Mediate Tumor Progression Research Articles

Multi-omics analysis identifies BCAT2 as a potential pan-cancer biomarker for tumor progression and immune microenvironment modulation.

Branched-chain amino acid transaminase 2 (BCAT2) encodes a crucial protein involved in the initial catalysis of branched-chain amino acid (BCAA) catabolism, with emerging evidence suggesting its association with tumor progression. This study explores BCAT2 in a pan-cancer multi-omics context and evaluates its prognostic significance. We utilized a multi-database approach, analyzing cBioPortal for genetic alterations, RNA-Seq data from TCGA and GTEx for expression patterns, and RSEM for transcript analysis. Protein expression and interaction networks were assessed using the Human Protein Atlas, UniProt, and STRING. Prognostic value was determined through Cox regression analysis of TCGA clinical survival data, while immune cell infiltration across various cancers was examined using TCGA data and the TIMER2 platform. Our results revealed that BCAT2 alterations are primarily amplifications and is upregulated in various tumors, correlating with poor survival rates in several tumor types, including GBMLGG, LGG, and UVM. Elevated BCAT2 protein levels were common in pan-cancer, interacting with a range of metabolic enzymes. Additionally, BCAT2 expression significantly influenced CD4+ T cells, CD8+ T cells, and Treg cells infiltration, with varied correlations across cancer types. These findings indicate BCAT2 as a potential biomarker for cancer diagnosis and therapy, potentially regulating key metabolic and immune factors to mediate tumor progression and the microenvironment.

Syntaxin6 contributes to hepatocellular carcinoma tumorigenesis via enhancing STAT3 phosphorylation

BackgroundSyntaxin6 (STX6) is a SNARE (Soluble N-ethylmaleimide-sensitive factor attachment protein receptors) protein complex located in the trans-Golgi network and endosomes, which is closely associated with a variety of intracellular membrane transport events. STX6 has been shown to be overexpressed in a variety of human malignant tumors such as esophageal, colorectal, and renal cell carcinomas, and participates in tumorigenesis and development.MethodsBased on clinical public database and clinical liver samples analysis, the expression of STX6 in hepatocellular carcinoma (HCC) tissues was investigated. The effects of STX6 on proliferation, migration and invasion of HCC cell in vitro and in vivo were evaluated through gain- and loss-of-function studies. We further performed RNA-seq analysis and protein interactome analysis, to further decifer the detailed mechanisms of STX6 in the regulation of the JAK-STAT pathway in HCC.ResultsSTX6 expression was upregulated in HCC tissues and its expression was highly correlated with the high histological grade of the tumor. STX6 promoted HCC cell proliferation, migration and invasion both in vitro and in vivo. Mechanistically, STX6 mediated tumor progression depending on promoting the activation of JAK-STAT signaling pathway. Receptor for activated protein kinase C (RACK1) as an essential adaptor protein mediating STX6 regulation of JAK-STAT pathway. Specifically, STX6 interacted with RACK1 and then recruited signal transducer and activator of transcription 3 (STAT3) to form a protein-binding complex and activates STAT3 transcriptional activity.ConclusionsThis study provided a novel concept that STX6 exerted oncogenic effects by activating the STAT3 signaling pathway, and STX6 might be a promising therapeutic target for HCC.

Features of HPV+ OPSCC predictive of recurrence identified by transcriptional profiling of a case-control cohort.

6055 Background: De-escalating therapy for HPV+ oropharyngeal cancers (OPSCCs) is hampered by poor ability to predict recurrence, and limited insight into the biologic traits predisposing to recurrence impedes personalizing therapy. We aimed to (1) define biologic features underlying therapy resistance by transcriptional profiling and (2) evaluate the features for prognostic utility. Methods: A single institution cohort of 851 HPV+ OPSCC patients undergoing transoral robotic surgery during 2007-2020 was used to identify 50 cases that recurred locoregionally in the adjuvant RT field (n=14) and/or at distant sites (n=43). As controls, we used 49 recurrence-free patients with long-term follow up and similar stage, smoking history, and adjuvant therapy. RNAseq of pretreatment tumors was used to compare individual gene expression and Hallmark/Kegg pathway activity between cases and controls using unpaired t-test (p<.05) and Gene Set Enrichment Analysis (p-adj<.05), respectively. Activity of the significant pathways was quantified in individual tumors using Gene Set Variation Analysis (GSVA), and a regression-based composite of GSVA scores was developed to distinguish cases from controls by ROC analysis. The composite score was used to test for stratification of recurrence free survival (RFS) in external cohorts using Youden Index and logrank test. Results: The21 Hallmark/Kegg pathways downregulated in cases indicated suppression of anti-tumor immunity, and the 20 upregulated ones revealed increased biosynthetic function and tumor cell proliferation. The 1472 upregulated mRNAs contained several components of the ATR-chk1 DNA repair pathway and trans-lesion synthesis polymerases, suggesting that mitigation of DNA replication stress enhanced tumor growth. The 958 downregulated mRNAs suggested reductions in the cytoplasmic dsDNA sensing and downstream NF-kB signals that lead to replication stress-driven anti-tumor immunity. Two GSVA scores were created to quantify the key tumor-intrinsic and immune suppressive features separately in each tumor. The scores correlated with each other (R=0.6, p<.001), and a regression-based combination of them distinguished cases from controls (AUC=.76, p<.001). This composite score also stratified RFS in three external cohorts: TCGA (n=52, p<0.001) and two single institution cohorts containing both surgical and nonsurgical cases (n=46, p=.002; n=81, p<0.001). Conclusions: HPV+ OPSCCs failing primary surgical therapy show evidence of reduced replication stress mediating tumor progression and low anti-tumor immunity. These features appear generalizable to heterogeneously treated external cohorts, where they predicted recurrence. Our results provide a new basis for creating transcriptomic predictors of treatment response and suggest targetable molecular mechanisms to overcome therapy resistance.

Dual roles of HK3 in regulating the network between tumor cells and tumor-associated macrophages in neuroblastoma

Neuroblastoma (NB) is the most common and deadliest extracranial solid tumor in children. Targeting tumor-associated macrophages (TAMs) is a strategy for attenuating tumor-promoting states. The crosstalk between cancer cells and TAMs plays a pivotal role in mediating tumor progression in NB. The overexpression of Hexokinase-3 (HK3), a pivotal enzyme in glucose metabolism, has been associated with poor prognosis in NB patients. Furthermore, it correlates with the infiltration of M2-like macrophages within NB tumors, indicating its significant involvement in tumor progression. Therefore, HK3 not only directly regulates the malignant biological behaviors of tumor cells, such as proliferation, migration, and invasion, but also recruits and polarizes M2-like macrophages through the PI3K/AKT-CXCL14 axis in neuroblastoma. The secretion of lactate and histone lactylation alterations within tumor cells accompanies this interaction. Additionally, elevated expression of HK3 in M2-TAMs was found at the same time. Modulating HK3 within M2-TAMs alters the biological behavior of tumor cells, as demonstrated by our in vitro studies. This study highlights the pivotal role of HK3 in the progression of NB malignancy and its intricate regulatory network with M2-TAMs. It establishes HK3 as a promising dual-functional biomarker and therapeutic target in combating neuroblastoma.

Benzo[a]pyrene promotes an epithelial‐to‐mesenchymal transition process in MCF10A cells and mammary tumor growth and brain metastasis in female mice

AbstractBreast cancer is the most frequent neoplasia in developed countries and the leading cause of death in women worldwide. Epithelial‐to‐mesenchymal transition (EMT) is a cellular process through which epithelial cells decrease or lose their epithelial characteristics and gain mesenchymal properties. EMT mediates tumor progression, because tumor cells acquire the capacity to execute the multiple steps of invasion and metastasis. Benzo[a]pyrene (B[a]P) is an environmental organic pollutant generated during the burning of fossil fuels, wood, and other organic materials. B[a]P exposition increases the incidence of breast cancer, and induces migration and/or invasion in MDA‐MB‐231 and MCF‐7 breast cancer cells. However, the role of B[a]P in the induction of an EMT process and metastasis of mammary carcinoma cells has not been studied in detail. In this study, we demonstrate that B[a]P induces an EMT process in MCF10A mammary non‐tumorigenic epithelial cells. In addition, B[a]P promotes the formation of larger tumors in Balb/cJ mice inoculated with 4T1 cells than in untreated mice and treated with dimethyl sulfoxide (DMSO). B[a]P also increases the number of mice with metastasis to brain and the total number of brain metastatic nodules in Balb/cJ mice inoculated with 4T1 cells compared with untreated mice and treated with DMSO. In conclusion, B[a]P induces an EMT process in MCF10A cells and the growth of mammary tumors and metastasis to brain in Balb/cJ mice inoculated with 4T1 cells.

Structure-Guided Discovery and Preclinical Assessment of Novel (Thiophen-3-yl)aminopyrimidine Derivatives as Potent ERK1/2 Inhibitors.

The RAS-RAF-MEK-ERK signaling cascade is abnormally activated in various tumors, playing a crucial role in mediating tumor progression. As the key component at the terminal stage of this cascade, ERK1/2 emerges as a potential antitumor target and offers a promising therapeutic strategy for tumors harboring BRAF or RAS mutations. Here, we identified 36c with a (thiophen-3-yl)aminopyrimidine scaffold as a potent ERK1/2 inhibitor through structure-guided optimization for hit 18. In preclinical studies, 36c showed powerful ERK1/2 inhibitory activities (ERK1/2 IC50 = 0.11/0.08 nM) and potent antitumor efficacy both in vitro and in vivo against triple-negative breast cancer and colorectal cancer models harboring BRAF and RAS mutations. 36c could directly inhibit ERK1/2, significantly block the phosphorylation expression of their downstream substrates p90RSK and c-Myc, and induce cell apoptosis and incomplete autophagy-related cell death. Taken together, this work provides a promising ERK1/2 lead compound for multiple tumor-treatment drug discovery.

Cancer-associated fibroblasts: a versatile mediator in tumor progression, metastasis, and targeted therapy.

Tumor microenvironment (TME) has been demonstrated to play a significant role in tumor initiation, progression, and metastasis. Cancer-associated fibroblasts (CAFs) are the major component of TME and exhibit heterogeneous properties in their communication with tumor cells. This heterogeneity of CAFs can be attributed to various origins, including quiescent fibroblasts, mesenchymal stem cells (MSCs), adipocytes, pericytes, endothelial cells, and mesothelial cells. Moreover, single-cell RNA sequencing has identified diverse phenotypes of CAFs, with myofibroblastic CAFs (myCAFs) and inflammatory CAFs (iCAFs) being the most acknowledged, alongside newly discovered subtypes like antigen-presenting CAFs (apCAFs). Due to these heterogeneities, CAFs exert multiple functions in tumorigenesis, cancer stemness, angiogenesis, immunosuppression, metabolism, and metastasis. As a result, targeted therapies aimed at the TME, particularly focusing on CAFs, are rapidly developing, fueling the promising future of advanced tumor-targeted therapy.

Abstract 4206: Regional transcriptomic profiling of the head and neck squamous cell carcinoma tumor microenvironment paired with murine models identify extracellular matrix components that mediate tumor progression and immune suppression

Abstract Purpose: A substantial contribution to the poor prognosis for head and neck squamous cell carcinoma (HNSCC) patients is the progression to distant metastasis. This occurs in 30% of patients over the course of their disease, and radically reduces the 5-year survival rate to under 50%. Investigations in recent years have begun to unveil the intricate interplay of the extracellular matrix (ECM) in both tumor-intrinsic signaling and immunoregulation. However, studies conducted in the context of metastatic have been limited. Here, we employ spatial transcriptomics on HNSCC patient biopsies to identify ECM components that may contribute to cancer progression and immune suppression. Experimental design: Comprehensive transcriptome profiling of HNSCC patient biopsies was conducted using the GeoMx Digital Spatial Profiler (DSP). This approach allowed us to characterize regions of the tumor microenvironment (TME) by immunofluorescent staining for subsequent bioinformatic analysis. To investigate the biological consequences of associations observed in our dataset, we performed in vivo murine studies. We have produced a series of SCC C57BL/6 murine cell lines derived from Smad4-/-KrasG12D Keratin15+ stem cells, which faithfully mimic the clinical progression of HNSCC tumors, including metastasis. Using these murine models, we applied relevant pharmacologic inhibitors and genetic knockdown of targets identified with our GeoMx DSP analysis. In vitro immune-based and functional assays were conducted to elucidate the underlying mechanisms contributing to SCC progression. Results: GeoMx DSP analysis revealed that laminins were enriched in stage IV HNSCC patient stroma, relative to lower stage patients. This coincided with TGFβ enrichment and inversely associated with signatures of anti-tumor immune infiltration. Analysis of The Cancer Genome Atlas data revealed similar inverse correlations in SCCs, distinct from breast cancer and lung adenocarcinoma. Our metastatic SCC murine lines likewise exhibited elevated levels of these laminin proteins, accompanied by high levels of laminin-binding integrins and TGFβ-Smad signaling. Independent of impacting cell proliferation, shRNA knockdown of laminin decreased tumor volume in an immune-competent setting. Targeting laminin-integrin signaling with the small molecule inhibitors Galunisertnib and Buparlisib also reduced the metastatic capacity of these cells in C57BL/6 mice. Conclusions: Spatial transcriptomic profiling of the HNSCC TME uncovers intricate associations and promising targets not readily discernible in bulk RNAseq analyses. Validation through genetic and pharmacological approaches in immune-competent models underscores their potential as strategies to reduce HNSCC burden. Citation Format: John D. Aleman, Khoa A. Nguyen, Erica Wong, Yao Ke, Hanne T. Lind, Clifford G. Tepper, Andrew C. Birkeland, Christian D. Young, Sana D. Karam, Xiao-Jing Wang. Regional transcriptomic profiling of the head and neck squamous cell carcinoma tumor microenvironment paired with murine models identify extracellular matrix components that mediate tumor progression and immune suppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4206.

Abstract 1224: 3D models of chemotherapy- and PARPi-resistant ovarian cancer indicate essential roles for JAK/STAT signaling in mediating drug resistance

Abstract Ovarian cancer is the deadliest gynecological cancer with a low 5-year overall survival rate that affects over 300,000 women globally each year. Given lack of routine screening procedures, ovarian cancer patients are frequently diagnosed with late-stage disease that rapidly progress despite initial responses to front line therapy. PARPi and VEGFi targeted therapies have improved progression free survival in small subsets of patients but have largely failed to improve overall survival. Since the robust uptake of PARPis in the front line setting, resistance to this class of drugs is a growing problem. The literature is filled with many novel treatment approaches, but the vast majority fail to gain traction in pre-clinical models and early phase trials. A contributing factor to these disappointments relates to the prevalent use of 2D monoculture model systems that do not recapitulate the tumor microenvironment. To address this issue, we utilized the Rastrum 3D cell printer to grow ovarian cancer cells and PDX models in bio-functional matrices which provide optimal stiffness and extracellular matrix components specific to the ovarian tumor microenvironment. We found that ovarian cancer cells grown under these conditions exhibit dramatic up-regulation of the JAK/STAT signaling pathway, including autocrine/paracrine signaling factors, which correlated with increased sensitivity to JAK/STAT inhibitors. This affect was lost when STAT1 and STAT3 knockout cell lines were grown in this 3D environment. Ovarian tumors are known to be rich in cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs). We incorporated primary CAFs into this 3D culture system and found they could promote ovarian cancer cell growth only in cancer cells with intact JAK/STAT signaling further indicating the essential role of this pathway in mediating tumor progression. Co-culture with M2 macrophages was also found to induce ovarian cancer cell growth, particularly in resistant models. Conversely, co-culture with M1 macrophages inhibited treatment naïve ovarian cancer cell growth but not resistant cell line growth. Single-cell RNAseq of the complex 3D cultures elucidated profound differences in the cross-talk between these cell types that is dependent on cancer cell sensitivity to standard-of-care agents. Finally, the addition of JAK/STAT inhibitors was found to substantially alter this cross-talk between cell types and inhibit cancer cell viability. These data further implicate the importance of JAK/STAT signaling, specifically in clinically relevant 3D models, in supporting ovarian cancer progression and response to chemotherapy and PARPi treatment. Ongoing studies are aimed at confirming these findings in pre-clinical mouse models and to elucidate the most effective JAK/STAT inhibitors in vivo with the goal of informing the design of future clinical trials. Citation Format: Esther Rodman, Xiyin Wang, Hannah Smith, Xiaonan Hou, Scott Kaufmann, John Weroha, John Hawse. 3D models of chemotherapy- and PARPi-resistant ovarian cancer indicate essential roles for JAK/STAT signaling in mediating drug resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1224.

CLEC4E upregulation in gastric cancer: A potential therapeutic target correlating with tumor-associated macrophages

BackgroundCLEC4E has been reported to promote lung cancer progression. Tumor-associated macrophages (TAMs) play an important role in tumorigenesis. Whether the expression of CLEC4E in TAMs is associated with gastric carcinogenesis remains unclear. MethodsThe TIMER, UALCAN, UCSC Xena, and KM plotter databases are used to examine the expression of CLEC4E and its prognostic significance in gastric cancer (GC). Additionally, GO, KEGG, and GSEA analysis were conducted, and single-cell RNA-seq (scRNA-seq) datasets were utilized. The Coremine medical database was used to predict therapeutic drugs, and molecular docking was performed. Human GC samples were obtained, and co-culture models were constructed to evaluate the effects of CLEC4E in TAMs on tumor growth, migration, and invasion in vitro. ResultsCLEC4E was significantly upregulated in GC, and high CLEC4E expression was associated with poor prognosis. Western blotting and immunostaining showed increased protein levels of CLEC4E in GC. GO, KEGG, and GSEA results indicated that CLEC4E is involved in immune response. Immune infiltration analysis demonstrated that CLEC4E expression positively correlated with multiple immune cell types. scRNA-seq analyses revealed that CLEC4E was predominantly expressed in myeloid cells specifically TAMs, in GC. In vitro experiments confirmed that MFC induced CLEC4E expression in TAMs to mediate tumor progression. Specifically targeting CLEC4E by si-CLEC4E or stigmasterol inhibited cancer cell migration and invasion. ConclusionCLEC4E is a potential prognostic biomarker and new therapeutic target for GC that can be specifically targeted by stigmasterol.

TLR2 and TLR9 Blockade Using Specific Intrabodies Inhibits Inflammation-Mediated Pancreatic Cancer Cell Growth.

Pancreatic cancer (pancreatic ductal adenocarcinoma, PDAC) remains a deadly cancer worldwide with a need for new therapeutic approaches. A dysregulation in the equilibrium between pro- and anti-inflammatory responses with a predominant immunosuppressive inflammatory reaction in advanced stage tumors seem to contribute to tumor growth and metastasis. The current therapies do not include strategies against pro-tumorigenic inflammation in cancer patients. We have shown that the upregulated cell surface expression of Toll-like Receptor (TLR) 2 and of TLR9 inside PDAC cells maintain chronic inflammatory responses, support chemotherapeutic resistance, and mediate tumor progression in human pancreatic cancer. We further demonstrated intracellular TLR2 and TLR9 targeting using specific intrabodies, which resulted in downregulated inflammatory signaling. In this study, we tested, for the first time, an intrabody-mediated TLR blockade in human TLR2- and TLR9-expressing pancreatic cancer cells for its effects on inflammatory signaling-mediated tumor growth. Newly designed anti-TLR2- and anti-TLR9-specific intrabodies inhibited PDAC growth. Co-expression analysis of the intrabodies and corresponding human TLRs showed efficient retention and accumulation of both intrabodies within the endoplasmic reticulum (ER), while co-immunoprecipitation studies indicated both intrabodies interacting with their cognate TLR antigen within the pancreatic cancer cells. Cancer cells with attenuated proliferation expressing accumulated TLR2 and TRL9 intrabodies demonstrated reduced STAT3 phosphorylation signaling, while apoptotic markers Caspases 3 and 8 were upregulated. To conclude, our results demonstrate the TLR2 and TLR9-specific intrabody-mediated signaling pathway inhibition of autoregulatory inflammation inside cancer cells and their proliferation, resulting in the suppression of pancreatic tumor cell growth. These findings underscore the potential of specific intrabody-mediated TLR inhibition in the ER relevant for tumor growth inhibition and open up a new therapeutic intervention strategy for the treatment of pancreatic cancer.

Diagnostic and therapeutic value of EVs in lungs diseases and inflammation.

Extracellular vesicles (EVs) are membrane-derived messengers which have been playing an important role in the inflammation and pathogenesis of lung diseases. EVs contain varieties of DNA, RNA, and membrane receptors through which they work as a delivery system for bioactive molecules as well as intracellular communicators. EV signaling mediates tumor progression and metastasis. EVs are linked with many diseases and perform a diagnostic role in lung injury and inflammation so are used to diagnose the severity of diseases. EVs containing a variety of biomolecules communicate with the recipient cells during pathophysiological mechanisms thereby acquiring the attention of clinicians toward the diagnostic and therapeutic potential of EVs in different lung diseases. In this review, we summarize the role of EVs in inflammation with an emphasis on their potential as a novel candidate in the diagnostics and therapeutics of chronic obstructive pulmonary disease, asthma, and sarcoidosis.

Crosstalk with lung fibroblasts shapes the growth and therapeutic response of mesothelioma cells

Mesothelioma is an aggressive cancer of the mesothelial layer associated with an extensive fibrotic response. The latter is in large part mediated by cancer-associated fibroblasts which mediate tumour progression and poor prognosis. However, understanding of the crosstalk between cancer cells and fibroblasts in this disease is mostly lacking. Here, using co-cultures of patient-derived mesothelioma cell lines and lung fibroblasts, we demonstrate that fibroblast activation is a self-propagated process producing a fibrotic extracellular matrix (ECM) and triggering drug resistance in mesothelioma cells. Following characterisation of mesothelioma cells/fibroblasts signalling crosstalk, we identify several FDA-approved targeted therapies as far more potent than standard-of-care Cisplatin/Pemetrexed in ECM-embedded co-culture spheroid models. In particular, the SRC family kinase inhibitor, Saracatinib, extends overall survival well beyond standard-of-care in a mesothelioma genetically-engineered mouse model. In short, we lay the foundation for the rational design of novel therapeutic strategies targeting mesothelioma/fibroblast communication for the treatment of mesothelioma patients.

MLC1 Overexpression Inhibits Tumor Progression through PI3K/AKT Signal Pathway in Prostate Cancer.

Prostate cancer (PC) is a prevalent malignancy in males, characterized by high morbidity and mortality. Despite MLC1 being established as a key mediator in tumor progression, its role in PC remains unexplored. This study aims to validate MLC1's anti-tumor effects and uncover potential mechanisms. MLC1's clinical significance is assessed using data from The Cancer Genome Atlas and the Genotype-Tissue Expression databases. MLC1 expression is significantly reduced in PC samples compared with the adjacent normal tissues. MLC1 expression correlates negatively with tumor metastasis and positively with the survival of patients with PC. In vitro, up-regulating MLC1 effectively inhibits tumor progression by curtailing proliferation, infestation, and migration through the deactivation of the PI3K/AKT signaling pathway. Conversely, down-regulating MLC1 promotes PC progression, a phenomenon alleviated by the PI3K/AKT inhibitor, Gefitinib. Furthermore, the anti-tumor function of MLC1 is corroborated by a reduction in tumor volume compared with the negative control in vivo. This study confirms the anti-tumor effects of MLC1 via in vitro and in vivo experiments, demonstrating its potential mechanism of inhibiting the PI3K/AKT signaling pathway.

Identification of Potential Protein Targets in Extracellular Vesicles Isolated from Chemotherapy-Treated Ovarian Cancer Cells.

Despite the ongoing clinical trials and the introduction of novel treatments over the past few decades, ovarian cancer remains one of the most fatal malignancies in women worldwide. Platinum- and paclitaxel-based chemotherapy is effective in treating the majority of patients with ovarian cancer. However, more than 70% of patients experience recurrence and eventually develop chemoresistance. To improve clinical outcomes in patients with ovarian cancer, novel technologies must be developed for identifying molecular alterations following drug-based treatment of ovarian cancer. Recently, extracellular vesicles (EVs) have gained prominence as the mediators of tumor progression. In this study, we used mass spectrometry to identify the changes in EV protein signatures due to different chemotherapeutic agents used for treating ovarian cancer. By examining these alterations, we identified the specific protein induction patterns of cisplatin alone, paclitaxel alone, and a combination of cisplatin and paclitaxel. Specifically, we found that drug sensitivity was correlated with the expression levels of ANXA5, CD81, and RAB5C in patients receiving cisplatin with paclitaxel. Our findings suggest that chemotherapy-induced changes in EV protein signatures are crucial for the progression of ovarian cancer.

Functional Characterization of Transforming Growth Factor-β Signaling in Dasatinib Resistance and Pre-BCR+ Acute Lymphoblastic Leukemia.

The multi-kinase inhibitor dasatinib has been implicated to be effective in pre-B-cell receptor (pre-BCR)-positive acute lymphoblastic leukemia (ALL) expressing the E2A-PBX1 fusion oncoprotein. The TGFβ signaling pathway is involved in a wide variety of cellular processes, including embryonic development and cell homeostasis, and it can have dual roles in cancer: suppressing tumor growth at early stages and mediating tumor progression at later stages. In this study, we identified the upregulation of the TGFβ signaling pathway in our previously generated human dasatinib-resistant pre-BCR+/E2A-PBX1+ ALL cells using global transcriptomic analysis. We confirm the upregulation of the TGFβ pathway member SMAD3 at the transcriptional and translational levels in dasatinib-resistant pre-BCR+/E2A-PBX1+ ALL cells. Hence, dasatinib blocks, at least partially, TGFβ-induced SMAD3 phosphorylation in several B-cell precursor (BCP) ALL cell lines as well as in dasatinib-resistant pre-BCR+/E2A-PBX1+ ALL cells. Activation of the TGFβ signaling pathway by TGF-β1 leads to growth inhibition by cell cycle arrest at the G0/G1 stage, increase in apoptosis and transcriptional changes of SMAD-targeted genes, e.g. c-MYC downregulation, in pre-BCR+/E2A-PBX1+ ALL cells. These results provide a better understanding about the role that the TGFβ signaling pathway plays in leukemogenesis of BCP-ALL as well as in secondary drug resistance to dasatinib.

Significance of Th17 and Treg in Treatment Efficacy and Outcome in Pediatric Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia represents a malignant proliferation of lymphoid cells blocked at an early stage of cell differentiation. It is the most common cancer occurring in children. Despite favorable prognosis, the survival rate of patients with poor treatment response or relapse remains dismal. The interaction between leukemic cells and the tumor immune microenvironment is pivotal in mediating tumor progression. In this study we evaluated associations between Treg and Th17 lymphocytes and the clinical presentation of ALL pediatric patients to validate their value in monitoring treatment outcome. The peripheral blood and bone marrow aspirates from 35 pediatric patients with ALL and 48 healthy control subjects were selected for the experiment. We demonstrated the numbers of Th17 lymphocytes and Tregs were increased in the bone marrow of ALL patients at the moment of diagnosis compared to the healthy control group, with the latter significantly decreasing during the course of ALL treatment. Patients with lower Th17 were found to demonstrate higher risk of blasts prevalence in bone marrow at day 33. ALL patients with lower WBC demonstrated higher frequency of Tregs. In summary, we identified a significant role of Th17 and Treg lymphocytes in ALL of pediatric patients and their contribution to disease-related parameters.

An integrated genomic approach identifies follistatin as a target of the p63-epidermal growth factor receptor oncogenic network in head and neck squamous cell carcinoma.

Although numerous putative oncogenes have been associated with the etiology of head and neck squamous cell carcinoma (HNSCC), the mechanisms by which these oncogenes and their downstream targets mediate tumor progression have not been fully elucidated. We performed an integrative analysis to identify a crucial set of targets of the oncogenic transcription factor p63 that are common across multiple transcriptomic datasets obtained from HNSCC patients, and representative cell line models. Notably, our analysis revealed FST which encodes follistatin, a secreted glycoprotein that inhibits the transforming growth factor TGFβ/activin signaling pathways, to be a direct transcriptional target of p63. In addition, we found that FST expression is also driven by epidermal growth factor receptor EGFR signaling, thus mediating a functional link between the TGF-β and EGFR pathways. We show through loss- and gain-of-function studies that FST predominantly imparts a tumor-growth and migratory phenotype in HNSCC cells. Furthermore, analysis of single-cell RNA sequencing data from HNSCC patients unveiled cancer cells as the dominant source of FST within the tumor microenvironment and exposed a correlation between the expression of FST and its regulators with immune infiltrates. We propose FST as a prognostic biomarker for patient survival and a compelling candidate mediating the broad effects of p63 on the tumor and its associated microenvironment.

G6PD activation in TNBC cells induces macrophage recruitment and M2 polarization to promote tumor progression.

Glucose-6-phosphate dehydrogenase (G6PD) is involved in triple-negative breast cancer (TNBC) progression. Metabolic crosstalk between cancer cells and tumor-associated macrophages mediates tumor progression in TNBC. Molecular biological methods were applied to clarify the mechanism of the crosstalk between TNBC cells and M2 macrophages. In the present study, we verified that G6PD overexpression drives M2 macrophage polarization by directly combining with phospho-STAT1 and upregulating CCL2 and TGF-β1 secretion in TNBC cells. In turn, M2-like TAMs activated TNBC cells through IL-10 secretion, providing feedback to upregulate G6PD and promote TNBC cell migration and proliferation in vitro. Furthermore, we found that 6-AN (a specific inhibitor of G6PD) not only suppressed the cancer-driven polarization of macrophages toward the M2 phenotype but also inhibited the inherent M2 polarization of macrophages. Targeting the G6PD-regulated pentose phosphate pathway restrained TNBC progression and M2-type polarization of macrophages in vitro and in vivo.

TAM receptor tyrosine kinase MERTK as a therapeutic target in the tumor immune microenvironment in a murine Krasmutant lung cancer model

Abstract TAM receptor tyrosine kinase MERTK is involved in apoptotic cell clearance to maintain tissue homeostasis. During efferocytosis, MERTK activation also promotes an immunosuppressive phenotype, which can be hijacked by tumor cells to escape immune attacking. Here, we determined the impact of MERTK inhibition on immune cell subsets in the tumor microenvironment in mice with subcutaneous murine CMT167 (Kras-G12V) lung cancer grafts. Although MERTK expression was not detected on tumor associated dendritic cells (DCs), there was a significant increase in the incidence of DCs in tumors from Mertk knockout (KO) mice compared to wild type mice by day 5 after inoculation of tumor cells. Characterization of tumor-associated macrophages by flow cytometry revealed three distinct populations expressing different levels of MHCII (CD11b +MHCII high, CD11b +MHCII +and CD11b highMHCII neg). Tumors from Mertk KO mice had significantly greater numbers of MHCII highmacrophages by day 10 after tumor cell inoculation and reduced numbers of MHCII negmacrophages by day 19. Together, the increased incidence of dendritic cells and the shift toward a more antigen-presenting macrophage phenotype are expected to promote anti-tumor immunity in Mertk KO mice. Indeed, CMT167 tumor growth was significantly reduced in Mertk KO mice and this effect was abrogated in Mertk KO scid mice, which lack functional B and T cells. Treatment with MRX-2843, a MERTK-selective inhibitor currently in Phase 1 clinical trials, also reduced tumor growth in mice. Together, these data demonstrate a critical role for MERTK as a mediator of tumor progression in the tumor immune microenvironment and implicate MRX-2843 as an effective strategy to exploit this finding in the clinic. The research was supported by the National Cancer Institute of the National Institutes of Health (Emory Lung Cancer Spore, Award Number P50CA217691). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.