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John D Rodriguez,1,2 Samantha Kerti,3 Adam Hamm,3 George W Ousler,1 Ethan Bensinger,1,2 Sadie Burnham,1,2 Mark B Abelson1,2,4,5 1Ora, Inc, Andover, MA, USA; 2Andover Eye Institute, Andover, MA, USA; 3Statistics and Data Corporation, Tempe, AZ, USA; 4Ophthalmology, Harvard Medical School, Cambridge, MA, USA; 5Mass Eye and Ear, Boston, MA, USACorrespondence: John D Rodriguez, Email jrodriguez@oraclinical.comPurpose: The absence of a standardized diagnostic method for clinical signs of Dry Eye Disease (DED) complicates clinical trials for future treatments. This paper evaluated Lissamine Green (LG) conjunctival staining as a valid, stable and modifiable endpoint for both clinical practice and clinical trials.Methods: Screening and pre-randomization data from two identically designed clinical trials for DED resulted in a pooled dataset of 494 subjects. Inclusion was based on reported symptoms, lissamine green (LG) conjunctival staining, Fluorescein (Fl) corneal and conjunctival staining, and Schirmer’s Test (ST). Outcome measures were assessed based on the modifiability of LG staining to exposure to a Controlled Adverse Environment (CAE®), correlation of LG to Fl staining, relative variation of LG staining scores and Schirmer test scores, and the correlation of LG staining with symptom scores.Results: The modifiability of LG conjunctival staining to environmental exposure was demonstrated, with nasal LG and FL staining displaying the most similar percent change. Nasal LG conjunctival staining scores for subjects with ST scores of less than 8mm were significantly higher than for subjects with ST greater than 8mm. LG staining scores were more consistent (25% change from baseline threshold) than ST scores. Finally, statistically significant correlations were found between LG staining and a number of symptom scores.Conclusion: This evaluation demonstrates the superiority of the utilization of a clinical endpoint focused on ocular surface damage. The reproducibility and modifiability of LG conjunctival staining to controlled adverse environment, coupled with its significant correlation with symptoms, positions it as an exemplary clinical sign endpoint for clinical management and in clinical trials. Our findings advocate for the adoption of LG conjunctival staining as a primary endpoint in both clinical research and drug development, offering a more effective means of identifying and addressing ocular surface damage in the realm of DED.Keywords: dry eye, conjunctiva, stain, lissamine green clinical trials, controlled adverse environment, disease models, drug screening, dry eye disease, efficacy endpoints

John D Rodriguez,1 Keith Lane,1 David A Hollander,1,2 Aron Shapiro,1 Sunita Saigal,1 Andrew J Hertsenberg,1 Garrick Wallstrom,3 Divya Narayanan,1 Endri Angjeli,1 Mark B Abelson1,4 1Ora, Inc., Andover, MA, USA; 2Jules Stein Eye Institute, University of California, Los Angeles, CA, USA; 3Statistics and Data Corporation, Tempe, AZ, USA; 4Department of Ophthalmology, Harvard Medical School, Boston, MA, USA Purpose: To identify parameters from cone function and recovery after photostress that detect functional deficits in early non-exudative age-related macular degeneration (AMD) and to determine the repeatability of these parameters. Methods: Cone-mediated visual function recovery after photostress was examined in three groups of subjects: young normal subjects (ages 20–29; N=8), older normal subjects (ages 50–90; N=9), and early non-exudative AMD subjects (ages 50–90; N=12). Eight AMD and four normal subjects were retested 1 year after the initial evaluation. Early Treatment Diabetic Retinopathy Study (ETDRS) visual acuity (VA) and parameters of cone function (baseline cone sensitivity and cone recovery half-life following photobleach) were measured and compared between AMD and normal subjects. Short-term repeatability was assessed for each subject’s initial evaluation. Long-term repeatability was assessed by comparing outcomes from the initial evaluation and 1-year follow-up. Results: The mean baseline cone threshold was significantly worse in subjects with early AMD compared to older normal subjects (-1.80±0.04 vs -1.57±0.06 log cd/m2 p=0.0027). Moreover, the baseline cone threshold parameter exhibited good short-term (intraclass correlation coefficient [ICC]=0.88) and long-term (ICC=0.85) repeatability in all subjects. The cone intercept parameter and ETDRS VA were not significantly different between AMD and older normal subject groups. Cone recovery half-life was significantly different between older normal and AMD subject groups (p=0.041). Neither ETDRS VA nor cone function parameters were significantly different for any group at the 1-year follow-up. Conclusion: The baseline cone threshold shows potential as a novel parameter to assess visual dysfunction in early AMD. This outcome consistently detected deficits in AMD subjects, and differentiated them from age-matched controls with high test–retest repeatability. Keywords: AMD, photobleach, photoreceptor, cone recovery, Ora LUX

John D Rodriguez,1 Patrick R Johnston,1 George W Ousler III,1 Lisa M Smith,1 Mark B Abelson1,21Ora, Inc, Andover, MA, USA; 2Department of Ophthalmology, Harvard Medical School, Boston, MA, USABackground: We have observed that dry eye redness is characterized by a prominence of fine horizontal conjunctival vessels in the exposed ocular surface of the interpalpebral fissure, and have incorporated this feature into the grading of redness in clinical studies of dry eye.Aim: To develop an automated method of grading dry eye-associated ocular redness in order to expand on the clinical grading system currently used.Methods: Ninety nine images from 26 dry eye subjects were evaluated by five graders using a 0–4 (in 0.5 increments) dry eye redness (Ora CalibraTM Dry Eye Redness Scale [OCDER]) scale. For the automated method, the Opencv computer vision library was used to develop software for calculating redness and horizontal conjunctival vessels (noted as "horizontality"). From original photograph, the region of interest (ROI) was selected manually using the open source ImageJ software. Total average redness intensity (Com-Red) was calculated as a single channel 8-bit image as R − 0.83G − 0.17B, where R, G and B were the respective intensities of the red, green and blue channels. The location of vessels was detected by normalizing the blue channel and selecting pixels with an intensity of less than 97% of the mean. The horizontal component (Com-Hor) was calculated by the first order Sobel derivative in the vertical direction and the score was calculated as the average blue channel image intensity of this vertical derivative. Pearson correlation coefficients, accuracy and concordance correlation coefficients (CCC) were calculated after regression and standardized regression of the dataset.Results: The agreement (both Pearson's and CCC) among investigators using the OCDER scale was 0.67, while the agreement of investigator to computer was 0.76. A multiple regression using both redness and horizontality improved the agreement CCC from 0.66 and 0.69 to 0.76, demonstrating the contribution of vessel geometry to the overall grade. Computer analysis of a given image has 100% repeatability and zero variability from session to session.Conclusion: This objective means of grading ocular redness in a unified fashion has potential significance as a new clinical endpoint. In comparisons between computer and investigator, computer grading proved to be more reliable than another investigator using the OCDER scale. The best fitting model based on the present sample, and usable for future studies, was C4 = –12.24 + 2.12C2HOR + 0.88C2RED :C4 is the predicted investigator grade, and C2HOR and C2REDare logarithmic transformations of the computer calculated parameters COM-Hor and COM-Red. Considering the superior repeatability, computer automated grading might be preferable to investigator grading in multicentered dry eye studies in which the subtle differences in redness incurred by treatment have been historically difficult to define.Keywords: conjunctival diseases, classification, diagnosis, humans, hyperemia, image processing, computer-assisted, observer variation, keratoconjunctivitis sicca

John D Rodriguez,1 George W Ousler III,1 Patrick R Johnston,1 Keith Lane,1 Mark B Abelson1,21Ora, Inc, Andover, MA, 2Schepens Eye Research Institute and Harvard Medical School, Boston, MA, USABackground: The purpose of this study was to investigate the occurrence and duration of extended blinks ≥ 70 msec and their associated interblink intervals in normal subjects and in subjects with mild to moderate dry eye.Methods: This single-center, prospective, double-blind study included 11 subjects with dry eye and eight subjects with normal eyes. Extended blinks were defined as lid closure in at least two successive video frames (≥ 70 msec). Digital video imaging of each subject's eyes was recorded while the subject viewed a 10-minute documentary. The subjects did not know that blink was the outcome being measured. Following capture, the videos were manually analyzed in a masked fashion for the occurrence of extended blinks. The length of the interblink interval (ie, time between blinks) before and after these extended blinks (the interblink interval ratio) was calculated, as well as differences in lid contact times.Results: The dry eye group had a median extended blink duration which was 2.53 times longer than that of the normal group. For subjects with dry eye, interblink intervals post-extended blink were significantly longer than interblink intervals pre-extended blink (P < 0.001). Interblink intervals did not lengthen significantly in normal subjects. In both groups, the duration of the extended blink was significantly (P = 0.001) and positively correlated with interblink interval ratio (post-extended to pre-extended blink interblink interval), such that for each doubling of extended blink duration, the interblink interval ratio increased by 10%. Blinks longer than one second in duration occurred almost exclusively in subjects with dry eye.Conclusion: This study reports three central findings: blink duration tended to be longer in subjects with dry eye; a lengthening of the interblink interval after an extended blink occurred in subjects with dry eye but not in those without dry eye; and a longer blink duration was associated with a significantly increased interblink interval ratio in all subjects.Keywords: dry eye, interblink intervals, visual function, visual tasks, diagnostic model

Myofibroblasts are the major source of extracellular matrix components that accumulate during tissue fibrosis, and hepatic stellate cells (HSCs) are believed to be the major source of myofibroblasts in the liver. To date, robust systems to genetically mani

This retrospective study assesses the feasibility of using an 80-lead body surface map (BSM) rather than the conventional 12-lead electrocardiogram (ECG) with exercise stress echocardiograms. A total of 122 patients comprised an 80-lead BSM group (n = 50) and a 12-lead ECG group (n = 72). Both groups had comparable mean maximum predicted heart rates (PHR) and image acquisition periods (P for both groups was < .0001). However, image acquisition in the 80-lead group had a time lapse twice the duration than the 12-lead group (P

Background: Research is needed to understand and address barriers to risk management for women at high (≥20% lifetime) risk for breast cancer, but recruiting this population for research studies is challenging., Objective: This paper compares a variety of recruitment strategies used for a cross-sectional, observational study of high-risk women., Methods: Eligible participants were assigned female at birth, aged 25-85 years, English-speaking, living in the United States, and at high risk for breast cancer as defined by the American College of Radiology. Individuals were excluded if they had a personal history of breast cancer, prior bilateral mastectomy, medical contraindications for magnetic resonance imaging, or were not up-to-date on screening mammography per American College of Radiology guidelines. Participants were recruited from August 2020 to January 2021 using the following mechanisms: targeted Facebook advertisements, Twitter posts, ResearchMatch (a web-based research recruitment database), community partner promotions, paper flyers, and community outreach events. Interested individuals were directed to a secure website with eligibility screening questions. Participants self-reported method of recruitment during the eligibility screening. For each recruitment strategy, we calculated the rate of eligible respondents and completed surveys, costs per eligible participant, and participant demographics., Results: We received 1566 unique responses to the eligibility screener. Participants most often reported recruitment via Facebook advertisements (724/1566, 46%) and ResearchMatch (646/1566, 41%). Community partner promotions resulted in the highest proportion of eligible respondents (24/46, 52%), while ResearchMatch had the lowest proportion of eligible respondents (73/646, 11%). Word of mouth was the most cost-effective recruitment strategy (US $4.66 per completed survey response) and paper flyers were the least cost-effective (US $1448.13 per completed survey response). The demographic characteristics of eligible respondents varied by recruitment strategy: Twitter posts and community outreach events resulted in the highest proportion of Hispanic or Latina women (1/4, 25% and 2/6, 33%, respectively), and community partner promotions resulted in the highest proportion of non-Hispanic Black women (4/24, 17%)., Conclusions: Although recruitment strategies varied in their yield of study participants, results overall support the feasibility of identifying and recruiting women at high risk for breast cancer outside of clinical settings. Researchers must balance the associated costs and participant yield of various recruitment strategies in planning future studies focused on high-risk women., (©Claire C Conley, Jennifer D Rodriguez, McKenzie McIntyre, Bethany L Niell, Suzanne C O'Neill, Susan T Vadaparampil. Originally published in the Journal of Medical Internet Research (https://www.jmir.org), 02.09.2024.)

Purpose: Women with greater than 20-25% lifetime breast cancer risk are recommended to have breast cancer screening with annual mammogram and supplemental breast MRI. However, few women follow these screening recommendations. The objective of this study was to identify barriers and facilitators of screening among women at high risk for breast cancer, guided by the Health Services Utilization Model (HSUM)., Methods: Unaffected high-risk women (N=63) completed semi-structured qualitative interviews exploring their experiences with breast cancer screening. Interviews were audio recorded, transcribed verbatim, and analyzed using a combined deductive and inductive approach., Results: Most participants (84%) had received a screening mammogram; fewer (33%) had received a screening breast MRI. Only 14% had received neither screening. In line with the HSUM, qualitative analysis identified predisposing factors, enabling factors, and need factors associated with receipt of breast cancer screening. Enabling factors - including financial burden, logistic barriers, social support, and care coordination - were most frequently discussed. Predisposing factors included knowledge, health beliefs, and self-advocacy. Need factors included healthcare provider recommendation, family history of breast cancer, and personal medical history. Although HSUM themes were consistent for both mammography and breast MRI, participants did highlight several important differences in barriers and facilitators between the two screening modalities., Conclusion: Barriers and enabling factors associated with supplemental screening for high-risk women represent possible intervention targets. Future research is needed to develop and test multilevel interventions targeting these factors, with the ultimate goal of increasing access to supplemental screening for high-risk women., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Aim: To investigate iron-deficiency anemia as a risk factor for dental pulp disease in children from the central Peruvian jungle., Methodology: A case-control study was carried out with 270 children, of which 90 referred to cases and 180, to controls. Patients with pulp disease were diagnosed according to the criteria of the Association of Endodontists and the American Board of Endodontics. A specific questionnaire was used to assess ferrous sulfate consumption, maternal education level, maternal age, occupation, and household income. Data were analyzed using Pearson's correlation coefficient and a binary logistic regression., Results: Iron deficiency anemia offers a risk factor for pulp disease in children (OR 7.44, IC 95% 4.0-13.8). According to multivariate analysis using binary logistic regression, ferrous sulfate consumption (OR 13.8, IC 95% 5.6.33.9), maternal education level (OR 2.4, IC 95% 1.1-5.3), maternal age (OR 7.5, IC 95% 2.9-19.4), household income (OR 4.0, IC 95% 1.6-9.6), and caries (OR 10.7, IC 95% 4.5-25.7) configured independent factors that were statistically associated with pulp disease., Conclusion: Iron deficiency anemia, ferrous sulfate consumption, maternal education level, maternal age, household income, and dental caries were positively associated with pulp disease in children.

There is a significant need to develop new environmentally friendly, extraction-free sample collection mediums that can effectively preserve and protect genetic material for point-of-care and/or self-collection, home-collection, and mail-back testing. Systematic evolution of ligands by exponential enrichment (SELEX) was used to create anti-ribonuclease (RNase) deoxyribonucleic acid (DNA) aptamers against purified RNase A conjugated to paramagnetic carboxylated beads. Following eight rounds of SELEX carried out under various stringency conditions, e.g., selection using Xtract-Free™ (XF) specimen collection medium and elevated ambient temperature of 28 °C, a panel of five aptamers was chosen following bioinformatic analysis using next-generation sequencing. The efficacy of aptamer inactivation of RNase was assessed by monitoring ribonucleic acid (RNA) integrity via fluorometric and real-time RT-PCR analysis. Inclusion of aptamers in reaction incubations resulted in an 8800- to 11,200-fold reduction in RNase activity, i.e., digestion of viral RNA compared to control. Thus, anti-RNase aptamers integrated into XF collection medium as well as other commercial reagents and kits have great potential for ensuring quality intact RNA for subsequent genomic analyses.

In this study, an alternative method to compendial analytical procedures with enhanced detection and separation capabilities was validated for the quality assessment of glutathione (GSH) drug substance. The related impurities A, B, C, and D present in GSH drug substance were characterized using a one-dimension proton nuclear magnetic resonance (1D 1 H NMR) method on a 600 MHz spectrometer equipped with a liquid nitrogen cryoprobe. Two sample preparations at different pH were optimized to ensure the unambiguous identification of different impurities in the GSH samples. Specifically, impurities A and C in a GSH sample can be tested at pH 3.0, while pH 7.4 is more suitable for testing impurities B and D. The quantitative NMR (qNMR) method was validated following International Council for Harmonisation (ICH) guidelines. The limit of detection (LOD) was less than 0.1% wt for an individual impurity, and the limit of quantitation (LOQ) ranged from 0.14 to 0.24% wt, using about 14 min experimental time per spectrum. Following validation, the qNMR method was applied to assess different commercial GSH bulk substance samples, an in-house compounded GSH drug product, and a GSH dietary supplement product. The method was also applied to monitor GSH degradation (hydrolysis and oxidation) over time to provide quantitative information on GSH degradation and stability. The results suggest that the qNMR method can serve as a highly specific and efficient orthogonal tool for assessing the quality of GSH pharmaceuticals, providing both qualitative and quantitative information on GSH and its related impurities A-D., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Qin Shu reports financial support was provided by US Food and Drug Administration, Center for Drug Evaluation and Research (CDER). Qin Shu reports a relationship with US Food and Drug Administration that includes: employment and funding grants. Mary Schleiff, Cynthia Sommers, Jingyue Yang, Xiaohui Shen, Jason D. Rodriguez, David A. Keire report a relationship with US Food and Drug Administration that includes: employment., (Published by Elsevier B.V.)

Recent treatment advances have resulted in significantly increased survival times following metastatic breast cancer (MBC) diagnosis. Novel treatment approaches-and their related side effects-have changed the landscape of MBC treatment decision-making. We developed a prototype of an online educational tool to prepare patients with MBC for shared decision-making with their oncologists. We describe the five phases of tool development: (1) in-depth, semi-structured qualitative interviews and (2) feedback on storyboards of initial content with patients with MBC and oncology providers. This was followed by three phases of iterative feedback with patients in which they responded to (3) initial, non-navigable website content and (4) a beta version of the full website. In the final phase (5), patients newly diagnosed with MBC (N = 6) used the website prototype for 1 week and completed surveys assessing acceptability, feasibility, treatment knowledge, preparation for decision-making, and self-efficacy for decision-making. Participants in Phase 1 characterized a cyclical process of MBC treatment decision-making and identified key information needs. Website content and structure was iteratively developed in Phases 2-4. Most participants in Phase 5 (n = 4) accessed the website 2-5 times. All participants who accessed the website at least once (n = 5) felt they learned new information from the website prototype and would recommend it to others newly-diagnosed with MBC. After using the website prototype, participants reported high preparation and self-efficacy for decision-making. This multiphase, iterative process resulted in a prototype intervention designed to support decision-making for MBC patients., (© 2024. The Author(s) under exclusive licence to American Association for Cancer Education.)

Introduction: Women with ≥20% lifetime breast cancer risk can receive supplemental breast cancer screening with MRI. We examined factors associated with recommendation for screening breast MRI among primary care providers (PCPs), gynecologists (GYNs), and radiologists. Methods: We conducted a sequential mixed-methods study. Quantitative: Participants ( N  = 72) reported recommendations for mammogram and breast MRI via clinical vignettes describing hypothetical patients with moderate, high, and very high breast cancer risk. Logistic regressions assessed the relationships of clinician-level factors (gender, specialty, years practicing) and practice-level factors (practice type, imaging facilities available) with screening recommendations. Qualitative: We interviewed a subset of survey participants ( n  = 17, 17/72 = 24%) regarding their decision-making about breast cancer screening recommendations. Interviews were audio-recorded, transcribed, and analyzed with directed content analysis. Results: Compared with PCPs, GYNs and radiologists were significantly more likely to recommend breast MRI for high-risk (ORs = 4.09 and 4.09, respectively) and very-high-risk patients (ORs = 8.56 and 18.33, respectively). Qualitative analysis identified two key phases along the clinical pathway for high-risk women. Phase 1 was "identifying high-risk women," which included three subthemes (systems for risk assessment, barriers to risk assessment, scope of practice issues). Phase 2 was "referral for screening," which included three subthemes (conflicting guidelines, scope of practice issues, legal implications). Frequency of themes differed between specialties, potentially explaining findings from the quantitative phase. Conclusions: There are significant differences between specialties in supplemental breast cancer screening recommendations. Multilevel interventions are needed to support identification and management of women with high breast cancer risk, particularly for PCPs.

Analyzing coeluting impurities with similar masses in synthetic oligonucleotides by liquid chromatography-mass spectrometry (LC-MS) poses challenges due to inadequate separation in either dimension. Herein, we present a direct method employing fully resolved isotopic envelopes, enabled by high resolution mass spectrometry (HRMS), to identify and quantify isobaric impurity ions resulting from the deletion or addition of a uracil (U) or cytosine (C) nucleotide from or to the full-length sequence. These impurities may each encompass multiple sequence variants arising from various deletion or addition sites. The method utilizes a full or targeted MS analysis to measure accurate isotopic distributions that are chemical formula dependent but nucleotide sequence independent. This characteristic enables the quantification of isobaric impurity ions involving sequence variants, a capability typically unavailable in sequence-dependent MS/MS methods. Notably, this approach does not rely on standard curves to determine isobaric impurity compositions in test samples; instead, it utilizes the individual isotopic distributions measured for each impurity standard. Moreover, in cases where specific impurity standards are unavailable, the measured isotopic distributions can be adequately replaced with the theoretical distributions (calculated based on chemical formulas of standards) adjusted using experiment-specific correction factors. In summary, this streamlined approach overcomes the limitations of LC-MS analysis for coeluting isobaric impurity ions, offering a promising solution for the in-depth profiling of complex impurity mixtures in synthetic oligonucleotide therapeutics.

Background: Annual screening breast MRI is recommended for women at high (≥ 20% lifetime) breast cancer risk, but is underutilized. Guided by the Health Services Utilization Model (HSUM), we assessed factors associated with screening breast MRI among high-risk women., Methods: From August 2020-January 2021, we recruited an online convenience sample of high-risk women ages 25-85 (N = 232). High-risk was defined as: pathogenic genetic mutation in self or first-degree relative; history of lobular carcinoma in situ; history of thoracic radiation; or estimated lifetime risk ≥ 20%. Participants self-reported predisposing factors (breast cancer knowledge, health locus of control), enabling factors (health insurance type, social support), need factors (perceived risk, screening-supportive social norms, provider recommendation), and prior receipt of screening breast MRI. Multivariable logistic regression analysis with backward selection identified HSUM factors associated with receipt of screening breast MRI., Results: About half (51%) of participants had received a provider recommendation for screening breast MRI; only 32% had ever received a breast MRI. Breast cancer knowledge (OR = 1.15, 95% CI = 1.04-1.27) and screening-supportive social norms (OR = 2.21, 95% CI = 1.64-2.97) were positively related to breast MRI receipt. No other HSUM variables were associated with breast MRI receipt (all p's > 0.1)., Conclusions: High-risk women reported low uptake of screening breast MRI, indicating a gap in guideline-concordant care. Breast cancer knowledge and screening-supportive social norms are two key areas to target in future interventions. Data were collected during the COVID-19 pandemic and generalizability of results is unclear. Future studies with larger, more heterogeneous samples are needed to replicate these findings., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Two decades ago, postmarket discovery of a second crystal form of ritonavir with lower solubility had major implications for drug manufacturers and patients. Since then, ritonavir has been reformulated via the hot-melt-extrusion process in an amorphous form. Here, quantitative low- and mid-frequency Raman spectroscopy methods were developed to characterize polymorphs, form I and form II, in commercial ritonavir 100 mg oral tablets as an alternate analysis approach compared to X-ray powder diffraction (XRPD). Crystallization in three lots of ritonavir products obtained from four separate manufacturers was assessed after storage under accelerated conditions at 40 °C and 75% relative humidity (RH). Results were compared with quantitative XRPD methods developed and validated according to ICH Q2 (R1) guidelines. In a four-week open-dish study, form I crystallization occurred in two of the four products and form II crystallization was detected in another ritonavir product. The limits of detection for XRPD, low-frequency Raman (LFR), and mid-frequency Raman (MFR) were determined to be 0.7, 0.8, and 0.5% for form I and 0.6, 0.6, and 1% for form II, respectively. Root-mean-squared-error of predictions were 0.6-1.0 and 0.6-2.5% for LFR- and MFR-based partial least-squares models. Further, ritonavir polymorphs could also be identified and detected directly from ritonavir tablets using transmission LFR. In summary, LFR was applied for the assessment of polymorphism in real-world samples. While providing analytical performance similar to conventional techniques, LFR reduced the single measurement time from 66 min (XRPD) to 10 s (LFR) without the need for tedious sample preparation procedures.

Background: The rapid detection of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is vital for patient care. The LumiraDx™ SARS-CoV-2 RNA Star Complete (RSC) is an Emergency Use Authorization -recognized molecular test using nasal/nasopharyngeal swabs immersed in a viral/universal transport medium (VTM/UTM). However, there is a critical need for an alternative medium for point-of-care testing (POCT). This study aimed to investigate Xtract-Free (XF), a novel collection medium for transport and direct (extraction-free) use with nucleic acid tests. Methods: Using serially diluted SARS-CoV-2 viral RNA (vRNA) in a routine UTM and XF, a limit of detection (LOD) was established via an RSC test and a quantitative reverse transcription PCR (RT-qPCR). Additionally, the results obtained from a panel of 108 clinical "car-side" nasal swabs collected in XF during the coronavirus pandemic and assessed using the "gold-standard" RT-qPCR assay were compared to Lumira's RSC assay. Results: The average replicate RT-qPCR cycle threshold (C T ) values for vRNA in XF and UTM were observed to be equivalent. An LOD for which five out of five replicates were detected using XF or VTM was approximately 2000 copies/mL. The nasal swabs collected in XF exhibited 93.9% positive percent agreement (sensitivity) and 100% negative percent agreement (specificity) compared to the RT-qPCR. Three specimens tested positive via an RT-qPCR were negative when tested via RSC; however, all three samples had C T values ≥ 36.4. Conclusions: XF is equivalent to VTM/UTM and is compatible for use with the RSC test. Furthermore, XF can be used directly with RT-qPCRs and rapid antigen testing without the requirement for separate nucleic acid extraction (an extraction-free process), making it ideal for cost-effective high-throughput and decentralized respiratory testing. Impact Statement: This study is the first to evaluate LumiraDx's SARS-CoV-2 RNA Star Complete assay in concert with Xtract-Free (XF), a novel collection medium containing a proprietary RNase-inactivating technology for the rapid, "extraction-free" detection of SARS-CoV-2 RNA from clinical nasal swabs. Specimens collected in XF combined with rapid LumiraDx detection provide a safe and sensitive alternative to VTM/UTM, and Molecular Transport medium (MTM) for high throughput, "extraction-free" molecular detection.

Maternal reprogramming of histone methylation is critical for reestablishing totipotency in the zygote, but how histone-modifying enzymes are regulated during maternal reprogramming is not well characterized. To address this gap, we asked whether maternal reprogramming by the H3K4me1/2 demethylase SPR-5/LSD1/KDM1A, is regulated by the chromatin co-repressor protein, SPR-1/CoREST, in Caenorhabditis elegans and mice. In C. elegans, SPR-5 functions as part of a reprogramming switch together with the H3K9 methyltransferase MET-2. By examining germline development, fertility, and gene expression in double mutants between spr-1 and met-2, as well as fertility in double mutants between spr-1 and spr-5, we find that loss of SPR-1 results in a partial loss of SPR-5 maternal reprogramming function. In mice, we generated a separation of function Lsd1 M448V point mutation that compromises CoREST binding, but only slightly affects LSD1 demethylase activity. When maternal LSD1 in the oocyte is derived exclusively from this allele, the progeny phenocopy the increased perinatal lethality that we previously observed when LSD1 was reduced maternally. Together, these data are consistent with CoREST having a conserved function in facilitating maternal LSD1 epigenetic reprogramming., (Published by Oxford University Press on behalf of the Genetics Society of America 2023.)

Nanomaterials have expanded the use of active pharmaceutical ingredients by improving efficacy, decreasing toxicity, and facilitating targeted delivery. To systematically achieve this goal, nanomaterial-containing drugs need to be manufactured with precision in attributes such as size, morphology, surface chemistry, and composition. Their physicochemical characterization is essential as their attributes govern pharmacokinetics yet can be challenging due to the nature of many nanomaterial-based formulations unless advanced sample fixation and in vitro characterization methods are utilized. Here, different cryogenic and other fixation strategies were assessed, and a novel physicochemical characterization method was developed using s canning e lectron R aman c ryo- m icroscopy (SERCM). A complex nanoparticle albumin bound paclitaxel ( nab -paclitaxel) formulation was chosen as a model drug. Plunge freezing (PF), high pressure freezing (HPF), freeze substitution (FS), and membrane filtration were compared for their influence on size and morphology measurements, and formulation-based variations were quantified. SERCM was introduced as a multiattribute physicochemical characterization platform, and the composition of nanoparticles was confirmed as albumin-paclitaxel complexes. By coupling image-based quantitative analysis with chemical analysis, SERCM has the potential to pave the way for the development of comprehensive tools for assessing injectable and ophthalmic nanomaterial-containing drugs in their native-like state.

Pharmaceutical dosage forms such as tablets and capsules are often coated with a functional polymer to modify the drug release. To obtain the drug release profiles, ensure quality control and predict in-vivo performance, dissolution studies are performed. However, dissolution tests are time-consuming, sample destructive and do not readily allow for at-line or in-line characterization. Rapid assessment of functional coatings is essential for products where a single capsule is comprised of hundreds of functionally-coated pellets and the collective drug release kinetics of the entire capsule depends on contributions from each pellet. Here, single Raman measurements were used to evaluate the coating thickness distributions of a dosage form comprised of small, functionally-coated pellets in capsules. First, the composition and physicochemical properties of pellets were characterized by multivariate analysis assisted Raman mapping of pellet cross-sections. Second, a method of collecting single Raman spectrum with spectral contributions from the coating and API layers was developed and optimized to estimate the thickness of coatings. The coating thicknesses obtained from single Raman measurements of pellets in each capsule revealed thickness distributions that correlated with the dissolution profiles (capsules with one distribution had single stage release and capsules with two distributions had a two-stage release). Finally, an unsupervised multivariate analysis method was demonstrated as a rapid and efficient way to correlate dissolution profiles of enterically coated pellets. In summary, this study presents a non-destructive and rapid characterization method for assessing coating thickness and has the potential to be applied in process analytical technologies to ensure coating uniformity and predict product dissolution rate performance., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Pregnenolone (PREG) is an endogenous steroid frequently sold as an over-the-counter dietary supplement touted to promote neurological and immunological health. While the PREG dietary supplement is added to the diet for health benefits, there are no FDA approved PREG drugs. However, compounded PREG drug products are available to U.S. patients. The FDA works with state regulatory authorities on the oversight of compounding activities, including developing 503A and 503B lists of bulk substances that compounders are permitted to use. PREG is one of the substances publicly nominated to be included on the 503B list. Compounded hormone therapies such as those using PREG are of interest given the lack of standardization in compounded drug products which may increase the possibility of underdosing, overdosing, or contamination. However, no USP monograph currently exists to evaluate the quality of PREG drug substance or product. To address knowledge gaps and assist in quality control, a simple and rapid quantitative proton nuclear magnetic resonance spectroscopy (qNMR) method for the identification and assay of PREG in different types of PREG products was developed and validated. PREG samples were characterized using 1D 1 H and 2D 1 H- 13 C HSQC NMR spectra. The qNMR assay method (taking approximately 10 min per NMR spectrum) was validated for precision, accuracy, specificity, robustness and linearity per ICH Q2(R1) guidance. The method was validated in a range from 0.032 to 3.2 mg/mL. As a proof of concept, seven PREG bulk substance samples, three tablet and two capsule PREG dietary supplements were assessed by the qNMR analytical procedure. NMR data from all tested samples met the expected criteria for identification and assay. The results demonstrate the potential of qNMR for the quality assessment of different types of PREG samples., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Qin Shu reports financial support was provided by US Food and Drug Administration, Center for Drug Evaluation and Research (CDER). Qin Shu reports a relationship with US Food and Drug Administration that includes: employment and funding grants. Mary Schleiff, Cynthia Sommers, Jingyue Yang, Xiaohui Shen, Jason D. Rodriguez reports a relationship with US Food and Drug Administration that includes: employment., (Published by Elsevier B.V.)

The last several years have seen an increasing number of examples of transgenerational epigenetic inheritance, in which phenotypes are inherited for three or more generations without changes to the underlying DNA sequence. One model system that has been particularly useful for studying transgenerational epigenetic inheritance is C. elegans. Their short generation time and hermaphroditic reproduction have allowed multiple transgenerational phenotypes to be identified, including aging, fertility, and behavior. However, it is still not clear how transgenerational epigenetic inheritance from the germline affects embryogenesis. Fortunately, the C. elegans embryo has a unique property that makes it ideal for addressing this question: they develop via an invariant lineage, with each cell undergoing stereotypical cell divisions to adopt the same cell fate in every individual embryo. Because of this invariant cell lineage, automated lineage tracing and single-cell RNA-seq can be employed to determine how transgenerational epigenetic inheritance from the germline affects developmental timing and cell fate. Unfortunately, difficulties with these techniques have severely limited their adoption in the community. Here, we provide a practical guide to automated lineage tracing coupled with single-cell RNA-seq to facilitate their use in studying transgenerational epigenetic inheritance in C. elegans embryos., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Delays in healthcare, including breast cancer screening, were documented during the coronavirus disease 2019 (COVID-19) pandemic. However, no studies have examined the impact of COVID-19 on healthcare among women at high (≥20 % lifetime) risk for breast cancer. This study fills that gap. Between August 2020 and January 2021, high-risk women (N = 225) living in the United States (US) completed an online survey assessing COVID-related healthcare disruptions. Descriptive statistics characterized the frequency of breast cancer screening (mammogram and breast magnetic resonance imaging [MRI]) since the beginning of the COVID-19 pandemic. Multivariable linear regression analysis with backward selection examined demographic characteristics associated with COVID-related healthcare disruptions. Since March 2020, 40 % of participants had received a mammogram and 12 % had received a screening breast MRI. On average, participants reported low levels of COVID-related healthcare disruptions ( M  = 1.97 on a 0-4 scale, higher = more disruptions). Participants who were younger (β = -0.21, p  = 0.002) and not working (β = 0.18, p  = 0.009) reported more COVID-related healthcare disruptions. Compared to non-Hispanic White participants, those from any other racial or ethnic group reported fewer COVID-related healthcare disruptions (β = -0.15, p  = 0.020). Although few high-risk women received breast cancer screening after the declaration of the COVID-19 pandemic, they reported overall low levels of COVID-related healthcare disruptions. Results identify subgroups of high-risk women whose healthcare may have been more affected by the pandemic. Efforts to encourage US women at high risk for breast cancer to return to routine preventive care (including breast cancer screening) may need to be targeted towards women who are younger, not working, and non-Hispanic White., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Drs. Conley and O’Neill have received research funding from Pfizer. The remaining authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Authors.)

The release and dissolution of an active pharmaceutical ingredient (API) from the solid oral formulation into the gastrointestinal (GI) tract is critical for the drug's absorption into systemic circulation. Extended-release (ER) solid oral dosage forms are normally subjected to physical shear and grinding forces as well as pressure exerted by peristaltic movements when passing through the GI tract. The complex physical contraction and sample friction exerted by the GI tract are not simulated well by compendial dissolution methods. These limitations render traditional in vitro dissolution testing unable to discriminate and predict a product's in vivo performance. The objective of this study was to develop a dissolution method that better simulates the GI environment that products are subject to when taken by patients. A newly designed Mechanical Apparatus under GI Conditions (MAGIC) was assembled with a dissolution platform and mechanical capabilities to allow in vitro dissolution testing under sample contractions and friction. The dissolution platform, with medium flow-through configuration, was manufactured by 3D printing. A 60 mg polymer matrix-based ER nifedipine product was tested. To simulate GI physiological conditions during the dissolution testing, the flow rate of the medium, and a combination of mechanical compression with rotation induced sample friction at various rotation frequencies were explored. The polymer matrix-based nifedipine ER formulation used here failed its controlled release functionality in the simulated GI environment under mechanical compression and sample friction. The results showed that the MAGIC system, with flow-through configuration under compression and sample friction, has advantages over compendial methods in testing ER solid oral formulations., (Published by Elsevier Inc.)

Tandem mass spectrometry (MS/MS) can provide direct and accurate sequence characterization of synthetic oligonucleotide drugs, including modified oligonucleotides. Multiple factors can affect oligonucleotide MS/MS sequencing, including the intrinsic properties of oligonucleotides (i.e., nucleotide composition and structural modifications) and instrument parameters associated with the ion activation for fragmentation. In this study, MS/MS sequencing of a thymidine (T)-rich and phosphorothioate (PS)-modified DNA oligonucleotide was investigated using two fragmentation techniques: trap-type collision-induced dissociation ("CID") and beam-type CID also termed as higher-energy collisional dissociation ("HCD"), preceded by a hydrophilic interaction liquid chromatography (HILIC) separation. A low to moderate charge state (-4), which predominated under the optimized HILIC-MS conditions, was selected as the precursor ion for MS/MS analysis. Comparison of the two distinctive ion activation mechanisms on the same precursor demonstrated that HCD was superior to CID in promoting higher sequence coverage and analytical sensitivity in sequence elucidation of T-rich DNA oligonucleotides. Specifically, HCD provided more sequence-defining fragments with higher fragment intensities than CID. Furthermore, the direct comparison between unmodified and PS-modified DNA oligonucleotides demonstrated a loss of MS/MS fragmentation efficiency by PS modification in both CID and HCD approaches, and a resultant reduction in sequence coverage. The deficiency in PS DNA sequence coverage observed with single collision energy HCD, however, was partially recovered by applying HCD with multiple collision energies. Collectively, this work demonstrated that HCD is advantageous to MS/MS sequencing of T-rich PS-modified DNA oligonucleotides., (Published 2022. This article is a U.S. Government work and is in the public domain in the USA. Journal of Mass Spectrometry published by John Wiley & Sons Ltd.)

In vitro dissolution testing is widely used to mimic and predict in vivo performance of oral drug products in the gastrointestinal (GI) tract. This literature review assesses the current in vitro dissolution methodologies being employed to simulate and predict in vivo drug dissolution under fasted and fed conditions, with emphasis on immediate release (IR) solid oral dosage forms. Notable human GI physiological conditions under fasted and fed states have been reviewed and summarized. Literature results showed that dissolution media, mechanical forces, and transit times are key dissolution test parameters for simulating specific postprandial conditions. A number of biorelevant systems, including the fed stomach model (FSM), GastroDuo device, dynamic gastric model (DGM), simulated gastrointestinal tract models (TIM), and the human gastric simulator (HGS), have been developed to mimic the postprandial state of the stomach. While these models have assisted in expanding physiological relevance of in vitro dissolution tests, in general, these models lack the ability to fully replicate physiological conditions/processes. Furthermore, the translatability of in vitro data to an in vivo system remains challenging. Additionally, physiologically based pharmacokinetic (PBPK) modeling has been employed to evaluate the effect of food on drug bioavailability and bioequivalence. Here, we assess the current status of in vitro dissolution methodologies and absorption PBPK modeling approaches to identify knowledge gaps and facilitate further development of in vitro dissolution methods that factor in fasted and fed states. Prediction of in vivo drug performance under fasted and fed conditions via in vitro dissolution testing and modeling may potentially help efforts in harmonizing global regulatory recommendations regarding in vivo fasted and fed bioequivalence studies for solid oral IR products., (© 2022. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

This article reviews soft tissue tumors of the head and neck following the 2020 revision of WHO Classification of Soft Tissue and Bone Tumours. Common soft tissue tumors in the head and neck and tumors are discussed, along with newly added entities to the classification system. Salient clinical and imaging features that may allow for improved diagnostic accuracy or to narrow the imaging differential diagnosis are covered. Advanced imaging techniques are discussed, with a focus on diffusion-weighted and dynamic contrast imaging and their potential to help characterize soft tissue tumors and aid in distinguishing malignant from benign tumors., Competing Interests: Disclosure None., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The COVID-19 pandemic created an increased demand for hygiene supplies such as hand sanitizers. In response, a large number of new domestic or imported hand sanitizer products entered the US market. Some of these products were later found to be out of specification. Here, to quickly assess the quality of the hand sanitizer products, a quantitative, through-container screening method was developed for rapid and non-destructive screening. Using spatially offset Raman spectroscopy (SORS) and support vector regression (SVR), active ingredients (e.g., type of alcohol) of 173 commercial and in-house products were identified and quantified regardless of the container material or opacity. Alcohol content in hand sanitizer formulations were predicted with high accuracy [Formula: see text] using SVR and [Formula: see text] of the substandard test samples were identified. In sum, a SORS-SVR method was developed and used for testing medical countermeasures used against COVID-19, demonstrating a potential for high-volume testing during public health threats., (© 2021. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

Introduction: To evaluate effects of peer-led study sessions on performance in a traditionally challenging course, Pharmacy Math, among first-year student pharmacists (P1s)., Methods: Peer-led study sessions were conducted throughout fall 2019 for P1s. Sessions were led by two second-year student pharmacists and focused on study skills and course-related strategies, principles, and content. P1s who attended the majority (at least five) of study sessions were compared to those who attended fewer sessions on student demographics, undergraduate science grade point average, and course outcome (pass/did not pass) using chi-square and independent samples t-tests. Relative risk (RR) was calculated. A sub-analysis of students considered at risk of failing was also conducted., Results: There were 200 P1 participants. Twenty-four students (12%) attended the majority of the sessions and 176 students (88%) attended fewer sessions. Of the 24 students who attended ≥ five study sessions, all passed Pharmacy Math, while 12 of the 176 students who attended fewer sessions failed Pharmacy Math. Students who attended ≥ five sessions had a 6.8% reduction in risk of failing compared to students who attended fewer sessions (RR = 0.93, 95% CI = 0.895, 0.97). More striking, at-risk students who attended ≥ five study sessions had a 17.1% reduction in risk of failing., Conclusions: Peer-led study sessions contribute to reduced risk of failing Pharmacy Math among students who attend a majority of study sessions. Improvements for the future were identified, including mandatory attendance, group structure, and creative ways to cover concepts., Competing Interests: Declaration of Competing Interest Marie Chisholm-Burns serves on the board of directors for the Accreditation Council for Pharmacy Education (ACPE). This manuscript does not represent ACPE or the boards' opinions or views., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Objective: 1) To compare vestibular schwannoma maximum linear dimensions and calculated volume with measured volume in accurately determining tumor volume and growth. 2) To determine natural growth history of vestibular schwannomas utilizing volumetric measurements in an observed patient population., Study Design: Retrospective chart review., Setting: Tertiary academic referral., Patients: One hundred fifty two adults with a vestibular schwannoma who underwent observational management with sequential magnetic resonance imaging (MRI) scans (496 scans)., Intervention: MRI scans., Main Outcome Measures: Tumor volume calculated from linear dimensions compared with measured volume. The percentage change in tumor size (linear or volume) between consecutive MRI scans., Results: The percentage change in tumor size between consecutive MRIs is significantly different between maximum linear dimension (MLD) and measured tumor volume (p = 0.03), but no difference exists in the percentage change between measured and calculated tumor volume (p = 0.882 for three linear measurements, p = 0.637 for two linear measurements). The overall number of growing tumors is 57.2% (n = 87) with an average growth rate of 62.6%. If a criterion for growth of 20% change is used, 32.2% of tumors monitored by linear volume would have demonstrated growth while 57.2% of tumors with measured volume demonstrated growth., Conclusion: Maximum linear dimensions are a significantly less sensitive measure of tumor growth compared with measured volumes. Calculated tumor volume utilizing three linear measurements is an accurate predictor of both measured tumor volume and tumor growth., Competing Interests: The authors disclose no conflicts of interest., (Copyright © 2020, Otology & Neurotology, Inc.)

Importance: A publication reported that N-nitrosodimethylamine (NDMA), a probable human carcinogen, was formed when ranitidine and nitrite were added to simulated gastric fluid. However, the nitrite concentrations used were greater than the range detected in acidic gastric fluid in prior clinical studies., Objective: To characterize NDMA formation following the addition of ranitidine to simulated gastric fluid using combinations of fluid volume, pH levels, and nitrite concentrations, including physiologic levels., Design, Setting, and Participants: One 150-mg ranitidine tablet was added to 50 or 250 mL of simulated gastric fluid with a range of nitrite concentrations from the upper range of physiologic (100 μmol/L) to higher concentrations (10 000 μmol/L) with a range of pH levels. NDMA amounts were assessed with a liquid chromatography-mass spectrometry method., Main Outcomes and Measures: NDMA detected in simulated gastric fluid 2 hours after adding ranitidine., Results: At a supraphysiologic nitrite concentration (ie, 10 000 μmol/L), the mean (SD) amount of NDMA detected in 50 mL simulated gastric fluid 2 hours after adding ranitidine increased from 222 (12) ng at pH 5 to 11 822 (434) ng at pH 1.2. Subsequent experiments with 50 mL of simulated gastric fluid at pH 1.2 with no added nitrite detected a mean (SD) of 22 (2) ng of NDMA, which is the background amount present in the ranitidine tablets. Similarly, at the upper range of physiologic nitrite (ie, 100 μmol/L) or at nitrite concentrations as much as 50-fold greater (1000 or 5000 μmol/L) only background mean (SD) amounts of NDMA were observed (21 [3] ng, 24 [2] ng, or 24 [3] ng, respectively). With 250 mL of simulated gastric fluid, no NDMA was detected at the upper physiologic range (100 μmol/L) or 10-fold physiologic (1000 μmol/L) nitrite concentrations, while NDMA was detected (mean [SD] level, 7353 [183] ng) at a 50-fold physiologic nitrite concentration (5000 μmol/L)., Conclusions and Relevance: In this in vitro study of ranitidine tablets added to simulated gastric fluid with different nitrite concentrations, ranitidine conversion to NDMA was not detected until nitrite was 5000 μmol/L, which is 50-fold greater than the upper range of physiologic gastric nitrite concentrations at acidic pH. These findings suggest that ranitidine is not converted to NDMA in gastric fluid at physiologic conditions.

Complex iron nanoparticle-based drugs are one of the oldest and most frequently administered classes of nanomedicines. In the US, there are seven FDA-approved iron nanoparticle reference drug products, of which one also has an approved generic drug product (i.e., sodium ferric gluconate (SFG)). These products are indicated for the treatment of iron deficiency anemia and are administered intravenously. On the molecular level, iron nanomedicines are colloids composed of an iron oxide core with a carbohydrate coating. This formulation makes nanomedicines more complex than conventional small molecule drugs. As such, these products are often referred to as nonbiological complex drugs (e.g., by the nonbiological complex drugs (NBCD) working group) or complex drug products (e.g., by the FDA). Herein, we report a comprehensive study of the physiochemical properties of the iron nanoparticle product SFG. SFG is the single drug for which both an innovator (Ferrlecit) and generic product are available in the US, allowing for comparative studies to be performed. Measurements focused on the iron core of SFG included optical spectroscopy, inductively coupled plasma mass spectrometry (ICP-MS), X-ray powder diffraction (XRPD), 57 Fe Mössbauer spectroscopy, and X-ray absorbance spectroscopy (XAS). The analysis revealed similar ferric-iron-oxide structures. Measurements focused on the carbohydrate shell comprised of the gluconate ligands included forced acid degradation, dynamic light scattering (DLS), analytical ultracentrifugation (AUC), and gel permeation chromatography (GPC). Such analysis revealed differences in composition for the innovator versus the generic SFG. These studies have the potential to contribute to future quality assessment of iron complex products and will inform on a pharmacokinetic study of two therapeutically equivalent iron gluconate products.

Formation of a zygote is coupled with extensive epigenetic reprogramming to enable appropriate inheritance of histone methylation and prevent developmental delays. In Caenorhabditis elegans , this reprogramming is mediated by the H3K4me2 demethylase SPR-5 and the H3K9 methyltransferase, MET-2. In contrast, the H3K36 methyltransferase MES-4 maintains H3K36me2/3 at germline genes between generations to facilitate re-establishment of the germline. To determine whether the MES-4 germline inheritance pathway antagonizes spr-5; met-2 reprogramming, we examined the interaction between these two pathways. We found that the developmental delay of spr-5; met-2 mutant progeny is associated with ectopic H3K36me3 and the ectopic expression of MES-4-targeted germline genes in somatic tissues. Furthermore, the developmental delay is dependent upon MES-4 and the H3K4 methyltransferase, SET-2. We propose that MES-4 prevents crucial germline genes from being repressed by antagonizing maternal spr-5; met-2 reprogramming. Thus, the balance of inherited histone modifications is necessary to distinguish germline versus soma and prevent developmental delay.This article has an associated 'The people behind the papers' interview., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

Objective: Management of small vestibular schwannomas has evolved to where observation with interval imaging is an accepted treatment strategy. Loss of residual hearing is a known complication of observation. Magnetic resonance imaging (MRI) may provide critical information to assist in determining which tumors are at highest risk of hearing loss. We wished to determine what effect fundal cap size and cochlear fluid-attenuated inversion recovery (FLAIR) signal had on the progression of hearing loss in a large cohort of observed subjects., Study Design: Retrospective chart review., Setting: Tertiary academic referral center., Patients: Three hundred ninety-three adults with a vestibular schwannoma who underwent expectant management with serial audiograms and MRI., Interventions: Audiogram and MRI., Main Outcome Measures: Hearing outcomes included pure-tone average and word discrimination score (WRS). Cochlear FLAIR signal was measured as a ratio between the affected and nonaffected cochlea. Cerebrospinal fluid fundal cap was measured from the most lateral aspect of the tumor to the fundus of the internal auditory canal., Results: An increased cochlear FLAIR ratio was associated with a worse initial WRS (p = 0.0001, β=-0.25). A multivariate regression analysis demonstrated the variables fundal cap and initial WRS to significantly predict change in WRS over time. The larger the fundal cap size, the smaller the change in the WRS (p = 0.047, β=-0.35)., Conclusions: Cerebrospinal fluid fundal cap size predicts the natural history of hearing in vestibular schwannoma patients. The presence of a smaller fundal cap is correlated with a greater risk of progression of hearing loss and should be a variable considered in the management of small vestibular schwannomas., Competing Interests: The authors disclose no conflicts of interest., (Copyright © 2020, Otology & Neurotology, Inc.)

Quality checks, assessments, and the assurance of food products, raw materials, and food ingredients is critically important to ensure the safeguard of foods of high quality for safety and public health. Nevertheless, quality checks, assessments, and the assurance of food products along distribution and supply chains is impacted by various challenges. For instance, the development of portable, sensitive, low-cost, and robust instrumentation that is capable of real-time, accurate, and sensitive analysis, quality checks, assessments, and the assurance of food products in the field and/or in the production line in a food manufacturing industry is a major technological and analytical challenge. Other significant challenges include analytical method development, method validation strategies, and the non-availability of reference materials and/or standards for emerging food contaminants. The simplicity, portability, non-invasive, non-destructive properties, and low-cost of NIR spectrometers, make them appealing and desirable instruments of choice for rapid quality checks, assessments and assurances of food products, raw materials, and ingredients. This review article surveys literature and examines current challenges and breakthroughs in quality checks and the assessment of a variety of food products, raw materials, and ingredients. Specifically, recent technological innovations and notable advances in quartz crystal microbalances (QCM), electroanalytical techniques, and near infrared (NIR) spectroscopic instrument development in the quality assessment of selected food products, and the analysis of food raw materials and ingredients for foodborne pathogen detection between January 2019 and July 2020 are highlighted. In addition, chemometric approaches and multivariate analyses of spectral data for NIR instrumental calibration and sample analyses for quality assessments and assurances of selected food products and electrochemical methods for foodborne pathogen detection are discussed. Moreover, this review provides insight into the future trajectory of innovative technological developments in QCM, electroanalytical techniques, NIR spectroscopy, and multivariate analyses relating to general applications for the quality assessment of food products.

Drug release plays a critical role in defining bioavailability for an extended release solid oral drug products and predictive dissolution tests are desired to establish clinically relevant quality standards for batch release. The objective of this study focuses on exploring the possible impacts of 1 gastrointestinal (GI) parameter for 1 drug: simulated GI contractions on nifedipine release (in 2 extended release solid oral formulations). The 60 mg nifedipine osmotic pump product A, and polymer matrix-based products B and C were examined in the study. An in-house dissolution system was used to simulate various levels of GI contractions on tested samples, and to monitor changes of sample mechanical properties during dissolution testing. The results show that the polymer matrix-based formulation failed to provide controlled release when simulated GI contraction was above 100 g of force. The method may be useful for polymer matrix-based products to assess potential formulation-related interactions with the GI tract during in vivo drug dissolution., (Published by Elsevier Inc.)

A private testing laboratory reported in a Citizen Petition (CP) to FDA that 16 of 38 metformin drug products they tested had N-nitrosodimethyl amine (NDMA) amounts above the allowable intake (AI) of 96 ng/day. Because the FDA had been monitoring drugs for nitrosamines, orthogonal analytical procedures had been developed, validated and applied to detect the following nitrosamines in metformin drug products (if present): (i) NDMA (with a dedicated method) or (ii) NDMA (with a second confirmatory method), N-nitroso-diethylamine (NDEA), N-ethyl-N-nitroso-2-propanamine (NEIPA), N-nitroso-diisopropylamine (NDIPA), N-nitroso-di-n-propylamine (NDPA), N-nitroso-methylphenylamine (NMPA), N-nitroso-di-n-butylamine (NDBA) and N-nitroso-N-methyl-4-aminobutyric acid (NMBA). In contrast to the private laboratory results, FDA testing on the same set of 38 samples with orthogonal procedures observed amounts over the AI in only 8 of the 38 products and generally observed lower values than reported by the private testing laboratory. As described here, the investigation into the cause of the discrepancy revealed that N,N-dimethylformamide (DMF) can interfere with NDMA measurements. The data showed that the use of sufficient mass accuracy in the data acquisition and appropriate mass tolerance setting in the data processing to assure the selectivity of mass spectrometry measurements of NDMA in the presence of co-eluting DMF was necessary to prevent overestimation of the level of NDMA in metformin drug products. Overall, care should be taken to assure the necessary specificity in analytical procedures for adequate assessment of the nitrosamine level in drug products that also contain DMF or other potential interfering substances.

The paclitaxel protein-bound particles for injectable suspension (marketed under the brand name Abraxane®) contains nanosized complexes of paclitaxel and albumin. The molecular interaction between paclitaxel and albumin within the higher-order nanostructure is analytically challenging to assess, as is any correlation of differences to differences in therapeutic effect. However, because the higher-order nanostructures may affect the paclitaxel release, a suitable in vitro assay to detect potential differences in paclitaxel release between comparator lots and products is desirable. Herein, solution NMR spectroscopy with a T 2 -filtering technique was developed to detect paclitaxel signal while suppressing albumin signals to follow the released paclitaxel in the NMR tube upon dilution. The non-invasive nature of NMR allows for precise measurement of a full range of dilution-induced drug release percentage from 14 to 92% without any sample extraction. The critical concentration of the drug product (DP) at 50% of release was 0.63 ± 0.04 mg/mL in PBS buffer. In addition, 2D diffusion ordered NMR spectroscopy (DOSY) results revealed that the released paclitaxel experiencing slightly slowed diffusion rates than free paclitaxel, which was attributed to paclitaxel in equilibrium with albumin-bound states. Collectively, the dilution-based NMR method offered an analytical approach to investigate physicochemical attributes of complex injectable products with minimal needed sample preparation and perturbation to nanoparticle formulation.

Objective: To document the outcome of radioiodine therapy (RIT) in differentiated thyroid cancer (DTC) patients with recent contrasted computed tomography (CCT)., Methods: Eighteen patients with DTC and recent thyroidectomy who underwent RIT within 90 days after a CCT were included. Disease status following RIT and whether the expected response to RIT was achieved were documented. Disease status was classified into one of three categories based on the patient's thyroglobuline level, radioiodine scan (RIS), and other imaging modalities: no evidence of disease (NED), microscopic residual disease (MRD), or gross residual disease (GRD). Expected response to RIT was based on the overall interpretation of the referring physicians of follow up thyroglobuline values, RIS findings and clinical assessment as reflected in progress notes. Follow-up stimulated thyroglobuline and (or) RIS was performed on average 10.8 months after RIT (median 12 months). The last progress note reviewed was on average 33.3 months after RIT (median 31 months)., Results: There were 12 patients with NED, two with MRD and four with GRD. Expected response to RIT was achieved in 17 patients. In one patient, the effectiveness of RIT could not be determined., Conclusion: RIT in postthyroidectomy setting can be successfully performed within 90 days after CCT. Further research is needed to confirm our findings.

Raman mapping is a powerful and emerging tool in characterization of pharmaceuticals and provides non-destructive chemical and structural identification with minimal sample preparation. One pharmaceutical form that is suitable but has not been studied in-depth with Raman mapping is transdermal delivery systems (TDS). TDS are dosage forms designed to deliver a therapeutically effective amount of active pharmaceutical ingredient (API) across a patient's skin. To enhance drug delivery through the skin, the API in the formulation is often close to a saturated or supersaturated state. Thus, improper use or off-label modifications can lead to occurrence of unwanted API changes, specifically, crystallization over time. Here, off-label modifications were mimicked on a set of fentanyl drug-in-adhesive TDS sold on the U.S. market by four different manufacturers via die cutting, and then the die cut TDS were investigated through confocal Raman mapping for structural and chemical changes. Using Multivariate Curve Resolution (MCR), not only was morphological and chemical characterization of transdermal systems provided, but also fentanyl crystals in certain products due to off-label modifications were identified. The chemometric model used in analysis of Raman maps allowed precise identification of fentanyl as the crystalline material as confirmed by the hit-quality-index correlation of component spectra from the chemometric model with library spectra of a fentanyl reference standard. The results show that confocal Raman mapping with MCR can be utilized in assessing pharmaceutical quality of TDS. This method has the potential to be widely used in characterization of such systems as an alternative to existing techniques.