Prototype and BA.5 protein nanoparticle vaccines protect against Omicron BA.5 variant in Syrian hamsters.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants with greater transmissibility or immune evasion properties has jeopardized the existing vaccine and antibody-based countermeasures. Here, we evaluated the efficacy of boosting pre-immune hamsters with protein nanoparticle vaccines (Novavax, Inc.) containing recombinant Prototype (Wuhan-1) or BA.5 S proteins against a challenge with the Omicron BA.5 variant of SARS-CoV-2. Serum antibody binding and neutralization titers were quantified before challenge, and viral loads were measured 3 days after challenge. Boosting with Prototype or BA.5 vaccine induced similar antibody binding responses against ancestral Wuhan-1 or BA.5 S proteins, and neutralizing activity of Omicron BA.1 and BA.5 variants. One and three months after vaccine boosting, hamsters were challenged with the Omicron BA.5 variant. Prototype and BA.5 vaccine-boosted hamsters had reduced viral infection in the nasal washes, nasal turbinates, and lungs compared to unvaccinated animals. Although no significant differences in virus load were detected between the Prototype and BA.5 vaccine-boosted animals, fewer breakthrough infections were detected in the BA.5-vaccinated hamsters. Thus, immunity induced by Prototype or BA.5 S protein nanoparticle vaccine boosting can protect against the Omicron BA.5 variant in the Syrian hamster model. IMPORTANCE: As SARS-CoV-2 continues to evolve, there may be a need to update the vaccines to match the newly emerging variants. Here, we compared the protective efficacy of the updated BA.5 and the original Wuhan-1 COVID-19 vaccine against a challenge with the BA.5 Omicron variant of SARS-CoV-2 in hamsters. Both vaccines induced similar levels of neutralizing antibodies against multiple variants of SARS-CoV-2. One and three months after the final immunization, hamsters were challenged with BA.5. No differences in protection against the BA.5 variant virus were observed between the two vaccines, although fewer breakthrough infections were detected in the BA.5-vaccinated hamsters. Together, our data show that both protein nanoparticle vaccines are effective against the BA.5 variant of SARS-CoV-2 but given the increased number of breakthrough infections and continued evolution, it is important to update the COVID-19 vaccine for long-term protection.

Diet-induced obesity and diabetes enhance mortality and reduce vaccine efficacy for SARS-CoV-2.

IMPORTANCE: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has caused a wide spectrum of diseases in the human population, from asymptomatic infections to death. It is important to study the host differences that may alter the pathogenesis of this virus. One clinical finding in coronavirus disease 2019 (COVID-19) patients is that people with obesity or diabetes are at increased risk of severe illness from SARS-CoV-2 infection. We used a high-fat diet model in mice to study the effects of obesity and type 2 diabetes on SARS-CoV-2 infection as well as how these comorbidities alter the response to vaccination. We find that diabetic/obese mice have increased disease after SARS-CoV-2 infection and they have slower clearance of the virus. We find that the lungs of these mice have increased neutrophils and that removing these neutrophils protects diabetic/obese mice from disease. This demonstrates why these diseases have increased risk of severe disease and suggests specific interventions upon infection.

Strong CD4+ T-Cell Responses to Ancestral and Variant Spike Proteins Are Established by NVX-CoV2373 Severe Acute Respiratory Syndrome Coronavirus 2 Primary Vaccination.

BACKGROUND: NVX-CoV2373 is an efficacious coronavirus disease 2019 (COVID-19) vaccine comprising full-length recombinant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (rS) glycoprotein and Matrix-M adjuvant. Phase 2 of a randomized, placebo-controlled, phase 1/2 trial in healthy adults (18-84 years of age) previously reported good safety/tolerability and robust humoral immunogenicity. METHODS: Participants were randomized to placebo or 1 or 2 doses of 5-µg or 25-µg rS with 50 µg Matrix-M adjuvant 21 days apart. CD4+ T-cell responses to SARS-CoV-2 intact S or pooled peptide stimulation (with ancestral or variant S sequences) were measured via enzyme-linked immunosorbent spot assay and intracellular cytokine staining. RESULTS: A clearly discernable spike antigen-specific CD4+ T-cell response was induced after 1 dose, but markedly enhanced after 2 doses. Counts and fold increases in cells producing Th1 cytokines exceeded those secreting Th2 cytokines, although both phenotypes were clearly present. Interferon-γ responses to rS were detected in 93.5% of 2-dose 5-µg recipients. A polyfunctional CD4+ T-cell response was cross-reactive and of equivalent magnitude to all tested variants, including Omicron BA.1/BA.5. CONCLUSIONS: NVX-CoV2373 elicits a moderately Th1-biased CD4+ T-cell response that is cross-reactive with ancestral and variant S proteins after 2 doses. CLINICAL TRIALS REGISTRATION: NCT04368988.

Immunogenicity and safety of heterologous Omicron BA.1 and bivalent SARS-CoV-2 recombinant spike protein booster vaccines: a phase 3, randomized, clinical trial.

BACKGROUND: Mutations present in emerging SARS-CoV-2 variants permit evasion of neutralization with prototype vaccines. A novel Omicron BA.1 subvariant-specific vaccine (NVX-CoV2515) was tested alone, or as a bivalent preparation in combination with the prototype vaccine (NVX-CoV2373), to assess antibody responses to SARS-CoV-2. METHODS: Participants aged 18 to 64 years immunized with 3 doses of prototype mRNA vaccines were randomized 1:1:1 to receive a single dose of NVX-CoV2515, NVX-CoV2373, or bivalent mixture in a phase 3 study investigating heterologous boosting with SARS-CoV-2 recombinant spike protein vaccines. Immunogenicity was measured 14 and 28 days after vaccination for the SARS-CoV-2 Omicron BA.1 sublineage and ancestral strain. Safety profiles of vaccines were assessed. RESULTS: Of participants who received trial vaccine (N = 829), those administered NVX-CoV2515 (n = 286) demonstrated superior neutralizing antibody response to BA.1 versus NVX-CoV2373 (n = 274) at Day 14 (geometric mean titer ratio [95% CI]: 1.6 [1.33, 2.03]). Seroresponse rates [n/N; 95% CI] were 73.4% [91/124; 64.7, 80.9] for NVX-CoV2515 versus 50.9% [59/116; 41.4, 60.3] for NVX-CoV2373. All formulations were similarly well-tolerated. CONCLUSIONS: NVX-CoV2515 elicited a superior neutralizing antibody response against the Omicron BA.1 subvariant compared with NVX-CoV2373 when administered as a fourth dose. Safety data were consistent with the established safety profile of NVX-CoV2373.

Phase 1-2 Trial of a SARS-CoV-2 Recombinant Spike Protein Nanoparticle Vaccine.

BACKGROUND: NVX-CoV2373 is a recombinant severe acute respiratory syndrome coronavirus 2 (rSARS-CoV-2) nanoparticle vaccine composed of trimeric full-length SARS-CoV-2 spike glycoproteins and Matrix-M1 adjuvant. METHODS: We initiated a randomized, placebo-controlled, phase 1-2 trial to evaluate the safety and immunogenicity of the rSARS-CoV-2 vaccine (in 5-µg and 25-µg doses, with or without Matrix-M1 adjuvant, and with observers unaware of trial-group assignments) in 131 healthy adults. In phase 1, vaccination comprised two intramuscular injections, 21 days apart. The primary outcomes were reactogenicity; laboratory values (serum chemistry and hematology), according to Food and Drug Administration toxicity scoring, to assess safety; and IgG anti-spike protein response (in enzyme-linked immunosorbent assay [ELISA] units). Secondary outcomes included unsolicited adverse events, wild-type virus neutralization (microneutralization assay), and T-cell responses (cytokine staining). IgG and microneutralization assay results were compared with 32 (IgG) and 29 (neutralization) convalescent serum samples from patients with Covid-19, most of whom were symptomatic. We performed a primary analysis at day 35. RESULTS: After randomization, 83 participants were assigned to receive the vaccine with adjuvant and 25 without adjuvant, and 23 participants were assigned to receive placebo. No serious adverse events were noted. Reactogenicity was absent or mild in the majority of participants, more common with adjuvant, and of short duration (mean, ≤2 days). One participant had mild fever that lasted 1 day. Unsolicited adverse events were mild in most participants; there were no severe adverse events. The addition of adjuvant resulted in enhanced immune responses, was antigen dose-sparing, and induced a T helper 1 (Th1) response. The two-dose 5-µg adjuvanted regimen induced geometric mean anti-spike IgG (63,160 ELISA units) and neutralization (3906) responses that exceeded geometric mean responses in convalescent serum from mostly symptomatic Covid-19 patients (8344 and 983, respectively). CONCLUSIONS: At 35 days, NVX-CoV2373 appeared to be safe, and it elicited immune responses that exceeded levels in Covid-19 convalescent serum. The Matrix-M1 adjuvant induced CD4+ T-cell responses that were biased toward a Th1 phenotype. (Funded by the Coalition for Epidemic Preparedness Innovations; ClinicalTrials.gov number, NCT04368988).

How do I save cell therapy product's storage space?

BACKGROUND: Availability of liquid nitrogen (LN2) freezer storage space is a major challenge for many transplant programs as they continue to grow and accumulate products. The recent trend of allogeneic grafts cryopreservation that started during the COVID-19 pandemic, made the situation even worse requiring an increase in storage capacity. Multi-compartment cryopreservation bags can help save storage space but can be tricky to use. Here, we describe the validation of muti-compartment cryopreservation bags for the purpose of donor lymphocyte infusion (DLI) aliquots. METHODS: We validated the use of five compartment cryobags for cryopreservation of cell therapy products. Four products were cryopreserved using these bags and each compartment was tested post-thaw for product volume distribution, total cell count recovery, and viability. Additionally, the integrity of both bag compartments and labels was assessed as well. RESULTS: All tested specimens met post-thaw viability and TNC recovery acceptability criteria. Fill volume was optimized at 24-25 mL for acceptable volume distribution between aliquots. With proper heat sealing between compartments, all aliquots retain their integrity and cryopreservation labels were adherent and legible. CONCLUSIONS: Muti-compartment bags can be used successfully for cryopreservation of cell therapy products and increase storage capacity.

Validation of automated fluorescent-based technology for measuring total nucleated cell viability of hematopoietic progenitor cell products.

BACKGROUND: A reliable rapid method for measuring total nucleated cell (TNC) viability is essential for cell-based products manufacturing. The trypan blue (TB) exclusion method, commonly used to measure TNC viability of hematopoietic progenitor cell (HPC) products, is a subjective assay, typically uses a microscope, and includes a limited number of cells. The NucleoCounter NC-200 is an automated fluorescent-based cell counter that uses pre-calibrated cartridges with acridine orange and DAPI dyes to measure cell count and viability. This study describes the validation of the NC-200 for testing HPC's viability. METHODS: Samples from 189 fresh and 60 cryopreserved HPC products were included. Fresh products were tested for viability after collection by both TB and NC-200. 7-aminoactinomycin D (7AAD) CD45+ cell viability results were obtained from a flow cytometry test. Cryopreserved products thawed specimens were tested for viability by both TB and NC-200. The NC-200 viability results were compared with the other methods. Acceptability criteria were defined as ≤10% difference between the NC-200 method and the other methods for at least 95% of the samples. RESULTS: Fresh products' mean viability difference between NC-200 and TB or 7AAD CD45+ method was 4.9% (95%CI 4.6-5.4) and 2.8% (95%CI 2.2-3.4), respectively. Thawed products' mean viability difference between NC-200 and TB was 3.0% (95%CI 0.4-5.6). CONCLUSION: The NC-200 automated fluorescent-based method can be used effectively to determine HPC's viability for both fresh and cryopreserved products. It can help eliminate human bias and provide consistent data and operational ease.

Induction of Cross-Reactive Hemagglutination Inhibiting Antibody and Polyfunctional CD4+ T-Cell Responses by a Recombinant Matrix-M-Adjuvanted Hemagglutinin Nanoparticle Influenza Vaccine.

BACKGROUND: Recurrent reports of suboptimal influenza vaccine effectiveness have renewed calls to develop improved, broadly cross-protective influenza vaccines. Here, we evaluated the safety and immunogenicity of a novel, saponin (Matrix-M)-adjuvanted, recombinant hemagglutinin (HA) quadrivalent nanoparticle influenza vaccine (qNIV). METHODS: We conducted a randomized, observer-blind, comparator-controlled (trivalent high-dose inactivated influenza vaccine [IIV3-HD] or quadrivalent recombinant influenza vaccine [RIV4]), safety and immunogenicity trial of qNIV (5 doses/formulations) in healthy adults ≥65 years. Vaccine immunogenicity was measured by hemagglutination-inhibition assays using reagents that express wild-type hemagglutination inhibition (wt-HAI) sequences and cell-mediated immune responses. RESULTS: A total of 1375 participants were randomized, immunized, and followed for safety and immunogenicity. Matrix-M-adjuvanted qNIV induced superior wt-HAI antibody responses against 5 of 6 homologous or drifted strains compared with unadjuvanted qNIV. Adjuvanted qNIV induced post-vaccination wt-HAI antibody responses at day 28 that were statistically higher than IIV3-HD against a panel of homologous or drifted A/H3N2 strains, similar to IIV3-HD against homologous A/H1N1 and B (Victoria) strains and similar to RIV4 against all homologous and drifted strains evaluated. The qNIV formulation with 75 µg Matrix-M adjuvant induced substantially higher post-vaccination geometric mean fold increases of influenza HA-specific polyfunctional CD4+ T cells compared with IIV3-HD or RIV4. Overall, similar frequencies of solicited and unsolicited adverse events were reported in all treatment groups. CONCLUSIONS: qNIV with 75 µg Matrix-M adjuvant was well tolerated and induced robust antibody and cellular responses, notably against both homologous and drifted A/H3N2 viruses. Further investigation in a pivotal phase 3 trial is underway. CLINICAL TRIALS REGISTRATION: NCT03658629.

A Pilot Study of Integration of Medical and Dental Care in 6 States.

INTRODUCTION: Poor oral health affects overall health. Chronic diseases and related risk factors such as tobacco use or consuming sugar-sweetened beverages can also increase a person's risk of periodontitis. Given the linkages between oral health and certain chronic diseases, we conducted a pilot study to facilitate intradepartmental collaborations between state chronic disease and oral health programs. METHODS: State health departments in 6 states (Alaska, Colorado, Georgia, Maryland, Minnesota, and New York) collaborated to develop and implement projects that addressed oral health and the following chronic diseases or risk factors: obesity, diabetes, heart disease, stroke, and tobacco use. States developed various projects, including media campaigns, clinical education, and screening and referrals. We used a mixed-methods approach to understand barriers to and facilitators of states' increasing collaboration and implementation of pilot projects. In-depth interviews were conducted with 12 staff (1 from oral health and 1 from chronic disease for each state). We also reviewed state-submitted documents and performance measures. RESULTS: All 6 states increased collaboration between their oral health and chronic disease programs and successfully implemented pilot projects. Collaboration was facilitated by investing in relationships, championing medical-dental integration, and meeting and communicating frequently. Barriers to collaboration included the perception of oral health in chronic disease programs as separate and distinct from other chronic diseases and the structure of funding. The pilot projects were facilitated by partner support, providing technical assistance to clinics, and working early on referral networks. Barriers to implementing the pilot projects included gaining clinician buy-in and establishing referral networks. CONCLUSION: This pilot study demonstrated that by fostering collaboration, state health departments are able to train dental and medical clinicians, deliver clinical preventive education to patients, implement referral systems, and deliver impressions via media campaigns.

Prevalence of Past-Year Dental Visit Among US Adults Aged 50 Years or Older, With Selected Chronic Diseases, 2018.

In this study, we used data from the Behavioral Risk Factor Surveillance System to conduct multivariable analyses to examine whether having selected chronic diseases was associated with lower past-year dental service utilization among US adults aged 50 years or older. We found consistent lower dental service utilization among older adults with diabetes, heart disease or stroke, and chronic obstructive pulmonary disease (COPD) compared with those without the disease after adjusting for sociodemographic characteristics. We also found lower dental service utilization among older adults with lower income, less education, and no health care coverage and among those who smoked. Effective interventions are needed to reduce disparities in access to dental care among at-risk and vulnerable populations.

Safety and Immunogenicity of a Respiratory Syncytial Virus Fusion (F) Protein Nanoparticle Vaccine in Healthy Third-Trimester Pregnant Women and Their Infants.

BACKGROUND: Respiratory syncytial virus (RSV) is the leading cause of infant lower respiratory tract disease and hospitalization worldwide. METHODS: Safety and immunogenicity of RSV fusion (F) protein nanoparticle vaccine or placebo were evaluated in 50 healthy third-trimester pregnant women. Assessments included vaccine tolerability and safety in women and infants, and RSV-specific antibody measures in women before and after vaccination, at delivery and post partum. RESULTS: The vaccine was well tolerated; no meaningful differences in pregnancy or infant outcomes were observed between study groups. RSV-specific antibody levels increased significantly among vaccine recipients, including responses competitive with well-described monoclonal antibodies specific for multiple RSV neutralizing epitopes. No significant antibody increase was seen among placebo recipients, although a shallow upward trend across the RSV season was noted. Transplacental antibody transfer was 90%-120% across assays for infants of vaccinated women. Women with an interval of ≥30 days between vaccination and delivery demonstrated higher placental antibody transfer rates than women with an interval <30 days. Half-lives of RSV-specific antibodies in infants approximated 40 days. There was no evidence of severe RSV disease in infants of vaccinated mothers. CONCLUSIONS: Data from this phase 2 study support a maternal immunization strategy to protect infants from RSV disease. CLINICAL TRIALS REGISTRATION: NCT02247726.

Bone Marrow Harvest in Pediatric Sibling Donors: Role of Granulocyte Colony-Stimulating Factor Priming and CD34+ Cell Dose.

To ensure optimal clinical outcomes for patients while retaining adequate protection for donors, the National Marrow Donor Program developed guidelines specifying that up to 20 mL/kg of bone marrow can be harvested from donors. These guidelines, originally developed for unrelated adult donors, are followed in children as well. We studied the impact of granulocyte colony-stimulating factor (G-CSF) priming on the cellular composition of harvested bone marrow, sought to develop an algorithm to optimize bone marrow harvest volume from pediatric matched sibling donors, and studied the impact of CD34+ cell dose on clinical outcomes. We analyzed data from 92 bone marrow harvests and clinical outcomes for 69 sibling recipient-donor duos, The mean age of recipients was 9.85 ± 5.90 years, and that of donors was 11.85 ± 6.36 years. G-CSF priming was not associated with higher yield of CD34+ cells/µL. The median CD34+ cell count obtained from donors was 700 cells/µL (range, 400-1700 cells/µL) in donors age <6 years, 360 cells/µL (range, 100-1100 cells/µL) in donors age 6 to 12 years, and 300 cells/µL (range, 80-800 cells/µL) in donors age >12 years (P < .001). The number of CD34+ cells infused had no impact on traditional clinical outcomes; however, it was significantly related to graft-versus-host disease/relapse/rejection-free survival. Our investigation revealed that ultimately, a CD34+ cell count of approximately 3 to 5 × 106/kg was a threshold beyond which increasing CD34+ cell dose did not impact outcome. In this study, we addressed the broad question of whether harvesting up to 20 mL/kg of bone marrow from a child donor is truly necessary for optimal outcomes in every pediatric case.

A Semi-Physiologically Based Pharmacokinetic Model Describing the Altered Metabolism of Midazolam Due to Inflammation in Mice.

PURPOSE: To investigate influence of inflammation on metabolism and pharmacokinetics (PK) of midazolam (MDZ) and construct a semi-physiologically based pharmacokinetic (PBPK) model to predict PK in mice with inflammatory disease. METHODS: Glucose-6-phosphate isomerase (GPI)-mediated inflammation was used as a preclinical model of arthritis in DBA/1 mice. CYP3A substrate MDZ was selected to study changes in metabolism and PK during the inflammation. The semi-PBPK model was constructed using mouse physiological parameters, liver microsome metabolism, and healthy animal PK data. In addition, serum cytokine, and liver-CYP (cytochrome P450 enzymes) mRNA levels were examined. RESULTS: The in vitro metabolite formation rate was suppressed in liver microsomes prepared from the GPI-treated mice as compared to the healthy mice. Further, clearance of MDZ was reduced during inflammation as compared to the healthy group. Finally, the semi-PBPK model was used to predict PK of MDZ after GPI-mediated inflammation. IL-6 and TNF-α levels were elevated and liver-cyp3a11 mRNA was reduced after GPI treatment. CONCLUSION: The semi-PBPK model successfully predicted PK parameters of MDZ in the disease state. The model may be applied to predict PK of other drugs under disease conditions using healthy animal PK and liver microsomal data as inputs.

Correction to: A Semi-Physiologically Based Pharmacokinetic Model Describing the Altered Metabolism of Midazolam Due to Inflammation in Mice.

One of the authors has his name incorrectly indexed in PubMed and SpringerLink as "Laird Forrest M" (last name "Laird Forrest"). His name should index as "Forrest M. Laird" with last name as "Forrest".

Characterization of a Prefusion-Specific Antibody That Recognizes a Quaternary, Cleavage-Dependent Epitope on the RSV Fusion Glycoprotein.

Prevention efforts for respiratory syncytial virus (RSV) have been advanced due to the recent isolation and characterization of antibodies that specifically recognize the prefusion conformation of the RSV fusion (F) glycoprotein. These potently neutralizing antibodies are in clinical development for passive prophylaxis and have also aided the design of vaccine antigens that display prefusion-specific epitopes. To date, prefusion-specific antibodies have been shown to target two antigenic sites on RSV F, but both of these sites are also present on monomeric forms of F. Here we present a structural and functional characterization of human antibody AM14, which potently neutralized laboratory strains and clinical isolates of RSV from both A and B subtypes. The crystal structure and location of escape mutations revealed that AM14 recognizes a quaternary epitope that spans two protomers and includes a region that undergoes extensive conformational changes in the pre- to postfusion F transition. Binding assays demonstrated that AM14 is unique in its specific recognition of trimeric furin-cleaved prefusion F, which is the mature form of F on infectious virions. These results demonstrate that the prefusion F trimer contains potent neutralizing epitopes not present on monomers and that AM14 should be particularly useful for characterizing the conformational state of RSV F-based vaccine antigens.

Safety of hematopoietic cell infusion in children with malignant and non-malignant diseases.

HPC infusions have been associated with a variety of adverse events related to either patient or HPC product-related factors. Studies documenting infusion-related AEs in children are limited. We reviewed HPC infusion records in 354 children. Infusion-related adverse events were classified as follows: grade 0-absent, grade I-mild, grade II-moderate, grade III-severe, grade IV-life-threatening, and grade V-death. The percentage of patients with grade 0, I, and II-IV AEs was as follows: 0 = 67%, I = 23.4%, and II-V = 9.6% (one patient had fatal anaphylactic reaction to dimethyl sulfoxide). The incidence of grade II-IV hypertension was 7.1%. There was a higher incidence of AEs with infusion of allogeneic bone marrow versus allogeneic PBSCs (47.4% vs 25.3%, P = .001). Cryopreserved products had a lower incidence of infusion-associated AEs compared with fresh HPC products (24% vs 39.4%, P = .003). Allogeneic HPC infusion volume (>100 mL) was a significant risk factor for infusion-associated AEs (P < .001). Patients >10 years who received autologous HPC infusions had higher risk of AEs when compared to patients <10 years (P = .01). Our study demonstrated that despite a high incidence of infusion-associated hypertension, HPC infusion is relatively safe in children. Investigating strategies to optimize management of hypertension in the setting of HPC infusion is warranted.

Evaluation and Optimization of Blood Micro-Sampling Methods: Serial Sampling in a Cross-Over Design from an Individual Mouse.

PURPOSE: Current practices applied to mouse pharmacokinetic (PK) studies often use large numbers of animals with sporadic or composite sampling that inadequately describe PK profiles.  The purpose of this work was to evaluate and optimize blood microsampling techniques coupled with dried blood spot (DBS) and LC-MS/MS analysis to generate reliable PK data in mice.  In addition, the feasibility of cross-over designs was assessed and recommendations are presented. METHODS: The work describes a comprehensive evaluation of five blood microsampling techniques (tail clip, tail vein with needle hub, submandibular, retro-orbital, and saphenous bleeding) in CD-1 mice.  The feasibility of blood sampling was evaluated based on animal observations, ease of bleeding, and ability to collect serial samples.  Methotrexate, gemfibrozil and glipizide were used as test compounds and were dosed either orally or intravenously, followed by DBS collection and LC-MS/MS analysis to compare PK with various bleeding methods. RESULTS: Submandibular and retro-orbital methods that required non-serial blood collections did not allow for inter-animal variability assessments and resulted in poorly described absorption and distribution kinetics.  The submandibular and tail vein with needle-hub methods were the least favorable from a technical feasibility perspective.  Serial bleeding was possible with cannulated animals or saphenous bleeding in non-cannulated animals. CONCLUSIONS:   Of the methods that allowed serial sampling, the saphenous method when executed as described in this report, was most practical, reproducible and provided for assessment of inter-animal variability.  It enabled the collection of complete exposure profiles from a single mouse and the conduct of an intravenous/oral cross-over study design.  This methodology can be used routinely, it promotes the 3Rs principles by achieving reductions in the number of animals used, decreased restraints and animal stress, and improved the quality of data obtained in mouse PK studies. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Eye Care Quality and Accessibility Improvement in the Community (EQUALITY) for adults at risk for glaucoma: study rationale and design.

BACKGROUND: Primary open angle glaucoma is a chronic, progressive eye disease that is the leading cause of blindness among African Americans. Glaucoma progresses more rapidly and appears about 10 years earlier in African Americans as compared to whites. African Americans are also less likely to receive comprehensive eye care when glaucoma could be detected before irreversible blindness. Screening and follow-up protocols for managing glaucoma recommended by eye-care professional organizations are often not followed by primary eye-care providers, both ophthalmologists and optometrists. There is a pressing need to improve both the accessibility and quality of glaucoma care for African Americans. Telemedicine may be an effective solution for improving management and diagnosis of glaucoma because it depends on ocular imaging and tests that can be electronically transmitted to remote reading centers where tertiary care specialists can examine the results. We describe the Eye Care Quality and Accessibility Improvement in the Community project (EQUALITY), set to evaluate a teleglaucoma program deployed in retail-based primary eye care practices serving communities with a large percentage of African Americans. METHODS/DESIGN: We conducted an observational, 1-year prospective study based in two Walmart Vision Centers in Alabama staffed by primary care optometrists. EQUALITY focuses on new or existing adult patients who are at-risk for glaucoma or already diagnosed with glaucoma. Patients receive dilated comprehensive examinations and diagnostic testing for glaucoma, followed by the optometrist's diagnosis and a preliminary management plan. Results are transmitted to a glaucoma reading center where ophthalmologists who completed fellowship training in glaucoma review results and provide feedback to the optometrist, who manages the care of the patient. Patients also receive eye health education about glaucoma and comprehensive eye care. Research questions include diagnostic and management agreement between providers, the impact of eye health education on patients' knowledge and adherence to follow-up and medication, patient satisfaction, program cost-effectiveness, and EQUALITY's impact on Walmart pharmacy prescription rates. DISCUSSION: As eye-care delivery systems in the US strive to improve quality while reducing costs, telemedicine programs including teleglaucoma initiatives such as EQUALITY could contribute toward reaching this goal, particularly among underserved populations at-risk for chronic blinding diseases.

Do children with specific language impairment and autism spectrum disorders benefit from the presence of orthography when learning new spoken words?

This experiment investigated whether children with specific language impairment (SLI), children with autism spectrum disorders (ASD), and typically developing children benefit from the incidental presence of orthography when learning new oral vocabulary items. Children with SLI, children with ASD, and typically developing children (n=27 per group) between 8 and 13 years of age were matched in triplets for age and nonverbal reasoning. Participants were taught 12 mappings between novel phonological strings and referents; half of these mappings were trained with orthography present and half were trained with orthography absent. Groups did not differ on the ability to learn new oral vocabulary, although there was some indication that children with ASD were slower than controls to identify newly learned items. During training, the ASD, SLI, and typically developing groups benefited from orthography to the same extent. In supplementary analyses, children with SLI were matched in pairs to an additional control group of younger typically developing children for nonword reading. Compared with younger controls, children with SLI showed equivalent oral vocabulary acquisition and benefit from orthography during training. Our findings are consistent with current theoretical accounts of how lexical entries are acquired and replicate previous studies that have shown orthographic facilitation for vocabulary acquisition in typically developing children and children with ASD. We demonstrate this effect in SLI for the first time. The study provides evidence that the presence of orthographic cues can support oral vocabulary acquisition, motivating intervention approaches (as well as standard classroom teaching) that emphasize the orthographic form.