Research IndicatorsGraph generated 27 February 2015 using data from PubMed using criteria.
Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic. Tag cloud generated 27 February, 2015 using data from PubMed, MeSH and CancerIndex
Specific Cancers (4)
Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.
Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).
OMIM, Johns Hopkin University
Referenced article focusing on the relationship between phenotype and genotype.
Cancer Genome Anatomy Project, NCI
COSMIC, Sanger Institute
Somatic mutation information and related details
Search the Epigenomics database and view relevant gene tracks of samples.
Latest Publications: CD59 (cancer-related)
Rösner T, Lohse S, Peipp M, et al.Epidermal growth factor receptor targeting IgG3 triggers complement-mediated lysis of decay-accelerating factor expressing tumor cells through the alternative pathway amplification loop.
J Immunol. 2014; 193(3):1485-95 [PubMed
] Related Publications
Binding of C1q to target-bound IgG initiates complement-mediated lysis (CML) of pathogens, as well as of malignant or apoptotic cells, and thus constitutes an integral part of the innate immune system. Despite its prominent molecular flexibility and higher C1q binding affinity compared with human IgG1, IgG3 does not consistently promote superior CML. Hence the aim of this study was to investigate underlying molecular mechanisms of IgG1- and IgG3-driven complement activation using isotype variants of the therapeutic epidermal growth factor receptor (EGFR) Ab cetuximab. Both IgG1 and IgG3 Abs demonstrated similar EGFR binding and similar efficiency in Fab-mediated effector mechanisms. Whereas anti-EGFR-IgG1 did not promote CML of investigated target cells, anti-EGFR-IgG3 triggered significant CML of some, but not all tested cell lines. CML triggered by anti-EGFR-IgG3 negatively correlated with expression levels of the membrane-bound complement regulatory proteins CD55 and CD59, but not CD46. Notably, anti-EGFR-IgG3 promoted strong C1q and C3b, but relatively low C4b and C5b-9 deposition on analyzed cell lines. Furthermore, anti-EGFR-IgG3 triggered C4a release on all cells but failed to induce C3a and C5a release on CD55/CD59 highly expressing cells. RNA interference-induced knockdown or overexpression of membrane-bound complement regulatory proteins revealed CD55 expression to be a pivotal determinant of anti-EGFR-IgG3-triggered CML and to force a switch from classical complement pathway activation to C1q-dependent alternative pathway amplification. Together, these data suggest human anti-EGFR-IgG3, although highly reactive with C1q, to weakly promote assembly of the classical C3 convertase that is further suppressed in the presence of CD55, forcing human IgG3 to act mainly through the alternative pathway.
Sebejova L, Borsky M, Jaskova Z, et al.Distinct in vitro sensitivity of p53-mutated and ATM-mutated chronic lymphocytic leukemia cells to ofatumumab and rituximab.
Exp Hematol. 2014; 42(10):867-74.e1 [PubMed
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Abnormalities in ATM and TP53 genes represent important predictive factors in chronic lymphocytic leukemia (CLL); however, the efficacy of CD20 targeting immunotherapy is only poorly defined in the affected patients. Therefore, we tested the in vitro response to ofatumumab (OFA) and rituximab (RTX) in 75 CLL samples with clearly defined p53 or ATM inactivation. Using standard conditions allowing complement-dependent cytotoxicity, i.e., 10 μg/mL of antibodies and 20% active human serum, we observed clear differences among the tested genetic categories: ATM-mutated samples (n = 17) represented the most sensitive, wild-type samples (n = 31) intermediate, and TP53-mutated samples (n = 27) the most resistant group (ATM-mut vs. TP53-mut: P = 0.0005 for OFA and P = 0.01 for RTX). The response correlated with distinct levels of CD20 and critical complement inhibitors CD55 and CD59; CD20 level median was the highest in ATM-mutated and the lowest in TP53-mutated samples (difference between the groups P < 0.01), while the total level of complement inhibitors (CD55 plus CD59) was distributed in the opposite manner (P < 0.01). Negligible response to both OFA and RTX was noted in all cultures (n = 10) tested in the absence of active serum, which strongly indicated that complement-dependent cytotoxicity was a principal cell death mechanism. Our study shows that (1) common genetic defects in CLL cells significantly impact a primary response to anti-CD20 monoclonal antibodies and (2) ATM-mutated patients with currently poor prognosis may potentially benefit from immunotherapy targeting CD20.
Kesselring R, Thiel A, Pries R, et al.The complement receptors CD46, CD55 and CD59 are regulated by the tumour microenvironment of head and neck cancer to facilitate escape of complement attack.
Eur J Cancer. 2014; 50(12):2152-61 [PubMed
] Related Publications
BACKGROUND: Membrane-bound complement restriction proteins (mCRPs) CD46, CD55 and CD59 enable tumour cells to evade complement dependent cytotoxicity and antibody-dependent killing mechanisms. But less is known about the role of these mCRPs in head and neck cancer.
METHODS: In this study we determined the expression of the mCRPs on head and neck squamous cell carcinoma (HNSCC) cell lines, on tumour tissue and TDLNs (tumour-draining lymph nodes) as well as on lymphocytes from HNSCC patients. The influence of the HNSCC microenvironment on the mCRP regulation was analysed using Flow Cytometry, Western blotting and small interfering RNAs (siRNA) transfection studies.
RESULTS: We examined the effects of the HNSCC tumour milieu on the expression levels of CD46, CD55 and CD59. We investigated the susceptibility of HNSCC cells to CDC (complement-dependent cytotoxicity) while silencing the mCRPs. Our results demonstrate a huge influence of the HNSCC tumour microenvironment on the regulation of mCRP expression and show a reciprocal regulation between the different mCRPs themselves.
CONCLUSIONS: In summary, our data indicate that HNSCC has evolved different strategies to evade complement attacks and that the tumour microenvironment leads to the enhancement of complement resistance of the surrounding tissue.
Modlin IM, Drozdov I, Kidd MGut neuroendocrine tumor blood qPCR fingerprint assay: characteristics and reproducibility.
Clin Chem Lab Med. 2014; 52(3):419-29 [PubMed
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BACKGROUND: We have developed a PCR-based tool that measures a 51-gene panel for identification of gastroenteropancreatic (GEP) neuroendocrine neoplasms (NENs) in peripheral blood. This manuscript assesses the robustness (performance metrics) of this tool with a specific focus on the effects of individual parameters including collection, storage, acid suppressive medication [proton pump inhibitor (PPI)], age, sex, race and food on accuracy.
METHODS: Performance metrics were evaluated using a gold standard (mRNA derived from three individual human neuroendocrine tumor cell lines) and clinical samples using qPCR.
RESULTS: One hundred percent of the 51 transcripts were amplified in the gold standard (NEN cell line-derived mRNA) (CQ<35, average efficiency 1.94). The inter- and intra-assay variations were 1%-2%. In clinical samples, 50 of 51 targets (98%) were amplified. The inter- and intra-assay reproducibility ranged between 0.4% and 1.2%. The coefficient of variation (CV) was 5.3%. Expression of the reference gene, ALG9, was robust [low variation, low M-value, high (99.5%) PCR efficiency] and unaffected by sample processing. Test meals, long-term PPI use (>1 year), age, sex and ethnicity had no effect on the signature. Expression of two genes, ALP2 and CD59 correlated strongly with RNA integrity (R=0.72, p<0.001) and could be used to assess storage and processing.
CONCLUSIONS: The 51 marker gene signature was robust and reproducible, exhibiting acceptable inter- and intra-assay metrics (<5%). Feeding, PPI intake, age, sex and ethnicity do not affect the signature. Expression levels of APLP2 and CD59 are effective surrogate markers of proper sample collection and processing.
Li B, Lin H, Fan J, et al.CD59 is overexpressed in human lung cancer and regulates apoptosis of human lung cancer cells.
Int J Oncol. 2013; 43(3):850-8 [PubMed
] Related Publications
CD59, belonging to membrane complement regulatory proteins (mCRPs), inhibits the cytolytic activity of complement and is overexpressed in many types of solid cancers. The aim of the present study was to detect the expression of CD59 in non-small cell lung cancer (NSCLC) and to investigate the relationship between decreased CD59 expression and tumorigenesis of NSCLC by transfecting recombinant retrovirus encoding shRNA targeting human CD59 into the human NSCLC cell line NCI-H157. CD59 expression in NSCLC was detected by immunocytochemistry (IHC). In the human NSCLC cell line NCI-H157, CD59 mRNA and protein expression suppressed with lentivirus-mediated RNAi was confirmed by using RT-PCR and western blotting, respectively. The proliferation and apoptosis of NCI-H157 cells was measured by using MTT assay and FACS. The resistance to complement cracking ability was detected by LDH assay. Caspase-3 expression in cells was assessed by IHC. Bcl-2 and Fas protein was determined by western blotting both in vitro and in vivo. CD59 is overexpressed in human NLCLC cancer. In NCI-H157 cells, lentivirus-mediated RNAi significantly reduced both CD59 mRNA and protein expression, which resulted in suppressing cell proliferation and increasing cell apoptosis. When incubated with fresh normal human serum (8%, v/v) for 1 h at 37˚C, the cell viability was decreased and cell apoptosis was increased in siCD59-infected NCI-H157 cells compared to siCD59-C-infected cells. Reduced CD59 expression led to increased expression of caspase-3 and Fas and decreased expression of Bcl-2. Furthermore, the nude mouse tumor graft weight was significantly decreased and survival rate was significantly increased in the siCD59 group. CD59 is overexpressed in human NLCLC. CD59 silencing in NSCLC cancer cells via retrovirus-mediated RNAi can enhance complement-mediated cell apoptosis, inhibiting the growth of NSCLC. CD59 may serve as a potential target for gene therapy in NSCLC.
Cellular heterogeneity of solid tumors represents a common problem in mass spectrometry (MS)-based analysis of tissue specimens. Combining immuno-laser capture microdissection (iLCM) and mass spectrometry (MS) provides a means to study proteins that are specific for pure cell subpopulations in complex tissues. CD24, as a cell surface marker for detecting pancreatic cancer stem cells (CSCs), is directly correlated with the development and metastasis of pancreatic cancer. Herein, we describe an in-depth proteomic profiling of frozen pancreatic CD24(+) adenocarcinoma cells from early stage tumors using iLCM and LC-MS/MS and a comparison with CD24(-) cells dissected from patient-matched adjacent normal tissues. Approximately 40 nL of tissue was procured from each specimen and subjected to tandem MS analysis in triplicate. A total of 2665 proteins were identified, with 375 proteins in common that were significantly differentially expressed in CD24(+) versus CD24(-) cells by at least a 2-fold change. The major groups of the differentially overexpressed proteins are involved in promoting tumor cell migration and invasion, immune escape, and tumor progression. Three selected candidates relevant to mediating immune escape, CD59, CD70, and CD74, and a tumor promoter, TGFBI, were further validated by immunohistochemistry analysis on tissue microarrays. These proteins showed significantly increased expression in a large group of clinical pancreatic adenocarcinomas but were negative in all normal pancreas samples. The significant coexpression of these proteins with CD24 suggests that they may play important roles in the progression of pancreatic cancer and could serve as promising prognosis markers and novel therapeutic targets for this deadly disease.
Sahu B, Laakso M, Pihlajamaa P, et al.FoxA1 specifies unique androgen and glucocorticoid receptor binding events in prostate cancer cells.
Cancer Res. 2013; 73(5):1570-80 [PubMed
] Related Publications
The forkhead protein FoxA1 has functions other than a pioneer factor, in that its depletion brings about a significant redistribution in the androgen receptor (AR) and glucocorticoid receptor (GR) cistromes. In this study, we found a novel function for FoxA1 in defining the cell-type specificity of AR- and GR-binding events in a distinct fashion, namely, for AR in LNCaP-1F5 cells and for GR in VCaP cells. We also found different, cell-type and receptor-specific compilations of cis-elements enriched adjacent to the AR- and GR-binding sites. The AR pathway is central in prostate cancer biology, but the role of GR is poorly known. We find that AR and GR cistromes and transcription programs exhibit significant overlap, and GR regulates a large number of genes considered to be AR pathway-specific. This raises questions about the role of GR in maintaining the AR pathway under androgen-deprived conditions in castration-resistant prostate cancer patients. However, in the presence of androgen, ligand-occupied GR acts as a partial antiandrogen and attenuates the AR-dependent transcription program. .
Kohler H, Rector K, Amick JGeneration of a potent recombinant homophilic chimeric anti-CD20 antibody.
Hybridoma (Larchmt). 2012; 31(6):395-402 [PubMed
] Related Publications
Previously we increased the potency of therapeutic antibodies in targeting, induction of apoptosis, and growth inhibition in vitro and in vivo by chemically conjugating a homophilic peptide to the antibody. Here, we describe the construction of a chimeric fusion gene derived from the murine anti-CD20 antibody (1F5) variable region, with an engineered homophilic domain at the C-terminus of the human IgG1 sequence. The construct was expressed in CHO suspension cells and purified. The potency of the homophilic anti-CD20 antibody was compared to a chimeric antibody without the engineered homophilic domain. In this comparison, the homophilic anti-CD20 antibody showed increased binding to a human CD20 cell line, and significantly more ADCC, CDC, and induction of apoptosis in three cell lines. In addition, the homophilic anti-CD20 antibody demonstrated increased inhibition of proliferation of two cell lines. These data show that homophilic fusion protein antibodies with enhanced therapeutic potency can be produced with industry-standard fermentation protocols.
Terp MG, Lund RR, Jensen ON, et al.Identification of markers associated with highly aggressive metastatic phenotypes using quantitative comparative proteomics.
Cancer Genomics Proteomics. 2012 Sep-Oct; 9(5):265-73 [PubMed
] Related Publications
BACKGROUND: The spread of cancer cells from a primary tumor to form metastases at distant sites is a complex process that remains poorly defined. Certain tumor cells are more aggressive and thus lead to rapid development of multiple distant metastases. Here, we identify proteins associated with these aggressive phenotypes.
MATERIALS AND METHODS: To identify proteins associated with cancer cell aggressiveness, we used comparative, quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteome analysis of a unique metastasis model comprised of three isogenic human breast cancer cell lines that are equally tumorigenic in mice, but display different metastatic potentials ranging from non-metastatic, intermediate-metastatic and highly-metastatic. The altered expression of selected proteins was subsequently confirmed by immunocyto- and immunohistochemistry.
RESULTS: The difference in metastatic capabilities was initially confirmed using live animal imaging. Comparative, quantitative proteomics identified 414 proteins, out of which 44 exhibited altered expression between the metastatic and non-metastatic cell lines. The proteins correlating with the aggressiveness of metastasis included leucine-rich repeat containing 59 (LRRC59), while CD59 and chondroitin sulfate proteoglycan 4 (CSPG4) exhibited an inverse correlation with metastatic capability. The altered expression levels of these proteins were biochemically confirmed, as well as demonstrated in xenografts generated from these cell lines. This analysis further demonstrated that the three proteins were associated with the aggressiveness of metastasis rather than metastasis colonization per se.
CONCLUSION: Our study provides novel insights into key proteins associated with the metastatic potential of breast cancer cells and identified LRRC59, CD59 and CSPG4 as candidates that merit further study.
Bellone S, Roque D, Cocco E, et al.Downregulation of membrane complement inhibitors CD55 and CD59 by siRNA sensitises uterine serous carcinoma overexpressing Her2/neu to complement and antibody-dependent cell cytotoxicity in vitro: implications for trastuzumab-based immunotherapy.
Br J Cancer. 2012; 106(9):1543-50 [PubMed
] Free Access to Full Article Related Publications
BACKGROUND: We evaluated the expression of CD46, CD55 and CD59 membrane-bound complement-regulatory proteins (mCRPs) in primary uterine serous carcinoma (USC) and the ability of small interfering RNA (siRNA) against these mCRPs to sensitise USC to complement-dependent cytotoxicity (CDC) and antibody (trastuzumab)-dependent cellular cytotoxicity (ADCC) in vitro.
METHODS: Membrane-bound complement-regulatory proteins expression was evaluated using real-time PCR (RT-PCR) and flow cytometry, whereas Her2/neu expression and c-erbB2 gene amplification were assessed using immunohistochemistry, flow cytometry and fluorescent in-situ hybridisation. The biological effect of siRNA-mediated knockdown of mCRPs on HER2/neu-overexpressing USC cell lines was evaluated in CDC and ADCC 4-h chromium-release assays.
RESULTS: High expression of mCRPs was found in USC cell lines when compared with normal endometrial cells (P<0.05). RT-PCR and FACS analyses demonstrated that anti-mCRP siRNAs were effective in reducing CD46, CD55 and CD59 expression on USC (P<0.05). Baseline complement-dependent cytotoxicity (CDC) against USC cell lines was low (mean ± s.e.m.=6.8 ± 0.9%) but significantly increased upon CD55 and CD59 knockdown (11.6 ± 0.8% and 10.7 ± 0.9%, respectively, P<0.05). Importantly, in the absence of complement, both CD55 and CD59, but not CD46, knockdowns significantly augmented ADCC against USC overexpressing Her2/neu.
CONCLUSION: Uterine serous carcinoma express high levels of the mCRPs CD46, CD55 and CD59. Small interfering RNA inhibition of CD55 and CD59, but not CD46, sensitises USC to both CDC and ADCC in vitro, and if specifically targeted to tumour cells, may significantly increase trastuzumab-mediated therapeutic effect in vivo.
Human amniotic mesenchymal stem cells (hAMSCs) demonstrated partially pluripotent characteristics with a strong expression of Oct4 and Nanog genes and immunomodulatory properties characterized by the absence of HLA-DR and the presence of HLA-G and CD59. The hAMSCs were reprogrammed into induced pluripotent stem cells (iPSCs) that generate a promising source of universal cardiac cells. The hAMSC-derived iPSCs (MiPSCs) successfully underwent robust cardiac differentiation to generate cardiomyocytes. This study investigated 3 key properties of the hAMSCs and MiPSCs: (1) the reprogramming efficiency of the partially pluripotent hAMSCs to generate MiPSCs; (2) immunomodulatory properties of the hAMSCs and MiPSCs; and (3) the cardiac differentiation potential of the MiPSCs. The characteristic iPSC colony formation was observed within 10 days after the transduction of the hAMSCs with a single integration polycistronic vector containing 4 Yamanaka factors. Immunohistology and reverse transcription-polymerase chain reaction assays revealed that the MiPSCs expressed stem cell surface markers and pluripotency-specific genes. Furthermore, the hAMSCs and MiPSCs demonstrated immunomodulatory properties enabling successful engraftment in the SVJ mice. Finally, the cardiac differentiation of MiPSCs exhibited robust spontaneous contractility, characteristic calcium transience across the membrane, a high expression of cardiac genes and mature cardiac phenotypes, and a contractile force comparable to cardiomyocytes. Our results demonstrated that the hAMSCs are reprogrammed with a high efficiency into MiPSCs, which possess pluripotent, immunomodulatory, and precardiac properties. The MiPSC-derived cardiac cells express a c-kit cell surface marker, which may be employed to purify the cardiac cell population and enable allogeneic cardiac stem cell therapy.
Carcinosarcomas of the female genital tract are rare tumors with an aggressive clinical behavior. Trastuzumab, a humanized monoclonal antibody, acts by binding to HER2/neu extracellular domain and exhibits therapeutic efficacy in HER2/neu-overexpressing cancers. Two uterine carcinosarcomas (UMMT-ARK-1, UMMT-ARK-2) and 2 ovarian carcinosarcomas (OMMT-ARK-1, OMMT-ARK-2) were established as primary tumor cell lines in vitro and evaluated for HER2/neu expression by immunohistochemistry, fluorescent in situ hybridization analysis, quantitative real-time polymerase chain reaction, and for membrane-bound complement regulatory proteins CD46, CD55, and CD59 by flow cytometry. Sensitivity to trastuzumab-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity was studied in 5-hr chromium release assays. HER2/neu expression was demonstrated in OMMT-ARK-1 and OMMT-ARK-2. OMMT-ARK-2 demonstrated an amplification of the c-erbB2 gene by fluorescent in situ hybridization analysis and a high sensitivity to ADCC (mean killing, 45.6%; range, 32.3%-72.6%). A lower level of killing was detected against the fluorescent in situ hybridization analysis-negative OMMT-ARK-1 cell line (mean, 26.5%; range, 21.0%-31.8%). CD46, CD55, and CD59 membrane-bound complement regulatory proteins were expressed at high levels in all primary mixed müllerian tumor cell lines, and all these tumors were found to be highly resistant to complement-dependent cytotoxicity with or without trastuzumab. Addition of untreated and heat-inactivated plasma did not significantly decrease ADCC against OMMT-ARK-2 cell line, suggesting that while the cell line is highly resistant to complement, irrelevant IgG does not significantly alter the ability of trastuzumab to mediate ADCC. Our results suggest that HER2/neu may represent a novel target for the immunotherapy of a subset of human carcinosarcomas refractory to salvage chemotherapy.
Xie XH, Gao MH, Zhang B, et al.Post-transcriptional CD59 gene silencing by siRNAs induces enhanced human T lymphocyte response to tumor cell lysate-loaded DCs.
Cell Immunol. 2012; 274(1-2):1-11 [PubMed
] Related Publications
CD59 is a complement regulatory protein known to prevent the membrane attack complex (MAC) from assembling. To investigate the role of CD59 molecules in human T cell activation in response to exogenous antigens, gene silencing via small interfering RNAs (siRNAs) was carried out. Subsequent T cell activation in response to both autologous dendritic cells (DCs) loaded with tumor lysate and beads coated with anti-CD3, anti-CD28 and anti-CD59 antibodies was investigated. The findings demonstrated that decreased CD59 expression on T cells significantly enhanced activation and proliferation of CD4(+) T cells and CD8(+) T cells while the expansion of CD4(+) CD25(+) regulatory T cells (Tregs) was not affected, and CD59 mediated inhibition of T cell activation requires the binding of CD59 with its ligand on antigen-presenting cells (APCs). The data support that CD59 down-regulates antigen-specific activation of human T lymphocytes in a ligand-dependent manner.
Li B, Chu X, Gao M, Xu YThe effects of CD59 gene as a target gene on breast cancer cells.
Cell Immunol. 2011; 272(1):61-70 [PubMed
] Related Publications
The retroviral-vector-targeted CD59 gene (pSUPER-siCD59) was constructed and transfected into breast cells (MCF-7). The results demonstrated that the retroviral vector-mediated RNAi successfully suppressed human CD59 gene. The expression of CD59 decreased at both mRNA and protein levels. Knockdown of CD59 abrogated its protective effect on complement-mediated cytolysis. Fas and caspase-3 were remarkably upregulated, which induced apoptosis and tumor growth suppression in MCF-7 cells. In addition, overexpression of CD59 promoted the proliferation of MCF-7 cells and inhibited anti-apoptotic Bcl-2 expression. In conclusion, CD59 may be a promising target in the gene therapy of breast cancer.
High androgen receptor (AR) level in primary tumour predicts increased prostate cancer-specific mortality. However, the mechanisms that regulate AR function in prostate cancer are poorly known. We report here a new paradigm for the forkhead protein FoxA1 action in androgen signalling. Besides pioneering the AR pathway, FoxA1 depletion elicited extensive redistribution of AR-binding sites (ARBs) on LNCaP-1F5 cell chromatin that was commensurate with changes in androgen-dependent gene expression signature. We identified three distinct classes of ARBs and androgen-responsive genes: (i) independent of FoxA1, (ii) pioneered by FoxA1 and (iii) masked by FoxA1 and functional upon FoxA1 depletion. FoxA1 depletion also reprogrammed AR binding in VCaP cells, and glucocorticoid receptor binding and glucocorticoid-dependent signalling in LNCaP-1F5 cells. Importantly, FoxA1 protein level in primary prostate tumour had significant association to disease outcome; high FoxA1 level was associated with poor prognosis, whereas low FoxA1 level, even in the presence of high AR expression, predicted good prognosis. The role of FoxA1 in androgen signalling and prostate cancer is distinctly different from that in oestrogen signalling and breast cancer.
Cocco E, Varughese J, Buza N, et al.Tissue factor expression in ovarian cancer: implications for immunotherapy with hI-con1, a factor VII-IgGF(c) chimeric protein targeting tissue factor.
Clin Exp Metastasis. 2011; 28(7):689-700 [PubMed
] Free Access to Full Article Related Publications
We evaluated the expression of tissue factor (TF) in ovarian cancer (EOC) and the potential of hI-con1, an antibody-like molecule targeting TF, as a novel form of therapy against chemotherapy-resistant ovarian disease. We studied the expression of TF in 88 EOC by immunohistochemistry (IHC) and real-time-PCR (qRT-PCR) and the levels of membrane-bound-complement-regulatory-proteins CD46, CD55 and CD59 in primary EOC cell lines by flow-cytometry. Sensitivity to hI-con1-dependent-cell-mediated-cytotoxicity (IDCC), complement-dependent-cell-cytotoxicity and inhibition of IDCC by γ-immunoglobulin were evaluated in 5-h (51)chromium-release-assays. Cytoplasmic and/or membrane TF expression was observed in 24 out of 25 (96%) of the EOC samples tested by IHC, but not in normal ovarian-tissue. EOC with clear cell histology significantly overexpress TF when compared to serous, endometrioid, or undifferentiated tumors by qRT-PCR. With a single exception, all primary EOC that overexpressed TF demonstrated high levels of CD46, CD55 and CD59 and regardless of their histology or resistance to chemotherapy, were highly sensitive to IDCC. The effect of complement and physiologic doses of γ-immunoglobulin on IDCC in ovarian cancer cell lines overexpressing TF was tumor specific and related to the overexpression of CD59 on tumor cells. Small-interfering-RNA-mediated knockdown of CD59 expression in ovarian tumors significantly increased hI-con1-mediated cytotoxic activity in vitro. Finally, low doses of interleukin-2 further increased the cytotoxic effect induced by hI-con1 (P < 0.01). hI-con1 molecule induces strong cytotoxicity against primary chemotherapy-resistant ovarian cancer cell lines overexpressing TF and may represent a novel therapeutic agent for the treatment of ovarian tumors refractory to standard treatment modalities.
Shan C, Zhang S, Cui W, et al.Hepatitis B virus X protein activates CD59 involving DNA binding and let-7i in protection of hepatoma and hepatic cells from complement attack.
Carcinogenesis. 2011; 32(8):1190-7 [PubMed
] Related Publications
Emerging evidence has shown that hepatitis B virus (HBV) X protein (HBx) plays a crucial role in the development of hepatocellular carcinoma. Complement regulatory proteins including CD46, CD55 and CD59 contribute to escape of tumor cells from complement-dependent cytotoxicity (CDC). However, little is known about the potential role of HBx in anti-CDC activity during hepatocarcinogenesis. In the present study, we for the first time report that HBx decreases the sensitivity of hepatoma and hepatic cells to CDC. Coincidentally, we demonstrated that HBx increased the promoter activity of CD59, as well as their messenger RNA and protein levels. Moreover, flow cytometry showed the increased expression level of CD59 protein on the surface of HBx-positive cells. Of interest, we found that HBx up-regulated CD59 by binding with cAMP response element-binding to the promoter region of the CD59 gene using chromatin immunoprecipitation assay. In addition, we showed that HBx up-regulated CD59 by let-7i at post-transcriptional regulation level. Our data showed that the deposition of C5b-9 were decreased on the cell surface in HepG2-X cells relative to HepG2 cells, suggesting that increased CD59 mediated by HBx prevents the formation of functional membrane attack complex. Furthermore, we demonstrated that down-regulation of CD59 was sufficient to abolish the resistance capability of CDC in HBx-positive cells by RNA interference (siRNA) in vitro and in vivo. Thus, we conclude that HBx contributes to cells resistance to CDC through CD59. Therapeutically, CD59 may serve as a target in HBV-associated hepatoma patients.
Geis N, Zell S, Rutz R, et al.Inhibition of membrane complement inhibitor expression (CD46, CD55, CD59) by siRNA sensitizes tumor cells to complement attack in vitro.
Curr Cancer Drug Targets. 2010; 10(8):922-31 [PubMed
] Related Publications
The efficacy of cancer-immunotherapy with complement-activating monoclonal antibodies is limited by over-expression of one or more membrane-bound complement regulatory proteins (mCRPs: CD46, CD55, CD59) on the surface of neoplastic cells. In this study we designed small interfering RNAs (siRNAs) for posttranscriptional gene knock down of CD46, CD55 and CD59 aiming at to sensitize tumor cells to complement attack and thereby to better exploit complement for tumor cell destruction. Tumor cell lines of different origin, such as Du145 (prostate), BT474 (breast) and K562 (erythroleukemia) were selected for the study. FACS-analysis demonstrated that siRNA anti-CD46(301) reduced CD46 protein expression up to 80%, siRNA anti-CD55(255) diminished CD55 protein expression up to 49%, and CD59 protein expression was inhibited up to 82% by siRNA anti-CD59(1339). Time course experiments revealed a long-lasting silencing effect with >50% complement regulator inhibition up to day 13. Upon mCRP knock down, complement-dependent cytotoxicity (CDC) was augmented by 20-30% for CD46, by up to 24% for CD55 and by up to 55% for CD59. The combined inhibition of all three inhibitors further enhanced CDC by up to 66%. Dependent on the cell line, CD46 and CD55 downregulation increased significantly C3 ospsonization, which is known to support cell-mediated defense mechanisms. mCRP blocking antibodies were only partly able to further augment the tumor cells' susceptibility to complement lysis. Thus, siRNA-induced inhibition of complement regulator expression clearly sensitizes malignant cells to complement attack and, if specifically targeted to the tumor, appears suited as adjuvant to improve antibody-based cancer immunotherapy.
Kolev MV, Ruseva MM, Morgan BP, Donev RMTargeting neural-restrictive silencer factor sensitizes tumor cells to antibody-based cancer immunotherapy in vitro via multiple mechanisms.
J Immunol. 2010; 184(11):6035-42 [PubMed
] Related Publications
Tumor cells escape clearance by complement by abundantly expressing CD59 and other membrane complement regulators. Recently, we designed a peptide derived from the neural-restrictive silencer factor (REST), REST68, which we showed to inhibit expression of CD59 in tumors lacking the full-length REST and proposed a detailed model for regulation of CD59 expression via interplay between REST and nucleolin (NCL) transcription factors. In this paper, we study in detail the mechanisms for sensitization of malignant cells to Ab-based cancer immunotherapy by the REST68 peptide and the implications of the REST/NCL model for the design of treatment resulting in higher tumor susceptibility. REST68 inhibited CD59 expression in malignant cells expressing either truncated or full-length REST, but not in nonmalignant cells. However, activation of protein kinase C (PKC) in nonmalignant cells, a process that contributes to cellular transformation, phosphorylated NCL and enabled suppression of CD59 expression by the REST68. Combined treatment of different tumor types with REST68 and PKC inhibitor synergized to further suppress CD59 expression and reduce resistance to complement lysis. The combined treatment also increased susceptibility of tumors expressing either of the REST isoforms to PBMC-mediated killing, which, at least in part, accounted for the strong promotion of apoptosis by the REST68/PKC inhibitor. These data demonstrate that REST68 sensitizes tumors to Ab-based cancer immunotherapy via multiple mechanisms. Furthermore, the REST/NCL interplay model for regulation of expression of cd59 and other genes involved in cell survival enables the design of treatments for different tumor types to achieve more efficient tumor clearance.
Tediose T, Kolev M, Sivasankar B, et al.Interplay between REST and nucleolin transcription factors: a key mechanism in the overexpression of genes upon increased phosphorylation.
Nucleic Acids Res. 2010; 38(9):2799-812 [PubMed
] Free Access to Full Article Related Publications
Non-malignant cells can be transformed via the activation of kinases that control degradation of neural-restrictive silencer factor (REST). Here, we identify a mechanism that contributes to the activation of genes, expression of which is controlled by responsive elements containing overlapping binding sites for REST and nucleolin. We demonstrate that both phosphorylated and non-phosphorylated nucleolin-bound DNA; however, only phosphorylated nucleolin successfully competed with either full-length REST or a REST-derived DNA-binding peptide, REST68, for binding to the overlapping binding sites. We show that this interplay between the two transcription factors regulates the activation of cell survival and immunomodulatory genes in tumors and non-malignant cells with activated protein kinase C, which is accompanied with alterations in cell proliferation and apoptosis. We propose a model for the regulation of these genes, which brings a new insight into the molecular mechanisms that control cellular transformation driven by activation of protein kinases.
BACKGROUND: T-cell large granular lymphocytic leukemia is a clonal proliferation of cytotoxic T-lymphocytes which often results in severe cytopenia. Current treatment options favor chronic immunosuppression. Alemtuzumab, a humanized monoclonal antibody against glycophosphatidylinositol-anchored CD52, is approved for patients refractory to therapy in other lymphoid malignancies.
DESIGN AND METHODS: We retrospectively examined treatment outcomes in 59 patients with CD8+ T-cell large granular lymphocytic leukemia, 41 of whom required therapy. Eight patients with severe refractory cytopenia despite multiple treatment regimens had been treated with subcutaneous alemtuzumab as salvage therapy. Flow cytometry was used to monitor expression of glycophosphatidylinositol-anchored CD52, CD55, and CD59 as well as to characterize T-cell clonal expansions by T-cell receptor variable beta-chain (Vbeta) repertoire.
RESULTS: Analysis of the effects of alemtuzumab revealed remissions with restoration of platelets in one of one patient, red blood cell transfusion independence in three of five patients and improvement of neutropenia in one of three, resulting in an overall response rate of 50% (4/8 patients). Clonal large granular lymphocytes exhibited decreased CD52 expression post-therapy in patients refractory to treatment. Samples of large granular lymphocytes collected prior to therapy also unexpectedly had a significant proportion of CD52-negative cells while a healthy control population had no such CD52 deficiency (p=0.026).
CONCLUSIONS: While alemtuzumab may be highly effective in large granular lymphocytic leukemia, prospective serial monitoring for the presence of CD52-deficient clonal cytotoxic T-lymphocytes should be a component of clinical trials investigating the efficacy of this drug. CD52 deficiency may explain lack of response to alemtuzumab, and such therapy may confer a survival advantage to glycophosphatidylinositol-negative clonal cytotoxic T-lymphocytes.
CD59, a broadly expressed GPI-anchored molecule, regulates formation of the membrane attack complex of the complement cascade. We previously demonstrated that mouse CD59 also down-modulates CD4(+) T cell activity in vivo. In this study, we explored the role of CD59 on human CD4(+) T cells. Our data demonstrate that CD59 is up-regulated on activated CD4(+) T cells and serves to down-modulate their activity in response to polyclonal and Ag-specific stimulation. The therapeutic potential of this finding was explored using T cells isolated from colorectal cancer patients. The findings were striking and indicated that blockade of CD59 significantly enhanced the CD4(+) T cell response to two different tumor Ags. These data highlight the potential for manipulating CD59 expression on T cells for boosting weak immune responses, such as those found in individuals with cancer.
CD59, belonging to membrane complement regulatory proteins (mCRPs), inhibits the cytolytic activity of complement and is over-expressed in solid cancers, including ovary cancer. The aim of the present study was to construct recombinant retrovirus encoding shRNA targeted human CD59 and infect A2780 cells in order to investigate the relationship between decreased CD59 expression and tumorigenesis of ovary cancer. siCD59 and siCD59-C were successfully constructed and identified by PCR, restriction endonuclease analyses and DNA sequencing, respectively. The siCD59 was able to efficiently infect A2780 cells, which was confirmed by Western blotting. When incubated with fresh normal human serum (8%, v/v) for 1 h at 37 degrees centigrade, the cell viability was decreased and cell damage was increased in siCD59 infected A2780 cells compared to siCD59-C infected cells. This led to the activation of caspase-3. The apoptosis in siCD59 infected cells was shown with hypercondensed nuclei using Hoechst staining. Meanwhile, the weight of ovary tumor graft in nude mice was significantly decreased in siCD59 group compared to that of siCD59-C group. And the expression of CD59 protein in tumor tissue in siCD59 group was significantly decreased. These results suggested that CD59 silencing in ovary cancer cells via retrovirus-mediated RNAi can enhance complement-mediated cell damage, inhibiting growth of ovary cancer. CD59 might be a potential target for gene therapy in ovary cancer.
Tang JH, He WJ, Huang H, et al.Important roles of glycosylphosphatidylinositol (GPI)-specific phospholipase D and some GPI-anchored proteins in the pathogenesis of hepatocellular carcinoma.
Clin Biochem. 2009; 42(4-5):400-7 [PubMed
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OBJECTIVE: To investigate the roles of glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) in the pathogenesis of hepatocellular carcinoma (HCC).
METHODS: The expression of the GPI-PLD in HCC was determined. The GPI-PLD gene was stably transfected in HepG2 cells and the proliferation of these cells was detected; CD55, CD59 and apoptotic cells were also analyzed.
RESULTS: The serum GPI-PLD activities, the protein and mRNA levels of GPI-PLD in HCC patients were decreased by 40%, 60% and 56%, respectively. The killing rate of CDC against the positive clone cells was significantly increased, but the proliferative capacity was obviously decreased. The apoptotic rate in positive clones was increased.
CONCLUSION: The expression of GPI-PLD decreases in HCC patients. The over-expression of GPI-PLD in HepG2 cells increases their sensitivity to CDC killing, impairs proliferative capacity and promotes apoptosis.
Multi-drug resistance (MDR) is a major obstacle in cancer chemotherapy. There are contrasting data on a possible correlation between the level of expression of the drug transporter P-glycoprotein (P-gp) and susceptibility to complement-dependent cytotoxicity (CDC). We therefore investigated the sensitivity of human ovarian carcinoma cells and their P-gp expressing MDR variants to complement. Chemoselected P-gp expressing MDR cells showed increased resistance to CDC associated with overexpression of membrane-bound complement regulatory proteins (mCRP) and increased release of the soluble inhibitors C1 inhibitor and factor I. MDR1 gene transfection alone did not alter the susceptibility of P-gp expressing A2780-MDR and SKOV3-MDR cells to CDC. However, subsequent vincristine treatment conferred an even higher resistance to complement to these cells, again associated with increased expression of mCRP. Blocking the function of P-gp with verapamil, cyclosporine A or the anti-P-gp-antibody MRK16 had no impact on their complement resistance, whereas blocking of mCRP enhanced their susceptibility to complement. These results suggest that enhanced resistance of chemoselected MDR ovarian carcinoma cells to CDC is not conferred by P-gp, but is due at least partly to overexpression of mCRP, probably induced by treatment with the chemotherapeutic agents.
Tumor cells escape clearance by complement by abundantly expressing CD59 and other membrane complement regulators. Existing strategies for blocking/knocking down these regulators can contribute to tumor immunoclearance in vitro; however, there are numerous difficulties restricting their use in vivo. Here, we report a new strategy for suppression of CD59 expression in neuroblastoma using peptides that target regulators of CD59 expression. We identified the neural-restrictive silencer factor (REST) as a target for modulation of CD59 expression in neuroblastoma. We next designed plasmids that encoded peptides comprising different DNA-binding domains of REST and transfected them into neuroblastoma cell lines. These peptides suppressed CD59 expression, sensitizing neuroblastoma to complement-mediated killing triggered by anti-GD2 therapeutic monoclonal antibody. These CD59-modulating peptides might be effective therapeutic adjuvants to therapeutic monoclonal antibodies used for treatment of neuroblastoma and other cancer types sharing the same mechanism for regulation of CD59 expression.
Membrane complement inhibitors (CD46, CD55 and CD59) are upregulated in some human cancers indicating that they play a role in immune evasion. We investigated complement inhibitor expression in bladder cancer and examined the hypothesis that selective pressure of an antibody response (anti-MUC1) results in the upregulated expression of complement inhibitors on tumor cells. Paired samples of tumor and normal tissue from 22 bladder cancer patients were analyzed for expression of MUC1, CD46, CD55 and CD59, and matched serum samples analyzed for anti-MUC1 IgM and IgG levels. Relationships between anti-MUC1 antibody levels and complement inhibitor expression were investigated. MUC1 mRNA was upregulated in 86% of tumor samples. CD46 was upregulated in 77%, CD55 in 55% and CD59 in 59% of tumors. Low titer anti-MUC1 IgM was detected in normal human sera, but was elevated in 41% of the bladder cancer patients. Anti-MUC1 IgG was virtually absent from normal sera, but present in 32% of the cancer patients. There was a direct relationship between anti-MUC1 antibody titer and expression level of complement inhibitors. Analysis of the correlation of each antibody with the expression of each complement inhibitor by Spearman's rank test revealed a strong correlation between both anti-MUC1 IgM and IgG levels and increased expression of CD46 and CD55, and combined anti-MUC1 IgM/IgG levels correlated with increased expression of all 3 complement inhibitors. In conclusion, the data demonstrate upregulated complement inhibitor expression and the presence of an anti-MUC1 antibody response in bladder cancer patients and support the hypothesis of antibody-mediated immune selection.
Tímár J, Mészáros L, Ladányi A, et al.Melanoma genomics reveals signatures of sensitivity to bio- and targeted therapies.
Cell Immunol. 2006; 244(2):154-7 [PubMed
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Most of the melanoma markers used today are melanocytic markers or pigmentation pathway-associated genes driven by the microphthalmia transcription factor, MITF, and include among others, tyrosinase, dopachrome tautomerase, DCT, melan-A and S100B. Genomic studies repeatedly revealed several novel melanoma marker genes including those of the transcription factor NOTCH2, WNT5A, proliferation-associated genes TOPO2A and CDC2, membrane receptors FGFR and EphA3, adhesion molecules N-cadherin, beta3 integrin and syndecan-4, and the cell surface antigens CD59/protectin and MIA. Other genomic analyses tried to define the gene signature of the metastatic disease but failed to find a consistent one except the gold standard genes of beta3 integrin, syndecan-4 and WNT5a. Studies on the gene signatures of chemoresistance and cytokine sensitivity of melanoma clearly defined apoptosis-resistance as one of the key elements of the above biological properties, but the data are controversial, mostly because of the use of inappropriate model systems and the lack of confirmation on clinical samples. Accordingly, application of genomic technologies must be more "translational" to provide breakthrough in melanoma diagnosis and therapy.
Pretargeted radioimmunotherapy (PRIT) using streptavidin (SA)-conjugated antibodies (Abs), followed by clearing agent and radiolabeled biotin is a promising method that can increase the effectiveness of RIT, while decreasing the toxicities associated with directly labeled Abs. Although CD20 has been the traditional target antigen for RIT of non-Hodgkin lymphoma (NHL), studies targeting HLA DR and CD22 have yielded promising results. Targeting all 3 antigens at once may further augment the effect of PRIT. This study compares the targeting of Ramos, Raji, and FL-18 lymphoma xenografts with either anti-CD20 Ab/SA (1F5/SA), anti-HLA DR Ab/SA (Lym-1/SA), anti-CD22 Ab/SA (HD39/SA), or all 3 conjugates in combination, followed 24 hours later by a biotin-N-acetyl-galactosamine clearing agent, and 3 hours after that by (111)In-DOTA-biotin. The Ab/SA conjugate yielding the best tumor uptake and tumor-to-normal organ ratios of radioactivity varied depending on the target antigen expression on the cell line used, with 1F5/SA and Lym-1/SA yielding the most promising results overall. Also, the best tumor-to-normal organ ratios of absorbed radioactivity were obtained using single conjugates optimized for target tumor antigen expression rather than the combination therapy. This study highlights the importance of screening the antigenic expression on lymphomas to select the optimal reagent for PRIT.
Gee JM, Shaw VE, Hiscox SE, et al.Deciphering antihormone-induced compensatory mechanisms in breast cancer and their therapeutic implications.
Endocr Relat Cancer. 2006; 13 Suppl 1:S77-88 [PubMed
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Breast cancer inhibition by antihormones is rarely complete, and our studies using responsive models reveal the remarkable flexibility of breast cancer cells in recruiting alternative signalling to limit maximal anti-tumour effects of oestrogen receptor alpha (ER) blockade. The recruited mechanism involves antihormone-induced expression of oestrogen-repressed signalling genes. For example, epidermal growth factor receptor gene (EGFR) is induced by antioestrogens and maintains residual kinase and ER phosphorylation, cell survival genes, and thereby allows incomplete antihormone response and emergence of resistance. Microarrays are revealing the breadth of antihormone-induced genes that may attenuate growth inhibition, including NFkappaB, Bag1, 14-3-3zeta and tyrosine kinases, such as HER2 and Lyn. Three concepts are emerging: first, some genes are induced exclusively by antioestrogens, while others extend to oestrogen deprivation; secondly, some are transiently induced, while others persist into resistance; finally, some confer additional adverse features when tumour cells are in an appropriate context. Among the latter is CD59 whose antioestrogen induction may permit evasion of immune surveillance in vivo. Also, induction of pro-invasive genes (including NFkappaB, RhoE and delta-catenin) may underlie our findings that antioestrogens can markedly stimulate migratory behaviour when tumour intercellular contacts are compromised. Based on our promising studies selectively inhibiting EGFR (gefitinib), NFkappaB (parthenolide) or CD59 (neutralising antibody) together with antioestrogens, we propose that co-targeting strategies could markedly improve anti-tumour activity (notably enhancing cell kill) during the antihormone-responsive phase. Furthermore, subverting those induced signalling genes that are retained into resistance (e.g. EGFR, NFkappaB, HER2) may prove valuable in this state. Alongside future deciphering and targeting of genes underlying antioestrogen-promoted invasiveness, embracing of intelligent combination strategies could significantly extend patient survival.