primary care models that include pharmacists, the extent to which pharmacists are working in primary care and the factors associated with colocation is unknown. OBJECTIVES: This study aimed to analyze the physical colocation of pharmacists with primary care providers (PCPs) and examine predictors associated with colocation. RESEARCH DESIGN: This is a retrospective cross-sectional study of pharmacists and PCPs with individual National Provider Identifiers in the National Plan and Provider Enumeration System's database. Pharmacist and PCP practice addresses of the health care professionals were geocoded, and distances less than 0.1 miles were considered physically colocated. SUBJECTS: In all, 502,373 physicians and 221,534 pharmacists were included. RESULTS: When excluding hospital-based pharmacists, 1 in 10 (11%) pharmacists were colocated with a PCP. Pharmacists in urban settings were more likely to be colocated than those in rural areas (OR=1.32, CI: 1.26-1.38). Counties with the highest proportion of licensed pharmacists per 100,000 people in the county had higher colocation (OR=1.38, CI: 1.32-1.45). Colocation was significantly higher in states with an expanded scope of practice (OR 1.37, CI: 1.32-1.42) and those that have expanded Medicaid (OR 1.07, CI: 1.03-1.11). Colocated pharmacists more commonly worked in larger physician practices. CONCLUSION: Although including pharmacists on primary care teams improves clinical outcomes, reduces health care costs, and enhances patient and provider experience, colocation appears to be unevenly dispersed across the United States, with lower rates in rural areas. As the integration of pharmacists in primary care continues to expand, knowing the prevalence and facilitators of growth will be helpful to policymakers, researchers, and clinical administrators.

Identification of structural motifs critical for human G6PC2 function informed by sequence analysis and an AlphaFold2-predicted model.

G6PC2 encodes a glucose-6-phosphatase (G6Pase) catalytic subunit, primarily expressed in pancreatic islet ß cells, which modulates the sensitivity of insulin secretion to glucose and thereby regulates fasting blood glucose (FBG). Mutational analyses were conducted to validate an AlphaFold2 (AF2)-predicted structure of human G6PC2 in conjunction with a novel method to solubilize and purify human G6PC2 from a heterologous expression system. These analyses show that residues forming a predicted intramolecular disulfide bond are essential for G6PC2 expression and that residues forming part of a type 2 phosphatidic acid phosphatase (PAP2) motif are critical for enzyme activity. Additional mutagenesis shows that residues forming a predicted substrate cavity modulate enzyme activity and substrate specificity and residues forming a putative cholesterol recognition amino acid consensus (CRAC) motif influence protein expression or enzyme activity. This CRAC motif begins at residue 219, the site of a common G6PC2 non-synonymous single-nucleotide polymorphism (SNP), rs492594 (Val219Leu), though the functional impact of this SNP is disputed. In microsomal membrane preparations, the L219 variant has greater activity than the V219 variant, but this difference disappears when G6PC2 is purified in detergent micelles. We hypothesize that this was due to a differential association of the two variants with cholesterol. This concept was supported by the observation that the addition of cholesteryl hemi-succinate to the purified enzymes decreased the Vmax of the V219 and L219 variants ∼8-fold and ∼3 fold, respectively. We anticipate that these observations should support the rational development of G6PC2 inhibitors designed to lower FBG.

G6PC1 and G6PC2 influence G6P flux but not HSD11B1 activity.

In the endoplasmic reticulum (ER) lumen, glucose-6-phosphatase catalytic subunit 1 and 2 (G6PC1; G6PC2) hydrolyze glucose-6-phosphate (G6P) to glucose and inorganic phosphate whereas hexose-6-phosphate dehydrogenase (H6PD) hydrolyzes G6P to 6-phosphogluconate (6PG) in a reaction that generates NADPH. 11ß-hydroxysteroid dehydrogenase type 1 (HSD11B1) utilizes this NADPH to convert inactive cortisone to cortisol. HSD11B1 inhibitors improve insulin sensitivity whereas G6PC inhibitors are predicted to lower fasting blood glucose (FBG). This study investigated whether G6PC1 and G6PC2 influence G6P flux through H6PD and vice versa. Using a novel transcriptional assay that utilizes separate fusion genes to quantitate glucocorticoid and glucose signaling, we show that overexpression of H6PD and HSD11B1 in the islet-derived 832/13 cell line activated glucocorticoid-stimulated fusion gene expression. Overexpression of HSD11B1 blunted glucose-stimulated fusion gene expression independently of altered G6P flux. While overexpression of G6PC1 and G6PC2 blunted glucose-stimulated fusion gene expression, it had minimal effect on glucocorticoid-stimulated fusion gene expression. In the liver-derived HepG2 cell line, overexpression of H6PD and HSD11B1 activated glucocorticoid-stimulated fusion gene expression but overexpression of G6PC1 and G6PC2 had no effect. In rodents, HSD11B1 converts 11-dehydrocorticosterone (11-DHC) to corticosterone. Studies in wild-type and G6pc2 knockout mice treated with 11-DHC for 5 weeks reveal metabolic changes unaffected by the absence of G6PC2. These data suggest that HSD11B1 activity is not significantly affected by the presence or absence of G6PC1 or G6PC2. As such, G6PC1 and G6PC2 inhibitors are predicted to have beneficial effects by reducing FBG without causing a deleterious increase in glucocorticoid signaling.

An Enhancer Within Abcb11 Regulates G6pc2 in C57BL/6 Mouse Pancreatic Islets.

G6PC2 is predominantly expressed in pancreatic islet ß-cells where it encodes a glucose-6-phosphatase catalytic subunit that modulates the sensitivity of insulin secretion to glucose by opposing the action of glucokinase, thereby regulating fasting blood glucose (FBG). Prior studies have shown that the G6pc2 promoter alone is unable to confer sustained islet-specific gene expression in mice, suggesting the existence of distal enhancers that regulate G6pc2 expression. Using information from both mice and humans and knowledge that single nucleotide polymorphisms (SNPs) both within and near G6PC2 are associated with variations in FBG in humans, we identified several putative enhancers 3' of G6pc2. One region, herein referred to as enhancer I, resides in the 25th intron of Abcb11 and binds multiple islet-enriched transcription factors. CRISPR-mediated deletion of enhancer I in C57BL/6 mice had selective effects on the expression of genes near the G6pc2 locus. In isolated islets, G6pc2 and Spc25 expression were reduced ∼50%, and Gm13613 expression was abolished, whereas Cers6 and nostrin expression were unaffected. This partial reduction in G6pc2 expression enhanced islet insulin secretion at basal glucose concentrations but did not affect FBG or glucose tolerance in vivo, consistent with the absence of a phenotype in G6pc2 heterozygous C57BL/6 mice.

The Distribution of Additional Residency Slots to Rural and Underserved Areas.

This study analyzes data from the Centers for Medicare & Medicaid Services to identify whether new residency training slots went to rural and underserved areas with the greatest need.

Postgraduate education to expand access to dental care: A roadmap to community dental program development.

Evidence indicates an increasing shortage of dentists in communities across the United States with potentially significant implications for oral health, as well as overall health and well-being. One strategy to increase access to dental care in rural and underserved communities is community-based postgraduate dental training. However, developing new dental programs requires navigating complex accreditation, financial and community governance, among other, barriers. The Roadmap for Teaching Health Center Dental Program Development presents a framework that guides institutions through the successive steps of developing new postgraduate training programs from identification of need to ultimate maintenance and sustainability. The tool assists programs in anticipating and understanding requirements, reducing time, expense, and uncertainty. While the framework was developed for community-based programs, the steps are applicable to postgraduate programs sponsored by academic institutions as well.

Molecular mechanisms of catalytic inhibition for active site mutations in glucose-6-phosphatase catalytic subunit 1 linked to glycogen storage disease.

Mediating the terminal reaction of gluconeogenesis and glycogenolysis, the integral membrane protein G6PC1 regulates hepatic glucose production by catalyzing hydrolysis of glucose-6-phosphate (G6P) within the lumen of the endoplasmic reticulum. Consistent with its vital contribution to glucose homeostasis, inactivating mutations in G6PC1 cause glycogen storage disease (GSD) type 1a characterized by hepatomegaly and severe hypoglycemia. Despite its physiological importance, the structural basis of G6P binding to G6PC1 and the molecular disruptions induced by missense mutations within the active site that give rise to GSD type 1a are unknown. Exploiting a computational model of G6PC1 derived from the groundbreaking structure prediction algorithm AlphaFold2 (AF2), we combine molecular dynamics (MD) simulations and computational predictions of thermodynamic stability with a robust in vitro screening platform to define the atomic interactions governing G6P binding as well as explore the energetic perturbations imposed by disease-linked variants. We identify a collection of side chains, including conserved residues from the signature phosphatidic acid phosphatase motif, that contribute to a hydrogen bonding and van der Waals network stabilizing G6P in the active site. Introduction of GSD type 1a mutations into the G6PC1 sequence elicits changes in G6P binding energy, thermostability and structural properties, suggesting multiple pathways of catalytic impairment. Our results, which corroborate the high quality of the AF2 model as a guide for experimental design and to interpret outcomes, not only confirm active site structural organization but also suggest novel mechanistic contributions of catalytic and non-catalytic side chains.

Bolstering the rural physician workforce in underserved communities: Are Rural Residency Planning and Development Programs finding the sweet spot?

PURPOSE: The purpose of this study is to describe the characteristics of Rural Residency Planning and Development (RRPD) Programs, compare the characteristics of counties with and without RRPD programs, and identify rural places where future RRPD programs could be developed. METHODS: The study sample comprised 67 rural sites training residents in 40 counties in 24 US states. Descriptive statistics were used to describe RRPD programs and logistic regression to predict the probability of a county being an RRPD site as a function of population, primary care physicians (PCP) per 10,000 population, and the social vulnerability index (SVI) compared to a control sample of nonmetro counties without RRPD sites. FINDINGS: Most RRPD grantees (78%) were family medicine programs affiliated with medical schools (97%). RRPD counties were more populous (P<.01), had a higher population density (P<.05), and a higher percent of the non-White or Hispanic population (P = .05) compared to non-RRPD counties. Both higher population (P<.001) and PCP ratio (P = .046) were strong predictors, while SVI (P = .07) was a weak predictor of being an RRPD county. CONCLUSIONS: RRPD sites appear to represent a "sweet spot" of rural counties that have the population and physician supply to support a training program but also are relatively more socially vulnerable with high-need populations. Additional counties fitting this "sweet spot" could be targeted for funding to address health disparities and health workforce maldistribution.

Responding to One Rural Community's Primary Care Needs: Investing in Workforce and Broad Community Stakeholder Engagement.

Lack of access to high-quality primary care has been shown to contribute to urban-rural health disparities. We describe a model in which an academic health system made targeted primary care investments to address rural health disparities while building the health workforce to ensure sustainability.

New Opportunities for Expanding Rural Graduate Medical Education to Improve Rural Health Outcomes: Implications of the Consolidated Appropriations Act of 2021.

Evidence shows that those living in rural communities experience consistently worse health outcomes than their urban and suburban counterparts. One proven strategy to address this disparity is to increase the physician supply in rural areas through graduate medical education (GME) training. However, rural hospitals have faced challenges developing training programs in these underserved areas, largely due to inadequate federal funding for rural GME. The Consolidated Appropriations Act of 2021 (CAA) contains multiple provisions that seek to address disparities in Medicare funding for rural GME, including funding for an increase in rural GME positions or "slots" (Section 126), expansion of rural training opportunities (Section 127), and relief for hospitals that have very low resident payments and/or caps (Section 131). In this Invited Commentary, the authors describe historical factors that have impeded the growth of training programs in rural areas, summarize the implications of each CAA provision for rural GME, and provide guidance for institutions seeking to avail themselves of the opportunities presented by the CAA. These policy changes create new opportunities for rural hospitals and partnering urban medical centers to bolster rural GME training, and consequently the physician workforce in underserved communities.

A proactive medical necessity review program reduces revenue loss associated with outpatient medical benefit drugs.

PURPOSE: A common denial trend that occurs with "outpatient medical benefit drugs" (ie, medications covered by a medical benefit plan and administered in an outpatient visit) is payers not requiring or permitting prior authorization (PA) proactively, yet denying the drug after administration for medical necessity. In this situation, a preemptive strategy of complying with payer-mandated requirements is critical for revenue protection. To address this need, our institution incorporated a medical necessity review into its existing closed-loop, pharmacy-managed precertification and denials management program. SUMMARY: Referrals for targeted payers and high-cost medical benefit drugs not eligible for PA and deemed high risk for denial were incorporated into the review. Payer medical policies were evaluated and clinical documentation assessed to confirm alignment. This descriptive report outlines the medical necessity workflow as a component of the larger precertification process, details the decision-making process when performing the review, and delineates the roles and responsibilities for involved team members. A total of 526 drug orders were evaluated from September 2018 to August 2019, with 146 interventions completed. Of the 761 individual claims affected by proactive medical necessity review, 99.2% resulted in payment and less than 1% resulted in revenue loss, safeguarding more than $5.3 million in annual institutional drug reimbursement. At the time of analysis, there were only 3 cases of revenue loss. CONCLUSION: Our institution's pharmacy-managed medical necessity review program for high-cost outpatient drugs safeguards reimbursement for therapies not eligible for payer PA. It is a revenue cycle best practice that can be replicated at other institutions.

Advancing Pharmacy Practice through an Innovative Ambulatory Care Transitions Program at an Academic Medical Center.

Hospital readmissions are common and often preventable, leading to unnecessary burden on patients, families, and the health care system. The purpose of this descriptive communication is to share the impact of an interdisciplinary, outpatient clinic-based care transition intervention on clinical, organizational, and financial outcomes. Compared to usual care, the care transition intervention decreased the median time to Internal Medicine Clinic (IMC) or any clinic follow-up visit by 5 and 4 days, respectively. By including a pharmacist in the hospital follow-up visit, the program significantly reduced all-cause 30-day hospital readmission rates (9% versus 26% in usual care) and the composite endpoint of 30-day health care utilization, which is defined as readmission and emergency department (ED) rates (19% versus 44% usual care). Over the course of one year, this program can prevent 102 30-day hospital readmissions with an estimated cost reduction of $1,113,000 per year. The pharmacist at the IMC collaborated with the Family Medicine Clinic (FMC) pharmacist to standardize practices. In the FMC, the hospital readmission rate was 6.5% for patients seen by a clinic-based pharmacist within 30 days of discharge compared to 20% for those not seen by a pharmacist. This transitions intervention demonstrated a consistent and recognizable contribution from pharmacists providing direct patient care and practicing in the ambulatory care primary care settings that has been replicated across clinics at our academic medical center.

Vitamin D Supplementation for Extraskeletal Indications in Older Persons.

Low levels of vitamin D have been implicated in a wide variety of conditions highly prevalent in the geriatric population, including fractures, functional limitations, cancer, cardiovascular disease, and depression. Vitamin D supplementation is often considered integral to the prevention of falls and fractures in the setting of osteoporosis. For other conditions, however, consensus is lacking, and the clinician may struggle to balance competing recommendations around screening, supplementation, and monitoring. This review seeks to provide an overview of the available evidence on the use of vitamin D supplementation to ameliorate sarcopenia, enhance cognition, treat depression, prevent cancer, and reduce mortality-outcomes that are common concerns in the geriatric population for which the merits of treatment are not always certain. Evidence suggests vitamin D supplementation may decrease mortality. Therefore, it may be reasonable to prescribe routine supplementation with oral cholecalciferol 800 to 1000 IU daily to all patients aged ≥65 years who do not have a contraindication. No screening or monitoring would be recommended for this population. We additionally recommend the use of oral cholecalciferol over ergocalciferol for any routine supplementation as this benefit was only observed with cholecalciferol. For patients with depression or cognitive disorders, we recommend screening for vitamin D deficiency, treating with oral cholecalciferol if present, and monitoring periodically to target a level of >30 ng/mL as an adjunct to usual care. The level of evidence certainly would not justify the use of vitamin D in place of more evidence-based therapies, but given the burden of these conditions in the geriatric population, we believe the potential benefit justifies the minimal risk.