Expression Of Tumor Progression Research Articles

Pan‑cancer analysis on the role of KMT2C expression in tumor progression and immunotherapy.

Histone lysine N-methyltransferase 2C (KMT2C) is involved in transcriptional regulation and DNA damage repair. Mutations in KMT2C have been implicated in the progression, metastasis, and drug resistance of multiple cancer types. However, the roles of KMT2C in the regulation of tumor prognosis, immune cell infiltration and the immune microenvironment in these multiple cancer types remain unclear. Therefore, in the present study, data from The Cancer Genome Atlas and Genotype-Tissue Expression databases were used for KMT2C expression analyses. Kaplan-Meier and univariate Cox regression analyses were also performed to investigate the prognostic role of KMT2C. In addition, Gene Set Enrichment Analysis (GSEA) was conducted to study the KMT2C-related signaling pathways. Tumor immune estimation resource 2 and single-sample GSEA were conducted to investigate the correlation between KMT2C expression and immune cell infiltrations, and Spearman's analysis was conducted to study the correlations among KMT2C, tumor mutational burden, microsatellite instability, immune regulators, chemokines and immune receptors. Immunohistochemistry of patient kidney tumor samples was performed to verify the correlation between KMT2C and programmed death-ligand 1 (PD-L1) expression. Finally, RNA interference, wound healing and colony formation assays were conducted to evaluate the effects of KMT2C expression on cell proliferation and metastasis. The results of the present study demonstrated that KMT2C was highly expressed in multiple cancer types, was a protective factor in kidney renal clear cell carcinoma and ovarian serous cystadenocarcinoma, and a risk factor for lung squamous cell carcinoma and uveal melanoma. In addition, KMT2C levels were negatively correlated with immune-activated pathways and the infiltration of immune cells, and positively correlated with inhibitory immune factors and tumor angiogenesis. Patients with low KMT2C expression had higher objective response rates to immunotherapy, and drug sensitivity analysis indicated that topoisomerase, histone deacetylase, DOT1-like histone H3K79 methyltransferase and G9A nuclear histone lysine methyltransferase inhibitors could potentially be used to treat tumors with high KMT2C expression levels. Finally, the KMT2C and PD-L1 expression levels were shown to be positively correlated, and KMT2C knockdown markedly promoted the proliferation and invasion capacities of A549 cells. In conclusion, the present study revealed that low KMT2C expression may be a promising biomarker for predicting the response of patients with cancer to immunotherapy. Conversely, high KMT2C expression was shown to promote tumor angiogenesis, which may contribute to the formation of the immunosuppressive tumor microenvironment.

Exploring aryl hydrocarbon receptor expression and distribution in the tumor microenvironment, with a focus on immune cells, in various solid cancer types.

Aryl hydrocarbon receptor (AhR) is a transcription factor that performs various functions upon ligand activation. Several studies have explored the role of AhR expression in tumor progression and immune surveillance. Nevertheless, investigations on the distribution of AhR expression, specifically in cancer or immune cells in the tumor microenvironment (TME), remain limited. Examining the AhR expression and distribution in the TME is crucial for gaining insights into the mechanism of action of AhR-targeting anticancer agents and their potential as biomarkers. Here, we used multiplexed immunohistochemistry (mIHC) and image cytometry to investigate the AhR expression and distribution in 513 patient samples, of which 292 are patients with one of five solid cancer types. Additionally, we analyzed the nuclear and cytosolic distribution of AhR expression. Our findings reveal that AhR expression was primarily localized in cancer cells, followed by stromal T cells and macrophages. Furthermore, we observed a positive correlation between the nuclear and cytosolic expression of AhR, indicating that the expression of AhR as a biomarker is independent of its localization. Interestingly, the expression patterns of AhR were categorized into three clusters based on the cancer type, with high AhR expression levels being found in regulatory T cells (Tregs) in non-small cell lung cancer (NSCLC). These findings are anticipated to serve as pivotal evidence for the design of clinical trials and the analysis of the anticancer mechanisms of AhR-targeting therapies.

Comparison of Tumor Control and Skin Damage in a Mouse Model after Ultra-High Dose Rate Irradiation and Conventional Irradiation.

Recent studies suggest ultra-high dose rate radiation treatment (UHDR-RT) reduces normal tissue damage compared to conventional radiation treatment (CONV-RT) at the same dose. In this study, we compared first, the kinetics and degree of skin damage in wild-type C57BL/6 mice, and second, tumor treatment efficacy in GL261 and B16F10 dermal tumor models, at the same UHDR-RT and CONV-RT doses. Flank skin of wild-type mice received UHDR-RT or CONV-RT at 25 Gy and 30 Gy. Normal skin damage was tracked by clinical observation to determine the time to moist desquamation, an endpoint which was verified by histopathology. Tumors were inoculated on the right flank of the mice, then received UHDR-RT or CONV-RT at 1 × 11 Gy, 1 × 15, 1 × 25, 3 × 6 and 3 × 8 Gy, and time to tumor tripling volume was determined. Tumors also received 1 × 11, 1 × 15, 3 × 6 and 3 × 8 Gy doses for assessment of CD8+/CD4+ tumor infiltrate and genetic expression 96 h postirradiation. All irradiations of the mouse tumor or flank skin were performed with megavoltage electron beams (10 MeV, 270 Gy/s for UHDR-RT and 9 MeV, 0.12 Gy/s for CONV-RT) delivered via a clinical linear accelerator. Tumor control was statistically equal for similar doses of UHDR-RT and CONV-RT in B16F10 and GL261 murine tumors. There were variable qualitative differences in genetic expression of immune and cell damage-associated pathways between UHDR and CONV irradiated B16F10 tumors. Compared to CONV-RT, UHDR-RT resulted in an increased latent period to skin desquamation after a single 25 Gy dose (7 days longer). Time to moist skin desquamation did not significantly differ between UHDR-RT and CONV-RT after a 30 Gy dose. The histomorphological characteristics of skin damage were similar for UHDR-RT and CONV-RT. These studies demonstrated similar tumor control responses for equivalent single and fractionated radiation doses, with variable difference in expression of tumor progression and immune related gene pathways. There was a modest UHDR-RT skin sparing effect after a 1 × 25 Gy dose but not after a 1 × 30 Gy dose.

Identifying the oncogenic roles of FAP in human cancers based on systematic analysis.

Fibroblast activation protein-α (FAP) is a specific marker of cancer-associated fibroblasts (CAFs) and plays a crucial role in tumor development. However, the biological processes underlying FAP expression in tumor progression and tumor immunity have not been fully elucidated. We utilized RNA-seq data from The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) to perform differential analysis of FAP expression in tumor tissues and matched-normal tissues. The relationship between FAP expression and clinical prognosis, DNA methylation, and tumor-infiltrating immune cells in pan-cancer was assessed using R Studio (version 4.2.1). Additionally, we employed gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) to investigate the biological functions and pathways associated with FAP expression. FAP exhibits high expression in most malignancies, albeit to a lesser extent in CESC, KICH, UCEC, SKCM, THCA, and UCS. Furthermore, FAP is either positively or negatively associated with the prognosis of several malignancies. In seven types of cancer, FAP expression is positively correlated with DNA methylation. CIBERSORT analysis revealed an inverse correlation between FAP expression and T cells, B cells, monocytes, and NK cells, while it exhibited a positive correlation with M0, M1, and M2 macrophages. Enrichment analysis further demonstrated that FAP modulates the cell cycle, epithelial-mesenchymal transition (EMT) process, angiogenesis, and immune-related functions and pathways. Our findings indicate a close relationship between FAP expression and tumorigenesis as well as tumor immunity. FAP has the potential to serve as a diagnostic, prognostic, and immunotherapy marker.

Long non-coding RNA SNHG16 decreased SMAD4 to induce gemcitabine resistance in pancreatic cancer via EZH2-mediated epigenetic modification.

Gemcitabine resistance (GR) in pancreatic cancer (PC) results in poor patient outcomes. SMAD family member (Smad4) dysregulation is a significant role of GR in PC, and EZH2 is involved in Smad4 expression in tumor progression. Interestingly, lncRNA small nucleolar RNA host gene 16 (SNHG16) might interact with EZH2, indicating a potential pathway to overcome gemcitabine-resistant PC progression. We investigated the role of the SNHG16/EZH2/Smad4 pathway in gemcitabine-resistant PC cells (PANC-1/GR and SW1990/GR). First, we found that SNHG16 was upregulated both in wild-type PC cells and in gemcitabine-resistant PC cells. SNHG16 overexpression reduced gemcitabine cytotoxicity and apoptosis in PC cells. Meanwhile, SNHG16 upregulation caused p-Akt elevation and Smad4 reduction. However, SNHG16 silencing induced the opposite trend. Then, we found that EZH2 was enriched in SNHG16 based on RIP and RNA pulldown. In particular, SNHG16 overexpression promoted the interaction between EZH2 and the Smad4 promoter according to Chromatin immunoprecipitation-quantitative polymerase chain reaction. Finally, both EZH2 inhibition and Smad4 upregulation increased gemcitabine cytotoxicity and apoptosis in PC cells during SNHG16 overexpression. Moreover, both treatments decreased p-Akt and increased Smad4. Collectively, lncRNA SNHG16 decreased Smad4 to induce GR in PC via EZH2-mediated epigenetic modification.

MHC heterogeneity and response of metastases to immunotherapy.

In recent years, immunotherapy has proven to be an effective treatment against cancer. Cytotoxic T lymphocytes perform an important role in this anti-tumor immune response, recognizing cancer cells as foreign, through the presentation of tumor antigens by MHC class I molecules. However, tumors and metastases develop escape mechanisms for evading this immunosurveillance and may lose the expression of these polymorphic molecules to become invisible to cytotoxic T lymphocytes. In other situations, they may maintain MHC class I expression and promote immunosuppression of cytotoxic T lymphocytes. Therefore, the analysis of the expression of MHC class I molecules in tumors and metastases is important to elucidate these escape mechanisms. Moreover, it is necessary to determine the molecular mechanisms involved in these alterations to reverse them and recover the expression of MHC class I molecules on tumor cells. This review discusses the role and regulation of MHC class I expression in tumor progression. We focus on altered MHC class I phenotypes present in tumors and metastases, as well as the molecular mechanisms responsible for MHC-I alterations, emphasizing the mechanisms of recovery of the MHC class I molecules expression on cancer cells. The individualized study of the HLA class I phenotype of the tumor and the metastases of each patient will allow choosing the most appropriate immunotherapy treatment based on a personalized medicine.

Effects of NRP1 on angiogenesis and vascular maturity inendothelial cells are dependent on the expression of SEMA4D.

Angiogenesis and vascular maturation play important roles in tumorigenesis and tumor development. The expression of neuropilin 1 (NRP1) is closely associated with angiogenesis in tumors; however, the molecular mechanisms of action in angiogenesis and tumor maturation, as well as the potential clinical value of NRP1 remain unclear. The importance of NRP1 expression in tumor progression was determined using The Cancer Genome Atlas (TCGA) database analysis. Gain- and loss-of-function experiments of NRP1 were performed in vascular endothelial cells (ECs) to investigate the functions in angiogenesis. CCK-8, flow cytometry, Transwell experiments and a series of in vitro experiments were used to detect cell functions. A combination of angiogenesis antibody arrays and RNA-Seq analyses were performed to reveal the proangiogenic mechanisms of action. The function of semaphorin 4D (SEMA4D) was also investigated separately. NRP1 mRNA levels were significantly increased in primary tumors compared with normal tissues based on TCGA data (P<0.01) and were associated with tumor development in patients. Gain- and loss-of-function experiments highlighted the function of NRP1 in promoting EC proliferation, motility and capillary-like tube formation and in reducing apoptosis. NRP1 overexpression led to significantly decreased EC markers (PECAM-1, angiogenin, PIGF and MMP-9) expression levels and reduced the vascular maturity. MAPK7, TPM1, RRBP1, PTPRK, HSP90A, PRKD2, PFKFB3, RGS4 and SPARC were revealed to play important roles in this process. SEMA4D was revealed to be a key protein associated with NRP1 in ECs. These data indicated that NRP1-promoted angiogenesis may be induced at the cost of reducing maturity of the ECs. NRP1 may also be a therapeutic target for antiangiogenic strategies and a candidate prognostic marker for tumors.

CD147 Is a Promising Target of Tumor Progression and a Prognostic Biomarker.

Microenvironment plays a crucial role in tumor development and progression. Cancer cells modulate the tumor microenvironment, which also contribute to resistance to therapy. Identifying biomarkers involved in tumorigenesis and cancer progression represents a great challenge for cancer diagnosis and therapeutic strategy development. CD147 is a glycoprotein involved in the regulation of the tumor microenvironment and cancer progression by several mechanisms—in particular, by the control of glycolysis and also by its well-known ability to induce proteinases leading to matrix degradation, tumor cell invasion, metastasis and angiogenesis. Accumulating evidence has demonstrated the role of CD147 expression in tumor progression and prognosis, suggesting it as a relevant tumor biomarker for cancer diagnosis and prognosis, as well as validating its potential as a promising therapeutic target in cancers.

Abstract 4398: KDM3A and DCLK1 interactions promote stemness and tumorigenesis in PDAC

Abstract Pancreatic ductal adenocarcinoma (PDAC) is the major leading cause of cancer related human death in USA. There is no possible treatment or target available in PDAC. It was proposed that the cancer stem cells (CSCs) can regulates malignancy in PDAC. DCLK1 is one of the quiescent cancer stem cell marker in PDAC that can regulate tumor progression. Identification of the key factors that influence stemness will help to target PDAC. Histone lysine demethylase KDM3A is an enzyme/protein which can regulate stem cell renewability and tumor progression, thereby KDM3A can interact with DCLK1 for tumor progression. KDM3A influences tumor growth and regulates stemness. Therefore, our goal is to find out the the role of KDM3A in tumor progression in PDAC through interactions with DCLK1. We observed expression of KDM3A in both PDAC patients’ samples and cells. Knockdown and overexpression of KDM3A were executed by using lentiviral vector. Tumor progression were also observed in orthotopic mice model. ChIP and RNA seq were performed to validate the data. KDM3A was overexpressed in human PDAC patient tissues and human pancreatic cancer cells with concomitant increase of CSC marker, DCLK1. Moreover, DCLK1 and KDM3A was found to be co-localized in patient tissue samples and identified binding sites of KDM3A with DCLK1 using ChIP-seq. Knockdown of KDM3A abrogates oncogenic potential whereas, overexpressed KDM3A in transformed HPNE cells showed malignant properties with enhanced invasive property, pancosphere formation, foci formation and tumor formation in mouse. Moreover, ChIP-seq and RNA seq suggested that KDM3A regulated DCLK1 expression in tumor progression. In conclusion, co-expression of DCLK1 with KDM3A influenced stemness and enhance tumor progression in PDAC. Citation Format: Chandrayee Ghosh, Kankaraj Palaniyandi, Santanu Paul, Prasad Dandawate, Sonia Rawal, Dharmalingam Subramaniam, Subhash Padhye, Sumedha Gunewardena, Sufi Thomas, Maura O'Neil, Roy Jensen, Danny Welch, Sally Milisky, Scott Weir, Tomoo Iwakuma, Shrikant Anant, Animesh Dhar. KDM3A and DCLK1 interactions promote stemness and tumorigenesis in PDAC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4398.

The prognostic significance of p63 cytoplasmic expression in colorectal cancer. An immunohistochemical study.

Objectives:To evaluate p63 expression pattern in Saudi colorectal cancer (CRC) patients and correlate that with clinicopathological parameters and its role in carcinogenesis and prognosis.Methods:Archival tumor samples were analyzed by immunohistochemistry for p63 expression in 324 consecutive Saudi patients diagnosed with CRC between January 2006 and December 2017 at the Pathology Department of a tertiary care Hospital, Madinah, Saudi Arabia.Results:P63 over-expression was absent in normal mucosa, while 12.5% cases of adenoma showed its over-expression. In CRC, p63 expression was high in 24.1% of cases. There were no significant correlations between p63 expression and gender, tumor location, tumor size, and tumor histologic differentiation. However, high p63 expression revealed a significant correlation with age (p=0.035), tumor type (p=0.004), American Joint Committee on Cancer stage (p=0.046), lymph node metastasis (p=0.006), lymphovascular invasion (p=0.006), distant metastasis (p=0.049) high Ki67 expression (p=0.000) and K-ras expression (p=0.002). The Kaplan-Meier analysis revealed a shorter period of survival with p63 over-expression (p<0.001). The Cox-regression model analysis showed that p63 over-expression was an independent prognostic marker in CRC (p=0.000).Conclusion:P63 expression increased from normal to adenoma to carcinoma sequence. Moreover, p63 cytoplasmic expression seems to be related to high Ki67 indexing, K-ras expression, advanced tumor stage and poor clinical outcome of CRC. These findings suggest a significant role of cytoplasmic p63 expression in tumor progression and prognosis.

MiR-508-3p inhibits cell invasion and epithelial-mesenchymal transition by targeting ZEB1 in triple-negative breast cancer.

Recently, studies have identified that microRNAs (miRNAs) are novel regulators for gene expression in tumor progression including breast cancer. The aim of the study is to investigate the clinical significance and underlying functions between miR-508-3p expression and triple-negative breast cancer (TNBC) development. Quantitative Real-time PCR (QRT-PCR) was performed to determine the expression of miR-508-3p in 54 pairs of TNBC specimens and adjacent non-tumor tissues. The association between miR-508-3p expression and clinicopathological factors was assessed using x2-test. Transwell invasion assays were used to assess cell invasion ability. Luciferase reporter assay, Western blot analyses and qRT-PCR were performed to demonstrate ZEB1 was a direct target of miR-508-3p. We demonstrated that miR-508-3p expression was remarkably decreased in TNBC tissues and cells. Lower miR-508-3p expression significantly associated with lymph node metastasis and distant metastasis in TNBC patients (p < 0.05). Ectopic expression of miR-508-3p significantly suppressed cell invasion ability of TNBC. MiR-508-3p overexpression suppressed cell epithelial-mesenchymal transition (EMT) phenomenon of TNBC by upregulating E-cadherin expression, but downregulating Vimentin expression. In addition, we revealed that ZEB1 was a direct target of miR-508-3p in TNBC cells. MiR-508-3p significantly suppressed cell EMT process by regulating ZEB1 expression. We found that miR-508-3p may be a potential therapeutic target of TNBC.

Lumican and versican protein expression are associated with colorectal adenoma-to-carcinoma progression.

BackgroundOne prominent event associated with colorectal adenoma-to-carcinoma progression is genomic instability. Approximately 85% of colorectal cancer cases exhibit chromosomal instability characterized by accumulation of chromosome copy number aberrations (CNAs). Adenomas with gain of chromosome 8q, 13q, and/or 20q are at high risk of progression to cancer. Tumor progression is also associated with expansion of the extracellular matrix (ECM) and the activation of non-malignant cells within the tumor stroma. The glycoproteins versican and lumican are overexpressed at the mRNA level in colon carcinomas compared to adenomas, and are associated with the formation of tumor stroma.PurposeThe aim of this study was to characterize versican and lumican protein expression in tumor progression and investigate their association with CNAs commonly associated with adenoma-to-carcinoma progression.MethodsTissue microarrays were constructed with colon adenomas and carcinomas that were characterized for MSI-status and DNA copy number gains of chromosomes 8q, 13q and 20q. Sections were immunohistochemically stained for lumican and versican. Protein expression levels were evaluated using digitized slides, and scores were finally dichotomized into a positive or negative score per sample.ResultsLumican and versican expression were both observed in neoplastic cells and in the tumor stroma of colon adenomas and carcinomas. Lumican expression was more frequently present in epithelial cells of carcinomas than adenomas (49% versus 18%; P = 0.0001) and in high-risk adenomas and carcinomas combined compared to low-risk adenomas (43% versus 16%; P = 0.005). Versican staining in the tumor stroma was more often present in high-risk adenomas combined with carcinomas compared to low-risk adenomas (57% versus 36%; P = 0.03) and was associated with the presence of gain of 13q (71% versus 44%; P = 0.04).ConclusionEpithelial lumican and stromal versican protein expression are increased during colorectal adenoma-to-carcinoma progression.

DPPIV/CD26: a tumor suppressor or a marker of malignancy?

Dipeptidyl peptidase IV (DPPIV/CD26) is a multifunctional protein with intrinsic peptidase activity that inactivates or degrades some bioactive peptides. It is the main cellular binding protein for ecto-adenosine deaminase and interacts with extracellular matrix proteins, besides participating in different signaling pathways. Due to these multiple functions, DPPIV/CD26 has been shown to be closely related to the tumor process. It has been reported that the progression of certain types of cancer is accompanied by a decrease in DPPIV/CD26 expression, and studies have shown that the malignant phenotype can be reverted when DPPIV/CD26 expression is induced in these cancer cells, characterizing this protein as a tumor suppressor. On the other hand, DPPIV/CD26 was described as a protein associated with invasion and metastatic spread, characterizing it as a marker of malignancy. Thus, this review explores the roles of DPPIV/CD26 expression in tumor progression in different types of cancer and demonstrates the importance of this protein as a promising therapeutic target and tumor biomarker.

Potential Involvement of Jagged1 in Metastatic Progression of Human Breast Carcinomas.

Jagged1, the ligand of Notch, has been shown to be involved in formation of bone metastases in an experimental study. Here, clinical relevance of Jagged1 expression in tumor progression was assessed in human breast carcinomas. Jagged1 expression was evaluated by immunohistochemistry in 228 tumor tissue samples and compared to clinicopathologic parameters and patients' outcomes. Furthermore, circulating tumor cells (CTCs) from peripheral blood of 100 unmatched metastatic cancer patients with progressive disease were enriched using Ficoll density gradient centrifugation and detected by pan-keratin/Jagged1/CD45 immunofluorescent staining. Jagged1 expression was detected in 50% of 228 tumors. Jagged1 expression was correlated with higher tumor grade (P = 0.047), vascular invasion (P = 0.026), luminal B subtype (P = 0.016), overexpression of Her-2 (P = 0.001), high Ki-67 expression (P = 0.035), and aldehyde dehydrogenase 1 (ALDH1) positivity (P = 0.013). Jagged 1 expression indicated shorter disease-free survival (DFS) (P = 0.040) and metastasis-free survival (P = 0.048) in lymph node-negative breast cancer for which it was the only independent predictor of DFS (multivariate analysis, P = 0.046). Tumors characterized by the strongest Jagged1 staining intensity (7.5% of cases) correlated with lymph node positivity (P = 0.037), metastatic relapse (P = 0.049), and higher number of disseminated tumor cells in bone marrow aspirates (P = 0.041). Twenty-one unmatched metastatic breast cancer patients with progressive disease were positive for CTCs, and 85.7% of the CTCs also expressed Jagged1. The presence of Jagged1(+) CTCs was significantly associated with shorter progression-free survival in patients treated with bisphosphonates (P = 0.013). Jagged1 expression characterizes more aggressive breast carcinoma and might be involved in tumor cell dissemination, metastatic progression, and resistance to bone-targeting therapy in breast cancer patients.

MMP3-mediated tumor progression is controlled transcriptionally by a novel IRF8-MMP3 interaction.

Interferon regulatory factor-8 (IRF8), originally identified as a leukemic tumor suppressor, can also exert anti-neoplastic activities in solid tumors. We previously showed that IRF8-loss enhanced tumor growth, which was accompanied by reduced tumor-cell susceptibility to apoptosis. However, the impact of IRF8 expression on tumor growth could not be explained solely by its effects on regulating apoptotic response. Exploratory gene expression profiling further revealed an inverse relationship between IRF8 and MMP3 expression, implying additional intrinsic mechanisms by which IRF8 modulated neoplastic behavior. Although MMP3 expression was originally linked to tumor initiation, the role of MMP3 beyond this stage has remained unclear. Therefore, we hypothesized that MMP3 governed later stages of disease, including progression to metastasis, and did so through a novel IRF8-MMP3 axis. Altogether, we showed an inverse mechanistic relationship between IRF8 and MMP3 expression in tumor progression. Importantly, the growth advantage due to IRF8-loss was significantly compromised after silencing MMP3 expression. Moreover, MMP3-loss reduced spontaneous lung metastasis in an orthotopic mouse model of mammary carcinoma. MMP3 acted, in part, in a cell-intrinsic manner and served as a direct transcriptional target of IRF8. Thus, we identified a novel role of an IRF8-MMP3 axis in tumor progression, which unveils new therapeutic opportunities.

P8.08 - Roles of Nectin-4 in tumor progression

ABSTRACT Nectin-4 is a transmembrane immunoglobulin weakly expressed in healthy tissues, in contrast with embryonic stage. Interestingly, an expression of nectin-4 is found in the breast, ovarian and lung tumors. This expression is correlated to the advanced stages of these cancerous pathologies. Furthermore, high levels of plasmatic form is also found at an important rate in the advanced stages of breast, ovarian and lung carcinomas. To identify the roles of nectin-4 expression in tumor progression, we used in our study MCF-7 & BT-474 cell lines expressing an endogenous nectin-4, but also MDA-MB213 cell lines transfected with native forms (MDA-MB213 R4. C1) or with a form of nectin-4 truncate of eight amino acids at juxta-membrane domain (MDA-MB231 Del8.6). We observed that the MDA-MB231 R4.C1 and MDA-MB231 Del8.6 cells, have high growth capacities. But this growth seems more important in drastic medium conditions, only for MDA-MB231 R4.C1 cells that produce a soluble form. Using monoclonal antibodies targeting the extracellular domain of nectin-4, we observe significant diminution of cell growth for BT-474 & MCF-7 cell lines. Similar results are obtained by using a specific siRNA. An implication of nectin-4 in the increase of the cellular migration is also observed. Indeed, cells expressed nectin-4 show a migration capability higher than about 66% compared with MDA-MB231 wt cells and it seems that the soluble form does not influence the intensity of cell migration.The use of a monoclonal antibody allowed to reduce of 23% the migration of MDA-MB231 R4.C1 cells. The cell clonogenicity test, demonstrate an effect on MDA-MB-231 R4.C1 cells compared with MDA-MB231 wt & MDA-MB231 Del8.6 cells. On MDA-MB231 wt & MDA-MB231 R4.C1 xenograft NOG mouse, the expression of nectin-4 increase significantly tumoral volume as well as the metastatic power. The use of anti-nectin-4 antibody does not affect the tumor growth, but seems to decrease the metastatic potential. These results, demonstrate that the cell surface expression of nectin-4 but also the soluble form are involved on tumorigenicity, by increasing proliferative and invasive potential. The monoclonal antibody anti-nectin-4 which we developed, can represent a promising therapeutic approach.

Correlation between SPARC (Osteonectin) expression with immunophenotypical and invasion characteristics of pituitary adenomas.

Pituitary adenomas are the third most common intracranial tumors. Invasive adenomas account for only 0.1-0.2% of pituitary tumors. SPARC is a matrix glycoprotein that plays a role in progression and invasiveness of neoplasms. In this study, we examined the potential role of SPARC in invasive pituitary adenomas. Forty pituitary adenomas have been examined with histopathological and immunohistochemical techniques. The cohort has been classified into two groups as invasive (n=25) and non-invasive (n=15) utilizing the Hardy classification. Formalin fixed tissues have been stained with hematoxylin eosin. Ki-67, p53, and SPARC monoclonal antibodies have been used. We did not detect any significant difference on Ki-67, SPARC, and p53 expression patterns correlating with the pathological subtype or invasiveness. Only 24% of invasive adenomas had Ki-67 levels over 1%. A total of 67.7% non-invasive adenomas had Ki-67 levels below 1%. We did not detect any relation between SPARC levels and invasiveness of pituitary adenomas. Absence of significant SPARC expression in tumor progression, sellar dilatation, erosion and destruction suggest that SPARC scores are not related with invasiveness or progressiveness of pituitary adenomas.

Abstract 2798: Targeting IL13R alpha2 heterogeneity in malignant glioma with engineered T cells

Abstract A majority of malignant glioma exclusively expresses Interleukin 13 receptor alpha chain variant 2 (IL13Rα2), making it an ideal candidate for targeted immunotherapy. A number of immunotherapeutic molecules have been developed against IL13Rα2 with varied levels of therapeutic efficacy. However, the importance of IL13Rα2 as a successful immunotherapy candidate is under scrutiny due to its heterogeneous expression on the tumor. We have used an IL13-chimeric antigen receptor (IL13CAR) expressing T cells to target the IL13Rα2 heterogeneity on malignant glioma. IL13Rα2 was over-expressed on U251MG human glioma cell line by stable transfection (IL13Rα2Hi) to evaluate the effects of its increased expression in tumor progression and immunotherapeutic efficacy, compared to the regular U251 cells (IL13Rα2Lo) and IL13Rα2-knocked down U251 cells (IL13Rα2KD). Tumor cells readily formed neurospheres under established culture conditions. Increased expression of IL13Rα2 was observed in IL13Rα2Hi-neurosphere derived tumor cells when compared to those derived from IL13Rα2Lo neurospheres, as well as the parental IL13Rα2Hi cells. IL13Rα2Hi-neurosphere derived tumor cells were also resistant to radiation therapy and temozolomide chemotherapy when compared to those IL13Rα2Lo neurospheres-derived tumor cells, as well as parental cell lines. Higher levels of IFN gamma and TNF alpha in the culture supernatants, increased T cell proliferation, and CAR+ TCR enrichment were observed when IL13CAR T cells were co-cultured with IL13Rα2Hi-neurosphere derived tumor cells, in comparison to co-culture with IL13Rα2Lo -neurosphere derived tumor cells, as well as the parental cells. IL13CAR T cells, upon co-culture with IL13Rα2Hi-neurosphere derived tumor cells exerted highest levels of cytotoxicity. Together, these results predict that IL13Rα2 heterogeneity can be addressed by a combination of established therapeutic regimen and targeted immunotherapy. While those cells expressing lower levels of IL13Rα2 are susceptible to radiation and chemotherapy, the high IL13Rα2-expressing tumor cells that escape these treatments can be candidates for IL13Rα2-targeted immunotherapy. Note: This abstract was not presented at the meeting. Citation Format: Sadhak Sengupta, Irina Balyasnikova, Seema Naheed, Maciej S. Lesniak, Richard P. Junghans, Prakash Sampath. Targeting IL13R alpha2 heterogeneity in malignant glioma with engineered T cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2798. doi:10.1158/1538-7445.AM2014-2798

Loss of ARID1A Expression Is an Early Molecular Event in Tumor Progression From Ovarian Endometriotic Cyst to Clear Cell and Endometrioid Carcinoma

ARID1A is a recently identified tumor suppressor participating in chromatin remodeling. Somatic inactivating mutations of ARID1A and loss of its expression occur frequently in ovarian clear cell and endometrioid carcinomas and in uterine endometrioid carcinomas. Because endometriotic epithelium is thought to be the cell of origin of most ovarian clear cell and endometrioid carcinomas, we undertook an analysis of ARID1A expression of these tumors arising within an endometriotic cyst (endometrioma). Our immunohistochemical study set consisted of 47 endometriotic cysts containing clear cell carcinoma in 24 cases, well-differentiated ovarian endometrioid carcinoma in 20 cases, and mixed clear cell and endometrioid carcinoma in 3 cases. ARID1A loss was observed in 31 (66%) of 47 carcinomas; and therefore, these cases were informative for determining the temporal sequence of loss of ARID1A expression in tumor progression. In 16 of the 47 cases, ARID1A immunoreactivity was retained in both the endometriotic cyst and the carcinoma; and thus, these cases were not informative. All of the 31 informative cases showed loss of ARID1A immunoreactivity in the carcinoma and in the endometriotic cyst epithelium in direct continuity with the carcinoma but not in the cyst epithelium that was not adjacent to the tumor. Loss of ARID1A function as shown by loss of expression, presumably due to mutations, is an early molecular event in the development of most ovarian clear cell and endometrioid carcinomas arising in endometriomas.

Abstract 1104: Nuclear translocalization of DICER correlates with high-risk clinical/histologic features in melanoma

Abstract Melanoma is an aggressive malignancy with a relatively high metastatic rate, responsible for almost 60% of lethal skin tumors. Despite numerous efforts, there is still a need to better understand the molecular pathways involved in malignant melanoma metastasis. In recent years, there have been several studies highlighting the role of deregulated microRNAs (miRNAs) expression in tumor progression and metastasis. DICER is a central regulator of miRNA maturation and recently, has been shown to be upregulated in cutaneous melanoma at both protein and mRNA level. Arur et al. [1], reported DICER to be one of the ERK substrate, and this activation results in its nuclear translocalization (personal communication). However, it is unknown whether differential cellular expression of DICER contributes to the pathogenesis of melanoma. In order to test the hypothesis that aberrant nucleo-cytoplasmic expression of DICER plays a significant role in melanoma progression, we have performed semiquantitative immunohistochemical analysis of DICER in benign nevi (n=17), primary melanomas without metastasis (n=38) and with metastasis (n=27), and melanoma metastases (n=34 cases). A significant positive correlation was observed between the percentage of positive cells (p=0.003) and staining intensity (p=0.01) for nuclear DICER with progression from benign nevi to primary melanoma. Interestingly, primaries without metastasis had significantly higher levels of DICER immunopositivity in tumor cells than primaries with metastasis (p=0.02). Furthermore, metastatic melanomas showed significantly larger number of cells having intense immunoreactivity for DICER protein in nucleus than the primary melanomas (p=0.02). Significantly larger percentage of tumor cells showing DICER expression in the nucleus were observed in those lesions having ulceration (p=0.007) and vertical growth phase, which are well-established markers of poor clinical outcomes in patients with cutaneous melanoma. On the other hand, lesions showing no vascular invasion expressed higher levels of DICER in the cytoplasm (p&amp;lt;0.0001). Disease-specific survival was significantly shorter in patients with increased nuclear expression of DICER (p=0.04). Thus, differential cellular localization of miRNA-regulating DICER protein correlates with disease progression and high-risk clinical and histologic features in melanoma. Therefore, it will be interesting to further delineate the role of nuclear DICER in melanoma.