Faculty Bibliography

OBJECTIVES/OBJECTIVE:The gut microbiome plays a crucial role in regulating systemic immunity and has been implicated in several chronic inflammatory diseases. Intestinal expansions of Ruminococcus gnavus (RG), a dominant gut commensal, correlate with disease flares in lupus nephritis (LN), but the underlying mechanism remains unknown. METHODS:In a Pilot cohort of patients with biopsy-proven LN, subsetted by gut microbiota community, immune status was characterised using bulk-blood RNA sequencing libraries, serum levels of representative host proteins, and levels of immunoglobulin (Ig)G antibodies to the novel lipoglycan (LG) produced by pathogenic RG strains. A Validation LN cohort was evaluated for blood transcriptomic profiles and levels of anti-LG antibodies. In murine models, mechanistic hypotheses were tested after RG gut colonisation or after intraperitoneal injection with an LG preparation, with outcomes determined by transcriptomic analyses, platelet functional readouts, and tissue histology. RESULTS:In a Pilot cohort of patients with LN, RG gut expansions were associated with high-level platelet, neutrophil, and monocyte activation. Serum levels of platelet factor 4 and release of neutrophil extracellular traps (NETs) were significantly higher in patients with high serum IgG antibody against the novel RG-specific LG, a marker of in vivo immune exposure. An LN Validation cohort confirmed these correlates and showed that anti-LG antibodies serve as a surrogate for thromboinflammatory profile in this LN-associated endotype. In mice, gut colonisation with LG-producing RG strains or a single LG injection caused megakaryocytosis and platelet activation; RG colonisation with LG-producing strains induced tubulointerstitial injury with NETosis. In vivo responses to LG toxin were Toll-like receptor 2-dependent. CONCLUSIONS:Gut expansions of the RG pathobiont may contribute to autoimmune pathogenesis through the LG toxin and cause LN flares through thromboinflammatory mechanisms in this previously unrecognised LN endotype.

BACKGROUND AND AIMS/OBJECTIVE:There are currently no brief quantitative assessments that capture the drug patterns of people who engage in use of more than one drug on the same day or simultaneously. The current study examined the retest reliability, acceptability and feasibility of a new quantitative assessment to measure polysubstance use. DESIGN/METHODS:A tool for assessing simultaneous and same-day polysubstance behaviors, the polysubstance assessment tool (PAT) was developed in interviewer-administered and electronic self-administered formats. Participants were allocated 1:1 to receive either version of the PAT and returned one to three days later to repeat the assessment. SETTING/METHODS:New York City, New York, USA. PARTICIPANTS/METHODS:Adults (18 + years, n = 115) who reported use of more than one drug per day in the last 30 days. MEASUREMENTS/METHODS:Test-retest reliability estimates for dichotomous items were assessed using Cohen's kappa, Gwet's Agreement Coefficient 1 (AC1) and percent agreement. Continuous items were assessed with two-way mixed effects intraclass correlations. Bivariate analyses examined acceptability using nine Likert-type survey questions. Feasibility was examined via time to completion. FINDINGS/RESULTS:Overall reliability was moderate to excellent [Gwet's AC1 range 0.70-0.96; intraclass correlation (ICC) range 0.62-0.88]. Reliability was higher for simultaneous polysubstance use (Gwet's AC1 = 0.90) as compared with same-day (Gwet's AC1 = 0.70). Acceptability was high, with no statistically significant difference between the self- and interviewer-administered versions of the tool. Median time to completion was 7 minutes, and was statistically significantly lower for the self-administered tool (median = 5 minutes) compared with the interviewer-administered version (median = 8 minutes) (P < 0.001). CONCLUSIONS:A new polysubstance assessment tool appears to have good reliability and can be considered by researchers seeking a quantitative measure of polysubstance use behaviors given its simplicity, high acceptability and quick completion time.

BACKGROUND/UNASSIGNED:Effective management of alcohol withdrawal syndrome during hospitalization is paramount to patient safety and quality care. NYC Health + Hospitals initiated a quality improvement project to pilot an electronic health record (EHR) integrated, nurse-driven CIWA-Ar symptom-triggered protocol, including recommendations for medications for alcohol use disorder (MAUD), in medical and surgical units at 3 public hospitals. OBJECTIVE/UNASSIGNED:To describe implementation processes and to report related implementation outcomes (appropriateness, feasibility, and adoption) of the updated CIWA-Ar protocol in a safety net hospital setting. METHODS/UNASSIGNED:NYC Health + Hospitals implemented a standardized CIWA-Ar symptom-triggered, nurse-driven EHR protocol on March 15, 2022. The protocol included order sets, practice advisories, task lists, and reminders for assessments and orders. We measured nursing perspectives on feasibility and appropriateness at 6 months via a survey. We measured provider adoption as the proportion of admissions with a CIWA-Ar protocol ordered among admissions that triggered a recommendation, and MAUD use as the proportion of admissions with a MAUD order during hospitalization among all patients with a protocol ordered. RESULTS/UNASSIGNED:= .249). CONCLUSIONS/UNASSIGNED:The CIWA-Ar protocol was appropriate, feasible, and adopted at NYC public hospitals. Quality improvements to ensure protocol fidelity with benzodiazepine dosing and MAUD prescribing are needed.

PURPOSE/OBJECTIVE:Pancreatic cystic lesions (PCL) commonly undergo surveillance using MRI with MR cholangiopancreatography (MRCP). Our objective is to compare the performance of a single-shot fat-saturated T2-weighted technique with deep-learning reconstruction (DL HASTE-FS) to a conventional T2-weighted Half fourier Single-shot Turbo spin-Echo (HASTE) sequence and to MRCP for the purpose of PCL detection, characterization, and surveillance. METHODS:In this retrospective study, 91 consecutive patients underwent 3T abdominal MRI with MRCP protocol including DL HASTE-FS and conventional HASTE between 8/2/2023 and 10/3/2023. Three abdominal radiologists rated overall and lesion-specific image quality on a 5-point Likert scale, including pancreatic margin and duct sharpness, and PCL conspicuity. A subset of 70 preselected index PCLs were evaluated for cyst features, confidence of diagnosing side-branch IPMN, and suitability of DL HASTE-FS in replacing MRCP for PCL surveillance. RESULTS:DL HASTE-FS received higher scores for pancreatic duct border sharpness (4.1 vs. 3.9; p = .004), pancreatic duct visibility compared to MRCP (2.0 vs. 1.9; p = .04), cyst conspicuity (4.4 vs. 3.9; p < .001), and sharpness of cyst wall and internal septations (4.3 vs. 3.7; p < .001) compared to conventional HASTE. In contrast, conventional HASTE received higher scores for pancreatic margin sharpness (4.2 vs. 3.8; p < .001) and peripancreatic vessel clarity (4.2 vs. 3.4; p < .001). For the 70 preselected index PCLs, readers visualized more PCLs and had higher confidence in diagnosing SB-IPMN on DL HASTE-FS than on conventional HASTE (3.6 vs. 3.4; p < .001). Finally, DL HASTE-FS was deemed a suitable replacement to MRCP for more cases than conventional HASTE (83% vs. 48%; p < .001). CONCLUSION/CONCLUSIONS:DL HASTE-FS outperforms conventional HASTE for PCL detection and characterization, and is a suitable alternative to 3D MRCP in the context of PCL surveillance, potentially reducing exam time and cost.

RAS proteins control signals required for cell growth and survival and, when constitutively activated by mutation, can drive oncogenesis. RAS proteins are primarily regulated by their GTP or GDP binding state, which is controlled by guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs). RAS proteins are also substrates for dozens of posttranslational modifications (PTMs) that target them to membranes and serve as a secondary means of regulation. Because the newly developed direct RAS inhibitors do not produce durable responses in RAS-dependent cancer, there is renewed interest in targeting the PTMs of RAS. These modifications are the subject of this review.

BACKGROUND/UNASSIGNED:The metabolic mechanisms underlying right ventricular (RV) dysfunction are poorly understood, particularly outside of group 1 pulmonary hypertension (PH). We aimed to identify metabolites and pathways associated with RV systolic function and explored whether associations differed by pulmonary vascular resistance, PH group 1 status, and sex. METHODS/UNASSIGNED:We analyzed data from the multicenter PVDOMICS (Pulmonary Vascular Disease Phenomics) cohort. RV systolic function metrics included fractional area change (echo), global longitudinal strain (echo), and ejection fraction (cardiac magnetic resonance). We used linear regression adjusted for age, sex, body mass index, and PH group to assess associations between metabolites and RV function. Pathway enrichment analyses were used to identify pathways significantly associated with RV function. Interaction terms were assessed to determine whether metabolite associations were modified by pulmonary vascular resistance, group 1 PH status, or sex. Least absolute shrinkage and selection operator regression was used to develop metabolite-based scores for RV function, and prognostic performance was assessed. RESULTS/UNASSIGNED:There were 979 participants with plasma metabolomics and RV function data. Linear regression identified 170 metabolites that were significantly associated with all 3 RV metrics. Androgenic steroid, gamma-glutamyl amino acid, polyamine, vitamin A, fatty acid, and sterol pathways are most strongly associated with RV systolic function. Two metabolites interacted with group 1 PH status, and 6 interacted with pulmonary vascular resistance. Four androgenic steroids are associated more strongly with RV systolic function in females compared with males. Metabolite-based scores were prognostically equivalent to RV systolic function metrics and less accurate than REVEAL Lite 2 scores. CONCLUSIONS/UNASSIGNED:We provide a blueprint of metabolites and metabolic pathways associated with RV systolic function across the spectrum of PH. Novel links to vitamin A and glutathione metabolites were observed. We detected few metabolites that associated with RV systolic function differentially by group 1 PH status or degree of pulmonary vascular resistance elevation. Androgenic steroids may associate more strongly with RV systolic function in females compared with males.

BLU-451 is a potent and selective tyrosine kinase inhibitor designed to target uncommon epidermal growth factor receptor (EGFR) mutations, spare wild-type EGFR, and be active in the central nervous system (CNS). In vitro EGFR enzyme assays and engineered cell line models revealed selectivity and anti-proliferative potency of BLU-451 against a wide range of EGFR mutations in non-small cell lung cancer, including common mutations, atypical mutations, and exon 20 insertions. Particularly, BLU-451 demonstrated robust antitumor activity in EGFR exon 20 insertion cell-derived and patient-derived xenograft models and was well tolerated in animal studies. Pharmacokinetic and pharmacodynamics analyses showed that BLU-451 suppressed exon 20 insertion phosphorylated EGFR expression levels within tumor tissues but not in skin and intestinal tissues, potentially indicative of lower toxicity associated with wild-type EGFR. In intracranial tumor models with imaging-based analyses, BLU-451 showed CNS antitumor efficacy. Early clinical data from patients enrolled in the phase 1 portion of the phase 1/2 CONCERTO trial (NCT05241873) demonstrated the overall clinical potential of BLU-451, including its CNS antitumor activity in patients with EGFR exon 20 insertions and atypical mutations. Initial data suggest that BLU-451 is an active molecule. Further evaluation of the safety profile and pharmacokinetic properties of BLU-451 is needed.

Small GTPases play important roles in cellular signaling. Due to their small sizes (∼21 kDa), structural studies of small GTPases have been predominantly performed using x-ray crystallography in which crystal lattice contacts made it challenging to define unperturbed conformations of the key switch regions. Here, we develop a protein-engineering strategy that enables cryo-EM analysis of small soluble proteins and applied to RAS. We fused the C-terminal α5 helix of the RAS globular domain to a small protein BRIL by forming a continuous helix, which leaves most RAS surfaces exposed to the solvent and unperturbed, followed by the complex formation with an anti-BRIL Fab. This engineered complex with an increased molecular weight, termed "RAS-lollipop", enabled single-particle cryo-EM of RAS. Using this approach, we determined the cryo-EM structure of NRAS, whose structural studies using crystallography have been the least successful among the RAS isoforms. We revealed the conformations of the switch region and α 5 helix that differ from those observed in published crystal structures, and also defined the binding site of an NRAS-specific monobody. We uncovered an unexpected surfactant-like property of this monobody, which reduces orientation biases of particles on cryo-EM grids. Together, this work establishes a platform for visualizing small GTPases and potentially other small proteins with minimal perturbation of their surfaces.