Significance Level Research Articles

Neuromuscular Responses to 5 K Time Trial Load Carried by Spanish Army Marines

One of the physical requirements for marines involves covering a specific distance while carrying individual combat gear, supplies, or other military equipment across challenging terrain. Training for this physically and mentally demanding task is a routine component of their preparation. However, further research is needed to better understand the neuromuscular impact of such demanding efforts, strenuous maneuvers, and the recovery process in the subsequent days. Twenty-nine marines completed a 5 km time trial while carrying 24 kg of combat gear, undergoing evaluations at four time points: pre exercise, immediately post exercise, 24 h post exercise, and 48 h post exercise. Repeated measures ANOVA, paired samples t-test, and effect size (ES) analysis were conducted, presenting the results as the mean ± standard deviation (SD). The significance level was set at p ≤ 0.05. Several variables and their corresponding p-values demonstrated changes over time, including the following: the isometric mid-thigh pull (IMTP) (p = 0.001); countermovement jump height (VJ CMJ) (p ≤ 0.001); rating of fatigue scale (ROF) (p ≤ 0.001); blood lactate (BL) levels (p ≤ 0.001); maximum pull-ups (PUmax) (p ≤ 0.001); body mass (BM) (p ≤ 0.001); dominant hand grip strength (DHGS) (p = 0.406); and non-dominant hand grip strength (NDHGS) (p = 0.805). Incident reports and perceived effort (IRPE) revealed a progressive and significant increase between the first and last kilometer of the test, specifically in perceived variables of fatigue, muscle pain, joint pain, shortness of breath, palpitations, excessive sweating, and muscle tremors (all p ≤ 0.001). These findings may contribute to optimizing training programs to better align with operational demands, thereby improving task performance and overall mission effectiveness. In conclusion, the military test had a significant neuromuscular impact on the body, initially resulting in the potentiation of absolute global isometric strength and lower-limb power. However, these values declined below the baseline levels at 24 and 48 h post test.

Meta 95-95-95 durante a infância e pré-adolescência na América Latina e Caribe Latina e Caribe

Highlights The 95-95-95 targets set by the Joint United Nations Program on HIV/AIDS (UNAIDS) aim to reduce new infections, discrimination and deaths from HIV to zero. The rates for achieving the 95-95-95 targets are much lower than expected, especially for the group of children and teenagers. In the countries analyzed, only Guatemala met the first target 95. The second target was met by ten countries, and the third target was met by only four countries. Despite the increase in strategies for controlling HIV, there are many inequalities between countries, contributing to the vulnerability of some population groups, such as children and teenagers. Introduction: The Joint United Nations Programme on HIV/AIDS (UNAIDS) has set the 95-95-95 targets. It is expected that 95% of HIV-positive individuals will know their HIV status and of these, 95% will be on antiretroviral therapy (ART) and 95% will achieve viral suppression. Objective: To analyze the distribution and spatial autocorrelation of the 95-95-95 targets among children and adolescents living with HIV/AIDS in Latin America and the Caribbean. Materials and Methods: An epidemiological ecological study using data from AIDSinfo by the Joint United Nations Programme on HIV/AIDS (UNAIDS) on the 95-95-95 targets among children and adolescents between 2015 and 2022. The analysis was performed using Moran's spatial autocorrelation coefficient (global and local), considering a 5% significance level. Results: A total of 52,000 and 42,000 cases of HIV/AIDS among children and adolescents in Latin America and the Caribbean were analyzed, respectively, for the period from 2015 to 2022. Disparities in the targets were identified between countries, with significant spatial autocorrelation for the second target of 95, showing a value of 0.375 (p-value 0.017). Discussion: Among the 27 countries included in the study, only Guatemala met the first 95 target. To reach the goal, strategies are needed to increase access to HIV testing, with more accessible counseling and testing services. Conclusion: There are discrepancies between countries in Latin America and the Caribbean in meeting and increasing the percentage of the 95-95-95 targets. The study allowed the identification of priority areas for attention, highlighting the need for new strategies and policies tailored to each locality, assisting in the achievement of the targets. How to cite this article: Piran, Camila Moraes Garollo; De Oliveira, Natan Nascimento; Mori, Mariana Martire; De Oliveira, Rosana Rosseto; Villarroel Yañez, Leslie; Carmona-Gutiérrez, Andrés Antonio; Furtado, Marcela Demitto . Target 95-95-95 during childhood and preadolescence in Latin America and the Caribbean. Revista Cuidarte. 2025;16(1): e4186. https://doi.org/10.15649/cuidarte.4186

Abstract 3300: A longitudinal, circulating tumor molecular response biomarker as a predictor of clinical outcomes in a real-world cohort of patients with advanced solid tumors treated with tyrosine kinase inhibitors

Abstract Studies have shown that early changes in quantitative measures of circulating tumor DNA (ctDNA) in response to immunotherapy can predict clinical outcomes and may be useful for identifying molecular responders (MRs) and molecular non-responders (nMRs) who may benefit from a change in therapy. Yet few studies have evaluated treatment with tyrosine kinase inhibitors (TKis). The cohort consisted of deidentified patients from the Tempus clinicogenomic database with stage IV solid tumors who underwent ctDNA NGS. Patients who had a pre-treatment baseline liquid biopsy (T0) and a longitudinal sample 21-180 days after initiating TKi therapy (T1) were included. TKis were classified by the Tempus Medical Ontology, which was validated using the NCI Metathesaurus TKi concept (NCI Code C1967). xM for treatment response monitoring estimates tumor fraction (TF) via an ensemble algorithm that incorporates pathogenic variant allele frequencies, copy number information, and germline information. Evaluable patients had at least one blood sample with a TF ≥ limit of blank of 0.09%. MR was quantified as a ≥ 50% decrease in TF from T0 to T1. Patients with consistently low TF < 1% at both T0 and T1 were also classified as MRs given previous data demonstrating that baseline low TF is correlated with prolonged clinical outcomes. All other patients were classified as nMRs. Real-world overall survival (rwOS) was defined from T1 to death or last known clinical record in event-free patients. Stratified Cox proportional hazards models were used to estimate the hazard ratio (HR) for MR status (MR vs. nMR), stratified by line of therapy (LOT, first line, 1L vs. ≥ second-line, 2L+). A 1-sided Wald test was conducted using a 5% significance level. Predicted rwOS estimates were obtained from the Cox models. The evaluable cohort consisted of 109 patients (1L=62, 57%, 2L+=47, 43%). Cancer types included non-small cell lung cancer (NSCLC, n=52, 48%), breast cancer (BC, n=35, 32%), and 10 other cancer types each comprising < 5% of the patient cohort. The most common therapies were osimertinib in NSCLC patients (n=34) and alpelisib + fulvestrant in BC patients (n=11). Median time from the start of TKi therapy to T1 was 93 days and median follow-up was 10.3 months. Sixty patients had MR and 49 patients were classified as nMRs. MR was significantly associated with longer rwOS than nMRs (HR = 0.45; P = 0.006). Median predicted rwOS among MRs receiving TKis in 1L (n=43) was 28.4 months vs. 16.5 months for nMRs (n = 19). The HR for death between MR vs nMR was consistent across different cancer subgroups and LOT: HR=0.38 for NSCLC patients, HR=0.57 for BC patients, HR=0.46 for 1L patients and HR=0.44 for 2L+ patients. These data demonstrate that longitudinal molecular treatment response can stratify rw-outcomes to TKi therapy in a heterogeneous, real-world pan cancer cohort. Citation Format: John Guittar, Akash Mitra, Rossin Erbe, Shannon Mulmat, Michelle Stein, Xifeng Wang, Seung Won Hyun, Chithra Sangli, Halla Nimeiri, Rotem Ben-Shachar, Wade T. Iams, Charu Aggarwal. A longitudinal, circulating tumor molecular response biomarker as a predictor of clinical outcomes in a real-world cohort of patients with advanced solid tumors treated with tyrosine kinase inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3300.

Abstract 4630: A novel approach to define ctDNA molecular response to immunotherapy

Abstract Molecular biomarkers that quantify changes in circulating tumor DNA (ctDNA) can predict clinical outcomes. However, there is no consensus on classification of molecular responders (MRs) and molecular non-responders (nMRs), who may benefit from early treatment intervention. We explored the classification of MRs vs. nMRs in a real-world pan-cancer cohort of patients treated with immune checkpoint inhibitor (ICI) therapy in two ways. (1) As previously described, a ≥ 50% decrease in ctDNA tumor fraction (TF) between baseline and on-treatment time points (MR50), and, (2) hypothesizing that consistently low TF may represent a favorable subgroup with prolonged outcomes despite small increases in TF, MRlow, we evaluated a “low TF” threshold, above the TF detection rate, classifying patients as MRlow if TF remains below the TF threshold at both timepoints, regardless of magnitude change. The cohort consisted of advanced cancer patients from the Tempus de-identified clinicogenomic database who received a liquid biopsy at pre-treatment baseline and within 21-180 days after starting ICI therapy. TF was quantified via an ensemble algorithm, xM for treatment response monitoring, that incorporates pathogenic variant allele frequencies, copy number information, and germline information. Evaluable patients had TF ≥ limit of blank in at least one blood sample. Real-world overall survival (rwOS) was defined as the time from on-treatment testing to death or last known clinical record in event-free patients. Cox proportional hazards models were used to estimate the hazard ratio (HR) for MR status (MR vs. nMR) and tests for significance were conducted using 1-sided Wald tests at a 5% significance level. The evaluable cohort consisted of 71 advanced pan-cancer patients with >10 cancer types; most commonly NSCLC (38.0%, n=27) and small cell lung cancer (19.7%, n=14). Twenty-five patients (35.2%) were treated with ICI monotherapy and 46 (64.8%) were treated with ICI-chemotherapy combination. Using a definition of MR of a 50% decrease in TF alone, 44 patients were MRs and 27 patients were nMRs, with MRs having significantly longer rwOS than nMRs (HR=0.44, P=0.012). We next evaluated various low TF thresholds and found that a 1% low TF threshold further improved the association between MR and rwOS (HR = 0.32, P = 0.0008). 53 patients were classified as MRs [18 MRlow + 35 MR50] and 18 patients were classified as nMRs. Of the MRlow patients, 6 had increasing TF. Median rwOS for nMRs was seven months and not reached at 18 months for both MRlow and MR50.This study suggests that patients with consistently low TF below 1% while on ICI therapy (MRlow), regardless of TF changes, represent a unique, biologically differentiated, prognostic group. Our study suggests that MRlow patients have prolonged rw-survival, similar to molecular responders based on a 50% decrease in TF (MR50). Prospective validation for this threshold is planned. Citation Format: John Guittar, Sandra Hui, Terri Driessen, Wei Zhu, Christine Lo, Akash Mitra, Michelle Stein, Halla Nimeiri, Rotem Ben-Shachar, Wade T. Iams. A novel approach to define ctDNA molecular response to immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4630.

Abstract 6842: LgBiT-HiBiT engineered cell lines for protein degradation detection: Comprehensive PROTACs evaluation platform with endpoint and real-time kinetic analysis

Abstract Small molecule drugs have long been integral to disease treatment, but but their limitations have become increasingly apparent over time. The advent of proteolysis-targeting chimeras (PROTACs) offers a promising solution to some of these challenges. PROTACs, also known as hetero-bifunctional compounds, represent a new therapeutic class that includes a specific ligand to bind the target protein, a suitable linker, and an E3 ubiquitin ligase substrate. Upon binding to target proteins, PROTACs recruit an E3 ligase and induce the degradation of target proteins through the ubiquitin-proteasome system. This mechanism allows PROTACs to address protein targets with small-molecule binding ligands that exhibit limited efficacy, broadening the scope of druggable proteins. Additionally, unlike traditional small molecules that merely inhibit protein activity, PROTACs eliminate both enzymatic and non-enzymatic functions of target proteins, reducing the likelihood of drug resistance. Currently, Western Blot (WB) technology is widely used to validate target protein degradation and evaluate the efficacy of PROTACs. While effective for detecting significant protein level changes, WB is time-consuming, lacks precise quantification, and is impractical for monitoring protein degradation at multiple time points due to its high cost and labor intensity. To efficiently and accurately evaluate PROTACs, Kyinno have developed a series of cell models based on the LgBiT-HiBiT technology. LgBiT and HiBiT are high-affinity protein subunits that together constitute NanoLuc luciferase, catalyzing the luminescence of a luciferin substrate detectable by a microplate reader. We have achieved stable overexpression of LgBiT protein in cells and utilized CRISPR-Cas9 gene editing technology to insert HiBiT tags at the N- or C-terminal of endogenous target genes. These cell models are suitable for both endpoint and pharmacokinetic studies. Using a luciferase substrate appropriate for live-cell detection, the luminescence value reflects the current level of HiBiT protein, which correlates with the level of the target protein. Additionally, employing a long-term luciferase substrate allows real-time monitoring of protein levels at each time point. Currently, these cell lines target proteins such as IRAK3, GSPT1, GSPT2, ELAVL1, and GRK2. They support high-throughput screening of PROTACs, molecular glues, lysosome-targeting chimeras (LYTACs), and antibody-based PROTACs (AbTACs), offering a robust platform for advancing drug discovery and development. Citation Format: Siyu Li, Shi Qin, Yao Peng, Yunpeng Zhai, Jinying Ning, Feng Hao. LgBiT-HiBiT engineered cell lines for protein degradation detection: Comprehensive PROTACs evaluation platform with endpoint and real-time kinetic analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6842.

Osteosarcopenia, osteoarthritis and frailty: a two-sample Mendelian randomization study.

Musculoskeletal disease, which has a complicated relationship with frailty, is a common clinical problem among elderly individuals. This study evaluated the potential causal relationships between osteosarcopenia, osteoarthritis and frailty by Mendelian Randomization (MR) analysis. This study employed a two-sample MR approach to investigate the causal relationships among osteosarcopenia, osteoarthritis and frailty. Published summary statistics were used to obtain instrumental variables at the genome-wide significance level. Among the age groups with osteoporosis, high total bone mineral density (TBMD) (45-60, OR = 0.966, 95% CI 0.940-0.993, P = 0.013) and TBMD (over 60, OR = 0.974, 95% CI 0.954-0.994, P = 0.011) reduced the risk of frailty. Similarly, high forearm BMD (FA-BMD), high ultradistal forearm BMD (UFA-BMD), and high Heel-BMD at different sites also reduced the risk of frailty (OR = 0.966, 95% CI 0.936-0.996, P = 0.028; OR = 0.975, 95% CI 0.953-0.997, P = 0.029; OR = 0.981, 95% CI 0.967-0.995, P = 0.008). Among the characteristics related to sarcopenia, grip strength in the left hand, grip strength in the right hand, appendicular lean mass, and walking pace were all protective factors for frailty (OR = 0.788, 95% CI 0.721-0.862, P < 0.001; OR = 0.800, 95% CI 0.737-0.869, P < 0.001; OR = 0.955, 95% CI 0.937-0.974, P = 0.000; OR = 0.480, 95% CI 0.388-0.593, P < 0.001), with low grip strength in those over 60 years of age significantly positively correlated with frailty (OR = 1.168, 95% CI 1.059-1.289, P = 0.002). The MR results of osteoarthritis and frailty revealed a causal relationship between specific joint sites and frailty, including KOA (OR = 1.086, 95% CI 1.017-1.160, P = 0.014), HOA (OR = 1.028, 95% CI 1.007-1.049, P = 0.009), and KOA/HOA (OR = 1.082, 95% CI 1.053-1.113, P = 0.000), increasing the risk of frailty. Osteosarcopenia, osteoarthritis and frailty exhibit significant causal effects, rendering them risk factors for frailty. Therefore, in clinical practice, patients with osteosarcopenia and osteoarthritis should be required to undergo relevant interventions to reduce the risk of frailty.

Abstract 6127: Genomic instability regulates STING mediated anti-tumor immunity through distinctive cytosolic DNA structures

Abstract Background. Cancer genomes are characterized by instability, with significant levels of DNA damage and mutation due to DNA repair defects. In colorectal cancer (CRC), defects in mismatch repair define the microsatellite instable (MSI) subtype, with stronger immunogenicity and better prognosis than the more common chromosomally instable (CIN) subtype. The mechanisms behind these differences are unknown, but their understanding could identify strategies to improve CIN CRC prognosis. The STING pathway detects foreign DNA in the cytosol, leading to induction of innate and adaptive immune responses. In cancer cells, STING also recognizes self cytosolic DNA (cyDNA) in the form of micronuclei or diffuse fragments generated by endogenous or treatment induced DNA damage. Our lab previously identified this pathway as a key player in the increased cytotoxic T cell infiltration characteristic of MSI CRCs. As MSI CRCs do not contain more cyDNA than CIN, we hypothesize that specific sequence or structural patterns exist within cyDNA from different CRC subtypes that lead to differential STING activation and thereby induction of overall anti-tumor immunity. Methodology. We first aimed to examine whether cyDNA produced in MSI vs CIN CRC cells differentially induce STING and thereby anti-tumor immunity. CyDNA isolated from MC38 CRC cells with mutations in Mlh1 (modeling MSI) or Kras (CIN) were used to stimulate dendritic cells (DCs), key mediators of anti-tumor immunity. We found equal concentrations of MSI cyDNA more strongly activates STING, and thereby cytotoxic T cells, than CIN cyDNA. CyDNA from MSI CRC patient organoids showed similar results. Next, we aimed to characterize specific changes in cyDNA composition in MSI vs CIN CRC cells and examine how these alterations affect STING signalling. Sequencing identified patterns prevalent in highly stimulatory cyDNAs, including microsatellites. Stimulation with oligos and cyDNA enriched for microsatellites indicate enhanced STING and cytotoxic T cell activation through improved cGAS binding. Likewise, adoptive transfer of DCs activated by microsatellite rich DNAs induced stronger DC and T cell activation in orthotopic CIN tumors. Preliminary data suggests secondary DNA structures may influence this process. Finally, we aim to evaluate the contribution of micronuclei on STING activation in MSI vs CIN CRCs. DC stimulations with micronuclei isolated from MSI and CIN cells indicate MSI micronuclei also lead to enhanced STING activation over CIN micronuclei. We will evaluate micronuclei composition to identify underlying mechanisms. Significance. CRC is the second most deadly cancer worldwide, and immune recognition is crucial to treatment efficacy. This project will further our understanding of immune differences between MSI and CIN CRCs that may be used to develop therapeutic strategies, such as STING agonists, to improve CIN patient prognosis. Citation Format: Shayla Mosley, Natalie Lapa, Courtney Mowat, Kristi Baker. Genomic instability regulates STING mediated anti-tumor immunity through distinctive cytosolic DNA structures [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6127.

Perception of Flipped Classroom Strategy for Physics Instruction in Secondary Students in Kauma Sub-County Kilifi County, Kenya

The flipped classroom strategy (FCS) has emerged as an innovative pedagogical approach that enhances active learning, particularly in science subjects like physics. This study investigated the students’ perception of the strategy towards physics instruction. The study adopted mixed research methods, incorporating both qualitative and quantitative data collection techniques. The targeted population of this study was 870 Form One students in secondary schools in Kauma Sub-County. Random sampling was used to sample the schools and students that were involved in the study. The research explored how FCS influences student engagement, conceptual understanding, and achievement in physics. Perception questionnaires were administered to the students after they had undergone the treatment. The questionnaire consisted of closed-ended statements and open-ended questions. The closed-ended section had two sets of statements. The first set was a general statement asking students to indicate whether they perceive the flipped classroom to be a good strategy for physics instruction. The second set consisted of statements that were framed into a Likert-scaled format with five choices illustrating the extent of their agreement with the statements provided. These statements acted as factors that helped to determine the learners’ perception of the flipped classroom towards physics instruction. A total of 84 students answered the questionnaire. MANOVA test was conducted between the perception responses of the students and the students’ responses on the Likert-scaled statements with the help of SPSS 16 and a confidence level of 0.05 to test H03. From the results, the significance level of the MANOVA test was 0.000 for Wilks’ lambda and Roy’s largest root. Since 0.000&lt;0.05, H01 was rejected and the study concluded that students perceived flipped classrooms as an effective strategy for physics instruction. The findings provide insights into the effectiveness of FCS in enhancing physics instruction and offer recommendations for its integration into the Kenyan secondary school curriculum.

Abstract 5284: Reduced occludin expression is related to unfavorable tumor phenotype and poor prognosis in many different tumor types: a tissue microarray study on 16,870 tumors

Abstract Occludin is a key component of tight junctions. Reduced occludin expression has been linked to cancer progression. To study the role of occludin in cancer, a tissue microarray containing 16, 870 samples from 148 different tumor types and 608 samples of 76 different normal tissue types was analyzed by immunohistochemistry. Occludin immunostaining was observed in 10, 746 (76.6%) of 14, 017 analyzable tumors, including 18.9% with weak, 16.2% with moderate, and 41.6% with strong staining intensity. Occludin positivity was found in 134 of 148 tumor categories and was most frequent in adenocarcinomas (37.5-100%) and neuroendocrine neoplasms (67.9-100%), less common in squamous cell carcinomas (23.8-93%) and in malignant mesotheliomas (up to 48.1%) and rare in Non-Hodgkin’s lymphomas (1-2%) and most mesenchymal tumors. Reduced occludin staining was associated with advanced pT stage (p&amp;lt;0.0001), L1 status (p=0.0384), and absence of microsatellite instability (p&amp;lt;0.0001) in colorectal adenocarcinoma, advanced pT stage in pancreatic adenocarcinoma (p=0.005), high ISUP grade (p&amp;lt;0.0001), advanced pT stage (p&amp;lt;0.0001), high UICC stage (p&amp;lt;0.0001), distant metastasis (p=0.0422), and shortened overall (p=0.0045) or recurrence-free survival (p=0.0116) in clear cell renal cell carcinoma (RCC), high pT stage (p&amp;lt;0.0001) and UICC stage (p=0.0228), distant metastasis (p=0.0338), and shortened recurrence-free survival (p=0.006) in papillary RCC, and advanced pT stage (p=0.0133) in serous high-grade ovarian cancer. Occludin staining was unrelated to parameters of tumor aggressiveness in breast, gastric, endometrial, and thyroidal cancer. Our data demonstrate significant levels of occludin expression in many different tumor entities and identify reduced occludin expression as a potentially useful prognostic feature in several tumor entities. Citation Format: Seyma Büyücek, Florian Viehweger, Viktor Reiswich, Natalia Gorbokon, Viktoria Chirico, Christian Bernreuther, Florian Lutz, Simon Kind, Ria Schlichter, Sören Weidemann, Till S Clauditz, Andrea Hinsch, Ahmed Abdulwahab Bawahab, Frank Jacobsen, Andreas M Luebke, David Dum, Elena Bady, Lisa Hornsteiner, Claudia Hube-Magg, Martina Kluth, Katharina Möller, Anne Menz, Andreas M Marx, Till Krech, Patrick Lebok, Christoph Fraune, Guido Sauter, Ronald Simon, Eike Burandt, Sarah Minner, Stefan Steurer, Maximilian Lennartz, Morton Freytag. Reduced occludin expression is related to unfavorable tumor phenotype and poor prognosis in many different tumor types: a tissue microarray study on 16, 870 tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5284.

Temporary Suspension of School-Based Management: its Effect on Learning Outcomes of Pupils at Sto. Niño District

Success in education is only enclosed within the classroom amidst teachers and students alone. School-based management (SBM) entails the delivery of success and the structure of concrete learning outcomes. The study was participated in by 268 Grade VI pupils from 10 schools in Sto. Nino District through random sampling. Schools were identified as SBM level 3 among the 13 schools in the last SBM Regional Validation, years back 2020-2021. The survey questionnaire comprised four sections: Part I comprised pupils’ profiles, part II was on temporary suspension of SBM, including the leadership and governance, curriculum and instruction, accountability and continuous improvement, and resources management, and part III was on researcher-made questionnaire on attitudes of pupils towards learning. Part IV was a researcher-made questionnaire on challenges encountered by pupils. The content experts validated the instrument, and it was pilot-tested. Results revealed that the p-value of .511 is greater than the standard significance level of 0.05, leading to the decision to accept the null hypothesis. Hence, it can be concluded that the temporary suspension of the SBM bears no effect on the attitude of pupils in Sto. Nino District. In this study, the efficiency of school-based management relies on the stakeholders’ capacity to actualize the need and identify the existing challenges that confront the implementation. Even further, one of the indicators of the success of School-Based Management implementation lies in the students’ learning outcomes. Learners’ learning outcome depends on their attitude toward learning and their challenges in the teaching-learning process. Like the conduct of SBM, its efficiency is seen in the learning outcomes, and learning outcomes are seen in learners’ efficiency in the academic efforts.

Abstract 5922: B cell subtype is a predictive and pharmacodynamic biomarkers for combination therapy in stage III-IV melanoma: a simon phase II trial

Background: Assessing biomarkers to predict immune-related adverse events (irAEs) is crucial for optimizing melanoma treatment with immune checkpoint inhibitors (ICIs). T and B lymphocytes are key mediators of autoimmunity and are implicated in the development of irAEs. Here we focus on analysis of specific B cell subsets, as predictive and pharmacodynamic biomarkers for irAEs in a phase II trial combining nivolumab (3 mg/kg), ipilimumab(1 mg/ml), and the IL-6 receptor inhibitor tocilizumab (4 mg/kg) in unresectable/advanced stage III/IV melanoma patients. Methods: We analyzed cryopreserved PBMC from melanoma patients collected at baseline and week 7, and categorized by irAE severity. Patients were categorized as either experiencing severe irAEs (grade 3-4) (TOX) or no toxicity or grade 1-2 irAEs (NT). We used multi-parameter flow cytometry panel to analyze B cells. PBMCs analyzed by a 35-color antibody B cell panel by spectral flow cytometry. The data underwent quality control, and normalization and initial clustering. CD19+ B cells were concatenated and clustered by FlowJo software. Fifteen B cell clusters was obtained and phenotyped using Cluster Explorer. Statistical analyses were performed using multiple logistic regression and Mann-Whitney U test, with a significance level set at 0.05. Results: Among the 70 patients administered treatment, we observed a best overall response rate of 57%. Grade 3-5 irAEs occurred in 22% of patients by week 24. We analyzed 58 quality-controlled baseline samples and 54 7-week samples. Fifteen distinct clusters were identified after clustering B cell populations using UMAP and re-clustering using PHATE. Two of these clusters showed significant differences in both multivariate logistic regression and Mann-Whitney U tests. TOX patients had significantly lower baseline naive B cell counts than NT patients (p=0.016), suggesting naïve B cells as a predictive marker for toxicity. A significant increase in late memory B cells at week 7 was a key pharmacodynamic marker (p=0.003). Conclusion: Our findings suggest that naive B cells could serve as a toxicity biomarker in combination therapy with ipilimumab, nivolumab, and tocilizumab. Late memory B cells emerged as a promising pharmacodynamic biomarker. Immunological monitoring may help elucidate the mechanisms of irAEs and predict irAE occurrence. Citation Format: Teruyuki Mizutani, Amrutesh Puranik, Tomoaki Muramatsu, Joanna Handzlik, Ankita Mitra, Diana Kimono, Stanzin Idga, Eddy Yang, Ramin Herati, Michelle Krogsgaard, Mehnert Janice. B cell subtype is a predictive and pharmacodynamic biomarkers for combination therapy in stage III-IV melanoma: a simon phase II trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5922.

Abstract 4990: A discovery of novel methylation profiles in head and neck squamous cell carcinoma in relation to pretreatment pain by cancer subtype

Abstract Pain is the initial symptom reported by nearly 75% of individuals diagnosed with head and neck squamous cell carcinoma (HNSCC), yet effective management remains elusive, with up to 80% of patients reporting inadequate pain management despite opioid use. Pretreatment pain is a key predictor of survival and recurrence, but the molecular mechanisms underlying pain in HNSCC are not well understood. This pilot study aims to explore the epigenetic mechanisms underlying pain perception in HNSCC patients by cancer subtype and identify potential biomarkers to improve pain management strategies. DNA methylation levels were generated for individuals newly diagnosed with HNSCC of oral cavity (OCSCC; n=18) or oropharynx (OPSCC; n=8) prior to definitive treatment using the Illumina Infinium MethylationEPIC v2.0 array. Perceived worst pain score was measured using the Brief Pain Inventory-Worst Pain item prior to treatment, categorized according to NCCN guidelines, with patients reporting no pain serving as the reference group. After quality control, M-values for 26 samples and 913, 962 methylation probes were evaluated. Differentially methylated positions (DMPs) associated with pain severity were identified combining subtypes, with hypo- (fold change (FC) &amp;lt; -2) and hypermethylated (FC &amp;gt; 2) sites determined after adjusting for age. Stratified analyses were performed by cancer type. All analyses set a significance level with a false discovery rate &amp;lt; 5%. Pain phenotype and blood collection occurred on the same day, after diagnosis but prior to definitive treatment. The cohort had a mean age of 65.1 ± 11.3 years, 42.3% of those reporting any level of pain were male. Pre-treatment pain was reported by 94.4% (N=17) of OCSCC and 25% (N=2) of OPSCC patients. DMP analysis revealed probe cg08573204_TC21 in LMO1 gene was hypermethylated in OCSCC patients with mild pain versus no pain (p=0.02, Log2FC=1.11). While LMO1 has not been previously linked to pain, its role in adrenergic cell identity suggests potential involvement in pain pathways. In patients with moderate/severe pain, cg18566479_BC21 located in CHDH gene was hypermethylated in OCSCC (p=1.09E-04, Log2FC=1.98). CHDH, associated with metabolic signatures in HNSCC and neuroinflammatory activation in neuropathic pain, showed significant hypermethylation, suggesting its potential as a biomarker for pain management in HNSCC. Differential DNA methylation may be associated and predictive of the development and severity of pain in HNSCC. With future work in larger samples, this research may provide insight into the biological underpinnings of pain and guide cancer symptom management by using DNA methylation profiles to identify patients at risk for pain who may benefit from preventative measures. Citation Format: Monica A. Wagner, Yanning Wu, Naji Ayyash, Fredrick R. Schumacher, Quintin Pan. A discovery of novel methylation profiles in head and neck squamous cell carcinoma in relation to pretreatment pain by cancer subtype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4990.

Metabolomic and functional analyses of small molecules secreted by intestinal nematodes in the activation of epithelial tuft cells.

Intestinal helminth parasites trigger the host immune response through epithelial sensory tuft cells, but helminth-derived molecules that may activate tuft cells are poorly characterized. The study aimed to identify small molecules released in vitro by two nematode parasites, that infect rodents (Nippostrongylus brasiliensis) and ruminants (Haemonchus contortus), and to test candidate ligands in an in vivo model of tuft cell differentiation. Small molecules were analyzed by hydrophilic interaction liquid chromatography (HILIC) of material released by adult parasites incubated in serum-free media, followed by mass spectrometry; selected molecules were administered to mice and tuft cell expansion enumerated after 5days. A range of different conditions (culture media, timing, oxygenation) were tested, and comparisons made between the conditions, and between the two nematode species at selected points. Common products across the conditions and species included carboxylic acids (malate, succinate), medium chain fatty acids (such as decanoic and undecanoic acids), purines (guanine, xanthine and their derivatives), and phosphocholine compounds. We selected 19 of the prominent molecules for in vivo testing by oral administration, including succinate, a known activator of tuft cell differentiation. Malate elicited a low but significant level of tuft cell expansion, while undecanoic acids with or without a bromine substitution were also able to induce significant differentiation comparable to succinate. Other molecules including phosphorylcholine had no effect. Multiple molecular species including decanoic and undecanoic acids released by helminths may contribute to activation of tuft cells in vivo.

Abstract 2276: Whole exome sequencing of immune-mediated colitis in cancer patients treated with immune checkpoint inhibitors

Abstract Background: Immune checkpoint inhibitors (ICIs) have transformed cancer treatment, providing survival benefits across multiple cancer types. Despite this remarkable advancement, ∼15-30% of patients develop immune-related adverse events (irAEs). One of the common irAEs is immune-mediated colitis, and the incidence ranges from 1% to 25% by type of therapy. Previously, we have shown that polygenic risk scores of ulcerative colitis predict colitis irAE. However, the role of rare coding variants on the development of colitis irAE remains largely unknown. Methods: We conducted whole exome sequencing for colitis irAE in 1, 215 cancer patients treated with ICIs. Association analyses were carried out for each gene separately for protein-truncating missense variants, predicted splice (Sp) variants, and non-synonymous predicted deleterious (deN; CADD≥20 or VEST≥0.8) or any non-synonymous (N) variants. The main association analyses were burden tests in which genotypes were collapsed to a 0/1 variable based on whether samples carried a variant of the given class. All models were adjusted for age at diagnosis, sex, type of ICI therapy, recruitment site, and five ancestry-informative principal components. The exome-wide threshold for significance was set at 2.5 × 10-6, with genes beyond this considered significant. Results: Among 1, 215 patients treated with immune checkpoint inhibitors (ICIs), 60 (4.9%) developed colitis. We identified rare coding variants in two genes associated with an increased risk of colitis at the exome-wide significance level: PT+Sp+N variants in LRRC47 (OR = 5.5, P = 2.4 × 10-6) and PT+S+deN variants in PFKP (OR = 65.9, P = 1.2 × 10-6). Additionally, PT+S+deN variants in PTPRH showed a suggestive association with colitis irAEs (OR = 8.8, P = 7.1 × 10-5). Conclusions: To our knowledge, this is the first study to assess the role of rare coding variants in colitis immune-related adverse events (irAEs) among cancer patients treated with ICIs. Our findings highlight two exome-wide significant associations, implicating variants in LRRC47 and PFKP in the development of colitis irAEs. LRRC47 has previously been linked to inflammatory bowel disease through differential methylation patterns. Furthermore, genes involved in glycolysis, including PFKFB3, PKM, and PFKP, have been shown to be upregulated in patients with ulcerative colitis. Abnormal activity of PTPRH has also been reported to disrupt the regulatory mechanisms of the intestinal mucosa, potentially contributing to inflammation. These findings provide novel insights into the genetic underpinnings of colitis irAEs in cancer immunotherapy. Citation Format: Pooja Middha, 1 Zoe Quandt, 2 Karmugi Balaratnam, 3 Eduardo Cardenas, 1 Christina J. Falcon, 4 Princess Margaret Lung Group, Matthew A. Gubens, 1 Scott Huntsman, 1 Khaleeq Khan, 3 Min Li, 1 Christine M. Lovly, 5 Devalben Patel, 3 Luna Jia Zhan, 3 Melinda C. Aldrich, 6 Geoffrey Liu, 7 Adam J. Schoenfeld, 4 Elad Ziv1. Whole exome sequencing of immune-mediated colitis in cancer patients treated with immune checkpoint inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2276.

Abstract 7466: Advancing tumor characterization and immune phenotyping through multiomic in situ profiling of RNA and proteins

Abstract Cancer continues to be one of the leading causes of death and financial burden on public health worldwide. Despite advances in detection and treatments, there is still a significant level of uncertainty in predicting how an individual's cancer will respond to treatment due to tumor heterogeneity among different patients and multiple subclones from the same individual. A more comprehensive understanding of cancer cell populations, their molecular interactions, and their microenvironment can offer valuable insights into optimal treatment strategies and patient stratification. This can be achieved through the integrated detection of both RNA and protein targets, enabling a deeper exploration of the molecular mechanisms driving cancer. Here, we analyzed formalin-fixed, paraffin-embedded (FFPE) human tissue sections from diverse tissue types using spatial multi-omics technologies from 10x Genomics. A new add-on capability enables visualization and analysis of protein expression for key immuno-oncology markers including markers for immune cell typing, cell structure, proliferation, and differentiation, as well as immune checkpoint markers (e.g., PD-1, PD-L1, LAG-3, and VISTA). Simultaneous spatial profiling of 480 genes and 28 proteins, integrated with H&amp;E imaging, provided detailed insights into tissue architecture, tumor origins, immune cell infiltration, and the immune status of the tumor microenvironment. The combined RNA and protein profiling method is a powerful approach for validating RNA-based findings and providing definitive evidence at the functional biological level. Protein data also offers critical insights into the expression and function of different isoforms that are challenging to discern at the RNA level. For instance, isoforms such as CD45RA and CD45RO, which arise from alternative splicing of the PTPRC gene, are often indistinguishable using RNA profiling alone due to the shared exonic sequences in their transcripts. However, protein-level analysis can differentiate these isoforms and enables a more precise understanding of their distinct roles in the immune response, including the delineation of naïve and memory T-cell subsets. Furthermore, RNA expression does not always directly correlate with protein levels, underscoring the importance of integrating both data types. For instance, discrepancies between RNA and protein expression were observed for β-catenin, encoded by the CTNNB1 gene, across tissues. β-catenin plays a pivotal role in tumor progression through its dual functions in cell adhesion and signaling. By integrating RNA and protein data, researchers can uncover the complexities of post-transcriptional regulation, subcellular localization, and their combined influence on cancer progression, providing deeper insights into disease mechanisms and potential therapeutic targets. Citation Format: Thanutra Zhang, Smritee Dadhwal, Joshua Thao, Anatalia Robles, Cheyenne Christopherson, Suyog Pathare, Kristen Pham, Minkyung Park, Hsin Wen Chang, Janine Hensel, Cedric Espenel, Hayato Ikoma, Gabriele Girelli, Paul Ryvkin, Veronica Gonzalez Muñoz. Advancing tumor characterization and immune phenotyping through multiomic in situ profiling of RNA and proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7466.

Antimicrobial effect of 2.5% sodium hypochlorite irradiated with the 445 Nm diode laser against bacterial biofilms in root canal - in vitro pilot study.

This research investigates the efficacy of a novel blue diode laser (445nm wavelength) in enhancing the antimicrobial action of 2.5% sodium hypochlorite against biofilms of two prevalent microbial species found in root canals- in vitro. The research also includes a comparative analysis with a more established diode laser protocol (970nm), assessing their respective effectiveness in eliminating root canal pathogens. 62 extracted single-rooted human teeth were prepared with rotary endodontic instruments, sterilized, contaminated with a combination of two microbial cultures (E. faecalis, C. albicans) and incubated for 14 days at 37 ◦C. The samples were randomly distributed into 4 groups: G1- negative control, G2-2.5% NaOCl rinse, G3-445nm laser boosted 2.5% NaOCl rinse, G4-970nm laser boosted 2.5% NaOCl rinse. SEM imaging was done to visualize effects in each group. The number of viable microbes in each sample was determined by the colony forming unit count technique (CFU). A statistically significant reduction of microbial numbers after all treatments was observed (Mann-Whitney U test, significance level set to 5%). G3 and G4 showed similar results, both better than G2. The 445nm diode laser NaOCl irradiation protocol has improved antimicrobial effectiveness when compared to sole 2.5% NaOCl rinse and equal effectiveness as the 970nm NaOCl irradiation protocol, ex vivo.

Extrusion degree of a new low-viscosity, injectable, light-cured endodontic filling material: an in-situ study in a cadaveric close-to-mouth experimental model.

To evaluate the ability of the liquid-injectable OdneFill sealer to stay within the boundaries of prepared root canals of posterior teeth using a cadaveric model. Eight maxillary bone blocks containing 3 to 5 posterior teeth were collected. Pre-operative radiographs were taken, and only maxillary premolars (n = 10) and molars (n = 16) were included. The canals were prepared with WaveOne Gold Primary and Large instruments, then randomly assigned to two groups (n = 33 canals). Group 1 used Warm Vertical Compaction with gutta-percha and AH Plus sealer, while Group 2 used only the OdneFill sealer to fill the canals. Post-operative radiographs were taken to assess sealer extrusion into the periapical tissues using a 4-point scale. Data analysis was conducted with a 5% significance level. Out of 26 teeth examined (5 teeth were excluded), 47.6% (8 molars and 2 premolars) showed filling extrusion, with 77.8% of teeth in Group 1 (12 root canals) and 25.0% in Group 2 (3 root canals). A significant difference was found in the frequency of sealer extrusion between the two groups (P = 0.001), but no significant difference was observed between premolars and molars in terms of sealer extrusion frequency (P = 0.144). The OdneFill filling protocol showed better control over extrusion compared to the Warm Vertical Compaction technique in posterior teeth of a cadaveric model. This study highlights the potential of a new liquid-injectable root canal sealer designed to remain confined within the boundaries ofthe root canal anatomy.

Theranostic performance of EGFR-targeted ceria-based nanoparticles on EGFR-positive cancers.

EGFR is overexpressed in several cancers and hence EGFR-targeted theranostics is a promising approach to manage cancers, with widespread applicability. When nanoceria, which possesses intrinsic anticancer properties, is conjugated with EGFR-targeted fluorophore-tagged ligands, this nanoformulation can both image tumors and kill them through ROS-mediated cell destruction. Further, targeting enhances the cellular uptake of nanoparticles through EGFR-mediated endocytosis. The present work evaluates the in vitro theranostic performance of FITC-tagged EGF-functionalized nanoceria on EGFR-positive cancers. Three EGFR-positive cell lines were used for the study: MDA-MB-231, PANC-1 and HeLa. The EGFR-binding specificity of the EGF-functionalized nanoparticles was confirmed using western blot analysis. The therapeutic and diagnostic activities of the theranostic nanoparticles were confirmed, the former by cell viability assays and ROS measurements and the latter by confocal imaging. The results demonstrate significant ROS elevation levels for the treated cells and hence the suitability of the particles for therapeutic applications. The nanoparticles also are capable of detection using fluorescence imaging following 5minutes of treatment, thus confirming the applicability for imaging. Hemolysis assay studies revealed excellent hemocompatibility of the nanoparticles, confirming their suitability for in vivo applications.

Genetic Architecture of Hock Joint Bumps in Pigs: Insights from ROH and GWAS Analyses

The genetic mechanisms underlying the formation of defects, such as bumps and growths on the hock joints in pigs, remain poorly understood. Therefore, the aim of this study was to investigate the relationship between runs of homozygosity (ROH) and the formation of hock joint bumps, with the goal of identifying associated SNPs and candidate genes involved in these defects. The study was conducted on a population of Large White breed pigs (n = 568) using runs of homozygosity (ROH) analysis and genome-wide association studies (GWAS). The results suggested that the predisposition to hock joint bumps in pigs may have arisen due to recent selective breeding pressure. Using GWAS, 27 SNPs were identified at the suggestive significance level, with one SNP (rs325478346) reaching genome-wide significance. These markers are localized in genes associated with various biological processes, including lipid metabolism (VIPR2 and CFTR), inflammatory processes (MTURN and ADCY2), connective tissue structural integrity (COL27A1), muscle regeneration (PAMR1), and ion exchange and cellular homeostasis (KCNIP4 and NALCN), as well as regulation of cell growth, extracellular matrix remodeling, and fibroblast differentiation (CEP120 and SCUBE3). Further research utilizing omics technologies will provide deeper insights into the pathogenesis of hock joint bumps and contribute to the development of strategies for the prevention and potential treatment of this defect.

The Effect of Group Counseling with Assertive Training Techniques on Truancy Behavior of Grade VIII Junior High School Students

This study aims to determine the effect of group counseling services with Assertive Training techniques on students' truancy behavior. This study uses a preexperimental method with a one group pretest-posttest design. The population of the study was grade VIII students who had a habit of truancy, with a sample of 8 students taken using the Purposive Sampling technique. The instruments used were pretest and posttest which had been tested for validity and reliability. The collected data were analyzed using the t-test with a significance level of 0.05. The results of the t-test calculation showed a t-count value of 1.173, while t-table with db N-1 = 7 and a significance level of 0.05 was 0.754. Based on these results, it can be concluded that group counseling services with Assertive Training techniques have a positive and significant effect on reducing students' truancy behavior. These findings are expected to contribute to improving the quality of guidance and counseling services, especially in the application of Assertive Training techniques in schools