Celecoxib-Induced Modulation of Colon Cancer CD133 Expression Occurs through AKT Inhibition and Is Monitored by 89Zr Immuno-PET

We developed an immuno-PET technique that monitors modulation of tumor CD133 expression, which is required for the success of CD133-targeted therapies. Methods. Anti-CD133 antibodies were subjected to sulfhydryl moiety-specific 89Zr conjugation. 89Zr-CD133 IgG was evaluated for specific activity and radiolabel stability. Colon cancer cells underwent binding assays and Western blotting. Biodistribution and PET studies were performed in mice. Results. 89Zr-CD133 IgG showed excellent target specificity with 97.2 ± 0.7 % blocking of HT29 cell binding by an excess antibody. Intravenous 89Zr-CD133 IgG followed biexponential blood clearance and showed CD133-specific uptake in HT29 tumors. 89Zr-CD133 IgG PET/CT and biodistribution studies confirmed high HT29 tumor uptake with lower activities in the blood and normal organs. In HT29 cells, celecoxib dose-dependently decreased CD133 expression and 89Zr-CD133 IgG binding that reached 19.9 ± 2.1 % ( P < 0.005 ) and 50.3 ± 10.9 % ( P < 0.001 ) of baseline levels by 50 μM, respectively. Celecoxib treatment of mice significantly suppressed tumor CD133 expression to 67.5 ± 7.8 % of controls ( P < 0.005 ) and reduced tumor 89Zr-CD133 IgG uptake from 15.5 ± 1.4 % at baseline to 12.3 ± 2.0 % ID / g ( P < 0.01 ). Celecoxib-induced CD133 reduction in HT29 cells and tumors was associated with substantial suppression of AKT activation. There were also reduced HIF-1α accumulation and IκBα/NFκB phosphorylation. Conclusion. 89Zr-CD133 IgG PET provides high-contrast tumor imaging and monitors celecoxib treatment-induced modulation of tumor CD133 expression, which was found to occur through AKT inhibition. This technique may thus be useful for screening drugs that can effectively suppress colon cancer stem cells.

LncRNA TCL6 Contributes to CSC-like Properties via Modulating TP53 in HCC

BackgroundHepatocellular carcinoma (HCC) is the most common malignant tumors, accounting for most of the adult primary liver cancer. Herein, we aimed to analyze the expression of long non-coding RNA-T cell leukemia/lymphoma 6 (lncRNA-TCL6) in HCC and elucidate its mechanism involved in the HCC progression. Methodse performed RNA extraction and quantitative real-time polymerase chain reaction assays, spheroid formation assays, flow cytometry and western blot assays to assess the effect of TCL6 on the liver CSCs marker CD133 expression rate, sphere-forming ability of liver stem cells, and the relationship between TCL6 expression and stem cell factor (TP53, P21, CD44, KLF4, OCT4, Nanog, and Sox2). In addition, we used a dual luciferase assay to verify the relationship between miR-106a-5p and TP53.ResultsKnockdown of TCL6 expression significantly improved the CD133 expression rate and the liver stem cells sphere-forming ability in HCC, while TCL6 overexpression in HCC showed the opposite effect. Knockdown of TCL6 upregulated the KLF4mRNA expression, while TCL6 overexpression in HCC inhibited the TP53 and CDKN1A expression. Western blot assays showed that TCL6 expression was positively correlated with TP53 and P21, while negatively correlated with stem cell factor. Dual luciferase assay showed that TP53 was a target of miR-106a-5p.ConclusionResults suggested that reprogramming-related TCL6 may be a novel tumor suppressor gene in HCC, which inhibits the self-renewal of liver CSCs, in part by promoting the TP53 expression.

CD133 Expression in the Nucleus Is Associated with Endometrial Carcinoma Staging and Tumor Angioinvasion

Background: (1) Endometrial cancer is one of the most common cancers affecting women, with a growing incidence. To better understand the different behaviors associated with endometrial cancer, it is necessary to understand the changes that occur at a molecular level. CD133 is one of the factors that regulate tumor progression, which is primarily known as the transmembrane glycoprotein associated with tumor progression or cancer stem cells. The aim of our study was to assess the impact of subcellular CD133 expression on the clinical course of endometrial cancer. (2) Methods: CD133 expression in the plasma membrane, nucleus, and cytoplasm was assessed by immunohistochemical staining in a group of 64 patients with endometrial cancer representing FIGO I-IV stages, grades 1–3 and accounting for tumor angioinvasion. (3) Results: Nuclear localization of CD133 expression was increased in FIGO IB-IV stages compared to FIGO IA. Furthermore, CD133 expression in the nucleus and plasma membrane is positively and negatively associated with a higher grade of endometrial cancer and angioinvasion, respectively. (4) Conclusions: Our findings suggest that positive nuclear CD133 expression in the tumor may be related to a less favorable prognosis of endometrial carcinoma patients and has emerged as a useful biomarker of a high-risk group.

A pan-cancer perspective analysis reveals the opposite prognostic significance of CD133 in lower grade glioma and papillary renal cell carcinoma

CD133 is a valuable prognostic marker in multiple types of cancer. However, the expression, methylation levels, and prognostic relevance of CD133 have not been evaluated in a pan-cancer perspective. The expression and methylation levels of CD133 across different types of cancer were determined using The Cancer Genome Atlas (TCGA) dataset. Univariate cox regression and Kaplan-Meier survival were used to determine the prognostic significance of CD133 expression and methylation. CD133 was highly expressed in papillary renal cell carcinoma (PRCC) or pancreatic adenocarcinoma (PAAD). Correspondingly, PAAD and PRCC had low CD133 methylation levels. Through pan-cancer perspective analysis, we found that CD133 high expression was a poor prognostic factor in lower grade glioma (LGG), while, CD133 high expression was a good prognostic factor in PRCC. Moreover, genes positively correlated with CD133 expression were associated with the poor clinical outcomes of LGG. In PRCC, genes negatively correlated with CD133 expression were correlated with the poor overall survival. Furthermore, CD133 expression levels were highly correlated with the CD133 methylation levels in LGG or PRCC. Correspondingly, CD133 hypermethylation was a good prognostic factor in LGG. On the contrary, CD133 hypomethylation was a good prognostic factor in PRCC. We also found that CD133 was highly expressed and hypomethylated in wild type IDH subgroup of LGG. CD133 was highly expressed and hypomethylated in low stages and type1 of PRCC. CD133 high expression and hypomethylation were bad prognostic factors in LGG, while, CD133 high expression and hypomethylation were good prognostic factors in PRCC.

An Fc-Optimized CD133 Antibody for Induction of NK Cell Reactivity against B Cell Acute Lymphoblastic Leukemia

In recent decades, antibody-dependent cellular cytotoxicity (ADCC)-inducing monoclonal antibodies (mAbs) have revolutionized cancer immunotherapy, and Fc engineering strategies have been utilized to further improve efficacy. A promising option is to enhance the affinity of an antibody’s Fc-part to the Fc-receptor CD16 by altering the amino acid sequence. Herein, we characterized an S239D/I332E-modified CD133 mAb termed 293C3-SDIE for treatment of B cell acute lymphoblastic leukemia (B-ALL). Flow cytometric analysis revealed CD133 expression on B-ALL cell lines and leukemic cells of 50% (14 of 28) B-ALL patients. 293C3-SDIE potently induced NK cell reactivity against the B-ALL cell lines SEM and RS4;11, as well as leukemic cells of B-ALL patients in a target antigen-dependent manner, as revealed by analysis of NK cell activation, degranulation, and cytotoxicity. Of note, CD133 expression did not correlate with BCR-ABL, CD19, CD20, or CD22, which are presently used as therapeutic targets in B-ALL, which revealed CD133 as an independent target for B-ALL treatment. Increased CD133 expression was also observed in MLL-AF4-rearranged B-ALL, indicating that 293C3-SDIE may constitute a particularly suitable treatment option in this hard-to-treat subpopulation. Taken together, our results identify 293C3-SDIE as a promising therapeutic agent for the treatment of B-ALL.

Correlation of CD133 and SOX2 Expression with Regional Lymph Nodes Metastatic Status in Invasive Breast Carcinoma of No Special Type

Background: CD133 overexpression can increase cell proliferation, migration, and epithelialmesenchymal transition that promotes metastasis. CD133 expression is induced by hypoxiainduced factor (HIF) which requires SOX2 binding in the promoter region. SOX2 is an embryonal transcription factor that plays a role in the development of malignancy. The study aimed to analyze the expression of CD133 and SOX2 with regional lymph nodes (LN) metastatic status in invasive breast carcinoma of no special type (NST). Methods: The study was a cross-sectional design. Forty-five samples were retrieved from pathology archives in Dr. Soetomo Hospital from January to December 2018. Samples were divided into 2 groups, with and without regional LN metastasis. Immunohistochemistry with CD133 and SOX2 was applied to all samples. CD133 expression was assessed by immunoreactive score, and SOX2 expression was assessed by the percentage of tumor positive cells.Results: There was no significant difference in CD 133 expression between invasive breast carcinoma of NST with and without regional LN metastases (P = .990). A positive correlation was found in SOX2 expression between breast carcinoma with and without regional LN metastasis (P = .000; rs = .518). There was no correlation between CD133 and SOX2 expression (P = .082), which means that the high expression of CD133 did not affect SOX2 expression.Conclusions: CD133 expression was not significantly different in breast carcinoma with and without LN metastasis. The high expression of SOX2 was found significantly correlated with regional LN metastasis. SOX2 expression may become a potential prognostic marker in invasive breast carcinoma of NST. regional LN metastasis