Early Stage Research Articles

Neurofilament heavy phosphorylated epitopes as biomarkers in ageing and neurodegenerative disease

AbstractFrom the day we are born, the nervous system is subject to insult, disease and degeneration. Aberrant phosphorylation states in neurofilaments, the major intermediate filaments of the neuronal cytoskeleton, accompany and mediate many pathological processes in degenerative disease. Neuronal damage, degeneration and death can release these internal components to the extracellular space and eventually the cerebrospinal fluid and blood. Sophisticated assay techniques are increasingly able to detect their presence and phosphorylation states at very low levels, increasing their utility as biomarkers and providing insights and differential diagnosis for the earliest stages of disease. Although a variety of studies focus on single or small clusters of neurofilament phosphorylated epitopes, this review offers a wider perspective of the phosphorylation landscape of the neurofilament heavy subunit, a major intermediate filament component in both ageing and disease.image

Abstract B022: Investigating how monocytes impair NK cell cytotoxicity in lung metastasis of triple-negative breast cancer

Abstract Triple-negative breast cancer (TNBC) accounts for 20% of breast cancer (BC) cases worldwide, yet due to its aggressive phenotype and lack of targeted therapies it has the worst prognosis amongst BC subtypes. Up to 40% of TNBC patients will relapse after treatment and develop distant metastasis. Metastasis is the complex process by which disseminated tumor cells (DTCs) leave the primary tumor and grow at distal sites. Since most metastases occur within 3 years of diagnosis, the rapid onset and lack of effective treatments in metastatic TNBC (mTNBC) means less than 20% of patients survive after 4 years. Even though it is the most “immune-activated” BC subtype, and immune checkpoint blockade (ICB) has shown clinical benefits in early TNBC, there is a clear need for novel immunotherapeutic approaches for patients with mTNBC. Natural killer (NK) cells are innate lymphocytes that function as the body’s first line defense against metastatic cancer cells. NK cells eliminate DTCs in the blood or at distal sites via NK cell cytotoxicity (NKCC) and this process is known to be impaired in successful metastasis. We hypothesized that other cells within the tumor microenvironment (TME) can inhibit NKCC at the earliest stages of metastatic outgrowth. We have used Cherry-niche, an in vivo labelling tool, to characterize how TME cells change in early and established lung micro-metastases by single-cell RNA sequencing (scRNA-seq) and flow cytometry. Our initial analyses showed that monocytes are highly enriched, not only in the entire metastatic lung, but specifically in the established niche surrounding DTCs. Although we detected a slight increase in total NK cells, there was a shift in maturation status with fewer mature NK cells (CD27-CD11b+) and more immature NK cells (CD27+/-CD11b-) in the metastatic lung. Furthermore, NK cells were largely absent from the metastatic niche and those that were present displayed a dysfunctional phenotype. CellChat and NicheNet analyses of our scRNA-seq data predicted that several receptor-ligand interactions, including the cytokine macrophage migration inhibitory factor (MIF), can mediate the crosstalk between NK cells and monocytes. Next, we developed an in vitro 3D co-culture system to study the effect of blood-derived monocytes on human NKCC against mTNBC cells. We found that monocytes can suppress NKCC against MDA-MB-231 cells and decrease the viability of mature NK cells. Furthermore, recombinant MIF was able to directly inhibit NKCC, whilst a selective MIF antagonist was able to rescue monocyte-induced impairment of NKCC. These data implicate MIF as a critical monocyte-derived factor in the suppression of NK cell function against mTNBC cells. Future work will aim to identify the exact mechanism by which MIF inhibits NKCC, whilst also evaluate the effect of monocyte depletion and MIF inhibition on metastatic outgrowth and NK cell activation/maturation in vivo. This study will reveal novel inter-cellular dynamics occurring in the metastatic lung, as well as immunotherapeutic targets that may restore NKCC in mTNBC. Citation Format: Christos Ermogenous, Luigi Ombrato, Gordon Beattie. Investigating how monocytes impair NK cell cytotoxicity in lung metastasis of triple-negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr B022.

Impact of Multidisciplinary Team Management on Survival and Recurrence in Stage I–III Colorectal Cancer: A Population-Based Study in Northern Italy

This study aimed to assess whether multidisciplinary team (MDT) management improves outcomes in patients with stage I–III colorectal cancer (CRC) compared to non-MDT care. This study included 460 CRC patients diagnosed from 2017 to 2018 in a northern Italian province, of whom 300 (65%) were managed by MDT. MDT patients were younger, more frequently had rectal cancers, were diagnosed at earlier stages, and were more likely to undergo surgery and chemotherapy. Disease-free survival (DFS) and overall survival (OS) were significantly better in the MDT group. The recurrence rate was 8.5%, with no significant differences between MDT and non-MDT patients, although MDT patients exhibited lower recurrence rates for early-stage tumors. A multivariable analysis showed that DFS and OS were adversely affected by older age, advanced stage, and lack of MDT management. Kaplan–Meier estimates demonstrated a 3-year DFS of 78% in the MDT group versus 65% in the non-MDT group, as well as an OS of 83% versus 69%, respectively. The MDT approach was associated with improved treatment adherence and better management of recurrences. While limited by a small sample size, this population-based study highlights the beneficial impact of MDT care on CRC outcomes. Further research with extended follow-up is warranted to confirm these findings.

Tropical Cyclone Rainfall Structure and Its Long-Term Trend over the Western North Pacific

Abstract The tropical cyclone (TC) rainfall structure over the western North Pacific is categorized into four typical types. Clusters I and IV are featured by the southward-shifted and northward-shifted rainfall centers, respectively. Meanwhile, clusters II and III are characterized by the weak and strong maximum rainfall rates near the TC center, respectively. Apart from rainfall, clusters I–IV show diverse features in locations, intensities, and lifetime stages. Both clusters I and III appear over the low-latitude regions. However, TCs in cluster III have the strongest intensity and are in the mature stage of their lifetime, while TCs in cluster I are relatively weaker and in earlier stages. On the contrary, cluster IV has the highest mean latitude positions and strong intensity, mainly in their later lifetime period. Cluster II shows the weakest TC intensity and mainly locates in the subtropics. Diverse TC locations lead to different effects of large-scale circulation on TC rainfall patterns in clusters I–IV. For clusters I and III, abundant moisture provided by the southwest monsoon causes heavy rainfall. In contrast, cluster IV is mainly affected by midlatitude westerly troughs, which is responsible for the northward-shifted rainfall center. Cluster II shows the weakest TC intensity, resulting in a small rainfall rate. Further analysis shows that the changes in the relative frequency contributed 36% of the decreasing trend of basin-mean inner-core rainfall during 1998–2019, and the shift of rainfall pattern from cluster III to cluster I due to the enhanced monsoon plays a central role.

Evaluation of the minor clinical features of Hanifin and Rajka amongst children with atopic dermatitis, visiting a tertiary care hospital in North Eastern India

Atopic Dermatitis (AD) is a commonly encountered, extremely itchy, chronic and relapsing skin disorder, which is characterized by inflammation in the skin. It typically has its onset in childhood, presenting with eczematous lesions; and can also be associated with personal or family history of “atopy”, usually most often. The criterion put by Hanifin and Rajka, encompassing 4 major plus 23 minor criteria, is not infrequently used as a diagnostic criterion for this disease in a hospital setup; however, the frequency of occurrence of the minor criteria usually varies in the population.The study aims at estimating the frequency of the minor diagnostic criteria of the Hanifin and Rajka in children with AD. This was a hospital-based cross-sectional study, that was conducted on 70 children (less than 18yrs of age) by taking their history, doing clinical and ophthalmological examination and some basic tests.The most common occurrence amongst the minor criteria estimated from our study was xerosis i.e., in 45(64.2%). The other common minor criteria observed were pityriasis alba (42.8%) facial erythema (40%), orbital darkening (37.14%) Early age of onset was seen in children (35.7%). There is variation in the clinical features of AD according to geography and population. The prevalence and severity of the disease are dependent on several factors like ethnicity/race and environment. Hence, it is important for dermatologists to know the usual and common ways the disease presents, in a given population for the diagnosis so as to diagnose and treat the patients at an earlier stage and before significant discomfort.

Improved management of Italian ryegrass (Lolium perenne spp. multiflorum) with mixtures of tiafenacil and ACCase or glutamine synthetase inhibitors

Abstract Herbicide-resistant Italian ryegrass [Lolium perenne L. spp. multiflorum (Lam.) Husnot] is a significant problem in multiple cropping systems because of its rapid growth, open pollination, prolific seed production, and multiple cases of resistance worldwide, except to protoporphyrinogen oxidase (PPO) inhibitors. This research evaluated tiafenacil, a new PPO inhibitor, in mixtures with glutamine synthetase or acetyl-CoA carboxylase (ACCase)-inhibiting herbicides to manage resistant L. multiflorum populations. Tiafenacil efficacy against L. multiflorum was growth stage-dependent, with increased efficacy at earlier stages in greenhouse studies. The LD90 was 41.06 g ai ha-1 at BBCH23, and increased to 9.0-fold at BBCH33. Field studies indicated that changes in carrier volume did not affect tiafenacil’s efficacy; the highest tested rate of tiafenacil (75 g ai ha-1) reduced L. multiflorum inflorescence weight by 50-90%. Mixtures of tiafenacil and glufosinate (1,150 g ai ha-1) improved L. multiflorum control (+24-43%) and reduced inflorescence weight (+15-34%), particularly at the highest tested rates (50 and 75 g ai ha-1), suggesting synergistic effects based on Colby’s test. Tiafenacil with ACCase inhibitors improved L. multiflorum control (+19-49%) and inflorescence weight reduction (+8-13%). These mixtures exhibited an additive effect when combined with fluazifop and a synergistic effect with clethodim. Herbicide mixtures and application strategies are critical to effective L. multiflorum management. Tiafenacil, especially when used with glufosinate or ACCase inhibitors, offers an effective solution to L. multiflorum management and is a strategic tool against herbicide resistance, as resistance to PPO inhibitors has not evolved. Further research should assess practices to ensure the long-term viability of these mixtures for resistance management.

Adolescents’ Knowledge and Misconceptions About Sexually Transmitted Infections: A Cross-Sectional Study in Middle School Students in Portugal

Background/Objectives: Adolescence represents a period of heightened vulnerability to risky sexual behaviours. In Portugal, adolescents are initiating sexual activity at increasingly younger ages, making it relevant to characterise their knowledge about sexually transmitted diseases and prevention methods at an earlier stage of their development. Methods: A cross-sectional observational study was conducted involving the application of a questionnaire to 750 ninth-grade students from Vila Nova de Gaia, Porto, Portugal. The questionnaire covered topics such as perceptions of sexuality, knowledge of sexuality and STIs, methods of transmission prevention, and sources of information. Results: The mean age was 14.4 years, with 19.9% reporting having already initiated sexual activity. Overall, the knowledge about sexually transmitted infections was low, with a score of 7.08 out of a maximum score of 18. Condoms and immunisation were well-recognised preventive measures, but many misconceptions persisted. Better knowledge is associated with better attitudes regarding the likelihood of having sexual intercourse. Conclusions: This study reveals adolescents’ limited knowledge about sexually transmitted infections, misconceptions about prevention, and reliance on peers and the internet over trusted sources, highlighting the need for comprehensive sexual education in middle school.

Abstract B004: Ovarian cancer long intergenic noncoding RNA 1 is a potential therapeutic target in advanced ovarian cancer by promoting glycolytic pathway and hypoxic response

Abstract Ovarian cancer (OvC) is often diagnosed as an advanced disease because symptoms do not develop at earlier stages. Besides current treatments, alternative efficient therapies for OvC need to be established. We have previously identified that a novel long intergenic noncoding RNA (lincRNA) originated from chromosome 10q21 was predominantly expressed in clinical ovarian cancer tissues, designated as ovarian cancer lincRNA 1 (OIN1). Silencing of OIN1 in OvC cell lines substantially exhibited the inhibition of in vitro and in vivo cell proliferation, by upregulating apoptosis-related genes. In the present study, we aimed to evaluate whether OIN1 can be a therapeutic target in advanced OvC and how OIN1 drives the progression of OvC. We established long-term culturable spheroid cultures of OvC patient-derived cells (OvC-PDCs) from ascites of OvC patients as an advanced OvC model. The OvC-PDC spheroid cultures abundantly expressed OIN1 and stemness-related genes. OIN1-targeted siRNAs could suppress the spheroid growth and upregulate apoptosis-related genes such as RASSF5 and ADORA1 in OvC-PDC cultures. In a bloody ascites-generating xenograft in vivo model of mice intraperitoneally injected with OvC-PDC, OIN1-targeted siRNA treatment engineered by drug delivery system alleviated ascites production and prolonged the mice survival time. Based on transcriptomic analysis of OIN1 silencing in OvC-PDC cultures, we found that OIN1 inhibition exerts the downregulation of genes involved in glycolytic pathway and hypoxic response. We further demonstrated that OIN1 modulates HIF1a-mediated transcriptional regulation in OvC- PDCs. The present findings provide promising evidence that OIN1 can be a potential therapeutic target for advanced OvC. Citation Format: Kuniko Horie, Wataru Sato, Kazuhiro Ikeda, Satoshi Inoue. Ovarian cancer long intergenic noncoding RNA 1 is a potential therapeutic target in advanced ovarian cancer by promoting glycolytic pathway and hypoxic response [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RNAs as Drivers, Targets, and Therapeutics in Cancer; 2024 Nov 14-17; Bellevue, Washington. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(11_Suppl):Abstract nr B004.

Identification of two transcription factors that work coordinately to regulate early development in Entamoeba.

The protozoan parasite Entamoeba has a life cycle that switches between infective cysts and invasive trophozoites. Encystation, a crucial process in parasite biology, is controlled by different mechanisms including transcriptional control. We identified two nuclear proteins in Entamoeba invadens, EIN_066100 and EIN_085620, that regulate parasite development by binding to a DNA motif (TCACTTTC) in the promoter regions of genes upregulated in the first 8 h of stage conversion. Overexpression of EIN_066100, a homolog of MAK16 protein, resulted in reduced amoebic proliferation without affecting encystation efficiency. Overexpression of EIN_085620, a protein with an RNA-recognition motif (RRM), led to increased encystation efficiency. Glutathione S-transferase (GST) pull down assays revealed that EIN_066100 interacts with EIN_085620 both in vivo and in vitro, and this interaction is mediated by the EIN_085620 RRM domain. By evaluating truncated proteins with deletions at either the N-terminal or C-terminal regions of EIN_066100, we elucidated the importance of its N-terminal region in proper protein localization, proliferation, encystation, and interaction with EIN_085620. Taken together, these results indicate a coordinated role of EIN_066100 and EIN_085620 in regulating Entamoeba development. This work sheds light on the molecular mechanisms in the earliest stages of Entamoeba encystation.IMPORTANCEAn important biological process in the biology of Entamoeba is stage conversion, which plays a crucial role in disease propagation, facilitating parasite survival outside the host and spreading to new hosts. Multiple mechanisms contribute to controlling the expression of amebic stage-specific genes such as epigenetic and transcriptional control. Identification of early transcriptional control regulators is crucial to understanding the initiation of the encystation cascade. We identified two nuclear proteins, EIN_066100 and EIN_085620, involved in the proliferation and developmental regulation of E. invadens. These proteins work by direct binding to each other and mediating encystation efficiency. Study of new regulators involved in Entamoeba development represents an important advance in a critical aspect of parasite biology.

Dissecting the causal role of early inferior frontal activation in reading.

Cognitive models of reading assume that speech production occurs after visual and phonological processing of written words. This traditional view is at odds with more recent magnetoencephalography studies showing that the left posterior inferior frontal cortex (pIFC) classically associated with spoken production responds to print at 100-150 ms after word-onset, almost simultaneously with posterior brain regions for visual and phonological processing. Yet the theoretical significance of this fast neural response remains open to date. We used transcranial magnetic stimulation (TMS) to investigate how the left pIFC contributes to the early stage of reading. In Experiment 1, 23 adult participants (14 females) performed three different tasks about written words (oral reading, semantic judgment and perceptual judgment) while single-pulse TMS was delivered to the left pIFC, fusiform gyrus or supramarginal gyrus at different time points (50 to 200 ms after word-onset). A robust double dissociation was found between tasks and stimulation sites - oral reading, but not other control tasks, was disrupted only when TMS was delivered to pIFC at 100 ms. This task-specific impact of pIFC stimulation was further corroborated in Experiment 2, which revealed another double dissociation between oral reading and picture naming. These results demonstrate that the left pIFC specifically and causally mediates rapid computation of speech motor codes at the earliest stage of reading and suggest that this fast sublexical neural pathway for pronunciation, although seemingly dormant, is fully functioning in literate adults. Our results further suggest that these left-hemisphere systems for reading overall act faster than known previously.Significance Statement Recent neuroimaging data suggest that left posterior inferior frontal cortex, classically associated with spoken production, responds to print simultaneously with left fusiform and supramarginal gyri, each responsible for visual and phonological processing, contrary to traditional serial cascade models of reading. While the region is now known to mediate different aspects of cognitive processing, the functional significance of this fast neural response remains unclear. Using transcranial magnetic stimulation, we show that early inferior frontal activation plays a specific and causal role in speeded oral reading at 100 ms after word-onset. This fast sublexical neural pathway for pronunciation, although seemingly dormant, is fully functioning in literate adults. We also propose that the left-hemisphere reading systems act differently and faster than known previously.

Abstract A050: Early detection of liver cancer from diverse populations using cfDNA fragmentome and protein biomarkers

Abstract Hepatocellular carcinoma (HCC) is a leading cause of cancer death world-wide. In the US, liver cancer has increased in incidence and mortality due to a growing population from South and Central America, which faces higher risk of disease in part due to aflatoxin exposure. Recently, we showed that the DELFI (DNA evaluation of fragments for early interception) approach using genome-wide cell free DNA (cfDNA) fragmentation profiles and machine learning can be used for detection of liver and other cancers. Having previously demonstrated that a DELFI classifier accurately detects HCC in populations from the US and Hong Kong, we evaluated the classifier in other clinically relevant cohorts worldwide. Here we show that approach generalizes to diverse populations, and that including other genomic and protein features from the same blood draw improves performance. We examined plasma samples from 377 individuals, including 244 individuals with HCC and 133 without cancer, including 85 with cirrhosis. Plasma samples were collected from individuals with or without HCC in a case-control study in Guatemala (n=203) and from a prospective collection in Romania (n=174) and cfDNA was analyzed by whole- genome sequencing at ∼10x coverage. The median DELFI scores using a locked classifier (Foda et al., Cancer Discovery, 2023) were higher in both cohorts for patients with cancer across all stages compared to individuals without cancer, regardless of the presence of cirrhosis. Using the locked model with a threshold that corresponded to 80% specificity in prior work, DELFI detected individuals with HCC with sensitivities of 90% and 69% in the two cohorts at specificities of 92% and 86%, respectively. For early-stage HCC within Milan Criteria for liver transplantation, sensitivities were 85% in the case-control cohort and 64% in the prospective cohort. In these cohorts, the fixed DELFI model outperformed AFP at the clinically used threshold of 20 ng/mL with a sensitivity in the combined cohorts of 78% at 91% specificity, compared to 63% sensitivity at 91% specificity for AFP. A combined approach using either DELFI at a threshold that corresponded to a 90% specificity in our previous study or AFP at the clinical threshold resulted in 82% sensitivity (74% in early stage) at 92% specificity and was superior to estimates of AFP and ultrasound for early-stage disease (63% sensitivity at 84% specificity). Individuals who tested positive with DELFI had a significantly shorter overall survival (p=0.002, log rank test), even amongst individuals at the earliest stage, while AFP alone did not stratify survival for patients with early-stage disease. Single-molecule analyses from low coverage WGS of cfDNA revealed genome-wide mutational profiles that were similar to those of HCC and in individuals from Guatemala that were characteristic of aflatoxin exposure. Overall, this work provides insight into the origins of cfDNA in populations at risk for HCC and validates our genome-wide fragmentome approach for non-invasive cancer detection that may facilitate liver cancer screening. Citation Format: Zachariah H Foda, Daniel Bruhm C Bruhm, Akshaya V Annapragada, Shashikant Koul, Sarah Short, Keerti Boyapati, Adrianna Bartolomucci, Vilmos Adleff, Nicholas A Vulpescu, Hope Orjuela, Andrei Sorop, Razvan Iacob, Liana Gheorghe, Simona Dima, Katherine A McGlynn, Manuel Ramírez-Zea, Jillian Phallen, John Groopman, Robert B Sharpf, Victor E Velculescu. Early detection of liver cancer from diverse populations using cfDNA fragmentome and protein biomarkers [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A050.

Abstract B058: Novel machine learning lung cancer classifier shows significant correlation with cancer-specific fragmentomics features

Abstract Lung cancer is the second most prevalent cancer, and the leading cause of cancer-related deaths, in the US. Since the survival rate of patients diagnosed with lung cancer at earlier stages is dramatically higher than those diagnosed at later stages, technologies that shift lung cancer diagnosis to an earlier stage would be expected to improve survival rates. Fragmentomics, the study of short circulating cell-free DNA (cfDNA) fragments in the blood, is an emerging field in cancer diagnostics that has the potential to catalyze such a shift. Studies have shown that fragmentomics features, such as fragment sizes and end motifs, can be used to discriminate normal cfDNA from circulating tumor DNA (ctDNA) generated through tumor necrosis and apoptosis. Using this knowledge, machine learning tools, trained to identify ctDNA-specific features, are being leveraged in novel liquid biopsy assays to improve early-stage lung cancer detection. To create a machine learning classifier that detects discrete signals of ctDNA in individuals with lung cancer, Genece Health utilized low pass (∼3x coverage) whole genome sequencing (Illumina) of plasma extracted cfDNA from >400 lung cancer samples and >1,000 negative controls. This data was used to generate our novel FEMS (Fragment End Motifs and Sizes) dataframe, containing >30,000 features, by combining fragment lengths with their corresponding 5’ 4bp end motifs. The FEMS dataframe was then used to train a neural network machine learning classifier. To evaluate the performance of our machine learning classifier, we investigated whether the classifier scores correlated with well-known hallmarks of ctDNA. First, consistent with published ctDNA characteristics, we observed significant (p<0.01) enrichment of fragment sizes ≤167bp in lung cancer samples. Concordantly, our classifier scores are significantly correlated with this fragment size enrichment. Also, both fragment size enrichment and our classifier scores are correlated with lung cancer stage, with higher enrichment and higher scores observed in later stages. Next, we show that our classifier scores are correlated with specific end motif enrichments or depletions in lung cancer samples. For example, in lung cancer samples, CCCA end motifs were significantly enriched (p<0.01) in fragment sizes 120-140bp (max at 128bp), while significantly depleted in fragment sizes 190-210bp (min at 204bp). Once again, our classifier scores were significantly correlated with both the enrichment of this end motif in short fragments and its depletion in long fragments. Last, we assessed the classifier’s performance in an independent cohort (46 lung cancer and 132 non-cancer samples) and show 89.2% sensitivity with 90.4% specificity (auROC=0.957). The correlation observed between our classifier scores and published ctDNA characteristics, combined with the strong performance in an independent cohort, suggests that Genece’s novel machine learning classifier is effectively leveraging ctDNA-specific fragmentomics features for the identification of lung cancer in patient samples. Citation Format: Carlos Guzman, Michael L Salmans, Mengchi Wang, Byung In Lee, Andrew R Carson. Novel machine learning lung cancer classifier shows significant correlation with cancer-specific fragmentomics features [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr B058.

Non-invasive testing in metabolic dysfunction-associated steatotic liver disease

Metabolic dysfunction-associated steatotic liver disease (MASLD), previously referred to as non-alcoholic fatty liver disease (NAFLD), is a leading cause of chronic liver disease, affecting up to 30% of the global population. MASLD is strongly associated with metabolic risk factors such as obesity and type 2 diabetes, and can progress to advanced stages including cirrhosis and hepatocellular carcinoma. Early diagnosis and accurate staging of fibrosis are critical in managing the disease and preventing complications. While liver biopsy has long been considered the gold standard for assessing fibrosis, it is invasive and carries associated risks. In response, non-invasive tests (NITs) have emerged as essential alternatives for the diagnosis and monitoring of MASLD. Key methods include blood-based biomarkers such as the Fibrosis-4 (FIB-4) score, NAFLD Fibrosis Score (NFS), and Enhanced Liver Fibrosis (ELF) test, as well as imaging modalities like vibration-controlled transient elastography (VCTE) and magnetic resonance elastography (MRE). These tests provide safer, more accessible methods for identifying liver fibrosis and guiding clinical management. They are integral in assessing disease severity, guiding treatment decisions, and monitoring disease progression, particularly in light of emerging therapies. NITs have become increasingly recommended by clinical guidelines as they reduce the need for invasive procedures like liver biopsy, improving patient care and outcomes. In conclusion, non-invasive testing plays a crucial role in the effective management of MASLD, offering reliable alternatives for diagnosis and monitoring while minimizing risks associated with traditional invasive methods.

Abstract A075: Circulating T-cell receptor repertoire analysis improves cancer early detection

Abstract Introduction: Cancer screening by liquid biopsy promises to detect cancer early when it can be cured. However, current ctDNA-based approaches detect only a minority of early-stage cancers. For liquid biopsy to realize its full potential for cancer early detection, sensitivity for earliest stage disease must be improved. Methods: We set out to improve sensitivity of liquid biopsy by exploiting tumor recognition by T cells through sequencing of the circulating T-cell receptor repertoire. Using gDNA extracted from blood buffy coats, we studied a cohort of 471 lung cancer patients (85% stage I) and 600 subjects without cancer enriched for older individuals with a history of smoking. We performed TCR beta chain sequencing to yield a median of 101,899 TCR clonotypes per sample (median 81,602 cancer / 112,966 controls) and built a TCR sequence similarity graph to cluster clonotypes into TCR repertoire functional units (RFUs). The TCR frequencies of RFUs were tested for association with cancer status and significantly associated RFUs were combined using an SVM model into a cancer score. Cancer RFU levels were also correlated to imputed subject HLA types for 32 HLA Class I and 38 Class II alleles. The model was evaluated by 10-fold cross-validation and compared to a ctDNA panel of 237 mutation hotspots in 154 lung cancer driver genes and to 17 lung cancer related protein biomarkers in 86 subjects. Results: We identified 372 cancer-associated TCR RFUs at FDR≤0.1, including 198 enriched in cancer samples (fold-change range 1.03 to 3.13) and 174 enriched in controls (fold- change 0.96 to 0.30). Levels of 265/372 (71%) RFUs were correlated to presence of an HLA allele at FDR ≤ 0.1. Of the total 2,770 significant HLA-RFU associations, 479 (17%) involved an HLA Class I allele, while 2,291 (83%) involved an HLA Class II allele, highlighting a potential role for CD4 helper or regulatory T cell antigen recognition in the cancer RFUs found. The RFU cancer score achieved a cross-validated test ROC AUC of 0.71 for cancer status prediction (0.71 Stage I / 0.70 Stage II-IV) with 49% of Stage I cancers detected at a specificity of 80%. Importantly, the cancer score was not dependent on sample source site, technical or biological variation in TCR repertoire depth, or lung cancer histology subtype. The TCR RFU score also distinguished lung cancer patients from control subjects with other conditions such as benign lung nodules and COPD. We observed an up to ∼20%-point gain in sensitivity for stage I cancer when TCR RFUs were added to ctDNA and proteins in a potential multi-analyte cancer screening test. By contrast, TCR analysis did not provide additional sensitivity for stage II-IV disease. Conclusion: We demonstrate detection of a significant fraction of early lung cancer cases from blood by analyzing the circulating TCR repertoire and show that this signal is complementary to established analytes such as proteins and ctDNA. A similar study for the detection of early colorectal cancer and advanced adenomas is under way. Citation Format: Yilong Li, Michelle Nahas, Dennis Stephens, Kate Froburg, Emma Hintz, Devin Champagne, Amaneet Lochab, Markus Brown, Jasper Braun, María Antonia Fortuño, María-del-Mar Ocón, Andrea Pasquier, Inés María Luque, Christopher W. Seder, Jeffrey A. Borgia, Luis Miguel Seijo, Luis M Montuenga, Roman Yelensky. Circulating T-cell receptor repertoire analysis improves cancer early detection [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A075.

Abstract B011: Circulating tumor DNA profiling in colorectal cancer to detect guideline-based targeted mutations at a Quebec health care center

Abstract Liquid biopsy is an emerging tool in oncology, providing minimally invasive detection of tumor- specific DNA in body fluids, known as circulating tumor DNA (ctDNA). Colorectal cancer (CRC) has shown a high rate of shedding of ctDNA, making it a promising prognostic and possibly predictive biomarker in the personalized management of patients with CRC. Therefore, the half-life of ctDNA in circulation is estimated to be between 16 minutes and several hours, indicating a need for rapid processing. This study aimed to explore the concordance between the mutational profiles in tumor tissue and ctDNA from plasma obtained using EDTA tubes as well as the correlation between ctDNA and disease stage, to evaluate the potential of implementing liquid biopsy testing in CRC cases, following biobank standards. In this retrospective cohort, 58 patients with CRC stage T1 to T4 were enrolled at the Research Institute McGill University Health Centre (RI-MUHC). Tumor tissue was obtained through surgical resection or biopsies of formalin-fixed paraffin-embedded tissue, and blood samples were drawn pre-surgery using EDTA tubes. Cell-free DNA (cfDNA) was extracted from 1-2mL plasma samples using QIAamp Circulating Nucleic Acids. Next-Generation Sequencing using a 52-gene (AmpliSeq for Illumina Focus Panel) was performed to identify the tumor mutations and subsequently droplet digital PCR (ddPCR) was performed to identify the plasma mutations. Among the 58 patients, the mean age was 62 years (range 29-86), and 29 were women. Forty- one patients had left-sided tumor. At least 1 tumor-specific mutation was detected in all tissue tumor samples. Overall, the most frequently identified mutations in this cohort were: BRAF V600E (22.4%), KRAS G12D (12.1%), KRAS G12V and KRAS G13D (8.6%). Cell-free DNA was obtained from 32 patients and showed a mean concentration of 975 ng/mL (range 400- 1,995) for stage T1, 584 ng/mL (range 228-1,300) for stage T2, 1,127 ng/mL (range 249-4,160) for stage T3 and 2,083 ng/mL (range 860-3,680) for stage T4. ddPCR revealed mutation concordance between ctDNA and matched tumor tissue of 28.1%. When tumors were stratified by stage, the concordance was 0% (n=0/3) in patients with T1 tumors, 16.7% (n=1/6) in patients with T2 tumors, 25% (n=5/20) in patients with T3 tumors and 100% (3/3) in patients with T4 tumors. This study showed that ctDNA detection of mutations has excellent concordance in stage T4 tumors, highlighting the potential of ctDNA in advanced stage disease. However, in T1-3 tumor stages the concordance was low, showing the importance of preanalytical steps such as the use of cell-free DNA preservative tubes for sensitive and accurate detection of ctDNA in patients with earlier stage disease. Citation Format: Monyse de Nobrega, Kyle Dickinson, Saba Alsaddah, Alexandra Bartolomucci, Tadhg Ferrier, Anne Mahalia Olivier Mahalia Olivier, Mina Farag, Gertuda Evaristo, Sophie Camilleri-Broet, Andrea Gomez, Thupten Tsering, Lawrence Lee, Pierre Olivier Fiset, Julia V. Burnier. Circulating tumor DNA profiling in colorectal cancer to detect guideline-based targeted mutations at a Quebec health care center [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr B011.

The effect of sevelamer on serum calcification propensity in patients with chronic kidney disease: the results of a multicentre, double-blind, placebo-controlled, randomized clinical trial

Abstract Background The serum calcification propensity test (or T50 test) might become a standard tool for the assessment of vascular calcification risk, and T50 might be a valuable biomarker in clinical trials of treatments intended to slow the progression of vascular calcification. Literature data suggest that non-calcium-containing phosphate binders can influence T50 in chronic dialyzed patients. However, it is not clear whether similar interventions are effective in patients at earlier stages of chronic kidney disease (CKD). Methods The “FGF23 Reduction efficacy of a new Phosphate Binder in CKD” (FRENCH) trial was a multi-centre, double-blind, placebo-controlled, randomized trial of sevelamer carbonate in participants with stage 3b/4 CKD. In this subanalysis of the FRENCH data, T50 and other laboratory variables (including fetuin A, and ionized and total magnesium) were measured centrally at baseline and after 12 weeks of treatment. Results A total of 96 patients were screened; 78 (55 men and 23 women) met the inclusion criteria and were randomized to receive placebo (n=39) or sevelamer carbonate (n=39). The median [interquartile range] patient age was 66 [56-72], the median eGFR was 25 [21-30] mL/min/1.73m², and the mean T50 was 335 (82) minutes. In a linear regression model, T50 was independently associated with serum ionized magnesium, fetuin-A and bicarbonate levels; and inversely associated with phosphate concentration. The within-group changes in the mean [95% confidence interval] T50 between week 0 and week 12 were not significant in the sevelamer group or the placebo group (4.6 [-13.6; 22.8] minutes (p=0.61) and 7.8 [-16.4; 32.1] minutes (p=0.51), respectively). Furthermore, we did not observe significant changes in fetuin-A and magnesium levels. Conclusion A 12-week course of the non-calcium-containing phosphate binder sevelamer carbonate was not associated with a significant change in T50 in patients with stage 3b/4 CKD. Phosphate binders might not be an effective strategy for modifying serum calcification propensity in non-dialysis-dependent patients with CKD.

A systematic review on feature extraction methods and deep learning models for detection of cancerous lung nodules at an early stage -The Recent trends and challenges.

Lung cancer is one of the most common life-threatening worldwide cancers affecting both the male and female populations. The appearance of nodules in the scan image is an early indication of the development of cancer cells in the lung. The Low Dose Computed Tomography screening technique is used for the early detection of cancer nodules. Therefore, with more Computed Tomography (CT) lung profiles, an automated lung nodule analysis system can be utilized through image processing techniques and neural network algorithms. A CT image of the lung consists of many elements such as blood vessels, ribs, nodules, sternum, bronchi, and nodules. These nodules can be both benign and malignant, where the latter leads to lung cancer. Detecting them at an earlier stage can increase life expectancy by up to 5 to 10 years. To analyze only the nodules from the profile, respected features are calculated using image processing techniques. Based on the review, textural features were promising ones in medical image analysis and for solving computer vision problems. The significance of extracting the hidden features (textural) enables the higher performance of deep learning algorithms (DL) predominantly in medical imaging, which has resulted in improvement in accuracy in recent years. Earlier detection of cancerous lung nodules is possible through the combination of multi-featured extraction and classification techniques using image data. In this paper, we discussed the overview of lung cancer along with publicly available datasets for research purposes. The primary objective of the paper is to provide the importance of textural features when combined with different deep-learning models. It gives insights into their advantages, disadvantages, and limitations regarding possible research gaps. The paper compared the recent studies of deep learning models with and without feature extraction and concluded that DL models that include feature extraction were better than the others.

Assessing the Impact of Mass Media in Investigation of Cases in the Criminal Justice System of Uganda

This article assesses the impact of mass media in investigation of cases in the criminal justice system of Uganda. The study reveals that the media, through its depictions of crime, significantly influences public perception and understanding of criminal justice. Moreover, we found that the evolution of technology, specifically social media, plays a substantial role in media criminology. On this note, the article calls for force media liaison officers should be formally members of management teams within serious crime investigations, at least for the initial phase of the investigation when press interest is likely to be at its height. Second, investigators could achieve this by participating in joint exercises with media liaison and exposing them to a variety of media handling scenarios and exercises. Third, we should also consider introducing investigators to the principles of media handling in criminal investigations at an earlier stage in their careers. Fourth, there should be more effort to improve communications between press and media officers, as well as CID. This could be achieved through a combination of workshops and short secondments of junior officers in media offices. Finally, as part of ongoing family liaison, we should monitor and address the media’s impact on the relatives of victims. At the earliest opportunity, family liaison officers should provide relatives with copies of all formal press statements. Keywords: Court trial, Crime, Criminal justice system, Investigation, Mass media.

The challenges in the utilization of marine genetic resources in national and international jurisdictions from a legal perspective

This article examines the legal challenges associated with the utilization of marine genetic resources (MGR) at both the national level and beyond national jurisdiction (BBNJ). The legal challenges addressed are as follows: 1) MGR are located across various jurisdictions, encompassing both national and international domains. The analysis starts with an overview of the international regulations that govern the utilization of genetic resources (GR) and their influence on national legislation. It emphasizes the principle of state sovereignty over natural resources while defining MGR and determining ownership; 2) It further highlights the intersection of national and international laws, particularly in transboundary contexts and within Indigenous and Afro-descendant peoples (IADP) territories, analyzing how these regulations are interpreted and applied in such scenarios; 3) The legal challenges related to the use of MGR in international waters are examined. Special emphasis is placed on the recent United Nations (UN) Agreement concerning this issue. This includes an analysis of its impact and specific provisions related to the utilization of MGR, such as the quantity to be collected, the methodology employed, collection sites, among others. The article concludes by asserting that the equitable distribution of benefits from the use of GR should begin at the earliest stages of access to these resources, including project planning and sample collection, rather than being delayed until the patenting and commercialization phases. Early benefit-sharing is essential for promoting fairness and equity in the use of MGR.

Leveraging Implementation Science at the Early-Stage Development of a Novel Telehealth-Delivered Fear of Exercise Program to Understand Intervention Feasibility and Implementation Potential: Feasibility Behavioral Intervention Study.

To increase real-world adoption of effective telehealth-delivered behavioral health interventions among midlife and older adults with cardiovascular disease, incorporating implementation science (IS) methods at earlier stages of intervention development may be needed. This study aims to describe how IS can be incorporated into the design and interpretation of a study assessing the feasibility and implementation potential of a technology-delivered behavioral health intervention. We assessed the feasibility and implementation potential of a 2-session, remotely delivered, home-based behavioral intervention composed of psychoeducation, interoceptive exposure through low-to-moderate intensity walking, interoceptive counseling, and homework (Reducing Exercise Sensitivity with Exposure Training; RESET) among patients with recent acute coronary syndrome (ACS) and some fear of exercise. To assess intervention feasibility, we measured patient protocol adherence, intervention delivery fidelity, and completion of intervention outcome assessments using direct observations, fidelity checklists, surveys, and device-measured physical activity. To assess implementation potential, we measured implementation outcomes (feasibility, acceptability, and appropriateness) using 4-item measures, each rated from the patient perspective on a 1 to 5 Likert scale (1=completely disagree and 5=completely agree; criteria: ≥4=agree or completely agree), and patient-perceived implementation determinants and design feedback using survey and interview data. Interview data underwent thematic analysis to identify implementation determinant themes, which were then categorized into Consolidated Framework for Implementation Research (CFIR) domains and constructs. Of 31 patients approached during recruitment, 3 (10%) were eligible, enrolled, and completed the study (mean age 46.3, SD 14.0 y; 2/3, 67% male; 1/3, 33% Black; and 1/3, 33% Asian). The intervention was delivered with fidelity for all participants, and all participants completed the entire intervention protocol and outcome assessments. On average, participants agreed that the RESET intervention was feasible and acceptable, while appropriateness ratings did not meet implementation criteria (feasibility: mean 4.2, SD 0.4; acceptability: mean 4.3, SD 0.7; and appropriateness: mean 3.7, SD 0.4). Key patient-perceived implementation determinants were related to constructs in the innovation (design, adaptability, and complexity), inner setting (available resources [physical space, funding, materials, and equipment] and access to knowledge and information), and innovation recipient characteristics (motivation, capability, opportunity, and need) domains of the CFIR, with key barriers related to innovation design. Design feedback indicated that the areas requiring the most revisions were the interoceptive exposure design and the virtual delivery modality, and reasons why included low dose and poor usability. The RESET intervention was feasible but not implementable in a small sample of patients with ACS. Our theory-informed, mixed methods approach aided our understanding of what, how, and why RESET was not perceived as implementable; this information will guide intervention refinement. This study demonstrated how integrating IS methods early in intervention development can guide decisions regarding readiness to advance interventions along the translational research pipeline.