PRMT5 inhibition shows in vitro efficacy against H3K27M-altered diffuse midline glioma, but does not extend survival in vivo.

H3K27-altered Diffuse Midline Glioma (DMG) is a universally fatal paediatric brainstem tumour. The prevalent driver mutation H3K27M creates a unique epigenetic landscape that may also establish therapeutic vulnerabilities to epigenetic inhibitors. However, while HDAC, EZH2 and BET inhibitors have proven somewhat effective in pre-clinical models, none have translated into clinical benefit due to either poor blood-brain barrier penetration, lack of efficacy or toxicity. Thus, there remains an urgent need for new DMG treatments. Here, we performed wider screening of an epigenetic inhibitor library and identified inhibitors of protein arginine methyltransferases (PRMTs) among the top hits reducing DMG cell viability. Two of the most effective inhibitors, LLY-283 and GSK591, were targeted against PRMT5 using distinct binding mechanisms and reduced the viability of a subset of DMG cells expressing wild-type TP53 and mutant ACVR1. RNA-sequencing and phenotypic analyses revealed that LLY-283 could reduce the viability, clonogenicity and invasion of DMG cells in vitro, representing three clinically important phenotypes, but failed to prolong survival in an orthotopic xenograft model. Together, these data show the challenges of DMG treatment and highlight PRMT5 inhibitors for consideration in future studies of combination treatments.

In Vitro Human Haematopoietic Stem Cell Expansion and Differentiation.

The haematopoietic system plays an essential role in our health and survival. It is comprised of a range of mature blood and immune cell types, including oxygen-carrying erythrocytes, platelet-producing megakaryocytes and infection-fighting myeloid and lymphoid cells. Self-renewing multipotent haematopoietic stem cells (HSCs) and a range of intermediate haematopoietic progenitor cell types differentiate into these mature cell types to continuously support haematopoietic system homeostasis throughout life. This process of haematopoiesis is tightly regulated in vivo and primarily takes place in the bone marrow. Over the years, a range of in vitro culture systems have been developed, either to expand haematopoietic stem and progenitor cells or to differentiate them into the various haematopoietic lineages, based on the use of recombinant cytokines, co-culture systems and/or small molecules. These approaches provide important tractable models to study human haematopoiesis in vitro. Additionally, haematopoietic cell culture systems are being developed and clinical tested as a source of cell products for transplantation and transfusion medicine. This review discusses the in vitro culture protocols for human HSC expansion and differentiation, and summarises the key factors involved in these biological processes.

Comparing Kidney Health Outcomes in Children, Adolescents, and Adults With Focal Segmental Glomerulosclerosis.

Importance: Focal segmental glomerulosclerosis (FSGS) is a common cause of end-stage kidney disease (ESKD) across the lifespan. While 10% to 15% of children and 3% of adults who develop ESKD have FSGS, it remains uncertain whether the natural history differs in pediatric vs adult patients, and this uncertainty contributes to the exclusion of children and adolescents in clinical trials. Objective: To examine whether there are differences in the kidney health outcomes among children, adolescents, and adults with FSGS. Design, Setting, and Participants: This cohort study used pooled and parallel analyses, completed July 5, 2022, from 3 complimentary data sources: (1) Nephrotic Syndrome Rare Disease Clinical Research Network (NEPTUNE); (2) FSGS clinical trial (FSGS-CT); and (3) Kidney Research Network (KRN). NEPTUNE is a multicenter US/Canada cohort study; FSGS-CT is a multicenter US/Canada clinical trial; and KRN is a multicenter US electronic health record-based registry from academic and community nephrology practices. NEPTUNE included 166 patients with incident FSGS enrolled at first kidney biopsy; FSGS-CT included 132 patients with steroid-resistant FSGS randomized to cyclosporine vs dexamethasone with mycophenolate; and KRN included 184 patients with prevalent FSGS. Data were collected from November 2004 to October 2019 and analyzed from October 2020 to July 2022. Exposures: Age: children (age <13 years) vs adolescents (13-17 years) vs adults (≥18 years). Covariates of interest included sex, disease duration, APOL1 genotype, urine protein-to-creatinine ratio, estimated glomerular filtration rate (eGFR), edema, serum albumin, and immunosuppressive therapy. Main Outcomes and Measures: ESKD, composite outcome of ESKD or 40% decline in eGFR, and complete and/or partial remission of proteinuria. Results: The study included 127 (26%) children, 102 (21%) adolescents, and 253 (52%) adults, including 215 (45%) female participants and 138 (29%) who identified as Black, 98 (20%) who identified as Hispanic, and 275 (57%) who identified as White. Overall, the median time to ESKD was 11.9 years (IQR, 5.2-19.1 years). There was no difference in ESKD risk among children vs adults (hazard ratio [HR], 0.67; 95% CI, 0.43-1.03) or adolescents vs adults (HR, 0.85; 95% CI, 0.52-1.36). The median time to the composite end point was 5.7 years (IQR 1.6-15.2 years), with hazard ratio estimates for children vs adults of 1.12 (95% CI, 0.83-1.52) and adolescents vs adults of 1.06 (95% CI, 0.75-1.50). Conclusions and Relevance: In this study, the association of FSGS with kidney survival and functional outcomes was comparable at all ages.

Niche space of corals along the Florida reef tract.

Over the last three decades corals have declined precipitously in the Florida Keys. Their population decline has prompted restoration effort. Yet, little effort has been invested in understanding the contemporary niche spaces of coral species, which could assist in prioritizing conservation habitats. We sought to predict the probability of occurrence of 23 coral species, including the critically endangered Acropora cervicornis, using observations at 985 sites from 2011-2015. We ran boosted regression trees to evaluate the relationship between the presence of these corals and eight potential environmental predictors: (i) bathymetry (m), (ii) mean of daily sea surface temperature (SST) (°C), (iii) variance of SST (°C), (iv) range of SST (°C), (v) chlorophyll-a concentration (mg m3), (vi) turbidity (m-1), (vii) wave energy (kJ m-2), and (viii) distance from coast (km). The Marquesas and the lower and upper Florida Keys were predicted to support the most suitable habitats for the 23 coral species examined. A. cervicornis had one of the smallest areas of suitable habitat, which was limited to the lower and upper Florida Keys, the Dry Tortugas, and nearshore Broward-Miami reefs. The best environmental predictors of site occupancy of A. cervicornis were SST range (4-5°C) and turbidity (K490 between 0.15-0.25 m-1). Historically A. cervicornis was reported in clear oligotrophic waters, although the present results find the coral species surviving in nearshore turbid conditions. Nearshore, turbid reefs may shade corals during high-temperature events, and therefore nearshore reefs in south Florida may become important refuges for corals as the ocean temperatures continue to increase.

Heterogeneity of Myc expression in breast cancer exposes pharmacological vulnerabilities revealed through executable mechanistic modeling.

Cells with higher levels of Myc proliferate more rapidly and supercompetitively eliminate neighboring cells. Nonetheless, tumor cells in aggressive breast cancers typically exhibit significant and stable heterogeneity in their Myc levels, which correlates with refractoriness to therapy and poor prognosis. This suggests that Myc heterogeneity confers some selective advantage on breast tumor growth and progression. To investigate this, we created a traceable MMTV-Wnt1-driven in vivo chimeric mammary tumor model comprising an admixture of low-Myc- and reversibly switchable high-Myc-expressing clones. We show that such tumors exhibit interclonal mutualism wherein cells with high-Myc expression facilitate tumor growth by promoting protumorigenic stroma yet concomitantly suppress Wnt expression, which renders them dependent for survival on paracrine Wnt provided by low-Myc-expressing clones. To identify any therapeutic vulnerabilities arising from such interdependency, we modeled Myc/Ras/p53/Wnt signaling cross talk as an executable network for low-Myc, for high-Myc clones, and for the 2 together. This executable mechanistic model replicated the observed interdependence of high-Myc and low-Myc clones and predicted a pharmacological vulnerability to coinhibition of COX2 and MEK. This was confirmed experimentally. Our study illustrates the power of executable models in elucidating mechanisms driving tumor heterogeneity and offers an innovative strategy for identifying combination therapies tailored to the oligoclonal landscape of heterogenous tumors.

Fostering open collaboration in drug development for paediatric brain tumours.

Brain tumours have become the leading cause of child mortality from cancer. Indeed, aggressive brainstem tumours, such as diffuse intrinsic pontine glioma (DIPG), are nearly uniformly fatal. These tumours display a unique set of driver mutations that distinguish them from adult gliomas and define new opportunity for the development of precision medicines. The specific association of ACVR1 mutations with DIPG tumours suggests a direct link to neurodevelopment and highlights the encoded bone morphogenetic protein receptor kinase ALK2 as a promising drug target. Beneficial effects of ALK2 inhibition have now been observed in two different in vivo models of DIPG. Nonetheless, such tumours present a huge challenge for traditional economic models of drug development due to their small market size, high failure rate, tumour location and paediatric population. Moreover, a toolkit of different investigational drugs may be needed to fully address the heterogeneity of these tumours in clinical trials. One new business model is suggested by M4K Pharma, a recent virtual start up that aims to align diffuse academic and industry research into a collaborative open science drug discovery programme. Fostering scientific collaboration may offer hope in rare conditions of dire unmet clinical need and provide an alternative route to affordable medicines.

Tumour cell invasiveness and response to chemotherapeutics in adipocyte invested 3D engineered anisotropic collagen scaffolds.

Breast cancers are highly heterogeneous and their metastatic potential and response to therapeutic drugs is difficult to predict. A tool that could accurately gauge tumour invasiveness and drug response would provide a valuable addition to the oncologist's arsenal. We have developed a 3-dimensional (3D) culture model that recapitulates the stromal environment of breast cancers by generating anisotropic (directional) collagen scaffolds seeded with adipocytes and culturing tumour fragments therein. Analysis of tumour cell invasion in the presence of various therapeutic drugs, by immunofluorescence microscopy coupled with an optical clearing technique, demonstrated the utility of this approach in determining both the rate and capacity of tumour cells to migrate through the stroma while shedding light also on the mode of migration. Furthermore, the response of different murine mammary tumour types to chemotherapeutic drugs could be readily quantified.

Contraception Update: Oral Contraception.

The oral contraceptive pill (OCP) is the most commonly used form of reversible contraception. The two types of OCPs are combination oral contraceptives (COCs), which contain estrogen and progesterone, and progestin-only pills (POPs). Both have failure rates of approximately 7.2% to 9% with typical use, and are safe for most patients. Because estrogen-containing contraceptives can increase the risk of venous thromboembolism, patients with conditions associated with a risk of cardiovascular events should not use COCs. Blood pressure level should be assessed before initiation of oral contraceptives. Noncontraceptive benefits of oral contraceptives include reduced risk of ovarian and endometrial cancers, more favorable bleeding patterns, and improvement in menstruation-related symptoms such as acne, migraine headaches, and premenstrual dysphoric disorder. OCPs can be initiated any time the physician can be reasonably certain that the patient is not pregnant. Extended cycle regimens may be preferred by some patients. After assessing need, physicians should present all methods that can be used safely using a tiered effectiveness approach. High-quality contraceptive counseling includes working collaboratively with patients to find the most effective and acceptable method for them and helping to identify factors that may assist in or hinder their ability to use the method correctly over time.

Contraception Update: Progestin-Only Implants and Injections.

Progestin-only contraception is a popular method of birth control in the United States and worldwide. Progestin-only implants and injections allow patients access to long-term contraception with simple options for reversal or removal. The implant is one of the most effective forms of contraception and there are few contraindications. Manufacturer-led training is required to become certified in insertion and removal. The most common adverse effect of the implant is a change in menstrual bleeding patterns. Little evidence has shown weight gain or decreased bone mineral density with use. The depot medroxyprogesterone acetate (DMPA) injection is used widely and is effective. Adverse effects that may limit use include changes in bleeding patterns and bone mineral density loss, which is reversible after discontinuation. The risk of weight gain with DMPA is greatest in obese adolescents and black patients. There is no significantly increased risk of cancer with either method. Both are safe for use in the postpartum period, during breastfeeding, and immediately after abortion.

Contraception Update: Sterilization.

Female sterilization procedures include postpartum partial salpingectomy via cesarean or minilaparotomy incision, interval laparoscopic procedures, or hysteroscopic placement of microinserts. Rates of failure and serious complications are low and comparable among the various methods. A hysteroscopic procedure requires a 3-month confirmatory hysterosalpingogram before it is considered effective for contraception. Hysteroscopic sterilization has been shown to be associated with a higher reoperation rate than laparoscopic procedures. For male sterilization, vasectomy is a noninvasive and highly effective method. Vasectomy is an outpatient procedure performed under local anesthesia. The procedure requires confirmation of azoospermia with a semen analysis 8 to 16 weeks after the procedure. Patients who are considering sterilization should be counseled about all the available options and the permanent nature of such procedures.

Contraception Update: Intrauterine Devices.

Copper-containing and hormonal intrauterine devices (IUD) are long-acting, highly effective contraceptive methods. They can be used safely by nulliparous patients, adolescents, patients with history of ectopic pregnancy, and patients with risk factors for sexually transmitted infections or a history of pelvic inflammatory disease (PID). These devices can be placed safely immediately postpartum and postabortion and should be inserted when physicians can be reasonably certain that the woman is not pregnant. If a woman with an IUD is shown to be pregnant, the device should be removed if strings are visible. Bleeding and cramping after insertion can be managed with nonsteroidal anti-inflammatory drugs. Perforation is rare, but may require surgical removal of the device. If a woman with an IUD is diagnosed with PID, the device can be left in place and antibiotic treatment initiated.

Altered juvenile fish communities associated with invasive Halophila stipulacea seagrass habitats in the U.S. Virgin Islands.

Caribbean seagrass habitats provide food and protection for reef-associated juvenile fish. The invasive seagrass Halophila stipulacea is rapidly altering these seascapes. Since its arrival in the Caribbean in 2002, H. stipulacea has colonized and displaced native seagrasses, but the function of this invasive seagrass as a juvenile fish habitat remains unknown. To compare diversity, community structure, and abundance of juvenile fish between H. stipulacea and native seagrass beds, fish traps were deployed in four nearshore bays around St. Thomas, U.S. Virgin Islands. Traps were deployed in Frenchman, Lindbergh, and Sprat Bays for 24 h intervals in patches of bare sand, patches of H. stipulacea and patches of the native Caribbean seagrasses Thalassia testudinum and Syringodium filiforme. Traps were then deployed in Brewers Bay for 12 h intervals in stands of H. stipulacea and S. filiforme. Relative and total abundances of juvenile fish, identified at least to family, were compared across treatment habitats for each trap deployment period. The catch from H. stipulacea, compared to native seagrasses, comprised a greater abundance of nocturnal carnivores Lutjanus synagris (family Lutjanidae) and Haemulon flavolineatum (family Haemulidae). Additionally, the herbivore species Sparisoma aurofrenatum (family Labridae) and Acanthurus bahianus (family Acanthuridae) and the diurnal carnivore species Pseudopeneus maculatus (family Mullidae) were relatively scarce in H. stipulacea. The catch from sand was much smaller, compared to vegetated habitats, and comprised only L. synagris, H. flavolineatum, and H. aurolineatum. These results provide evidence of reduced family diversity and altered juvenile fish assemblages in H. stipulacea, driven by an abundance of some nocturnal carnivores and scarcity of herbivores and diurnal carnivores. The findings from the present work underpin the need for further investigation and mitigation of this invasion, particularly where H. stipulacea is driving seascape-alterations of key juvenile fish habitats.

The Role of Community Health Centers in Reducing Racial Disparities in Spatial Access to Primary Care.

INTRODUCTION: Racial minorities are more likely to live in primary care shortage areas. We sought to understand community health centers' (CHCs) role in reducing disparities. METHODS: We surveyed all primary care practices in an urban area, identified low access areas, and examined how CHCs influence spatial accessibility. RESULTS: Census tracts with higher rates of public insurance (≥40% vs <10%, odds ratio [OR] = 31.06, P < .001; 30-39% vs 10%, OR = 7.84, P = 0.001) were more likely to be near a CHC and those with moderate rates of uninsurance (10%-19% vs <10%, OR = 0.42, P = .045) were less likely. Racial composition was not associated with proximity. Tracts close to a CHC were less likely (OR = 0.11, P < .0001) to be in a low access area. This association did not differ based on racial composition. DISCUSSION: Although CHCs were more likely to be in areas with a greater fraction of racial minorities, location was more strongly influenced by public insurance rates. CHCs reduced the likelihood of being in low access areas but the effect did not vary by tract racial composition.

Racial Disparities In Geographic Access To Primary Care In Philadelphia.

Primary care is often thought of as the gateway to improved health outcomes and can lead to more efficient use of health care resources. Because of primary care's cardinal importance, adequate access is an important health policy priority. In densely populated urban areas, spatial access to primary care providers across neighborhoods is poorly understood. We examined spatial variation in primary care access in Philadelphia, Pennsylvania. We calculated ratios of adults per primary care provider for each census tract and included buffer zones based on prespecified drive times around each tract. We found that the average ratio was 1,073; the supply of primary care providers varied widely across census tracts, ranging from 105 to 10,321. We identified six areas of Philadelphia that have much lower spatial accessibility to primary care relative to the rest of the city. After adjustment for sociodemographic and insurance characteristics, the odds of being in a low-access area were twenty-eight times greater for census tracts with a high proportion of African Americans than in tracts with a low proportion of African Americans.

Mice with mutant Inf2 show impaired podocyte and slit diaphragm integrity in response to protamine-induced kidney injury.

Mutations in the INF2 (inverted formin 2) gene, encoding a diaphanous formin family protein that regulates actin cytoskeleton dynamics, cause human focal segmental glomerulosclerosis (FSGS). INF2 interacts directly with certain other mammalian diaphanous formin proteins (mDia) that function as RhoA effector molecules. FSGS-causing INF2 mutations impair these interactions and disrupt the ability of INF2 to regulate Rho/Dia-mediated actin dynamics in vitro. However, the precise mechanisms by which INF2 regulates and INF2 mutations impair glomerular structure and function remain unknown. Here, we characterize an Inf2 R218Q point-mutant (knockin) mouse to help answer these questions. Knockin mice have no significant renal pathology or proteinuria at baseline despite diminished INF2 protein levels. INF2 mutant podocytes do show impaired reversal of protamine sulfate-induced foot process effacement by heparin sulfate perfusion. This is associated with persistent podocyte cytoplasmic aggregation, nephrin phosphorylation, and nephrin and podocin mislocalization, as well as impaired recovery of mDia membrane localization. These changes were partially mimicked in podocyte outgrowth cultures, in which podocytes from knockin mice show altered cellular protrusions compared to those from wild-type mice. Thus, in mice, normal INF2 function is not required for glomerular development but normal INF2 is required for regulation of the actin-based behaviors necessary for response to and/or recovery from injury.

Teaching Health Center Graduate Medical Education Locations Predominantly Located in Federally Designated Underserved Areas.

Background The Teaching Health Center Graduate Medical Education (THCGME) program is an Affordable Care Act funding initiative designed to expand primary care residency training in community-based ambulatory settings. Statute suggests, but does not require, training in underserved settings. Residents who train in underserved settings are more likely to go on to practice in similar settings, and graduates more often than not practice near where they have trained. Objective The objective of this study was to describe and quantify federally designated clinical continuity training sites of the THCGME program. Methods Geographic locations of the training sites were collected and characterized as Health Professional Shortage Area, Medically Underserved Area, Population, or rural areas, and were compared with the distribution of Centers for Medicare and Medicaid Services (CMS)-funded training positions. Results More than half of the teaching health centers (57%) are located in states that are in the 4 quintiles with the lowest CMS-funded resident-to-population ratio. Of the 109 training sites identified, more than 70% are located in federally designated high-need areas. Conclusions The THCGME program is a model that funds residency training in community-based ambulatory settings. Statute suggests, but does not explicitly require, that training take place in underserved settings. Because the majority of the 109 clinical training sites of the 60 funded programs in 2014-2015 are located in federally designated underserved locations, the THCGME program deserves further study as a model to improve primary care distribution into high-need communities.

Dental Hygienist Providers in Long-Term Care: Meeting the Need.

UNLABELLED: Dental hygienists with expertise in the policies, protocols, and practices of long-term care settings can provide oral health care services that complement the health complexities of residents. BACKGROUND: Adults in the United States are living longer and retaining more teeth, creating an increased demand for oral health care within long-term care facilities. Oral health has, in the past, been perceived as less important than other aspects of daily care and focused more on comfort than control of potential pathogens of oral origin. The concept in medicine of a care continuum from cradle to grave has not generally included provisions for oral health because historically few private dental practices have had provisions for continuation of care once an individual is unable to access the private office. This article supports the inclusion of dental hygienist care team members in long-term care settings. METHODS: The author has provided oral health care services in long-term care for many years, modeling this important career path for dental hygienists. Examples of the personal contributions of the dental hygienist author in helping to meet the extreme needs of this population illustrate this viable career option for dental hygienists. Resources used include federal and state health care publications, data from the American Dental Hygienists Association and selected state dental hygiene associations, published literature, and interviews with dental hygiene business owners, as well as the author's years of expertise. CONCLUSION: Oral health is a significant health concern for elders and others in long-term care settings. Daily oral care practices and optimal oral health are related to the well-being and quality of life that long-term care residents deserve. Pathways to providing preventive oral health services will gain momentum as dental hygienists become an integral part of the long-term care setting health care team.

The facilitators and barriers associated with implementation of a patient-centered medical home in VHA.

BACKGROUND: The patient-centered medical home (PCMH) is a team-based, comprehensive model of primary care. When effectively implemented, PCMH is associated with higher patient satisfaction, lower staff burnout, and lower hospitalization for ambulatory care-sensitive conditions. However, less is known about what factors contribute to (or hinder) PCMH implementation. We explored the associations of specific facilitators and barriers reported by primary care employees with a previously validated, clinic-level measure of PCMH implementation, the Patient Aligned Care Team Implementation Progress Index (Pi(2)). METHODS: We used a 2012 survey of primary care employees in the Veterans Health Administration to perform cross-sectional, respondent-level multinomial regressions. The dependent variable was the Pi(2) categorized as high implementation (top decile, 54 clinics, 235 respondents), medium implementation (middle eight deciles, 547 clinics, 4537 respondents), and low implementation (lowest decile, 42 clinics, 297 respondents) among primary care clinics. The independent variables were ordinal survey items rating 19 barriers to patient-centered care and 10 facilitators of PCMH implementation. For facilitators, we explored clinic Pi(2) score decile both as a function of respondent-reported availability of facilitators and of rating of facilitator helpfulness. RESULTS: The availability of five facilitators was associated with higher odds of a respondent's clinic's Pi(2) scores being in the highest versus lowest decile: teamlet huddles (OR = 3.91), measurement tools (OR = 3.47), regular team meetings (OR = 2.88), information systems (OR = 2.42), and disease registries (OR = 2.01). The helpfulness of four facilitators was associated with higher odds of a respondent's clinic's Pi(2) scores being in the highest versus lowest decile. Six barriers were associated with significantly higher odds of a respondent's clinic's Pi(2) scores being in the lowest versus highest decile, with the strongest associations for the difficulty recruiting and retaining providers (OR = 2.37) and non-provider clinicians (OR = 2.17). Results for medium versus low Pi(2) score clinics were similar, with fewer, smaller significant associations, all in the expected direction. CONCLUSIONS: A number of specific barriers and facilitators were associated with PCMH implementation, notably recruitment and retention of clinicians, team huddles, and local education. These findings can guide future research, and may help healthcare policy makers and leaders decide where to focus attention and limited resources.