Immune System Research Articles

Research progress on the mechanism of exercise against depression

The substantial global health burden of depression encourages the development of innovative and broadly effective interventions. This paper aimed to examine recent advancements by which exercise works as an antidepressant and recommends optimal types and quantity of exercise as supplemental therapies in treating depression. Sedentary behavior and low physical activity significantly influence the onset of depression. Being an effective treatment option, exercise can significantly reduce depression risk. Exercise exerts antidepressant effects as it modulates neurotransmitters, neuroplasticity, the immune system, and hormone levels. Effective exercise forms include yoga, strength training, and walking/jogging. Tailored exercise regimens that consider individual preferences and tolerability can improve outcomes. Regular exercise enhances general well-being and reduces depressive symptoms. Additional research is needed to understand the complex basis of exercise's effects on depression. Exercise is a cost-effective and accessible intervention for depression management that needs additional exploration. Thus, customized exercise programs, as per each patient’s needs, are essential for their successful implementation clinically.

Review of Thymic Peptides and Hormones: From Their Properties to Clinical Application

Abstract Background The thymus is the main lymphoid organ that regulates the functions of the immune system, protecting against pathogens, tumors, antigens, and mediators of tissue damage. It produces a family of hormone-like peptides that can modulate physiological processes such as stimulation or suppression of immune responses, angiogenesis, and wound healing. Objective This review aims to comprehensively characterize the properties of thymic peptides and their clinical applications. Methodology This article discusses the structure, biological properties, mechanism of action, and therapeutic applications of the most important thymic hormones (thymosin alpha 1, thymosin beta-4, thymulin, and thymopoietin), as well as preparations that are purified thymic extracts. Results Thymic peptides and extracts act in multiple manners on the immune system: they stimulate the differentiation and maturation of T cells, regulate the activity of natural killer cells and dendritic cells, and induce the release of proinflammatory cytokines, and their immunomodulatory effects have been confirmed in numerous clinical studies. An important feature of thymus preparations is their therapeutic safety—even long-term use does not cause side effects. Conclusions A wide range of therapeutic uses, i.e., from defensive and immunomodulatory tasks to participation in tissue regeneration processes, has led to the use of thymic peptides in the treatment of neoplastic diseases, viral infections, autoimmune diseases or immunodeficiencies. Further investigation of the mechanisms of action of thymic peptides may contribute to the discovery of new therapeutic targets.

Sleep & Nutrition for Athletes

Abstract Sleep is vital for the maintenance of physical and mental health, recovery and performance in athletes. Sleep also has a restorative effect on the immune system and the endocrine system. Sleep must be of adequate duration, timing, and quality to promote recovery following training and competition. Inadequate sleep adversely impacts carbohydrate metabolism, appetite, energy intake and protein synthesis affecting recovery from the energy demands of daily living and training/competition related fatigue. Sleep’s role in overall health and wellbeing has been established. Athletes have high sleep needs and are particularly vulnerable to sleep difficulties due to high training and competition demands, as such the implementation of the potential nutritional interventions to improve sleep duration and quality is commonplace. The use of certain nutrition strategies and supplements has an evidence base i.e. carbohydrate, caffeine, creatine, kiwifruit, magnesium, meal make-up and timing, protein and tart cherry. However, further research involving both foods and supplements is necessary to clarify the interactions between nutrition and the circadian system as there is potential to improve sleep and recovery. Additional research is necessary to clarify guidelines and develop products and protocols for foods and supplements to benefit athlete health, performance and/or recovery. The purpose of this review is to highlight the potential interaction between sleep and nutrition for athletes, and how these interactions might benefit sleep and/or recovery.

Using GeoMx DSP Spatial Proteomics to Investigate Immune Infiltration of NOD Mouse Islet and Exocrine Compartments

Abstract Purpose Type 1 Diabetes (T1D) pathogenesis involves immune cells infiltrating pancreatic Islets of Langerhans, leading to T cell activation, beta cell destruction, and impaired insulin production. However, infiltration has a heterogenic nature that isn’t described in detail, as not all islets are infiltrated. The aim of this study was to investigate if the observed heterogeneity is coupled to differences in immune and/or dysfunctional status of islets or exocrine cells, and if specific markers could elucidate mechanistic details of T1D pathogenesis. Procedures The GeoMx platform was used to spatially quantify protein levels in pancreatic islets and exocrine tissue in Non-Obese Diabetic (NOD) mice. The protein panel included 17 immune activity markers and nine dysfunction markers. Immunohistochemical (IHC) staining and digital image analysis was used to analyze select marker proteins. Results Use of the GeoMx platform to investigate T1D was shown to be possible, as Granzyme B protein levels were found to be lower in distal islet areas when compared to proximal areas. Smooth Muscle Actin protein levels were higher in exocrine areas proximal to immune-infiltrated islets, when compared to distally located exocrine areas. Findings from GeoMx were however not observed in IHC-stained sections. Conclusions This study demonstrates that investigating T1D is possible with spatial proteomics, as the assays revealed presence of heterogenic islet areas in NOD mice, which may play a role in T1D progression and escape from immune recognition. This study highlights the potential of spatial technologies for elucidating T1D pathogenesis and future treatment strategies.

Challenges for the human immune system after leaving Earth

AbstractFrom the start of life on Earth, several immune defense mechanisms have evolved to guarantee cellular integrity, homeostasis, and host survival. All these sophisticated balances as shaped by and towards the environmental needs have occurred over hundreds of millions of years. Human spaceflight involves various health hazards, such as higher levels of radiation, altered gravity, isolation and confinement, living in tight quarters, and stress associated with being away from home. A growing body of evidence points towards immunological changes in astronauts, including heightened pro-inflammatory responses, reactivation of latent viruses, and cell-mediated alterations, reflecting a dysbalanced state in astronauts. Simultaneously, enhanced pathogenicity, virulence, and drug resistance properties of microorganisms tip the scale out of favor for prolonged stay in space. As we have learned from the past, we see potential for the human immune system, forged and maintained throughout evolutionary history, to adapt to the space exposome. It is unlikely that this will happen in the short time frames set for current space exploration missions. Instead, major risks to astronaut health need to be addressed first, before humans can safely evolve into the space environment.

An update on the pathogenesis and ethnopharmacological therapeutic approaches of vulvovaginal candidiasis

AbstractIn the era of globalization, the concept of world health needs to be practiced where a significant consideration should be given to women health. Of many issues, the most common and under-rated is vulvovaginal candidiasis (VVC); a superficial to invasive mucosal infection of lower reproductive tract in women, primarily caused by commensal yeast Candida. The inability of conventional drugs in triumphing the VVC has made researchers to review the potential of traditional medicines. The present review aimed to highlight the problem and concerns of women related to VVC, covering all aspects of disease development, host response, conventional drug failure, and possibilities of formulating phytomolecule based VVC therapy. The health and disease issues in women are considerably severe, especially for those living in impoverished conditions. Any dysbiosis in vaginal microbiome disturbs the harmony between residing microbes and leads to commencement of VVC. Lactobacillus is key care-taker microbe that keeps check on over growth of Candida by various means. However, Candida have evolved numerous virulence traits to overcome the governance of Lactobacillus as well as host immune system. The present review comprehensively outlined the virulence, and pathogenicity of Candida along with its interaction with host immune system. The review has also highlighted the current conventional drugs and their drawbacks, along with potential of ethnopharmacological phytoactive molecules as future therapies for defining the therapeutic management of the disease. Graphical Abstract

Profiling serum immunodominance following SARS-CoV-2 primary and breakthrough infection reveals distinct variant-specific epitope usage and immune imprinting

Over the course of the COVID-19 pandemic, variants have emerged with increased mutations and immune evasive capabilities. This has led to breakthrough infections (BTI) in vaccinated individuals, with a large proportion of the neutralizing antibody response targeting the receptor binding domain (RBD) of the SARS-CoV-2 Spike glycoprotein. Immune imprinting, where prior exposure of the immune system to an antigen can influence the response to subsequent exposures, and its role in a population with heterogenous exposure histories has important implications in future vaccine design. Here, we develop an accessible approach to map epitope immunodominance of the neutralizing antibody response in sera. By using a panel of mutant Spike proteins in a pseudotyped virus neutralization assay, we observed distinct epitope usage in convalescent donors infected during wave 1, or infected with the Delta, or BA.1 variants, highlighting the antigenic diversity of the variant Spikes. Analysis of longitudinal serum samples taken spanning 3 doses of COVID-19 vaccine and subsequent breakthrough infection, showed the influence of immune imprinting from the ancestral-based vaccine, where reactivation of existing B cells elicited by the vaccine resulted in the enrichment of the pre-existing epitope immunodominance. However, subtle shifts in epitope usage in sera were observed following BTI by Omicron sub-lineage variants. Antigenic distance of Spike, time after last exposure, and number of vaccine boosters may play a role in the persistence of imprinting from the vaccine. This study provides insight into RBD neutralizing epitope usage in individuals with varying exposure histories and has implications for design of future SARS-CoV-2 vaccines.

Epigenetic Alterations in Post-Traumatic Stress Disorder: Comprehensive Review of Molecular Markers

Background: Post-traumatic stress disorder (PTSD) can occur after a traumatic event. PTSD is characterized by nightmares, flashbacks and avoidance of stressors. It currently affects 2–8% of the population, with military personnel particularly susceptible. Studies show that environmental stressors can induce various epigenetic changes that shape the PTSD phenotype. Despite the significant impact of epigenetic factors on PTSD symptoms and susceptibility, they have not been widely discussed in the literature. This review focuses on describing epigenetic mechanisms in PTSD, especially DNA methylation, chromatin regulation, and noncoding RNA. Summary: The article includes relevant studies published from 2013 to 2023, excluding non-English-language studies or studies with insufficient data. This review investigated gene methylation changes in association with PTSD, including those related to the hypothalamic-pituitary-adrenal axis, brain-derived neurotrophic factor, neurotransmitters, and immune system functioning, as well as the role of histones and regulatory noncoding RNAs. Key Messages: Epigenetic alterations play a crucial role in shaping PTSD susceptibility, symptomatology, and long-term outcomes, highlighting their potential as important markers and therapeutic targets. Understanding these alterations can aid in developing clinical strategies to better predict, prevent, and treat PTSD. However, further large-scale longitudinal studies are needed to establish the temporal relationship between epigenetic changes and the onset of PTSD, as well as to classify other potential epigenetic mechanisms.

Chlorella vulgaris as a food substitute: Applications and benefits in the food industry

AbstractChlorella vulgaris, a freshwater microalga, is gaining attention for its potential as a nutritious food source and dietary supplement. This review aims to provide a comprehensive discussion on C. vulgaris, evaluating its viability as a food substitute in the industry by exploring the nutritional value and application of C. vulgaris in the food industry. Rich in protein, lipids, carbohydrates, vitamins, and minerals, Chlorella offers substantial nutritional benefits, positioning it as a valuable food substitute. Its applications in the food industry include incorporation into smoothies, snacks, and supplements, enhancing the nutritional profile of various food products. The health benefits of Chlorella encompass antioxidant activity, immune system support, and detoxification, contributing to overall well‐being. Despite these advantages, the commercialization of Chlorella faces significant challenges. These include variability in antibacterial activity due to strain and growth conditions, high production costs, contamination risks, and sensory issues such as unpleasant taste and smell. Additionally, Chlorella can accumulate heavy metals from its environment, necessitating stringent quality control measures. Future prospects involve improving Chlorella strains through genetic manipulation to enhance nutrient content, developing cost‐effective culture systems, and exploring advanced processing techniques like pulsed electric fields for better digestibility. Addressing sensory issues through flavor‐masking strategies and employing environmental management practices will further support Chlorella's integration into the food industry. Although C. vulgaris shows great potential as a nutritious food ingredient, overcoming existing challenges and optimizing production methods would be crucial for its successful adoption and widespread use.

Abstract B039: Exploring the impact of neoantigen expression on lung tumor development

Abstract Lung adenocarcinoma (LUAD) stands as a formidable challenge in oncology, despite the emergence of immunotherapy as a promising treatment avenue. The dynamic interplay between the immune system and cancer plays a pivotal role in shaping cancer development and progression. Central to this process are neoantigens, tumor-specific antigens recognized by cytotoxic T cells, which trigger the elimination of tumor cells. It has been implicated that the presence of T cells also decreases tumor grade. However, the precise impact of neoantigen expression on LUAD development and progression remains poorly understood. Our lab previously developed a genetically engineered mouse model called the iNversion-INduced Joined neoAntigen (NINJA) system with the ability to induce expression of a fluorescent neoantigen in oncogenic KRAS, P53-null, tdTomato-expressing lung adenocarcinoma cells (KPT-NINJA). Using this system, we track tumor initiation, progression, and advanced LUAD development across 20 weeks in both immune-deficient and -sufficient settings. Further, tumors initiated in this system give rise to heterogeneous (neoantigen+ and -) tumors, enabling the study of T cell-driven tumor selection. Through multiplexed immunofluorescence, we identified that neoantigen+ tumors experience increased immune infiltration (CD3+, CD4+, CD8+, B220+) and reduced volume, ultimately leading to prolonged animal survival. Surprisingly, advanced lung tumors maintain neoantigen+ cells, indicating that endogenous lung T cells are ineffective at clearing neoantigen-expressing tumors. Yet, we detected that infiltrated tumors exhibit reduced p-ERK expression, suggesting that T cells may slow tumor progression through a non-cytotoxic mechanism. Transferring stimulated neoantigen-specific T cells to tumor-bearing mice largely eliminated neoantigen+ tumor cells. All these data together suggest complex mechanisms for the role of endogenous T cells in controlling lung tumors. Through molecular characterization of tumor progression and T cell-mediated tumor suppression, this study reveals specific pathways that LUAD cells undergo during the process of development and progression, providing valuable insights into the immune response against neoantigen-expressing tumors. The characterization of early neoantigen-expressing tumor cells is pivotal in validating its utility for studying cancer immunotherapy and identifying key molecular changes during tumor development. Ultimately, by unraveling the intricate interplay between neoantigen expression, immune surveillance, and tumor progression, this project aims to contribute significantly to our understanding of how neoantigens shape the dynamics of tumor biology. Citation Format: Jennifer L. Loza, Ishan Bansal, Brian Hunt, Kelli Connolly, Advait Jeevanandam, Srividhya Venkatesan, Nikhil Joshi. Exploring the impact of neoantigen expression on lung tumor development [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 B039.

Abstract B029: Mapping the vascular walls of colorectal cancer

Abstract Introduction: Properties of tumor angiogenesis impact on tumor growth and composition of the tumor microenvironment. Tumor vessels are composed of pericytes and other mural cells. Detailed characterization of the cellular composition of the microvasculature can suggest functional vessel and case properties relevant for outcome, response to treatment and immune surveillance. Results: Based on single cell RNA-seq from three human stage II/III colon cancers and pilot in-situ multi-antibody staining of six tumors two main clusters of perivascular cells were identified; B1 (MCAM+, MYH11-) and B2 (MCAM+, MYH11+) cells. B1 cells displayed pericyte features including expression of RGS5, whereas B2 cells showed high expression of smooth muscle cell markers like ACTG2 and Desmin. Desmin-high B2 cells were predominantly located in muscular layers, whereas Desmin-low/MYH11-high B2 cells showed perivascular enrichment. An extended novel analyses workflow of digital image analysis was used to profile peri-endothelial composition in 19 intervals; 11 1-µm intervals from 0 to 10 µm and 8 5-µm intervals from 10 to 50 µm from endothelial areas. Each of these expansion areas was quantitated regarding density of PDGFRB+/- cells including B1/B2 cells, and a MYH11+/MCAM- cells. Density of macrophage subsets (CD68-positive subsets defined by CD163 and CD11c) were also quantitated. In general, B1 cells showed stronger spatial association with endothelial cells than B2 cells, and this spatial association was stronger for the PDGFRB+ B1 subset. Survival analysis indicated that prognostic relevance was strongest when PDGFRB+ (favorable prognosis) cells were closest to endothelial cells. Regarding perivascular macrophage density, M0 and transit M1/M2 macrophages were predominantly enriched in the peri-endothelial regions, whereas density of M1 and M2 increased gradually as distance from vessels increased. M2 density was overall associated with poor prognosis, whereas high density of other macrophage subsets overall was associated with good prognosis. Ongoing studies are using a clustering-based approach to identify vessel subsets, and prognosis associations are being consolidated in additional cohorts. Summary: This study provides a novel approach for high resolution profiling of perivascular status in CRC, and possibly other tumor types, that could be of specific importance to identify tumor features associated with response to angiogenesis-directed therapies. Citation Format: Linglong( 凌 龙 ) Huang( 黄 ), Mercedes Herrera, Jonas Sjölund, Vladimir Chocoloff, Simon Joost, Rasul M. Tabiev, Lina Wik Leiss, Carina Strell, Luis Nunes, Artur Mezheyeuski, David Edler, Anna Martling, Fredrik Pontén, Bengt Glimelius, Tobias Sjöblom, Maria Kasper, Kristian Pietras, Arne Östman. Mapping the vascular walls of colorectal 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 B029.

Abstract PR014: DNA associated with EVs is uniquely chromatinized and prevents metastasis by enhancing anti-tumor immunity

Abstract Cells release extracellular vesicles (EVs) loaded with functional biomolecules that facilitate intercellular communication in health and disease. These EVs contain DNA (EV-DNA), mirroring the mutational profile of parental cells, yet the mechanism of DNA packaging onto EVs and its role in diseases like cancer remain unclear. Using advanced imaging and biochemical techniques, we discovered that EV-DNA predominantly resides on the vesicle surface, bound to modified histones. Through genome-wide CRISPR knockout screens in cancer cell lines, we identified immune-related pathways and genes crucial for EV-DNA loading, such as APAF1 and NCF1. In several cancer models, loss of APAF1 reduced EV-DNA packaging, and enhancing metastasis, thus implicating EV-DNA in immune surveillance against cancer spread. We demonstrated that EV-DNA uptake by liver macrophages triggers DNA damage response, altering cytokine production and promoting immune activation in pre-metastatic sites. Moreover, quantifying EV-DNA in colorectal cancer biopsies revealed its potential as a predictive metastatic risk biomarker. Our findings shed light on EV-DNA packaging mechanisms and its significance, suggesting that elevated levels of tumor-derived EV-DNA bolster anti-tumor immunity and inhibit metastasis. Citation Format: Inbal Wortzel, Han-Sang Kim, David C. Lyden. DNA associated with EVs is uniquely chromatinized and prevents metastasis by enhancing anti-tumor immunity [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 PR014.

Abstract B021: Aneuploidy-associated cohesin RAD21 gains promote immune evasion in prostate cancers

Abstract Aneuploidy, characterized by imbalanced chromosome numbers, is a hallmark of cancer and is strongly correlated with lethal progression in prostate cancer. Our recent work identified that gains of chromosome 8q, the most frequently gained chromosome in aneuploidy prostate cancers, drive cancer progression, partially through the cohesin RAD21 gene located on the chr8q24 region. We demonstrated that increased RAD21 expression accelerates lethal progression and oncogenesis in both prostate cancer and Ewing sarcoma by alleviating oncogenic stress during early-stage oncogenesis in both cell line and organoid models. Therefore, we hypothesize that elevated RAD21 levels could enable cancer cells to evade immune surveillance by mitigating oncogenic stress and DNA damage. In this study, we developed isogenic prostate and breast cancer cell culture models with varying levels of RAD21. Using a natural killer (NK) cell co-culture assay, we observed that cells with elevated RAD21 expression were significantly less susceptible to NK-mediated cytotoxicity and exhibited increased survivorship when co-cultured with NK cells. These findings align with publicly available data showing that tumors with high RAD21 levels have reduced NK cell infiltration in prostate cancer. Our results suggest that increased RAD21 expression, commonly associated with chr8q gains, facilitates immune evasion in cancer cells, thereby promoting early-stage oncogenesis. Understanding the role of RAD21 in immune evasion could reveal novel therapeutic strategies targeting immune surveillance mechanisms in prostate cancer. Citation Format: Elise G. DeArment, Ruoxi W. Wang, Kate Lu, Xiaofeng A. Su, Andrew Elliott, Nicholas A. Zorko, Justin H. Hwang, Xiaofeng A. Su. Aneuploidy-associated cohesin RAD21 gains promote immune evasion in prostate cancers [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 B021.

Abstract B001: Intra- and inter-tumoral heterogeneity of melanoma across different metastatic sites

Abstract Melanoma, a highly metastatic skin cancer, exhibits variations in prognosis and response to therapy based on the site of metastasis. Despite the success of immunotherapy and targeted therapies in melanoma, over half of metastatic melanoma patients will experience disease progression due to therapy resistance. The heterogeneity and plasticity of melanoma cells contribute to metastatic dissemination and therapy resistance. Our aim is to determine whether distinct clones and/or their transcriptional cell states can predict the formation of tumors in various organs and assess how these clones change over time. To identify melanoma clones across different metastatic sites, NOD scid gamma (NSG) mice and C57BL/6 mice were injected subcutaneously, intravenously or intracranially with the same pool of cells of barcoded luciferase expressing YUMMER1.7 murine melanoma cells. Transduction conditions ensured that one DNA barcode integrated into cell genomes at one barcode per cell, serving as a lineage tag. Bioluminescence imaging was performed once a week to monitor tumor growth of mice injected intravenously and intracranially. Subcutaneous tumors were measured by calliper. Mice were euthanized at different time points; day 8, 15, 22, 29 post-implantation and at ethical endpoints. Tumors were harvested and DNA sequencing was performed to identify barcodes expressed by the tumor cells. All mice developed tumors, with 100% penetrance in NSG mice. However, in C57BL/6 mice, 10% of mice intravenously injected and 27% of subcutaneously implanted mice showed complete lesion regression, suggesting that the immune system may be responsible for tumor regression. Analysis of barcodes allowed us to assess the heterogeneity of melanoma tumors at different metastatic sites and their evolution over time. Lineage tracing and clonal heterogeneity will be evaluated using the state of art technology, SPLINTR (Single-cell Profiling and LINeage Tracing), enabling us to match a cells evolution with changes in transcriptional states. Barcode analysis performed before implanting the cells and, at different timepoints in subcutaneous and lung tumors showed that dominant subclones at the baseline were also dominant in subcutaneous and lung tumors in immunocompromised mice. In contrast, dominant subclones in immunocompetent mice was those present in lower proportion at baseline. Additionally, greater variability of subclones was observed especially in lung and brain tumors, across immunocompetent mice, likely as a mechanism of resistance, enabling these tumors to overcome immunoediting. Understanding the variability of clonality between different metastatic sites and over time will improve our comprehension of the role of different subclones in organ- specific metastasis and their transcriptional cell states. Citation Format: Veronica L. Aedo Lopez, Reem Saleh, Benjamin Blyth, Xin Du, Dane Vassiliadis, Katie Fennell, Davide Moi, Roberta Mazzieri, Riccardo Dolcetti, Karen E. Sheppard, Grant A. McArthur. Intra- and inter-tumoral heterogeneity of melanoma across different metastatic sites [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 B001.

Abstract A023: Arachidonic acid metabolism promoting cross resistance in melanoma cells

Abstract Melanoma patients initially respond well to first line immunotherapy. However, cross resistance to immunotherapy and BRAF/MEK inhibitor is a clinical challenge for melanoma care. To investigate drug tolerant and resistant mechanisms promoting cross resistance, we analyzed RNA-sequencing and gene set enrichment targeted therapy resistant melanoma cells revealing an enriched arachidonic acid gene signature. Intracellular arachidonic acid levels were increased in drug resistant and drug tolerant melanoma models. Arachidonic acid is enzymatically converted by COX1 and COX2 to prostaglandins, which is subsequently secreted into the microenvironment. Drug tolerant and resistant cells had increased protein expression of COX1 and COX2 concurrently with increased secretion of prostaglandin E2. COX1/2 inhibition and COX2 knockdown in vitro had no effect on cell growth in drug tolerant or resistant cells compared to parental cells. However, secreted prostaglandins are known immunomodulators, thus we explored how COX1/2 inhibition in combination with BRAF inhibitor plus MEK inhibitor affected immune cell infiltration. COX1/2 inhibition increased CD4+ and CD8+ T cell infiltration as compared to BRAF inhibitor plus MEK inhibitor alone. Under constant BRAF inhibitor plus MEK inhibitor treatment, we found that COX1/2 inhibition upon reaching MRD status prevented tumor relapse in vivo and increased survival dependent on an immune system. Future studies will evaluate the importance of arachidonic acid metabolism in promoting cross resistance to immune checkpoint blockade and whether T cell depletion prevents the delayed tumor relapse. Overall, COX1/2 inhibition delayed tumor relapse and survival in an immune-dependent manner and demonstrated a potential cross resistant mechanism. Citation Format: Casey D. Stefanski, Daniel A. Erkes, Timothy J. Purwin, Glenn L. Mersky, Erica L. Kitterman, Manal Mustafa, Jelan Haj, Diana Melissaratos, Andrew E. Aplin. Arachidonic acid metabolism promoting cross resistance in melanoma cells [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 A023.

Abstract A031: ECM architecture drives distinct evolutionary patterns of tumor immune escape and elimination in breast cancer

Abstract Cancer progression poses a clinical hurdle due to tumor heterogeneity that contributes to treatment resistance and immune evasion. While the adaptive immune system can effectively eliminate cancer in certain instances, tumor escape remains challenging. The immune microenvironment plays an important role in this process. One unanswered question is the role of ECM geometry, also known as Tumor Associated Collagen Signatures (TACS), and its effects on tumor immune recognition. In solid malignancies, ECM geometry has been associated with disease stage and T cell infiltration. Specifically, empirically observed ECM topologies are frequently categorized based on fiber arrangement: random fibers (TACS1), circumferentially aligned fibers (TACS2), and radially arranged fibers (TACS3). Even though a clear negative correlation between TACS and patient survival has been established, the specific roles and extent of TACS in T cell-driven cancer evolution remain uncharacterized. The precise mechanisms underlying how TACS influences cell movement are still not fully elucidated, with divergent opinions on whether and how the ECM mediates immune cell infiltration. At present, we currently lack a physical model relating the impact of TACS on the spatial co-evolution between an adaptive immune repertoire and a heterogeneous population of evading cancer cells. Here, we developed the EVO-ACT (EVOlutionary Agent-based Cancer T cell interaction) model, which is based on the Gillespie algorithm, to study the effects of TACS on tumor evolution and dynamical tumor-T cell interactions. Our analysis of tumor-T cell interactions across three TACS types reveals distinct migration patterns, with TACS3 showing the highest efficiency, TACS1 intermediate, and TACS2 the lowest. TACS impacts T cells more than tumors, with stronger chemokine gradients aiding T cell infiltration. Our model suggests that despite TACS2's limited efficiency, it cannot fully impede T cell infiltration. Varied migration efficiencies lead to diverse T cell infiltration patterns and immunoediting levels. TACS3 shows a benefit to immune recognition and higher survival, contrary to lower survival in TACS3 patients observed clinically in breast cancer. Our results indicate that only after introducing phenotypic changes, such as Epithelial-Mesenchymal Transition (EMT), and noting that EMT occurs exclusively after the TACS2-TACS3 progression, can our findings successfully explain the clinically observed trends. Considering mesenchymal tumor cells' ability to upregulate PD-1/PD-L1, we explore TACS-specific tumor evolution's impact on PD-1/PD-L1 inhibitor responses and patient survival. We observe higher tumor escape rates and lower survival in TACS3 compared to TACS2. Thus, TACS3 alone does not account for lower survival in breast cancer; TACS3-associated late-stage cancer byproducts like EMT and elevated checkpoint expression significantly decrease survival. Citation Format: Yijia Fan, Jason Tomas George. ECM architecture drives distinct evolutionary patterns of tumor immune escape and elimination in 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 A031.

Abstract C004: Glucocorticoids establish a metastatic-promoting tumor microenvironment in a PDAC mouse model

Abstract Disseminated cancer cells (DCCs) arriving to a new organ may either proliferate and form metastasis, get eliminated by immune cells, or enter a quiescent stage. The factors that determine the DCCs’ fate, including getting quiescent DCCs to re-initiate proliferation and successfully evade the immune system, remain poorly understood, in part due to the lack of robust mouse models. We established a pancreatic ductal adenocarcinoma (PDAC) mouse model of dormant DCCs in the liver. An “immunization protocol” was used to mimic PDAC resection in patients: KPC1199 PDAC cells were injected to establish primary tumors and allow an adaptive immune response to develop against the cancer cells; the tumors were surgically resected; and new KPC1199 cells were intrasplenically injected to establish experimental liver metastases. The overwhelming majority of the KPC1199 cells were eliminated. No metastases spontaneously formed during 20 months of observation, but about 3-8 sporadic, single DCCs were found per liver cross section, and 94% of these DCCs were negative for proliferation markers. Although the model is based on entraining an adaptive immune response, T cell depletion resulted in only 25-30% of mice developing just 1-3 metastases each. This suggests that T cell depletion results in metastasis from the small percentage of DCCs that were proliferating. The DCC-hosting mice therefore represents of model to identify factors that can trigger further DCC proliferation and metastatic recurrence. Patients with PDAC often experience significant stress, leading to excess of endogenous glucocorticoids (GCs). Additionally, synthetic GCs are used during e.g., chemotherapy to manage side effects. To mimic elevated GC levels in patients, we treated DCC-hosting mice with synthetic GCs resulting in an increased percentage of proliferating DCCs (increasing from <6% of the examined cells in control mice to about 36%). Yet, no metastases formed. GC treatment also decreased T cell and increased neutrophil liver infiltration. Neutrophils can form protease-containing neutrophil extracellular traps (NETs), which we previously showed trigger DCC proliferation by proteolytic remodeling of the extracellular matrix protein laminin. We found that neutrophils from GC-treated mice spontaneously formed more NETs, and additionally, NET-generated laminin remodeling was observed in the liver metastasis. Together, the data suggest, that GC treatment drive quiescent DCCs to proliferate via neutrophils but alone this is insufficient to cause metastases. Consistent with this interpretation, T cell depletion combined with GC treatment resulted in multiple metastases in all mice examined. Of note, deleting the GC receptor in the DCCs did not reduce metastases, supporting that GCs act on the host and not the cancer cells. In conclusion, our data suggest that for quiescent DCCs to form metastases, they need signals that trigger proliferation and simultaneously, they must overcome immune surveillance. Elevated GC levels or GC treatment in immune suppressed individuals represents such a scenario. Citation Format: Xiao Han, Xueyan He, Yuan Gao, Sicheng Pan, Mikala Egeblad. Glucocorticoids establish a metastatic-promoting tumor microenvironment in a PDAC mouse model [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 C004.

Abstract C047: AGX101: Preclinical evaluation of a TM4SF1-directed tubulin inhibitor conjugate with ongoing first-in-human trial

Abstract Introduction: TM4SF1 (Transmembrane-4 L-Six-Family-Member-1) is an endothelial marker with critical roles in angiogenesis, as well as a tumor cell antigen that contributes significantly to invasion and metastasis. TM4SF1 is upregulated 20-fold in angiogenic tumor vascular endothelium compared to normal vasculature endothelium and exhibits a unique nuclear internalization pathway. AGX101 is a novel tubulin inhibitor conjugate specifically directed against TM4SF1, delivering a potent maytansinoid payload directly to the nucleus of cells within the tumor microenvironment. Delivery to both the vascular and tumor cell compartments of the tumor results in three mechanisms of action (MoAs): (1) activation of tumor immune surveillance, (2) tumor blood supply deprivation, and (3) direct tumor cell killing. Methods: TM4SF1 expression was assessed across solid tumor samples using immunohistochemistry. Safety and pharmacokinetics of AGX101 were evaluated in non-human primates, with escalating doses to determine the highest non-severely toxic dose (HNSTD). Efficacy studies were also conducted in mouse models, enabling assessment of the minimum effective dose (MED) needed to engage each of the three MoAs. The combination of HNSTD and MED enables calculation of a therapeutic index (TI). Preclinical efficacy studies used either AGX101 (in human tumor xenograft models, to study the tumor cell killing MoA) or AGXB01, a rodent surrogate of AGX101, to study vascular and immune MoAs and all three MoAs in syngeneic mouse cancer models. The potential for synergy with immune checkpoint inhibitors (ICIs) was also investigated. A first-in-human study of AGX101 in patients with advanced solid tumors with the primary objective of safety and dose-limiting toxicities (DLTs) has initiated. Results: TMFS41 is broadly expressed across solid tumors with increasing scoring intensity in higher grade tumors. In non-human primates, AGX101 exhibited a favorable safety profile, being tolerated at relatively high doses and with dose dependent, monitorable, and reversible effects. In preclinical efficacy studies, AGXB01 and AGX101 as monotherapies demonstrated robust efficacy through each of the three MoAs in multiple syngeneic, xenograft, and orthotopic tumor models in mice and rats as well as non-tumor models of angiogenesis. Measured by exposure, TI was large. Additionally, AGXB01 was shown to potentiate the effects of ICIs in syngeneic mouse models (CT26, Renca, and B16-F10), suggesting a potential synergistic approach in cancer therapy. The first dose level of AGX101 monotherapy in humans has cleared without DLTs. Conclusion: AGX101, targeting the TM4SF1 antigen, represents a promising new approach in cancer therapy. The preclinical data suggest that AGX101 could provide a significant therapeutic benefit by novel and differentiated mechanisms of action, namely selectively targeting the tumor vasculature and potentiating immune-based therapies. Further clinical development of AGX101 is ongoing. Citation Format: Ildefonso Ismael Rodriguez Rivera, Meredith S. Pelster, Shou-Ching Jaminet, Paul Jaminet, Glen J. Weiss. AGX101: Preclinical evaluation of a TM4SF1-directed tubulin inhibitor conjugate with ongoing first-in-human trial [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 C047.

Abstract A032: Uncovering the impact of current therapies on LUAD dormant metastasis

Abstract Despite recent therapeutic advances, Lung Adenocarcinoma (LUAD) remains a leading cause of mortality worldwide, with most deaths attributed to metastasis. Metastasis is mediated by disseminated metastasis initiating cells (MICs) that can persist in a dormant state in distant sites for months to years and ultimately drive metastatic relapse, which is clinically incurable. Adjuvant therapy seeks to prevent cancer relapse, however, efforts to improve adjuvant treatments are hindered by an insufficient understanding of the molecular mechanisms supporting the long-term viability of dormant MICs. We have established that primitive LUAD cells characterized by the spontaneous expression of the pluripotency transcription factor SOX2 can enter an immune-evasive dormant state with downregulation of immune response mediators (STING, NK ligands, MHC-I) to evade immune surveillance during dormancy. Now, we hypothesize that dormant and immune evasive progenitor MICs may also escape therapies targeting growth pathways. We have generated latency competent cells (LCCs) that can remain in a latent state for months by isolating progenitor cells from the Kras LSL-G12D/+ ;Trp53 flox/flox mouse model and screening their metastatic potential and progenitor state in the B6 background. LCCs entering a slow-cycling state in response to dormancy signals remain sensitive to targeted KRAS inhibitors but contain a resistant subpopulation that survives. To specifically characterize the progenitor state in these LCC populations, we have developed and validated two strategies to fluorescently mark SOX2 expression and activity, respectively. As speculated, the resistant population is mostly composed of SOX2+ progenitor LCCs after treatment with KRAS inhibitor under dormancy signaling. Furthermore, induced-dormancy also drives an accumulation of cells in the SOX2+ progenitor state in these LCC-rich populations. Ongoing studies are focused on delineating the emergence of the SOX2+ progenitor state in LCCs during dormancy and the biological mechanisms driving their persistence and resistance to targeted therapies. We aim to define (1) the dependence of dormant LCCs on oncogenic signaling, the (2) effect of targeted therapy on MICs, and (3) strategies to overcome potential resistance mechanisms in dormant MICs. Our ultimate goal is to delineate strategies to clear latent MICs and prevent LUAD recurrence. Citation Format: Inês Godet, Zhenghan Wang, Joan Massagué. Uncovering the impact of current therapies on LUAD dormant metastasis [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 A032.

Abstract B030: Productive T-cell-mediated immunoediting of developing sarcomas by distinct mechanisms

Abstract Immunoediting is the process where developing tumors are shaped by cells of the immune system. Tumors with low immunogenicity are likely to emerge after neoantigen-specific T cells eliminate the most immunogenic cell populations. However, direct interrogation of mechanisms by which this occurs has not possible due to the pre-emergent nature before the disease is verifiably established. By modifying a mouse model of soft-tissue sarcoma where emergent tumors are consistently edited, we developed a system to examine mechanisms underlying how T cells interact with pre-emergent tumor cell populations to suppress tumor emergence. This system involves production of neoantigen-expressing autochthonous tumors with a fluorescent reporter to define neoantigen-expression status of individual cells. Tumors emerge with lower penetrance and delayed onset in T cell-sufficient mice, while every tumor from T cell-deficient mice contain a significant fraction of neoantigen-negative tumor cells. Yet, ∼53% of T cell-sufficient mice never developed, which suggests that neoantigen silencing was necessary, but not sufficient for tumor emergence. Consistent with this, genetic removal of the neoantigen after malignant transformation shows that tumor development is only rescued when neoantigen loss occurs prior to the peak T cell response, occurring around 8 days post transformation. Moreover, clonality studies reveal the presence of T cells reduces the number clones that contribute to tumor emergence down to 1-2 clones per emerged tumor. Mechanistically, blockade of the T cell effector cytokine, IFNgamma, rescues the neoantigen-negative, but not neoantigen-expressing tumor cells, demonstrating that T cells control early tumors via at least two distinct mechanisms: IFNgamma-dependent suppression of neoantigen-negative tumors cells and IFNgamma-independent suppression of neoantigen-expressing cells. Thus, our findings support that neoantigen-negative cells are subjected to cytokine-mediated suppression from T cells responding to the neoantigen-positive population in the pre-emergent tumor microenvironment. This bystander suppression can prevent tumor emergence despite non-homogeneity in neoantigen expression. Citation Format: Brian G Hunt, Julie F. Cheung, Srividhya Venkatesan, Jennifer L. Loza, Kelli A. Connolly, Shudipto Wahed, Elaine Cheng, Ishan Bansal, Emily Borr, Wei Wei, Ivana William, Brittany Fitzgerald, Nikhil Joshi. Productive T-cell-mediated immunoediting of developing sarcomas by distinct mechanisms [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 B030.