Citations

AbstractNucleic acid amplification testing (NAAT) remains one of the most reliable methods for pathogen identification. However, conventional bulk NAATs may not be sufficiently fast or sensitive enough for the detection of clinically‐relevant pathogens in point‐of‐care testing. Here, we have developed a digital droplet RT‐LAMP (ddRT‐LAMP) assay that rapidly and quantitatively detects the SARS‐CoV‐2 viral E gene in microfluidic drops. Droplet partitioning using ddRT‐LAMP significantly accelerates detection times across a wide range of template concentrations compared to bulk RT‐LAMP assays. We discover that a reduction in droplet diameter decreases assay times up to a certain size, upon which surface adsorption of the RT‐LAMP polymerase reduces reaction efficiency. Optimization of drop size and polymerase concentration enables rapid, sensitive, and quantitative detection of the SARS‐CoV‐2 E gene in only 8 min. These results highlight the potential of ddRT‐LAMP assays as an excellent platform for quantitative point‐of‐care testing.

e23252 Background: The UPMC-Kline Health Center (KHC) in Harrisburg, PA, shows a lower cervical cancer screening rate compared to the general population in the USA (57.08% vs 78%, respectively, based on the Medical Group Fiscal Year 2023 Internal Quality Measures). To understand this gap, we distributed a comprehensive questionnaire to patients, aiming to pinpoint factors contributing to the lower screening rate and plan targeted interventions for improvement. Methods: We randomly distributed questionnaires to female patients aged 21 to 61 during appointment check-in at the KHC, considering our linguistically diverse patient population with surveys available in Spanish, English, and Nepali. Given the diverse linguistic landscape of our patient population, we prepared questionnaires in Spanish, English, and Nepali. The survey aims to identify and address specific clinic-related problems, with a follow-up assessment after a few months to gauge improvements following interventions targeting the underlying causes of screening gaps. Results: Over a three-month span, 65 questionnaires were completed. Among the participants, 38% (25 patients - 7 white/Caucasian, 11 black/African American, 1 Asian, 7 others; 12 Hispanic, 13 non-Hispanic) revealed that they had not undergone screening within the past three years. The analysis of this subgroup unveiled significant issues: 84% (21/25) did not receive paperwork to schedule the test, 88% (22/25) were not contacted to schedule the test, 76% (19/25) did not receive reminder notifications before appointments, and 72%(18/25) expressed concerns about the lack of insurance coverage for costs. Furthermore, 76% (19/25) indicated the requirement to pay a co-pay, and an equal percentage reported difficulties in obtaining the screening test. Additionally, 72%(18/25) mentioned that their healthcare providers did not discuss the screening's importance, while 76%(19/25) noted their providers did not explain the purpose of the test. Conclusions: These findings underscore critical gaps in communication, accessibility, and cost-related barriers to cervical cancer screening at KHC. This ongoing quality improvement project, scheduled for data recollection in a few months, encompasses proposed interventions such as automated reminders, technology integration for communication, and distributing flyers in clinic and waiting areas to foster open communication and address inadequate discussions about the importance of screening. The aim is to improve patient awareness, engagement, and ultimately prevent delayed cancer diagnoses. KHC is committed to enhancing the cervical cancer screening rate to minimize the likelihood of delayed cancer diagnoses.

e23244 Background: The UPMC-Kline Health Center (KHC) located in Harrisburg, PA, has a lower colon cancer screening rate compared to the general patient population in the USA (26.03% vs 78%, respectively, according to the Medical Group Fiscal Year 2023 Internal Quality Measures). To understand the reasons behind this disparity, a comprehensive questionnaire was conducted aiming to identify factors contributing to the lower colonoscopy screening rate and to implement targeted interventions to improve the situation. Methods: Surveys were distributed to eligible patients aged 45 to 75 at the clinic, considering linguistic diversity by preparing questionnaires in Spanish, English, and Nepali. Distribution, randomized and conducted by medical assistants, aims to identify and address specific clinic-related issues. Subsequent assessment, after a few months, will determine improvements resulting from interventions targeting the underlying causes of screening gaps. Results: Over a three-month period, a total of 77 questionnaires were completed and collected. We analyzed responses from 33 patients (42%, 14 females, and 19 males; 16 white/Caucasian, 14 black/African American, 2 Asian, 1 other; 8 Hispanic/Latino, 25 non-Hispanic/Latino) who had not undergone a colonoscopy in the past 10 years. Findings revealed that 72% (24/33) of patients reported not receiving paperwork for scheduling, while 72% (24/33) were not contacted to schedule the test. Additionally, 78% (26/33) of patients did not receive a reminder call or message before their scheduled appointments. 51% (17/33) of patients highlighted a lack of insurance coverage for costs, with 63% (21/33) indicating the need for a co-pay. 57% (19/33) of patients reported difficulties in obtaining the screening test. 51% (17/33) of patients mentioned that their healthcare provider discussed the importance of the screening test, and 54% (18/33) of patients noted that their providers explained the purpose of the test. Conclusions: The survey identifies scheduling, communication, and cost-related barriers to colon cancer screening at KHC. Proposed interventions, including the implementation of automated reminders, integrating technology to facilitate communication, and incorporating flyers within the clinic and patient waiting areas. This quality improvement project is ongoing, with data scheduled for recollection in a few months. These measures are anticipated to foster improvements, aiding more patients in preventing delayed cancer diagnoses. KHC aims to create a patient-friendly environment by addressing scheduling inefficiencies, improving communication, and tackling financial concerns for timely and accessible colon cancer screenings.

538 Background: The phase 3 PALLAS trial compared 2 years of the CDK4/6 inhibitor palbociclib with endocrine therapy of provider choice, versus endocrine therapy alone, as adjuvant treatment for patients (pts) with Stage II-III hormone receptor-positive HER2-negative (HR+/HER2-) breast cancer. The SET2,3 index adjusts the measurement of SETER/PR index of endocrine receptor-related transcriptional activity with a baseline prognostic index (BPI) derived from tumor size (T), number of involved lymph nodes (N) and gene expression levels of ESR1, PGR, ERBB2 and AURKA (RNA4 subtype score). High SET2,3 (index ≥ 2.10) represents high endocrine sensitivity with favorable prognosis. Clinical outcome results from PALLAS were previously published, and the trial now has 5-year median follow-up. Methods: Participants in PALLAS provided a tumor tissue block prior to randomization. RNA was extracted from samples with sufficient tumor tissue and cellularity (>25mm2 with ≥20% cancer nuclei). SET2,3 index was measured from 300ng total RNA using the QuantiGene Plex platform (ThermoFisher, Waltham, MA) following the manufacturer’s protocol and using pre-defined quality control (QC) parameters and cut point. Results from primary tumor samples prior to systemic therapy were analyzed. Invasive disease-free survival (iDFS) was summarized with Kaplan-Meier plots using log-rank test. Cox models were adjusted for treatment arm assignment, with a treatment arm by SET index value interaction term to test for prediction of palbociclib benefit. The pre-defined level of significance was a two-sided 0.05. Results: There were 4075 pts from the total PALLAS population (N=5796) who provided a primary tumor sample that met pre-analytical requirements to perform the SET assay, from which 3388 (83%) results passed quality control, including 3093 from a sample obtained prior to receiving systemic therapy (1559 in palbociclib arm and 1534 in the control arm). SET2,3 index could not be calculated for 12 pts due to missing tumor or nodal information. There was no evidence of an interaction between treatment arm and SET indices (continuous) for iDFS: SET2,3 index interaction p=0.12 and SETER/PR index interaction p=0.92. Estimated 5-year iDFS was superior for those with high SET2,3: iDFS was 91.4% (95%CI 89.9–92.8) in 1719 (55.8%) pts with high SET2,3, versus 78.5% (95%CI 76.0–80.8) in 1362 (44.2%) pts with low SET2,3 (HR 0.38, 95%CI 0.31–0.47, p<0.0001). The SETER/PR index was also significantly associated with iDFS (HR 0.70 per 0.5 units, 95%CI 0.65–0.77, p<0.0001). Conclusions: There was no evidence that the SET2,3 or SETER/PR index of endocrine-related transcriptional activity predicts benefit from palbociclib treatment. SET2,3 index was strongly prognostic for pts with Stage II-III breast cancer, with excellent 5-year IDFS for pts classified as SET2,3 high. Clinical trial information: NCT02513394 .

: Cerebral Edema (CE) is the final common pathway of brain death. In severe neurological disease, neuronal cell damage first contributes to tissue edema, and then Increased Intracranial Pressure (ICP) occurs, which results in diminishing cerebral perfusion pressure. In turn, anoxic brain injury brought on by decreased cerebral perfusion pressure eventually results in neuronal cell impairment, creating a vicious cycle. Traditionally, CE is understood to be tightly linked to elevated ICP, which ultimately generates cerebral hernia and is therefore regarded as a risk factor for mortality. Intracranial hypertension and brain edema are two serious neurological disorders that are commonly treated with mannitol. However, mannitol usage should be monitored since inappropriate utilization of the substance could conversely have negative effects on CE patients. CE is thought to be related to bloodbrain barrier dysfunction. Nonetheless, a fluid clearance mechanism called the glial-lymphatic or glymphatic system was updated. This pathway facilitates the transport of cerebrospinal fluid (CSF) into the brain along arterial perivascular spaces and later into the brain interstitium. After removing solutes from the neuropil into meningeal and cervical lymphatic drainage arteries, the route then directs flows into the venous perivascular and perineuronal regions. Remarkably, the dual function of the glymphatic system was observed to protect the brain from further exacerbated damage. From our point of view, future studies ought to concentrate on the management of CE based on numerous targets of the updated glymphatic system. Further clinical trials are encouraged to apply these agents to the clinic as soon as possible.

AbstractDNA damage plays a significant role in the tumorigenesis and progression of the disease. Abnormal DNA repair affects the therapy and prognosis of cancer. In this study, it is demonstrated that the deubiquitinase USP25 promotes non‐homologous end joining (NHEJ), which in turn contributes to chemoresistance in cancer. It is shown that USP25 deubiquitinates SHLD2 at the K64 site, which enhances its binding with REV7 and promotes NHEJ. Furthermore, USP25 deficiency impairs NHEJ‐mediated DNA repair and reduces class switch recombination (CSR) in USP25‐deficient mice. USP25 is overexpressed in a subset of colon cancers. Depletion of USP25 sensitizes colon cancer cells to IR, 5‐Fu, and cisplatin. TRIM25 is also identified, an E3 ligase, as the enzyme responsible for degrading USP25. Downregulation of TRIM25 leads to an increase in USP25 levels, which in turn induces chemoresistance in colon cancer cells. Finally, a peptide that disrupts the USP25‐SHLD2 interaction is successfully identified, impairing NHEJ and increasing sensitivity to chemotherapy in PDX model. Overall, these findings reveal USP25 as a critical effector of SHLD2 in regulating the NHEJ repair pathway and suggest its potential as a therapeutic target for cancer therapy.

Background: Dietary nitrate consumption is associated with cardiometabolic health in many but not all studies. The enterosalivary nitrate-nitrite-nitric oxide (NO 3 - NO 2 - NO) pathway requires the oral microbiome for NO production (a signaling molecule beneficial to cardiometabolic health) following nitrate intake. We hypothesize that the oral microbiome modifies the cardiometabolic benefits of nitrate intake. Methods: We analyzed cross-sectional data from 657 ORGINS participants. Dietary nitrate intakes and alternative healthy eating index (AHEI) scores were calculated from food frequency questionnaire responses and a US food database. Subgingival 16S rRNA microbial genes (Illumina, MiSeq) were sequenced, and PICRUSt2 estimated metagenomic content. The Microbiome Induced Nitric oxide Enrichment Score (MINES) was calculated as a microbial gene abundance ratio representing enhanced net capacity for NO generation. Cardiometabolic biomarker (systolic and diastolic blood pressure, HbA1C, glucose, insulin, and HOMA-IR) Z-scores were averaged to form a cardiometabolic Z-score (CMZ). Adjusted multivariable linear models regressed CMZ and individual Z-scores on nitrate intake tertiles. Results: Participants were mean age 31±9, 72% female, 31% White, 14% Black, and 28% Hispanic. Mean nitrate intake was 189±171 mg/day. Higher nitrate intake was significantly associated with lower CMZ, glucose, insulin, and HOMA-IR. Mean CMZ(SE) across dietary nitrate tertiles were 0.53(0.30), 0.35(0.31), and -0.11(0.30), p for trend=0.04; findings were attenuated after adjustment for AHEI (p for trend=0.12). In subgroup analysis by MINES level, higher nitrate intake was only associated with lower cardiometabolic biomarker levels among participants with a <median MINES even after AHEI adjustment (Table). Conclusion: Higher dietary nitrate intake was associated with lower cardiometabolic risk biomarker levels only among those with reduced potential for oral microbiome-dependent NO generation.

Abstract Background: Low levels of HER2 expression (HER2-low) have recently emerged as a therapeutic target in patients (pts) with breast cancer (BC). The aim of this study was to evaluate clinical and demographic variables associated with HER2 expression, as well as the value of HER2-low as a prognostic and predictive biomarker for palbociclib benefit in pts enrolled in the PALLAS trial. Methods: The phase III PALLAS trial investigated the addition of palbociclib to adjuvant endocrine therapy (ET) in pts with stage II-III hormone receptor-positive BC. The aims of this analysis are (1) to assess the association between the presence of low levels of HER2 expression with demographic and clinicopathological parameters, (2) to test the prognostic value of HER2-low status on invasive disease-free survival (IDFS), distant relapse-free survival (DRFS), and overall survival (OS) and (3) to assess the value of HER2 expression as a predictive biomarker of response to palbociclib. HER2-low was defined as HER2 immunohistochemistry (IHC) 1+ or IHC 2+ with negative in situ hybridization; all pathologic evaluation was performed locally. Pts with HER2-0 and Her2-low BC were included in this analysis; those with HER2-positive BC or missing HER2 status were excluded. Association of HER2 with other baseline covariates were tested with Chi-squared tests. Prognostic and predictive power of HER2 were assessed with Cox Models. Multivariable models included age, T-stage, N-stage, grade and progesterone receptor (PR) expression. Hazard ratios (HR) and 95% Confidence intervals (CI) are reported. Results: From the original PALLAS intention-to-treat population (N = 5,753), 5304 pts (92.2%) were included in this analysis. Among these, 2,254 pts (42.5%) were classified as HER2 IHC 0 (HER2-0) and 3,050 (57.5%) as HER2-low (1,838 with IHC 1+ and 1,212 with IHC 2+). For this analysis, median follow-up was 59.8 months. Compared to HER2-0, pts with HER2-low tumors had statistically higher estrogen receptor expression levels (mean expression 89.2% vs 88.2%, p = 0.019), lower PR expression levels (65.9% vs 68.2%, p = 0.007). Importantly, HER2-low status varied significantly across 21 participating countries (range 16.7% to 75.6%; p&lt; 0.001) and was more frequent in pts enrolled in North America (63.1%) than in Europe (53.4%) and other regions (53.4%) (p &lt; 0.001). No differences in HER2 expression were observed according to anatomic stage, T-stage, N-stage, histological grade, age, menopausal status, primary surgery type, prior radiation, prior chemotherapy, or baseline performance status. There were no statistically significant differences in IDFS, DRFS or OS according to HER2 status in univariate or multivariable Cox models (prognostic value, HR provided in the Table). There was no significant interaction between the HER2 status and the benefit to palbociclib in IDFS, DRFS or OS (Table). Similar results were obtained when HER2-0 was compared to HER2 1+ and HER2 2+ separately. Conclusions: In this large, prospective and international cohort, no differences were observed in clinical parameters, prognosis, or differential benefit from palbociclib (predictive value) between HER2-0 and HER2-low tumors. Significant geographic variability was observed in the prevalence of HER2-low status, suggesting a high degree of variation in pathologic assessment of HER2 expression without impact on outcomes. Support: AFT, Abbvie Pfizer; Clinicaltrials.gov: NCT02513394 https://acknowledgments.alliancefound.org Citation Format: Guilherme Nader-Marta, Christian F. Singer, Dominik Hlauschek, Angela DeMichele, Paolo Tarantino, Georg Pfeiler, Miguel Martín, Justin Balko, Zbigniew Nowecki, Marija Balic, Adam Brufsky, Arlene Chan, Patrick G. Morris, Tufia Haddad, Sibylle Loibl, Yuan Liu, Lidija Sölkner, Christian Fesl, Erica Mayer, Michael Gnant. Clinical characterization, prognostic and predictive values of HER2-low in early breast cancer in the PALLAS trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-01-13.