Hu8F4-CAR T cells with mutated Fc spacer segment improve target-specificity and mediate anti-leukemia activity in vivo.

Hu8F4 is a T cell receptor (TCR)-like antibody with high affinity for leukemia-associated antigen PR1/HLA-A2 epitope. Adapted into a chimeric antigen receptor (CAR) format, Hu8F4-CAR is comprised of the Hu8F4 scFv, the human IgG1 CH2CH3 extracellular spacer domain, a human CD28 costimulatory domain, and the human CD3ζ signaling domain. We have demonstrated high efficacy of Hu8F4-CAR-T cells against PR1/HLA-A2-expressing cell lines and leukemic blasts from AML patients in vitro. Previous studies have shown that modification of the Fc domains of IgG4 CH2CH3 spacer regions can eliminate activation-induced cell death and off-target killing mediated by mouse Fc gamma receptor (FcgR)-expressing cells. We generated Hu8F4-CAR(PQ) with mutated Fc receptor binding sites on the CH2 domain of Hu8F4-CAR to prevent unwanted interactions with FcgR-expressing cells in vivo. The primary human T cells transduced with Hu8F4-CAR(PQ) can specifically lyse HLA-A2+ PR1-expressing leukemia cell lines in vitro. Furthermore, both adult donor-derived and cord blood-derived Hu8F4-CAR(PQ)-T cells are active and can eliminate U937 leukemia cells in NSG mice. Herein, we demonstrate that modification of the IgG1-based spacer can eliminate Fc receptor-binding-induced adverse effects and Hu8F4-CAR(PQ)-T cells can kill leukemia in vivo.

Community-Engaged Use of Low-Cost Sensors to Assess the Spatial Distribution of PM2.5 Concentrations across Disadvantaged Communities: Results from a Pilot Study in Santa Ana, CA.

PM2.5 is an air pollutant that is widely associated with adverse health effects, and which tends to be disproportionately located near low-income communities and communities of color. We applied a community-engaged research approach to assess the distribution of PM2.5 concentrations in the context of community concerns and urban features within and around the city of Santa Ana, CA. Approximately 183 h of one-minute average PM2.5 measurements, along with high-resolution geographic coordinate measurements, were collected by volunteer community participants using roughly two dozen low-cost AtmoTube Pro air pollution sensors paired with real-time GPS tracking devices. PM2.5 varied by region, time of day, and month. In general, concentrations were higher near the city's industrial corridor, which is an area of concern to local community members. While the freeway systems were shown to correlate with some degree of elevated air pollution, two of four sampling days demonstrated little to no visible association with freeway traffic. Concentrations tended to be higher within socioeconomically disadvantaged communities compared to other areas. This pilot study demonstrates the utility of using low-cost air pollution sensors for the application of community-engaged study designs that leverage community knowledge, enable high-density air monitoring, and facilitate greater health-related awareness, education, and empowerment among communities. The mobile air-monitoring approach used in this study, and its application to characterize the ambient air quality within a defined geographic region, is in contrast to other community-engaged studies, which employ fixed-site monitoring and/or focus on personal exposure. The findings from this study underscore the existence of environmental health inequities that persist in urban areas today, which can help to inform policy decisions related to health equity, future urban planning, and community access to resources.

A Nationwide Study of the Prevalence and Initial Management of Atypical Genitalia in the Newborn in Scotland.

Provision of optimum healthcare for infants with atypical genitalia requires a clear understanding of the occurrence of this condition. The objective of this study was to determine the prevalence of atypical genitalia and its initial management. A prospective, electronic survey of clinicians within managed clinical networks in Scotland was undertaken between 2013 and 2019. Notification from clinicians was sought for term neonates requiring specialist input for atypical genitalia. Additional information was also sought from the 4 regional genetics laboratories that provided details for neonates who had an urgent karyotype performed for atypical genitalia or sex determination. In total, the study identified 171 term infants who required some investigation for atypical genitalia in the neonatal period, providing a birth prevalence of 1:1,881 term births. Of the 171 infants, 97 (57%) had specialist input over the first 3 months of life, providing a birth prevalence of 1:3,318 term births that received specialist input for atypical genitalia. A total of 92 of these 97 cases had complete 3-month follow-up data, 62 (67%) presented within 24 h of birth, and age at presentation ranged from birth to 28 days. Age at sex assignment ranged from birth to 14 days, and in 63 cases (68%), sex assignment occurred at birth. Thus, the birth prevalence of a case of atypical genitalia where sex assignment was reported to be delayed beyond birth was estimated at 1:11,097 births. In 1 case sex was re-assigned at 3 months. Atypical genitalia requiring specialist input within the first month of life are rare in term newborns, and in only a third of these cases, sex assignment is delayed beyond birth. This study provides new clinical benchmarks for comparing and improving the delivery of care in centres that manage these conditions.

Novel myeloperoxidase-derived HLA-A2-restricted peptides as therapeutic targets against myeloid leukemia.

BACKGROUND AIMS: Human myeloperoxidase has been shown to be overexpressed in many types of leukemia, such as chronic myeloid leukemia, acute myeloid leukemia and myelodysplastic syndrome. The authors identified two myeloperoxidase-derived HLA-A2-restricted peptides, MY4 and MY8, as novel leukemia-associated antigens. METHODS: Ex vivo-elicited MY4- and MY8-specific cytotoxic T lymphocytes were generated, and tested for leukemia cell lysis in vitro and in NOD/SCID AML xenograft model. RESULTS: These MY4- and MY8-specific cytotoxic T lymphocytes killed leukemic blasts while sparing healthy donor bone marrow cells. In addition, co-injection of MY4- and MY8-specific cytotoxic T lymphocytes into nonobese diabetic/severe combined immunodeficiency mice with acute myeloid leukemia drastically reduced tumor burden in vivo. The authors also found that MY4- and MY8-specific T cells could be detected in the peripheral blood mononuclear cells of allogeneic stem cell transplant recipients. CONCLUSIONS: These antigen-specific T cells were significantly increased in blood samples from patients compared with healthy donors, suggesting that both MY4 and MY8 are immunogenic and that MY4- and MY8-specific cytotoxic T lymphocytes may play a role in reducing leukemia in vivo. Thus, the discovery of MY4 and MY8 as novel leukemia-associated antigens paves the way for targeting these antigens in immunotherapy against myeloid leukemia.

Two unique HLA-A*0201 restricted peptides derived from cyclin E as immunotherapeutic targets in leukemia.

Immunotherapy targeting leukemia-associated antigens has shown promising results. Because of the heterogeneity of leukemia, vaccines with a single peptide have elicited only a limited immune response. Targeting several peptides together elicited peptide-specific cytotoxic T lymphocytes (CTLs) in leukemia patients, and this was associated with clinical responses. Thus, the discovery of novel antigens is essential. In the current study, we investigated cyclin E as a novel target for immunotherapy. Cyclin E1 and cyclin E2 were found to be highly expressed in hematologic malignancies, according to reverse transcription polymerase chain reaction and western blot analysis. We identified two HLA-A*0201 binding nonameric peptides, CCNE1M from cyclin E1 and CCNE2L from cyclin E2, which both elicited the peptide-specific CTLs. The peptide-specific CTLs specifically kill leukemia cells. Furthermore, CCNE1M and CCNE2L CTLs were increased in leukemia patients who underwent allogeneic hematopoietic stem cell transplantation, and this was associated with desired clinical outcomes. Our findings suggest that cyclin E1 and cyclin E2 are potential targets for immunotherapy in leukemia.

What Attracts Medical Students to Primary Care? A Nominal Group Evaluation.

OBJECTIVE: To examine the perceptions of first-year medical students on their experiences in primary care. METHODS: Nominal group technique sessions were conducted with first-year medical students for 5 years. Questions were designed to evaluate primary care experiences and the role of primary care physicians. The questions explored what would make them consider primary care, what would detract from it, and what primary care has to offer that no other specialty can. Responses were weighted and ranked. The main outcome was the top five responses to three questions that were obtained at each session. RESULTS: Thirty-four students generated 280 responses to 3 questions. The top 5 responses for each year resulted in 29 experiences that strengthen enthusiasm: patient interactions (weighted sum, 43%), physician interactions/role modeling (22%), community interactions (20%), healthcare system/finances (8%), and other (6%). The top 5 responses resulted in 26 experiences that weaken enthusiasm, including hidden curriculum (45%), poor role models (29%), uncertainties about the healthcare system such as finances and documentation (20%), and patient interactions (6%). The top 5 responses regarding the uniqueness of primary care resulted in 37 experiences, including patient interactions (38%), continuity of care (20%), knowledge base (13%), community impact (10%), lifestyle benefits (10%), and education/prevention (9%). CONCLUSIONS: Medical students highlighted unique relationships with patients and continuity of care as experiences that increase their enthusiasm for primary care. Negative experiences that weakened enthusiasm for primary care included hidden curriculum and poor role models. Programs that provide experiences in primary care can increase student interest in primary care careers.

Defining and implementing a model for pharmacy resident research projects.

OBJECTIVE: To describe a standard approach to provide a support structure for pharmacy resident research that emphasizes self-identification of a residency research project. METHODS: A subcommittee of the residency advisory committee was formed at our institution. The committee was initially comprised of 2 clinical pharmacy specialists, 1 drug information pharmacist, and 2 pharmacy administrators. The committee developed research guidelines that are distributed to residents prior to the residency start that detail the research process, important deadlines, and available resources. Instructions for institutional review board (IRB) training and deadlines for various assignments and presentations throughout the residency year are clearly defined. Residents conceive their own research project and emphasis is placed on completing assignments early in the residency year. RESULTS: In the 4 years this research process has been in place, 15 of 16 (94%) residents successfully identified their own research question. All 15 residents submitted a complete research protocol to the IRB by the August deadline. Four residents have presented the results of their research at multi-disciplinary national professional meetings and 1 has published a manuscript. Feedback from outgoing residents has been positive overall and their perceptions of their research projects and the process are positive. CONCLUSION: Pharmacy residents selecting their own research projects for their residency year is a feasible alternative to assigning or providing lists of research projects from which to select a project.

Novel associations in disorders of sex development: findings from the I-DSD Registry.

CONTEXT: The focus of care in disorders of sex development (DSD) is often directed to issues related to sex and gender development. In addition, the molecular etiology remains unclear in the majority of cases. OBJECTIVE: To report the range of associated conditions identified in the international DSD (I-DSD) Registry. DESIGN, SETTING, AND PATIENTS: Anonymized data were extracted from the I-DSD Registry for diagnosis, karyotype, sex of rearing, genetic investigations, and associated anomalies. If necessary, clarification was sought from the reporting clinician. RESULTS: Of 649 accessible cases, associated conditions occurred in 168 (26%); 103 (61%) cases had one condition, 31 (18%) had two conditions, 20 (12%) had three conditions, and 14 (8%) had four or more conditions. Karyotypes with most frequently reported associations included 45,X with 6 of 8 affected cases (75%), 45,X/46,XY with 19 of 42 cases (45%), 46,XY with 112 of 460 cases (24%), and 46,XX with 27 of 121 cases (22%). In the 112 cases of 46,XY DSD, the commonest conditions included small for gestational age in 26 (23%), cardiac anomalies in 22 (20%), and central nervous system disorders in 22 (20%), whereas in the 27 cases of 46,XX DSD, skeletal and renal anomalies were commonest at 12 (44%) and 8 (30%), respectively. Of 170 cases of suspected androgen insensitivity syndrome, 19 (11%) had reported anomalies and 9 of these had confirmed androgen receptor mutations. CONCLUSIONS: Over a quarter of the cases in the I-DSD Registry have an additional condition. These associations can direct investigators toward novel genetic etiology and also highlight the need for more holistic care of the affected person.

An electronic surveillance system for monitoring the hospital presentation of nutritional vitamin D deficiency in children in Scotland.

BACKGROUND: Routine surveillance would be valuable for vitamin D deficiency as symptomatic vitamin D deficiency may be common in Scotland. AIM: To assess the effectiveness of an electronic surveillance system to determine the current incidence of hospital-based presentation of childhood vitamin D deficiency in Scotland. METHODS: Active surveillance was performed for 2 years as part of an electronic web-based surveillance programme by the Scottish Paediatric Surveillance Unit. Notifications were followed by completion of a questionnaire. To further examine the validity of the system, cases with severe vitamin D deficiency in Glasgow and Edinburgh were identified from the regional laboratory and their clinical details were checked against those identified through the surveillance system. RESULTS: Between September 2009 and August 2011, 109 cases of vitamin D deficiency were notified. The majority of cases (n=82) were reported in Glasgow with an annual incidence of 41 cases per year. Fourteen cases were reported in Edinburgh during the first year of the study and two cases during the second year. At the time of clinical diagnosis, the median age of the children was 2 years (range 3 months-16 years). Cross-validation of data showed that among symptomatic cases that had a measured serum vitamin D of <14 nmol/L, 89% of eligible cases had been reported in Glasgow and 33% of cases had been reported in Edinburgh. CONCLUSION: The incidence of vitamin D deficiency remains high in Scotland. An electronic surveillance system can provide data for studying the epidemiology of vitamin D deficiency but may underestimate the number of positive cases.

Paracrine regulation of matrix metalloproteinase expression in endometriosis.

Following retrograde menstruation, shed endometrial tissue fragments attach to and invade the peritoneal surface to form established endometriotic lesions. With disease progression, the biochemically active lesions undergo remodeling and become fibrotic. Matrix metalloproteinase enzymes (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) play a significant role in normal endometrial remodeling during menses. Anomalous expression of MMPs and TIMPs has been identified in endometriotic lesions as compared to their highly regulated expression in eutopic endometrium. The paracrine mechanisms regulating misexpression of MMPs and TIMPs by endometriotic lesions are, however, not well defined. Misexpression of the MMPs and TIMPs may be due to innate anomalies in the eutopic endometrium from women with endometriosis, in the resident immune cells and peritoneal cells that juxtapose the ectopic endometrium, and/or numerous substances present in peritoneal fluid of women with endometriosis. The majority of MMPs are under strict transcriptional regulation. Steroid hormones and cytokines appear to act on the MMP promoter, either independently or in consort, to provide both positive and negative regulation of these genes. Misregulated expression of MMPs and TIMPs is associated with a more aggressive phenotype and a cascade of events facilitating peritoneal extracellular matrix degradation and establishment or remodeling of endometriotic lesions. The mechanisms by which MMP and TIMP expression are misregulated warrant further investigation as such information may provide insight into novel therapeutic modalities for endometriosis.