Unique Tumor Microenvironment Research Articles

Immunotherapy in Esophagogastric Cancer: Treatment Landscape, Challenges, and New Directions.

Cancers of the upper gastrointestinal tract represent a lethal disease entity comprising the esophagus, gastroesophageal junction, and stomach. The backbone of therapy in esophagogastric cancers has predominantly been chemotherapy-based. However, over the last decade, with the debut of immune checkpoint inhibitors, sophisticated molecular testing, and a more comprehensive understanding of the tumor microenvironment, immunotherapy has been incorporated into the treatment of localized and advanced esophagogastric cancers with promising results. This study aimed to review the unique tumor microenvironment and role of immunotherapy in esophagogastric cancers. We conducted a systematic review of clinical and translational research for immunotherapy in esophagogastric cancers. This article will explore the unique tumor microenvironment in gastroesophageal cancers, the role of immunotherapy in localized and advanced disease, challenges in management, and new therapeutic approaches in clinical trials. With further exploration into targeted therapy and immunotherapy, we anticipate the emergence of novel treatments that will improve survival and quality of life in patients with esophagogastric cancers.

Host-microbiota interactions contributing to the heterogeneous tumor microenvironment in colorectal cancer.

Colorectal cancer (CRC) exhibits pronounced heterogeneity and is categorized into four widely accepted consensus molecular subtypes (CMSs) with unique tumor microenvironments (TMEs). However, the intricate landscape of the microbiota and host-microbiota interactions within these TMEs remains elusive. Using RNA-sequencing data from The Cancer Genome Atlas, we analyzed the host transcriptomes and intratumoral microbiome profiles of CRC samples. Distinct host genes and microbial genera were identified among the CMSs. Immune microenvironments were evaluated using CIBERSORTx and ESTIMATE, and microbial coabundance patterns were assessed with FastSpar. Through LASSO penalized regression, we explored host-microbiota associations for each CMS. Our analysis revealed distinct host gene signatures within the CMSs, which encompassed ferroptosis-related genes and specific immune microenvironments. Moreover, we identified 293, 153, 66, and 109 intratumoral microbial genera with differential abundance, and host-microbiota associations contributed to distinct TMEs, characterized by 829, 1,270, 634, and 1,882 robust gene-microbe associations for each CMS in CMS1-CMS4, respectively. CMS1 featured inflammation-related HSF1 activation and gene interactions within the endothelin pathway and Flammeovirga. Integrin-related genes displayed positive correlations with Sutterella in CMS2, whereas CMS3 spotlighted microbial associations with biosynthetic and metabolic pathways. In CMS4, genes involved in collagen biosynthesis showed positive associations with Sutterella, contributing to disruptions in homeostasis. Notably, immune-rich subtypes exhibited pronounced ferroptosis dysregulation, potentially linked to tissue microbial colonization. This comprehensive investigation delineates the diverse landscapes of the TME within each CMS, incorporating host genes, intratumoral microbiota, and their complex interactions. These findings shed light on previously uncharted mechanisms underpinning CRC heterogeneity and suggest potential therapeutic targets.NEW & NOTEWORTHY This study determined the following: 1) providing a comprehensive landscape of consensus molecular subtype (CMS)-specific tumor microenvironments (TMEs); 2) constructing CMS-specific networks, including host genes, intratumoral microbiota, and enriched pathways, analyzing their associations to uncover unique patterns that demonstrate the intricate interplay within the TME; and 3) revealing a connection between immune-rich subtypes and ferroptosis activation, suggesting a potential regulatory role of the microbiota in ferroptosis dysregulation of the colorectal cancer TME.

Exosomal Cargo in Ovarian Cancer Dissemination.

Ovarian cancer (OC) has the highest mortality rate among all gynecologic cancers and is characterized by early peritoneal spread. The growth and development of OC are associated with the formation of ascitic fluid, creating a unique tumor microenvironment. Understanding the mechanisms of tumor progression is crucial in identifying new diagnostic biomarkers and developing novel therapeutic strategies. Exosomes, lipid bilayer vesicles measuring 30-150 nm in size, are known to establish a crucial link between malignant cells and their microenvironment. Additionally, the confirmed involvement of exosomes in carcinogenesis enables them to mediate the invasion, migration, metastasis, and angiogenesis of tumor cells. Functionally active non-coding RNAs (such as microRNAs, long non-coding RNAs, circRNAs), proteins, and lipid rafts transported within exosomes can activate numerous signaling pathways and modify gene expression. This review aims to expand our understanding of the role of exosomes and their contents in OC carcinogenesis processes such as epithelial-mesenchymal transition (EMT), angiogenesis, vasculogenic mimicry, tumor cell proliferation, and peritoneal spread. It also discusses the potential for utilizing exosomal cargo to develop novel "liquid biopsy" biomarkers for early OC diagnosis.

Dual-Targeting Biomimetic Semiconducting Polymer Nanocomposites for Amplified Theranostics of Bone Metastasis.

Bone metastasis is a type of metastatic tumors that involves the spreads of malignant tumor cells into skeleton, and its diagnosis and treatment remain a big challenge due to the unique tumor microenvironment. We herein develop osteoclast and tumor cell dual-targeting biomimetic semiconducting polymer nanocomposites (SPFeNOC ) for amplified theranostics of bone metastasis. SPFeNOC contain semiconducting polymer and iron oxide (Fe3 O4 ) nanoparticles inside core and surface camouflaged hybrid membrane of cancer cells and osteoclasts. The hybrid membrane camouflage enables their targeting to both metastatic tumor cells and osteoclasts in bone metastasis through homologous targeting mechanism, thus achieving an enhanced nanoparticle accumulation in tumors. The semiconducting polymer mediates near-infrared (NIR) fluorescence imaging and sonodynamic therapy (SDT), and Fe3 O4 nanoparticles are used for magnetic resonance (MR) imaging and chemodynamic therapy (CDT). Because both cancer cells and osteoclasts are killed synchronously via the combinational action of SDT and CDT, the vicious cycle in bone metastasis is broken to realize high antitumor efficacy. Therefore, 4T1 breast cancer-based bone metastasis can be effectively detected and cured by using SPFeNOC as dual-targeting theranostic nanoagents. This study provides an unusual biomimetic nanoplatform that simultaneously targets osteoclasts and cancer cells for amplified theranostics of bone metastasis.

Dual‐Targeting Biomimetic Semiconducting Polymer Nanocomposites for Amplified Theranostics of Bone Metastasis

AbstractBone metastasis is a type of metastatic tumors that involves the spreads of malignant tumor cells into skeleton, and its diagnosis and treatment remain a big challenge due to the unique tumor microenvironment. We herein develop osteoclast and tumor cell dual‐targeting biomimetic semiconducting polymer nanocomposites (SPFeNOC) for amplified theranostics of bone metastasis. SPFeNOC contain semiconducting polymer and iron oxide (Fe3O4) nanoparticles inside core and surface camouflaged hybrid membrane of cancer cells and osteoclasts. The hybrid membrane camouflage enables their targeting to both metastatic tumor cells and osteoclasts in bone metastasis through homologous targeting mechanism, thus achieving an enhanced nanoparticle accumulation in tumors. The semiconducting polymer mediates near‐infrared (NIR) fluorescence imaging and sonodynamic therapy (SDT), and Fe3O4 nanoparticles are used for magnetic resonance (MR) imaging and chemodynamic therapy (CDT). Because both cancer cells and osteoclasts are killed synchronously via the combinational action of SDT and CDT, the vicious cycle in bone metastasis is broken to realize high antitumor efficacy. Therefore, 4T1 breast cancer‐based bone metastasis can be effectively detected and cured by using SPFeNOC as dual‐targeting theranostic nanoagents. This study provides an unusual biomimetic nanoplatform that simultaneously targets osteoclasts and cancer cells for amplified theranostics of bone metastasis.

Engineering Clinically Relevant Probiotics with Switchable "Nano-Promoter" and "Nano-Effector" for Precision Tumor Therapy.

Probiotics have the potential as biotherapeutic agents for cancer management in preclinical models and human trials by secreting antineoplastic or immunoregulatory agents in the tumor microenvironment (TME). However, current probiotics lack the ability to dynamically respond to unique TME characteristics, leading to limited therapeutic accuracy and efficacy. Although progress has been made in customizing controllable probiotics through synthetic biology, the engineering process is complex and the predictability of production is relatively low. To address this, here, for the first time, this work adopts pH-dependent peroxidase-like (POD-like) artificial enzymes as both an inducible "nano-promoter" and "nano-effector" to engineer clinically relevant probiotics to achieve switchable control of probiotic therapy. The nanozyme initially serves as an inducible "nano-promoter," generating trace amounts of nonlethal reactive oxygen species (ROS) stress to upregulate acidic metabolites in probiotics. Once metabolites acidify the TME to a threshold, the nanozyme switches to a "nano-effector," producing a great deal of lethal ROS to fight cancer. This approach shows promise in subcutaneous, orthotopic, and colitis-associated colorectal cancer tumors, offering a new methodology for modulating probiotic metabolism in a pathological environment.

Multi-omic profiling reveals discrepant immunogenic properties and a unique tumor microenvironment among melanoma brain metastases

Melanoma brain metastases (MBM) are clinically challenging to treat and exhibit variable responses to immune checkpoint therapies. Prior research suggests that MBM exhibit poor tumor immune responses and are enriched in oxidative phosphorylation. Here, we report results from a multi-omic analysis of a large, real-world melanoma cohort. MBM exhibited lower interferon-gamma (IFNγ) scores and T cell-inflamed scores compared to primary cutaneous melanoma (PCM) or extracranial metastases (ECM), which was independent of tumor mutational burden. Among MBM, there were fewer computationally inferred immune cell infiltrates, which correlated with lower TNF and IL12B mRNA levels. Ingenuity pathway analysis (IPA) revealed suppression of inflammatory responses and dendritic cell maturation pathways. MBM also demonstrated a higher frequency of pathogenic PTEN mutations and angiogenic signaling. Oxidative phosphorylation (OXPHOS) was enriched in MBM and negatively correlated with NK cell and B cell-associated transcriptomic signatures. Modulating metabolic or angiogenic pathways in MBM may improve responses to immunotherapy in this difficult-to-treat patient subset.

Integrative Multi-Omics Analysis of Tumor-Immune Microenvironment in Multiple Myeloma: Novel Insights and Therapeutic Implications

Despite notable therapeutic advances that have improved the survival of multiple myeloma (MM) patients, development of drug resistance remains a major problem. Transcriptomic analysis provides an opportunity to dissect the complexity of tumors, including the surrounding microenvironment, which has a significant impact on MM tumor progression and patients' response to treatment, as demonstrated by the effectiveness of immunomodulatory therapies. To improve the tailoring of targeted and immune based therapeutic strategies, it is crucial to decipher the tumor-immune microenvironment profile in MM patients. We used a multi-omics data integration approach, including RNAseq-based gene expression for MM tumor cells (MMCs) (n=196) and for the corresponding purified tumor microenvironment (TME) (n=124), single nucleotide variant data from whole exome sequencing of MMCs (n=100), deconvolution of TME immune subtypes (n=124), and relevant clinical metadata. This allowed us to thoroughly characterize both the tumor and its TME. Applying Multiomics Factor Analysis (MOFA) identified 13 principal factors capturing the diversity of the disease, encompassing both MM malignant and microenvironment components. Factors 1 and 2 specifically represented the heterogeneity in the TME, incorporating gene expression profiles and deconvolution-based immune cell subtypes. On the other hand, Factors 6, 9, 11, and 12 were specific to variations in the gene expression profile of the malignant component. Importantly, the analysis of the variables linked to these main factors uncovers not only genes with established functions in myeloma but also novel candidates with potential importance. Based on the top-weight in the gene expression and deconvolution data, Factor 1 showed a positive association with MMP8 expression by TME cells and a negative correlation with the presence of CD8 positive T cells in the TME. Factor 2, on the other hand, exhibited positive alignments with the expression of BHLHA15, TNFRSF17 (BCMA), KLF15, and IGF1 by TME cells. Regarding Factor 6 and Factor 11, which capture tumor fraction heterogeneity, Factor 6 showed alignment with the expression of cancer testis antigens ( MAGEA6, MAGEA3), CXCL12, and KLF4. Meanwhile, Factor 11 was associated with the expression of genes such as SULF2, as well as CRP levels, percentage of CD138+ cells, and the percentage of cells in cell cycle phase S. Next, we investigated the predictive potential of the latent factors inferred by MOFA in models of clinical outcomes. Interestingly, 6 out of the 13 factors identified by MOFA showed significant associations with patients' overall survival (cox regression, p <= 0.05). These factors include Factor 1 and 2, which are related to the composition of the TME, and Factor 6 and 9, which are associated with myeloma progression status, osteolysis, t(11;14), t(4;14), and MM molecular classification. Finally, performing unsupervised clustering using the latent MOFA factors, we identified two main groups with distinct TME subtypes. These groups can be further divided into nine subgroups, each exhibiting unique TME characteristics. Notably, the TME subtypes showed correlations with essential gene expression signatures related to immune response regulation, T cell activation, and natural killer mediated immunity. Additionally, the significant presence of deconvolution-based immune cell types, including T cell CD8+, T cell regulatory, T cell CD4+ (non-regulatory), NK cells and Monocytes was observed. Moreover, the TME subtypes were associated with patients' overall survival and response to daratumumab. Patients presenting TME enriched in immune cells were associated with a significant better outcome ( p = 0.0026). These findings highlight the importance of TME analysis in predicting treatment response and MM patients' outcome in the context of immune-based therapies. Our integrative multi-omics analysis revealed comprehensive MM heterogeneity and distinct immune subtypes that could be of therapeutic interest for personalized therapeutic approaches.

The Crosstalk between the EGFR and IFN-γ Pathways and Synergistic Roles in Survival Prediction and Immune Escape in Gliomas.

Glioma is the most common primary malignant brain tumor. The poor prognosis of gliomas, especially glioblastoma (GBM), is associated with their unique molecular landscape and tumor microenvironment (TME) features. The epidermal growth factor receptor (EGFR) gene is one of the frequently altered loci in gliomas, leading to the activation of the EGFR signaling pathway and thus, promoting the genesis of gliomas. Whether there exist factors within the TME that can lead to EGFR activation in the context of gliomas is currently unexplored. In total, 702 samples from The Cancer Genome Atlas (TCGA) and 325 samples from The Chinese Glioma Genome Atlas (CGGA) were enrolled in this study. Gene signatures related to EGFR signaling and interferon-γ (IFN-γ) response were established via the LASSO-COX algorithm. Gene Set Enrichment Analysis (GSEA) and Gene Ontology (GO) analysis were applied for function exploration. Kaplan-Meier (KM) curves and single sample GSEA (ssGSEA) of immune cell subpopulations were performed to analyze the prognosis and TME characteristics of different subgroups. Moreover, Western blotting (WB) and flow cytometry (FCM) demonstrated the correlation between IFN-γ and EGFR signaling activation and the subsequent induction of programmed death ligand 1 (PD-L1) expression. An EGFR signaling-related risk score was established, and a higher score was correlated with poorer prognosis and a more malignant phenotype in gliomas. Biological function analysis revealed that a higher EGFR-related score was significantly associated with various cytokine response pathways, especially IFN-γ. Long-term (7 days) exposure to IFN-γ (400 ng/mL) induced the activation of EGFR signaling in the u87 cell line. Next, an IFN-γ response-related risk score was established; the combination of these two scores could be used to further reclassify gliomas into subtypes with different clinical features and TME features. Double high-risk samples tended to have a poorer prognosis and more immunosuppressive TME. Additionally, FCM discovered that the activation of EGFR signaling via EGF (100 ng/mL) could trigger PD-L1 protein expression. This research indicates that IFN-γ, an inflammatory cytokine, can activate the EGFR pathway. The combination of EGFR signaling and IFN-γ response pathway can establish a more precise classification of gliomas.

A 12-gene panel in estimating hormone-treatment responses of castration-resistant prostate cancer patients generated using a combined analysis of bulk and single-cell sequencing data

Background Castration-resistant prostate cancer (CRPC) represents one type of advanced prostate cancer (PCa) with a median survival time of 1–2 years. Currently, there is a lack of reliable gene panels in predicting hormone treatment (HT) responses due to limited knowledge of CRPC-specific tumor-microenvironment (TME) characteristics. Methods In this study, we first screened for up-regulated genes in CRPC samples using bulk-sequencing data retrieved from TCGA online database, and further investigated the expression status of these genes in four sets of downloaded single-cell RNA sequencing (scRNAseq) data: GSE117403 containing 16 normal human prostate samples; GSE141445 containing 13 PCa samples; GSE176031 containing 11 PCa samples and GSE137829 containing 6 CRPC samples. Results We identified a series of CRPC-specific TME characteristics including an enriched number of PEG10 + neuroendocrine cells, elevated expression of PPIB/CCDC74A/GAPDH/AR genes in tumor cells, increased expression of FAP/TGFB1 in cancer-associated fibroblasts (CAFs), suppressed immune environment featured by enhanced M2 macrophage polarization, T cell exhaustion and increased number of regulatory B cells. We further established a 12-gene panel using these characteristics and showed that this panel could separate CRPC samples from PCa samples (AUC of 0.78), and CRPC patients with higher panel scores tended to have treatment failure or progression (R = −0.47, p = 0.019). Conclusions Based on these unique TME characteristics of CRPC, we established a prediction tool for estimating the duration of HT responses in PCa treatment. Our results suggest mechanisms by which prostate cancer becomes castrate resistant. Further study of PEG10 (and/or others) to evaluate therapeutic efficacy should be considered.

Low-Temperature Photothermal Therapy Platform Based on Pd Nanozyme-Modified Hydrogenated TiO2.

In photothermal treatments (PTTs), normal tissues around cancerous tumors get injured by excessive heat, whereas damaged cancer cells are easily restored by stress-induced heat shock proteins (HSPs) at low temperatures. Therefore, to achieve a unique tumor microenvironment (TME), it is imperative to increase PTT efficiency and reduce normal tissue injury by adopting appropriate reactive oxygen species (ROS) and lipid peroxides (LPO) cross-linked with HSPs. In the present research, a potential strategy for mild photothermal treatments (mPTTs) was proposed by initiating localized catalytic chemical reactions in TME based on Pd nanozyme-modified hydrogenated TiO2 (H-TiO2@Pd). In vitro and in vivo evaluations demonstrated that H-TiO2@Pd had good peroxidase-like activities (POD), glutathione oxidase-like activities (GSHOx), and photodynamic properties and also satisfactory biocompatibility for 4T1 cells. Localized catalytic chemical reactions in H-TiO2@Pd significantly depleted GSH to downregulate the protein expression of GPX4 and promoted the accumulation of LPO and ROS, which consumed HSP70 or inhibited its function in 4T1 cells. Hence, the as-constructed low-temperature photothermal therapeutic platform based on Pd nanozyme-modified H-TiO2 can be a promising candidate to develop a safe and effective mPTT for cancer treatments.

P02.05.A DISSECTING THE DIFFUSE MIDLINE GLIOMA TUMOUR MICROENVIRONMENT COMMUNICATIONS USING MULTI-OMICS APPROACHES

Abstract BACKGROUND Diffuse midline glioma (DMG) represents a highly aggressive pediatric brain tumour with no chance of survival. Treatment options are urgently needed, and efforts have been put towards the development of T cell immunotherapy. However, this should consider the unique tumour microenvironment (TME) of DMG, both in terms of anatomical location and developmental stage. MATERIAL AND METHODS To gain knowledge on this unique tumour niche, we implemented an immuno-competent, syngeneic mouse model based on in-utero electroporation (IUE) of the key driver-mutations to induce tumour formation at embryonic stage. By combining snRNAseq, Spatial Transcriptomics and state-of-the-art imaging, we characterized the spatial and molecular landscape of DMG. RESULTS AND CONCLUSION We observed a high intratumoral heterogeneity with spatially restricted cell populations. Importantly, their molecular profiles correlate with patient data, illustrating the clinical relevance of the IUE model. Analysis of cell-cell communications in the TME revealed a novel signalling between DMG and immunosuppressive myeloid cells, that could impact T-cell response. Using an advanced patient-derived DMG organoid co-culture model with macrophages and engineered T-cells, we are currently investigating the role of this pathway on T-cells mediated killing of DMG organoids. Thus, this project fills a critical need to delineate and overcome the immune-suppressive environment in DMG and potentiate T cell targeting.

Electrolyte imbalance causes suppression of NK and T cell effector function in malignant ascites

BackgroundMalignant ascites commonly occurs in advanced or recurrent stages of epithelial ovarian cancer during peritoneal carcinomatosis and is correlated with poor prognosis. Due to its complex composition of cellular and acellular components malignant ascites creates a unique tumor microenvironment, which mediates immunosuppression and promotes progression of disease. However, the immunosuppressive mechanisms remain poorly understood.MethodsIn the present study, we explored the antitumor activity of healthy donor NK and T cells directed against ovarian cancer cells in presence of malignant ascites derived from patients with advanced or recurrent peritoneal carcinomatosis. A wide range of methods was used to study the effect of ascites on NK and T cells (FACS, ELISA, EliSpot, qPCR, Live-cell and confocal microscopy, Western blot and electrolyte flux assays). The ascites components were assessed using quantitative analysis (nephelometry, potentiometry and clinical chemistry) and separation methods (dialysis, ultracentrifugal filtration and lipid depletion).ResultsAscites rapidly inhibited NK cell degranulation, tumor lysis, cytokine secretion and calcium signaling. Similarly, target independent NK and T cell activation was impaired in ascites environment. We identified imbalanced electrolytes in ascites as crucial factors causing extensive immunosuppression of NK and T cells. Specifically, high sodium, low chloride and low potassium content significantly suppressed NK-mediated cytotoxicity. Electrolyte imbalance led to changes in transcription and protein expression of electrolyte channels and impaired NK and T cell activation. Selected inhibitors of sodium electrolyte channels restored intracellular calcium flux, conjugation, degranulation and transcript expression of signaling molecules. The levels of ascites-mediated immunosuppression and sodium/chloride/potassium imbalance correlated with poor patient outcome and selected molecular alterations were confirmed in immune cells from ovarian cancer patients.ConclusionOur data suggest a novel electrolyte-based mechanism of immunosuppression in malignant ascites of patients with peritoneal carcinomatosis. We show for the first time that the immunosuppression of NK cytotoxicity in coculture assays is correlated to patient poor survival. Therapeutic application of sodium channel inhibitors may provide new means for restoring immune cell activity in ascites or similar electrolyte imbalanced environments.

Primary and hTERT-Transduced Mesothelioma-Associated Fibroblasts but Not Primary or hTERT-Transduced Mesothelial Cells Stimulate Growth of Human Mesothelioma Cells.

Pleural mesothelioma (PM) is an aggressive malignancy that develops in a unique tumor microenvironment (TME). However, cell models for studying the TME in PM are still limited. Here, we have generated and characterized novel human telomerase reverse transcriptase (hTERT)-transduced mesothelial cell and mesothelioma-associated fibroblast (Meso-CAF) models and investigated their impact on PM cell growth. Pleural mesothelial cells and Meso-CAFs were isolated from tissue of pneumothorax and PM patients, respectively. Stable expression of hTERT was induced by retroviral transduction. Primary and hTERT-transduced cells were compared with respect to doubling times, hTERT expression and activity levels, telomere lengths, proteomes, and the impact of conditioned media (CM) on PM cell growth. All transduced derivatives exhibited elevated hTERT expression and activity, and increased mean telomere lengths. Cell morphology remained unchanged, and the proteomes were similar to the corresponding primary cells. Of note, the CM of primary and hTERT-transduced Meso-CAFs stimulated PM cell growth to the same extent, while CM derived from mesothelial cells had no stimulating effect, irrespective of hTERT expression. In conclusion, all new hTERT-transduced cell models closely resemble their primary counterparts and, hence, represent valuable tools to investigate cellular interactions within the TME of PM.

Tumor microenvironment responsive metal nanoparticles in cancer immunotherapy.

Malignant tumors have a unique tumor microenvironment (TME), which includes mild acidity, hypoxia, overexpressed reactive oxygen species (ROS), and high glutathione (GSH) levels, among others. Recently, TME regulation approaches have attracted widespread attention in cancer immunotherapy. Nanoparticles as drug delivery systems have ability to modulate the hydrophilicity of drugs to affect drug uptake and efflux in tumor. Especially, the metal nanoparticles have been extensive applied for tumor immunotherapy due to their unique physical properties and elaborate design. However, the potential deficiencies of metal nanoparticles due to their low biodegradability, toxicity and treatment side effects restrict their clinical application. In this review, we briefly introduce the feature characteristics of the TME and the recent advances in tumor microenvironment responsive metal nanoparticles for tumor immunotherapy. In addition, nanoparticles could be combined with other treatments, such as chemotherapy, radiotherapy and photodynamic therapy also is presented. Finally, the challenges and outlook for improving the antitumor immunotherapy efficiency, side effect and potential risks of metal nanoparticles has been discussed.

Tumor Microenvironment Can Predict Chemotherapy Response of Patients with Triple-Negative Breast Cancer Receiving Neoadjuvant Chemotherapy.

Triple-negative breast cancer (TNBC) is a breast cancer subtype that has poor prognosis and exhibits a unique tumor microenvironment. Analysis of the tumor microbiome has indicated a relationship between the tumor microenvironment and treatment response. Therefore, we attempted to reveal the role of the tumor microbiome in patients with TNBC receiving neoadjuvant chemotherapy. We collected TNBC patient RNA-sequencing samples from the Gene Expression Omnibus and extracted microbiome count data. Differential and relative abundance were estimated with linear discriminant analysis effect size. We calculated the immune cell fraction with CIBERSORTx and conducted survival analysis using the Cancer Genome Atlas patient data. Correlations between the microbiome and immune cell compositions were analyzed and a prediction model was constructed to estimate drug response. Among the pathological complete response group (pCR), the beta diversity varied considerably; consequently, 20 genera and 24 species were observed to express a significant differential and relative abundance. Pandoraea pulmonicola and Brucella melitensis were found to be important features in determining drug response. In correlation analysis, Geosporobacter ferrireducens, Streptococcus sanguinis, and resting natural killer cells were the most correlated factors in the pCR, whereas Nitrosospira briensis, Plantactinospora sp. BC1, and regulatory T cells were key features in the residual disease group. Our study demonstrated that the microbiome analysis of tumor tissue can predict chemotherapy response of patients with TNBC. Further, the immunological tumor microenvironment may be impacted by the tumor microbiome, thereby affecting the corresponding survival and treatment response.

Decoding Metabolic Symbiosis between Pancreatic Cancer Cells and Cancer-Associated Fibroblasts Using Cultured Tumor Microenvironment

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer with a poor prognosis, largely due to its unique tumor microenvironment (TME) and dense fibrotic stroma. Cancer-associated fibroblasts (CAFs) play a crucial role in promoting tumor growth and metastasis, contributing to the metabolic adaptation of PDAC cells. However, the metabolic interactions between PDAC cells and CAFs are not well-understood. In this study, an in vitro co-culture model was used to investigate these metabolic interactions. Metabolomic analysis was performed under monoculture conditions of Capan−1 PDAC cells and CAF precursor cells, as well as co-culture conditions of PDAC cells and differentiated inflammatory CAF (iCAF). Co-cultured Capan−1 cells displayed significant metabolic changes, such as increased 2-oxoglutaric acid and lauric acid and decreased amino acids. The metabolic profiles of co-cultured Capan−1 and CAFs revealed differences in intracellular metabolites. Analysis of extracellular metabolites in the culture supernatant showed distinct differences between Capan−1 and CAF precursors, with the co-culture supernatant exhibiting the most significant changes. A comparison of the culture supernatants of Capan−1 and CAF precursors revealed different metabolic processes while co-culturing the two cell types demonstrated potential metabolic interactions. In conclusion, this study emphasizes the importance of metabolic interactions between cancer cells and CAFs in tumor progression and highlights the role of TME in metabolic reprogramming.

Identification of Tissue-Resident Natural Killer and T Lymphocytes with Anti-Tumor Properties in Ascites of Ovarian Cancer Patients.

Women with ovarian cancer have limited therapy options, with immunotherapy being unsatisfactory for a large group of patients. Tumor cells spread from the ovary or the fallopian tube into the abdominal cavity, which is commonly accompanied with massive ascites production. The ascites represents a unique peritoneal liquid tumor microenvironment with the presence of both tumor and immune cells, including cytotoxic lymphocytes. We characterized lymphocytes in ascites from patients with high-grade serous ovarian cancer. Our data reveal the presence of NK and CD8+ T lymphocytes expressing CD103 and CD49a, which are markers of tissue residency. Moreover, these cells express high levels of the inhibitory NKG2A receptor, with the highest expression level detected on tissue-resident NK cells. Lymphocytes with these features were also present at the primary tumor site. Functional assays showed that tissue-resident NK cells in ascites are highly responsive towards ovarian tumor cells. Similar results were observed in an in vivo mouse model, in which tissue-resident NK and CD8+ T cells were detected in the peritoneal fluid upon tumor growth. Together, our data reveal the presence of highly functional lymphocyte populations that may be targeted to improve immunotherapy for patients with ovarian cancer.

DIPG-11. DISSECTING THE DIFFUSE MIDLINE GLIOMA TUMOR MICROENVIRONMENT COMMUNICATIONS USING MULTI-OMICS APPROACHES

Abstract Diffuse midline glioma (DMG) represents a highly aggressive pediatric brain tumor with no chance of survival. Treatment options are urgently needed, and efforts have been put towards developing T-cell immunotherapy. However, this should consider DMG’s unique tumor microenvironment (TME), both in terms of anatomical location and developmental stage. To gain knowledge on this unique tumor niche, we implemented an immuno-competent, syngeneic mouse model based on in-utero electroporation (IUE) of the key driver mutations to induce tumor formation at an embryonic stage. We characterized DMG’s spatial and molecular landscape by combining snRNAseq, spatial transcriptomics, and state-of-the-art imaging. We observed a high intratumoral heterogeneity with spatially restricted cell populations. Importantly, their molecular profiles correlate with patient data, illustrating the clinical relevance of the IUE model. Analysis of cell-cell communications in the TME revealed novel signaling between DMG and immunosuppressive myeloid cells that could impact T-cell response. Using an advanced patient-derived DMG organoid co-culture model with macrophages and engineered T-cells, we are currently investigating the role of this pathway on T-cells mediated killing of DMG organoids. Thus, this project fills a critical need to delineate and overcome the immune-suppressive environment in DMG and potentiate T cell targeting.

Abstract B039: Influence of tumor-associated fibroblasts and their exosomes in the development and progression of prostate cancer

Abstract The unique tumor microenvironment (TME) of the prostate and in particular fibroblasts are known to play an essential role in the growth and progression of prostate cancer (PCa). Earlier studies from our group have shown that co-injection of patient-derived prostate cancer-associated fibroblasts (CAFs) and non-cancer associated fibroblasts (NCAFs) with tumor cells stimulated tumor growth in xenograft models compared to the injection of tumor cells alone. Our present study aims to investigate the effect of patient-derived prostate CAFs and NCAFs on the functional aspects of tumor cells in-vitro and their role in epigenetic regulation. Three prostate CAFs and NCAFs pairs were isolated from tissues of patients who had undergone radical prostatectomy and cultivated in-vitro. We then examined the influence of these fibroblast pairs on the viability, proliferation and migration of prostate cancer cell lines LNCaP and LNCaP C4-2. Cells were co-cultured in a transwell system with tumor cells in the lower chamber and fibroblasts in the upper chamber for 2 and 4 days and the viability of the tumor cells was measured using WST-1 assay and proliferation using BrdU colorimetric assay. For migration assay, the tumor cells were pretreated with serum-deficient medium and were co-cultured in a transwell system as mentioned above for 4 days. The tumor cells were collected after co-culture and the migration was measured using the transwell migration system as well as Xcelligence real-time assays in parallel. Further, extracellular vesicles (EV) were enriched from fibroblasts by ultracentrifugation. The expression of miRNAs isolated from cells and EVs was studied by microarray analysis and RT-qPCR. Patient-derived fibroblasts significantly increased the viability and proliferation of the tumor cells compared to the tumor cells alone. The influence of NCAFs on both viability and proliferation was more pronounced compared to CAFs on the LNCaP cells. However, the influence of the fibroblasts was vice versa on the LNCaP-C4-2 cells. The LNCaP cells exhibited no migration whereas the LNCaP C4-2 cells exhibited slight migration. The effect of different fibroblast pairs on the migration of LNCaP C4-2 cells varied among the pairs. 103 miRNAs and 6 miRNAs were found to be differentially expressed between CAFs and NCAFs and between their EVs, respectively. miR-10b-5p and miR-210-3p were confirmed by RT-qPCR to be upregulated in the CAFs. Our in-vitro data confirmed the in-vivo results. Fibroblasts modulate viability, proliferation and migration of PCa. However, the effects are heterogeneous due to patient and cell line-specific characteristics. Further studies will focus on the possible effects of fibroblasts on apoptosis in correlation to viability and invasion. CAFs and their EVs are characterized by specific miRNA expression supporting an epigenetic regulation of fibroblast characteristics and functions. Therefore, we will further investigate the mechanism of tumor progression induced by fibroblasts and their EVs via miRNA regulation. Citation Format: Aishwarya Tagat, Hiresh Ayoubian, Angela Zaccagnino, Michael Stöckle, Johannes Linxweiler, Kerstin Junker. Influence of tumor-associated fibroblasts and their exosomes in the development and progression of prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B039.