Regulating Tumor Cell Proliferation Research Articles

Ligilactobacillus salivarius LZZAY01 accelerated autophagy and apoptosis in colon cancer cells and improved gut microbiota in CAC mice.

Colorectal cancer (CRC) is one of the malignant tumors globally, with high morbidity and mortality rates. The mainstay treatment of CRC includes surgery, radiotherapy, and chemotherapy. However, these treatments are associated with a high recurrence rate, poor prognosis, and highly toxic side effects. The probiotics have the potential to prevent CRC, and they display a favorable safety performance. Probiotics could provide a potential strategy to prevent and treat CRC. The impact of LZZAY01 on cancer cell lines CT-26, HCT-116, and SW-620 was evaluated by conducting cytotoxicity and clonogenicity tests. A model of colitis-associated cancer (CAC) was established in C57BL/6j mice following induction with AOM/DSS. The levels of autophagy and apoptosis proteins, tight junction proteins, and inflammatory factors were detected by western blotting, immunofluorescence assay, and enzyme-linked immunosorbent assay. High-throughput sequencing of gut 16S rRNA was performed to analyze the abundance and diversity of the gut microbiome. LZZAY01, a new strain of Ligilactobacillus salivarius, was certified by an evolutionary tree and average nucleotide identity. LZZAY01 enhanced autophagy and apoptosis in CT-26, HCT-116, and SW-620 cell lines. It preserved the integrity of the intestinal barrier by regulating the tight junction protein ZO-1 and claudin-1. The tumor necrosis factor-α and interleukin-6 were reduced by LZZAY01. The abundance and diversity of the intestinal microbiota were enhanced, especially the beneficial bacterial species maintaining the balance of the intestinal flora such as Bifidobacterium and Lactobacillus. L. salivarius LZZAY01 improved CAC via suppressing the growth of colon cancer cells, promoting autophagy and apoptosis, enhancing intestinal tight junctions, reducing intestinal barrier degradation, modifying the gut microbiota abundance, and decreasing inflammatory reactions.IMPORTANCEAlthough similar probiotics have been shown to have anticancer potential in colorectal cancer (CRC), there is a paucity of research related to the preventive function of probiotics against CRC. And there are fewer studies about the mechanism of probiotics' preventive effects on CRC. The regulation of tumor cell proliferation and apoptosis by the active ingredients of probiotics may be one of the mechanisms of their prevention of CRC. In this study, we explored the effects of L. salivarius LZZAY01 on autophagy and apoptosis of colon cancer cells in vitro and in vivo and proposed a possible mechanism for the prevention of CRC by probiotics.

E2F1-driven CENPM expression promotes glycolytic reprogramming and tumorigenicity in glioblastoma

Centromere protein M (CENPM), traditionally associated with chromosome segregation, is now recognized for its significant role in cancer biology. Particularly in glioblastoma (GBM), where less is known about CENPM compared to other centromere proteins (CENPs), it appears crucially involved in regulating tumor cell proliferation, invasion, and metabolic reprogramming-key factors in GBM's aggressiveness. Initial analyses using the GEPIA database (TCGA/GTEx datasets) reveal distinct patterns of CENPM expression in GBM, suggesting its potential as a therapeutic target. Our study manipulated CENPM expression through shRNA-mediated knockdown and vector-based overexpression in GBM cell lines LN229 and U251. Knockdown resulted in a 50% reduction in cell proliferation and a 70% decrease in invasion, accompanied by diminished glycolytic markers such as glucose consumption, lactate production, and ATP levels. Conversely, overexpression of CENPM enhanced both metabolic activity and invasive capacities. The introduction of the glycolytic inhibitor 2-DG effectively reversed the effects of CENPM modulation, highlighting a dependency on glycolytic pathways. Moreover, we identified E2F1 as a key regulator of CENPM, linking it to GBM’s metabolic alterations. In vivo studies using a BALB/c nude mouse xenograft model demonstrated that CENPM knockdown significantly inhibits tumor growth, with treated groups showing a 60% reduction in tumor volume over four weeks. These findings underscore the E2F1-CENPM axis as a promising target for therapeutic strategies, aiming to disrupt the metabolic and invasive pathways facilitated by CENPM in GBM. These insights establish a foundation for targeting the metabolic dependencies of tumor cells, potentially leading to innovative treatments for GBM.Graphical abstractE2F1 combined with the promoter of CENPM, and positively regulated CENPM expression. CENPM accelerated glycolysis to aggravate cell proliferation, invasion, and EMT process in GBM progression. And, CENPM facilitated tumor growth in vivo.

Study on the anti-colon cancer effect of Renshen Yangrong Decoction based on network pharmacology, molecular docking, and in vivo and in vitro experiments

Renshen Yangrong Decoction (RSYRD) is a traditional Chinese medicinal compound widely used as an adjuvant in colon cancer treatment. However, the specific role of RSYRD in anti-colon cancer remains unclear. This study aimed to investigate the anti-colon cancer effects of RSYRD using network pharmacology, molecular docking, and in vitro and in vivo experiments. The active ingredients and targets of RSYRD were identified utilizing databases, and RSYRD-ingredient-target and protein-protein interaction (PPI) networks were constructed. Gene Ontology (GO) functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were performed on the anti-colon cancer targets of RSYRD, followed by molecular docking using AutoDockTools 1.5.6. In vitro and in vivo experiments were conducted to explore the mechanism of RSYRD and evaluate its inhibitory efficacy on colon cancer. A total of 233 active ingredients and 88 colon cancer-related targets were identified, including 9 core targets. GO functional analysis revealed that RSYRD exerts anti-tumor effects through processes such as oxidative reactions, closely associated with intracellular reactive oxygen species (ROS). KEGG enrichment analysis indicated that the anti-tumor effect of RSYRD involved the interaction of cancer-related and viral disease-associated pathways, as well as the IL-17, TNF, and PI3K-AKT signaling pathways. Molecular docking showed stable ligand-receptor binding, with binding energies below −5.0 kcal/mol. In vitro experiments demonstrated that RSYRD increased ROS levels, reduced mitochondrial membrane potential (MMP), and promoted apoptosis in SW620 cells, thereby inhibiting cell proliferation and migration. In vivo experiments showed that RSYRD significantly suppressed colon cancer proliferation. This study identified Quercetin, Kaempferol, 7-Methoxy-2-methyl-isoflavone, Licochalcone-a, and Isorhamnetin as core ingredients in RSYRD, likely exerting anti-colon cancer effects by inhibiting proteins such as ESR1, HSP90AA1, NCOA2, and MAPK14, and suppressing the PI3K-AKT signaling pathway, thereby regulating tumor cell proliferation, migration, and apoptosis.

Integrating multi-omics techniques and in vitro experiments reveals that GLRX3 regulates the immune microenvironment and promotes hepatocellular carcinoma cell proliferation and invasion through iron metabolism pathways.

Hepatocellular carcinoma (HCC) is a common malignancy worldwide, and its development is closely related to abnormalities in iron metabolism. This study aims to systematically analyze changes in iron metabolism in the tumor microenvironment of HCC using single-cell sequencing technology, and investigate the potential mechanisms by which iron metabolism regulation affects the survival of liver cancer patients. Single-cell sequencing data from hepatocellular carcinoma patients were obtained from the GEO database. By iron metabolism genomic scoring, we assessed differences in iron metabolism levels in hepatocellular carcinoma samples. By cell communication analysis as well as GO and KEGG enrichment analysis, we determined the functional role of iron metabolism in different cell types. We used survival analysis and Kaplan-Meier curves to assess the impact of iron metabolism levels on patient prognosis. In addition, we identified and analyzed the expression profile of the GLRX3 gene, investigated its key regulatory role in iron metabolism, and validated its clinical value as a prognostic marker. Finally, we explored the effect of GLRX3 on hepatocellular carcinoma phenotype by in vitro experiments such as PCR, transwell, CCK8, and wound healing assay. Bioinformatics results and experimental validation confirmed the dysregulation of iron metabolism in the development of hepatocellular carcinoma, revealing iron's regulatory influence across various cell types. Additionally, GLRX3 was identified as a key regulatory factor in iron metabolism, and the mechanism by which GLRX3 regulates tumor cell proliferation and immune evasion was determined. Furthermore, experiments verified GLRX3's role in facilitating tumor cell proliferation and invasion. This study highlights the critical role of iron metabolism in the progression of hepatocellular carcinoma, particularly the regulatory mechanism of the GLRX3 gene in tumor cell proliferation and immune evasion. Iron metabolism abnormalities are not only drivers of liver cancer development but also key indicators of patient prognosis.

Recent Advances of Small Extracellular Vesicles for the Regulation and Function of Cancer-Associated Fibroblasts.

Cancer-associated fibroblasts (CAFs) are a heterogeneous cell population in the tumor microenvironment (TME) that critically affect cancer progression. Small extracellular vesicles (sEVs) act as information messengers by transmitting a wide spectrum of biological molecules, including proteins, nucleic acids, and metabolites, from donor cells to recipient cells. Previous studies have demonstrated that CAFs play important roles in tumor progression by regulating tumor cell proliferation, metastasis, therapeutic resistance, and metabolism via sEVs. In turn, tumor-derived sEVs can also regulate the activation and phenotype switch of CAFs. The dynamic crosstalk between CAFs and cancer cells via sEVs could ultimately determine cancer progression. In this review, we summarized the recent advance of the biological roles and underlying mechanisms of sEVs in mediating CAF-tumor cell interaction and its impact on cancer progression. We also reviewed the clinical applications of tumor- and CAF-derived sEVs, which could identify novel potential targets and biomarkers for cancer diagnosis, therapy, and prognosis.

CD73 promotes non–small cell lung cancer metastasis by regulating Axl signaling independent of GAS6

As catabolic enzyme, CD73 dephosphorylates adenosine monophosphate (AMP) and can also regulate tumor cell proliferation and metastasis. To date, very few studies have explored the role of CD73 in mediating non-small cell lung cancer (NSCLC) metastasis, and the underlying transducing signal has not been elucidated. In the present study, we demonstrated that the CD73/Axl axis could regulate Smad3-induced epithelial-to-mesenchymal transition (EMT) to promote NSCLC metastasis. Mechanically, CD73 can be secreted via the Golgi apparatus transport pathway. Then secreted CD73 may activate AXl by directly bind with site R55 located in Axl extracellular domain independently of GAS6. In addition, we proved that CD73 can stabilize Axl expression via inhibiting CBLB expression. We also identified the distinct function of CD73 activity in adenocarcinoma and squamous cell carcinoma. Our findings indicated a role of CD73 in mediating NSCLC metastasis and propose it as a therapeutic target for NSCLC.

Targeting CBP and p300: Emerging Anticancer Agents.

CBP and p300 are versatile transcriptional co-activators that play essential roles in regulating a wide range of signaling pathways, including Wnt/β-catenin, p53, and HIF-1α. These co-activators influence various cellular processes such as proliferation, differentiation, apoptosis, and response to hypoxia, making them pivotal in normal physiology and disease progression. The Wnt/β-catenin signaling pathway, in particular, is crucial for cellular proliferation, differentiation, tissue homeostasis, and embryogenesis. Aberrant activation of this pathway is often associated with several types of cancer, such as colorectal tumor, prostate cancer, pancreatic and hepatocellular carcinomas. In recent years, significant efforts have been directed toward identifying and developing small molecules as novel anticancer agents capable of specifically inhibiting the interaction between β-catenin and the transcriptional co-activators CBP and p300, which are required for Wnt target gene expression and are consequently involved in the regulation of tumor cell proliferation, migration, and invasion. This review summarizes the most significant and original research articles published from 2010 to date, found by means of a PubMed search, highlighting recent advancements in developing both specific and non-specific inhibitors of CBP/β-catenin and p300/β-catenin interactions. For a more comprehensive view, we have also explored the therapeutic potential of CBP/p300 bromodomain and histone acetyltransferase inhibitors in disrupting the transcriptional activation of genes involved in various signaling pathways related to cancer progression. By focusing on these therapeutic strategies, this review aims to offer a detailed overview of recent approaches in cancer treatment that selectively target CBP and p300, with particular emphasis on their roles in Wnt/β-catenin-driven oncogenesis.

Phosphatase LHPP confers prostate cancer ferroptosis activation by modulating the AKT-SKP2-ACSL4 pathway

LHPP, a novel, recognized tumor suppressor, exerts a critical influence on the regulation of tumor cell proliferation and survival by modulating various signaling pathways with its phosphatase activity. Here, we unveil a robust correlation between reduced LHPP expression and adverse prognosis in prostate cancer. We demonstrate that LHPP interacts with AKT, thereby dampening AKT phosphorylation and subsequently inhibiting ACSL4 phosphorylation at the T624 site. This interaction impedes phosphorylation-dependent ubiquitination, thwarting SKP2 from recognizing and binding to ACSL4 at the K621 site. As a result, ACSL4 is spared from lysosomal degradation, leading to its accumulation and the promotion of lipid peroxidation, and ferroptosis. Moreover, our findings reveal that Panobinostat, a potent histone-deacetylase inhibitor, intricately regulates LHPP expression at multiple levels through the inhibition of HDAC3. This complex modulation enhances the ferroptosis pathway, offering a novel mechanism for curtailing the growth of prostate tumors and highlighting its significant translational potential for clinical application.

Jaranol Loaded in Ferroferric Oxide Nanoparticles Inhibits the Activities of Liver Cancer Cell Through Mitogen-Activated Protein Kinase/Extracellular Signal-Regulated Kinase Signaling Pathway

Liver cancer is highly aggressive and the MEK/ERK signaling regulates tumor cell proliferation and metastasis. Jaranol is a natural product with anti-proliferative and anti-metastatic effects in several tumors. However, its interaction with the MEK/ERK pathway in liver cancer is unclear. This study explores whether ferroferric oxide nanoparticles-loaded Jaranol inhibits proliferation and metastasis in liver cancer cell Hep3B by regulating MEK/ERK signaling, and its underlying mechanism. Fe3O4-Jaranol nanoparticles were prepared and used in Hep3B experiments to observe the biological efficacy of Fe3O4-Jaranol, and further explore its effect and mechanism on the MEK/ERK pathway using PCR, WB, etc. Fe3O4-Jaranol were successfully prepared with a certain tumor suppressor effect in liver cancer. The expression of MEK/ERK was increased in liver cancer. Inhibiting its pathway activity suppressed the development of liver cancer. Trametinib and C16-PAF were used to inhibit Hep3B respectively. MEK expression in cells treated with Trametinib was reduced accompanied by a low expression of ERK, while the expression of MEK and ERK levels in the C16-PAF group showed an opposite trend, indicating that Trametinib, C16-PAF successfully intervened on MEK and ERK. Further analysis of the activity of Hep3B cells found that the proliferation ability of the Trametinib group was significantly inhibited. Fe3O4-Jaranol significantly inhibited liver cancer cell Hep3B and this effect was accomplished by inhibiting MEK/ERK signaling, causing tumor cell proliferation to be restricted and reducing the ability to metastasize. This research result provides strong evidence for a deep understanding of the mechanism of Jaranol in treating liver cancer, so as to better guide clinical practice.

Overexpression of mir-489–3p inhibits proliferation and migration of non-small cell lung cancer cells by suppressing the HER2/PI3K/AKT/Snail signaling pathway

BackgroundLung cancer is a highly prevalent malignancy with significant morbidity and mortality rates. MiR-489–3p, a microRNA, has been identified as a regulator of tumor cell proliferation and invasion. Its expression is downregulated in non-small cell lung cancer (NSCLC). Elucidating the molecular mechanisms underlying miR-489–3p′s role in NSCLC pathogenesis is crucial for identifying potential diagnostic and therapeutic targets. MethodsTo investigate the molecular mechanism of miR-489–3p in NSCLC, this study utilized A549, a commonly used NSCLC cell line. MiR-489–3p mimics and inhibitors were transfected into A549 cells. Additionally, co-transfection experiments using wortmannin, an inhibitor of the PI3K/AKT pathway, were performed. Expression of miR-489–3p and related proteins was analyzed by Western blotting and quantitative real-time PCR (qRT-PCR). Cell migration and proliferation were assessed by wound healing and colony formation assays, respectively. ResultsOverexpression of miR-489–3p significantly inhibited the proliferation and migration of A549 cells. This inhibitory effect was further enhanced upon co-transfected with wortmannin. Analysis of human lung specimens showed increased expression of HER2, PI3K, and AKT in lung adenocarcinoma tissues compared to adjacent non-cancerous tissues. ConclusionsThese findings suggest that miR-489–3p overexpression may inhibit NSCLC cell proliferation and migration by suppressing the HER2/PI3K/AKT/Snail signaling pathway. This study elucidates miR-489–3p′s molecular mechanisms in NSCLC and provides experimental basis for identifying early diagnostic markers and novel therapeutic targets.

Single-cell transcriptomic sequencing analysis of mechanistic insights into the IFN-γ signaling pathway in different tumor cells.

This study aimed to investigate the relationship between the interferon-gamma (IFN-γ) pathway in different tumor microenvironments (TME) and patients' prognosis, as well as the regulatory mechanisms of this pathway in tumor cells. Using RNA-seq data from the TCGA database, we analyzed the predictive value of the IFN-γ pathway across various tumors. We employed a univariate Cox regression model to assess the prognostic significance of IFN-γ signaling in different tumor types. Additionally, we analyzed single-cell RNA sequencing (scRNA-seq) data from the Gene Expression Omnibus (GEO) database to examine the distribution characteristics of the IFN-γ pathway and explore its regulatory mechanisms, highlighting how IFN-γ influenced cellular interactions within the TME. Our analysis revealed a significant association between the IFN-γ pathway and adverse prognosis in pan-cancer tissues (P < 0.001). Interestingly, this correlation varied regarding positive and negative regulation across different tumor types. Through a detailed examination of scRNA-seq data, we found that the IFN-γ pathway exerted substantial regulatory effects on stromal and immune cells. In contrast, its expression and regulatory patterns in tumor cells exhibited diversity and heterogeneity. Further analysis indicated that the IFN-γ pathway not only enhanced the immunogenicity of tumor cells but also inhibited their proliferation. Cell-cell interaction analysis confirmed the pivotal role of the IFN-γ pathway within the overall regulatory network. Moreover, we identified HMGB2 (high mobility group box 2) in T cells as a potential key regulator of tumor cell proliferation. The IFN-γ pathway exhibited a dual function by both suppressing tumor cell proliferation and enhancing their immunogenicity, positioning it as a pivotal target for refined cancer diagnosis and cancer strategies.

Berberine and berberine nanoformulations in cancer therapy: Focusing on lung cancer.

Lung cancer is the second most prevalent cancer and ranks first in cancer-related death worldwide. Due to the resistance development to conventional cancer therapy strategies, including chemotherapy, radiotherapy, targeted therapy, and immunotherapy, various natural products and their extracts have been revealed as alternatives. Berberine (BBR), which is present in the stem, root, and bark of various trees, could exert anticancer activities by regulating tumor cell proliferation, apoptosis, autophagy, metastasis, angiogenesis, and immune responses via modulating several signaling pathways within the tumor microenvironment. Due to its poor water solubility, poor pharmacokinetics/bioavailability profile, and extensive p-glycoprotein-dependent efflux, BBR application in (pre) clinical studies is restricted. To overcome these limitations, BBR can be encapsulated in nanoparticle (NP)-based drug delivery systems, as monotherapy or combinational therapy, and improve BBR therapeutic efficacy. Nanoformulations also facilitate the selective delivery of BBR into lung cancer cells. In addition to the anticancer activities of BBR, especially in lung cancer, here we reviewed the BBR nanoformulations, including polymeric NPs, metal-based NPs, carbon nanostructures, and others, in the treatment of lung cancer.

Long non-coding RNA SIX1-1 promotes proliferation of cervical cancer cells via negative transcriptional regulation of RASD1.

Cervical cancer poses a significant health burden for women globally, and the rapid proliferation of cervical cancer cells greatly worsens patient prognosis. Long non-coding RNAs (lncRNAs) play a crucial role in regulating tumor cell proliferation. However, the involvement of lncRNAs in cervical cancer cell proliferation remains unclear. In this study, we investigated the lncRNA SIX1-1, which was found to be upregulated in cervical cancer tissues and cell lines. Functional assays revealed that knockdown of SIX1-1 inhibited cell proliferation in vitro and reduced tumor growth in vivo. Mechanistically, SIX1-1 was predominantly localized in the nucleus and could bind with DNMT1 protein. The expression of SIX1-1 enhanced the interaction of DNMT1 with RASD1 promoter, leading to the methylation of the promoter and decreased mRNA transcription. Then RASD1 downregulation activated the cAMP/PKA/CREB signaling pathway, promoting cell proliferation. Rescue experiments showed that knockdown of RASD1 restored the inhibited cell proliferation caused by decreased expression of SIX1-1, indicating that RASD1 acted as the functional mediator of SIX1-1. In conclusion, SIX1-1 promoted cervical cancer cell proliferation by modulating RASD1 expression. This suggests that targeting the SIX1-1/RASD1 axis could be a potential antitumor strategy for cervical cancer.

The YY1-CPT1C signaling axis modulates the proliferation and metabolism of pancreatic tumor cells under hypoxia

Carnitine palmitoyltransferase 1C (CPT1C) is an enzyme that regulates tumor cell proliferation and metabolism by modulating mitochondrial function and lipid metabolism. Hypoxia, commonly observed in solid tumors, promotes the proliferation and progression of pancreatic cancer by regulating the metabolic reprogramming of tumor cells. So far, the metabolic regulation of hypoxic tumor cells by CPT1C and the upstream mechanisms of CPT1C remain poorly understood. Yin Yang 1 (YY1) is a crucial oncogene for pancreatic tumorigenesis and acts as a transcription factor that is involved in multiple metabolic processes. This study aimed to elucidate the relationship between YY1 and CPT1C under hypoxic conditions and explore their roles in hypoxia-induced proliferation and metabolic alterations of tumor cells. The results showed enhancements in the proliferation and metabolism of PANC-1 cells under hypoxia, as evidenced by increased cell growth, cellular ATP levels, up-regulation of mitochondrial membrane potential, and decreased lipid content. Interestingly, knockdown of YY1 or CPT1C inhibited hypoxia-induced rapid cell proliferation and vigorous cell metabolism. Importantly, for the first time, we reported that YY1 directly activated the transcription of CPT1C and clarified that CPT1C was a novel target gene of YY1. Moreover, the YY1 and CPT1C were found to synergistically regulate the proliferation and metabolism of hypoxic cells through transfection with YY1 siRNA to CRISPR/Cas9-CPT1C knockout PANC-1 cells. Taken together, these results indicated that the YY1-CPT1C axis could be a new target for the intervention of pancreatic cancer proliferation and metabolism.

Cyclin L1 participates in Adriamycin resistance and progression of osteosarcoma via PI3K/AKT-mTOR pathway.

Chemoresistance is a common and thorny problem in the treatment of osteosarcoma (OS), which obstructs the response of relapse or metastasis of OS to chemotherapy and leads to the unfavorable prognosis of OS patients. Cyclin L1 (CCNL1) is a non-canonical cyclin that plays an important role in the regulation of tumor cell proliferation and lymph node metastasis. In this work, we explored the impact of CCNL1 expression levels on proliferation, migration, and Adriamycin (ADM) resistance in OS and related mechanisms. We found that CCNL1 expression levels were significantly associated with clinical prognosis of patients with OS and CCNL1 could promote OS proliferation and migration. In addition, we also revealed that cellular CCNL1 was significantly increased in ADM-resistant OS cells and promoted ADM resistance. The PI3K/AKT-mTOR pathway is involved in CCNL1-mediated ADM resistance in OS. In summary, CCNL1 is involved in the progression of ADM resistance and OS through the PI3K/AKT-mTOR pathway, which will provide a new clue to the mechanism of ADM resistance and a potential target for the treatment of ADM-resistant OS.

MiR-504 knockout regulates tumor cell proliferation and immune cell infiltration to accelerate oral cancer development

miR-504 plays a pivotal role in the progression of oral cancer. However, the underlying mechanism remains elusive in vivo. Here, we find that miR-504 is significantly down-regulated in oral cancer patients. We generate miR-504 knockout mice (miR-504−/−) using CRISPR/Cas9 technology to investigate its impact on the malignant progression of oral cancer under exposure to 4-Nitroquinoline N-oxide (4NQO). We show that the deletion of miR-504 does not affect phenotypic characteristics, body weight, reproductive performance, and survival in mice, but results in changes in the blood physiological and biochemical indexes of the mice. Moreover, with 4NQO treatment, miR-504−/− mice exhibit more pronounced pathological changes characteristic of oral cancer. RNA sequencing shows that the differentially expressed genes observed in samples from miR-504−/− mice with oral cancer are involved in regulating cell metabolism, cytokine activation, and lipid metabolism-related pathways. Additionally, these differentially expressed genes are significantly enriched in lipid metabolism pathways that influence immune cell infiltration within the tumor microenvironment, thereby accelerating tumor development progression. Collectively, our results suggest that knockout of miR-504 accelerates malignant progression in 4NQO-induced oral cancer by regulating tumor cell proliferation and lipid metabolism, affecting immune cell infiltration.

Exploring the association of POSTN+ cancer-associated fibroblasts with triple-negative breast cancer

Triple-negative breast cancer (TNBC) is a subtype of breast cancer with a poor prognosis. Cancer-associated fibroblasts (CAFs) play a critical role in regulating TNBC tumor development. This study aimed to identify and characterize a specific subtype of CAFs associated with TNBC. Initially, using high-throughput bulk transcriptomic data in two cohorts, we identified three CAF-related subtypes (CS1, CS2, CS3) in TNBC samples. These three CAFs subtypes were closely linked to the tumor microenvironment. The CS1 subtype exhibited a relatively immune-rich microenvironment and a favourable prognosis, whereas the CS3 subtype displayed an immune-deprived tumor microenvironment and an unfavourable prognosis. Through WGCNA analysis, POSTN was identified as a key biomarker for CAFs associated with TNBC. Then, POSTN+CAFs was identified and characterized. Both POSTN and POSTN+CAFs showed significant positive correlations with stromal molecules HGF and MET at both the transcriptional and protein levels. Specifically co-localized with CAFs in the tumor stromal area, POSTN, produced by POSTN+CAFs, could modulate the HGF-MET axis, serving as a bypass activation pathway to regulate tumor cell proliferation in response to EGFR inhibitor and MET inhibitor. This study underscores the significance of POSTN and POSTN+CAFs as crucial targets for the diagnosis and treatment of TNBC.

Tropomyosin 2 Regulates Tumor Cell Proliferation, Immune Suppression, and Activation of the JNK Signaling Pathway in Colitis-Associated Cancer (CAC).

Tropomyosin 2 (TPM2) has been linked to the advancement of various tumor types, exhibiting distinct impacts on tumor progression. In our investigation, the primary objective was to identify the potential involvement of TPM2 in the development of colitis-associated cancer (CAC) using a mice model. This study used lentiviral vector complex for TPM2 knockdown (sh-TPM2) and the corresponding negative control lentiviral vector complex (sh-NC) for genetic interference in mice. CAC was induced in mice using azoxymethane (AOM) and dextran sulfate sodium salt (DSS). This study included 6 groups of mice models: Control, Control+sh-NC, Control+sh-TPM2, CAC, CAC+sh-NC, and CAC+sh-TPM2. Subsequently, colon tissues were collected and assessed using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for TPM2 mRNA levels and flow cytometry for infiltrating immune cells. Tumor number, size, and weight within colon tissues from CAC mice were measured and recorded. The hematoxylin-eosin staining was used for observing tissue pathology changes. The intestinal epithelial cells (IECs) were isolated and analyzed for cell proliferation. This analysis included examining the levels of 5-bromo-2-deoxyuridine (BrdU) and Ki-67 using immunohistochemistry. Additionally, the mRNA levels of proliferating cell nuclear antigen (PCNA) and Ki-67 were detected by qRT-PCR. This study also investigated the activation of the c-Jun N-terminal kinase (JNK) pathway using western blot analysis. Immunogenicity analyses were conducted using immunohistochemistry for F4/80 and flow cytometry. In 8-week-old mice, AOM injections and three cycles of DSS treatment induced TPM2 upregulation in tumor tissues compared to normal tissues (p < 0.05). Fluorescence-activated cell sorting (FACS)-isolated lamina CAC adenomas revealed macrophages and dendritic cells as primary TPM2 contributors (p < 0.001). Lentiviral TPM2 gene knockdown significantly reduced tumor numbers and sizes in CAC mice (p < 0.01, and p < 0.001), without invasive cancer cells. TPM2 suppression resulted in decreased IEC proliferation (p < 0.001) and reduced PCNA and Ki-67 expression (p < 0.05). Western blot analysis indicated reduced JNK pathway activation in TPM2-knockdown CAC mice (p < 0.05, p < 0.001). TPM2 knockdown decreased tumor-associated macrophage infiltration (p < 0.01) and increased CD3+ and CD8+ T cells (p < 0.01, and p < 0.001), with increased levels of regulator of inflammatory cytokines (CD44+, CD107a+) (p < 0.01, and p < 0.001), decreased levels of PD-1+ and anti-inflammatory factor (IL10+) (p < 0.01, and p < 0.001). Our results demonstrated that TPM2 knockdown suppressed the proliferation of CAC IECs, enhanced immune suppression on CAC IECs, and inhibited the JNK signaling pathway within the framework of CAC. These findings suggest TPM2 can serve as a potential therapeutic target for CAC treatment.

Abstract 3919: Targeting tissue-resident macrophage progenitors by anti-IL34 restores anti-tumor immunity and suppresses tumor growth

Abstract Macrophages play important roles in tumor microenvironment, where they can promote immune suppression, tumor growth and resistance to checkpoint inhibitor therapy and are the focus of efforts to develop novel cancer therapeutics. Strategies to inhibit macrophage accumulation to suppress tumor growth by blocking their recruitment from circulation have been only partially effective, leading to the identification of roles in tumor progression for tissue resident macrophages that are seeded during embryonic development. We examined the origins of lung tumor associated macrophages using lineage tracing, flow cytometry, and single cell sequencing. By comparing gene expression profiles of resident F4/80hi Lyve1+ Ly6C- macrophages and Ly6C+ F4/80lo CCR2+ recruited macrophages in subcutaneous, lung, central nervous and renal normal and tumor tissues, we identified generic tissue resident macrophage and recruited macrophage transcriptomic signatures, revealing that resident macrophages are characterized by signatures of Myc driven proliferation, fatty acid and cholesterol metabolism and immune suppression, while recruited macrophages are characterized by signature of inflammation and immune stimulation. Resident macrophages were associated with worse outcomes in cancer patients than recruited macrophages. Single cell sequencing and flow cytometry of genetically engineered mouse models of CC10-Cre KrasG12D;p53-/- and SPC-Cre KrasG12D lung cancer revealed that lung tumor progression was characterized by extensive expansion of immune suppressive, Ki67+ resident alveolar macrophages and loss of T cells in the lung. Single cell transcriptomics identified IL-34 expressed exclusively by tumor cells as a possible driver of alveolar macrophage proliferation and IGF1 expressed by tumor associated alveolar macrophages as a possible regulator of tumor cell proliferation. Accordingly, blockade of IL-34 profoundly reduced tissue resident macrophage proliferation, stimulated recruitment of activated T cells, and suppressed growth of KrasG12D driven tumors. Analysis of TCGA data sets revealed that expression of IGF1 or IL34 was associated with reduced survival in lung, kidney, liver, head and neck and ovarian cancer. These studies identify key roles for IL-34 and IGF1 in solid tumor progression and suggest new therapeutic targets for the development of therapeutic strategies to stimulate anti-cancer immunity. Citation Format: Hui Chen, Jaroslav Zak, Mark Paradise, Guangfang Wang, John Teijaro, Mark Onaitis, Judith Varner. Targeting tissue-resident macrophage progenitors by anti-IL34 restores anti-tumor immunity and suppresses tumor growth [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 3919.

Abstract 5274: Targeting retinoblastoma protein in tumor-associated macrophages suppresses ovarian cancer progression

Abstract Ovarian cancer is characterized by an immunosuppressive tumor microenvironment (TME) maintained by tumor-associated M2-like macrophages (TAMs) hindering anti-tumor responses and immunotherapy efficacy. An effective approach to target TAMs in ovarian cancer treatment is currently not available, but would be expected to release immunosuppressive pressure, enable a robust T cell response and improve immunotherapy outcomes. Retinoblastoma protein (Rb) is a well-known tumor suppressor and regulator of tumor cell proliferation. Accumulating evidence demonstrates its role in immune cells, in myeloid cells, in particular. However, these mechanisms remain poorly understood. Based on our previous data on Rb role in myeloid cell viability, we decided to test the effects of Rb modulation in macrophages. We used the small molecule, AP-3-84 compound, and newly developed analogs which bind the LxCxE motif of Rb and disrupt Rb interaction with its binding adaptor proteins. AP-3-84 induced cell death preferentially in macrophages, but not in T cells or tumor cells (without effects on cell proliferation). Gene and protein expression analysis revealed that Rb targeting induced major intracellular stress response programs and p53/mitochondria-related cell death pathways in TAMs. Moreover, we found that M2 type polarized macrophages expressed higher levels of Rb compared to M1 macrophages. In agreement with that, M2 type macrophages were significantly more sensitive to AP-3-84 treatment than M1 polarized cells. Next, we demonstrated that low dose therapeutic use of AP-3-84 in mice bearing ovarian cancer significantly re-shaped the immune composition of the TME by depleting TAMs and inducing remarkable T cell infiltration. Importantly, these AP-3-84 effects were immune-mediated, since identical AP-3-84 treatment in NSG immunodeficient mice had no effect on tumor growth. Similarly, anti-CD4/CD8 depletion also reverted AP-3-84 effects on cancer inhibition. An analysis of activation and differentiation markers in myeloid and T cell subsets in the TME showed a shift of the remaining macrophages towards an M1-like cell type accompanied by activation of T cells. Using ovalbumin-overexpressing ovarian cancer cell line, we documented the accumulation of antigen-specific T cells in the TME associated with a substantial delay of ovarian cancer upon AP-3-84 therapeutic Rb targeting. Ex vivo, we observed an analogous cell death induction by AP-3-84 treatment in TAMs from post-surgery ascites from ovarian cancer patients. Using available datasets, we further documented that an increase in Rb expression in the TME myeloid cells is associated with poorer prognosis. Overall, our data supports that therapeutic targeting of the Rb LxCxE motif is a promising approach for ovarian cancer treatment due to depletion of TAMs and re-shaping the ovarian cancer TME. Citation Format: Evgenii N. Tcyganov, Taekyoung Kwak, Xue Yang, Adi N. Poli, Colin Hart, Avishek Bhuniya, Joel Cassel, Andrew Kossenkov, Noam Auslander, Paridhima Sharma, Mark G. Cadungog, Stephanie Jean, Sudeshna Chatterjee-Paer, David Weiner, Laxminarasimha Donthireddy, Bryan Bristow, Rugang Zhang, Joseph M. Salvino, Luis J. Montaner. Targeting retinoblastoma protein in tumor-associated macrophages suppresses ovarian cancer progression [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 5274.