Preferentially Expressed Antigen Of Melanoma Expression Research Articles

PRAME Expression in Melanocytic Proliferations in Special Sites.

The subset of nevi occurring at special sites (eg, acral skin, anogeni-tal region, breast, ear, flexural surfaces) have normal histologic variations that preclude the use of routinely used diagnostic criteria for malignancy. Suggested criteria for differentiating malignant special-site lesions from benign lesions have been described, but there is an unmet need for a validated test aiding in the delineation of benign and malignant lesions at special sites. Preferentially expressed antigen of melanoma (PRAME) expression has been characterized as a relatively specific marker of melanoma, but not within the specific population of special-site lesions. This study aimed to determine if PRAME may serve as a specific marker of melanoma within the population of special-sites lesions.

Immunohistochemical expression of PRAME in 485 cases of epithelial tubo-ovarian tumors.

Preferentially expressed antigen of melanoma (PRAME) is a cancer/testis antigen selectively expressed in somatic tissues and various solid malignant tumors and is associated with poor prognostic outcome. Our research aimed to comprehensively compare its expression in a large cohort of tubo-ovarian epithelial tumors and examine its correlation with our clinico-pathologic data, as well as to assess its potential use in diagnostics and therapy.We examined 485 cases of epithelial tubo-ovarian tumors including 107 clear cell carcinomas (CCC), 52 endometroid carcinomas (EC), 103 high grade serous carcinomas (HGSC), 119 low grade serous carcinomas (LGSC)/micropapillary variant of serous borderline tumors (mSBT), and 104 cases of mucinous carcinomas (MC)/mucinous borderline tumors (MBT). The immunohistochemical analysis was performed using TMAs.The highest levels of expression were seen in EC (60%), HGSC (62%), and CCC (56%), while expression in LGSC/mSBT (4%) and MC/MBT (2%) was rare. The clinico-pathologic correlations and survival analysis showed no prognostic significance.The results of our study showed that PRAME is neither prognostic nor a suitable ancillary marker in the differential diagnosis of tubo-ovarian epithelial tumors. Nevertheless, knowledge about the PRAME expression may be important concerning its potential predictive significance, because targeting PRAME as a potential therapeutic option is currently under investigation.

Clinical features and prognosis of childhood B-lineage acute lymphoblastic leukemia expressing the PRAME gene

To study the clinical and prognostic significance of the preferentially expressed antigen of melanoma (PRAME) gene in the absence of specific fusion gene expression in children with B-lineage acute lymphoblastic leukemia (B-ALL). A total of 167 children newly diagnosed with B-ALL were enrolled, among whom 70 were positive for the PRAME gene and 97 were negative. None of the children were positive for MLL-r, BCR/ABL, E2A/PBX1, or ETV6/RUNX1. The PRAME positive and negative groups were analyzed in terms of clinical features, prognosis, and related prognostic factors. Compared with the PRAME negative group, the PRAME positive group had a significantly higher proportion of children with the liver extending >6 cm below the costal margin (P<0.05). There was a significant reduction in the PRAME copy number after induction chemotherapy (P<0.05). In the minimal residual disease (MRD) positive group after induction chemotherapy, the PRAME copy number was not correlated with the MRD level (P>0.05). In the MRD negative group, there was also no correlation between them (P>0.05). The PRAME positive group had a significantly higher 4-year event-free survival rate than the PRAME negative group (87.5%±4.6% vs 73.5%±4.6%, P<0.05), while there was no significant difference between the two groups in the 4-year overall survival rate (88.0%±4.4% vs 85.3%±3.8%, P>0.05). The Cox proportional-hazards regression model analysis showed that positive PRAME expression was a protective factor for event-free survival rate in children with B-ALL (P<0.05). Although the PRAME gene cannot be monitored as MRD, overexpression of PRAME suggests a good prognosis in B-ALL.

Prognostic significance of PRAME (preferentially expressed antigen of melanoma) expression in breast cancer

Purpose: The aim of this study is to detect the PRAME (Preferentially Expressed Antigen of Melanoma) in 54 patients with breast cancer and 37 patients with benign breast lesions. Materials and Methods: PRAME expressions in 54 breast cancer, 20 benign breast lesions and 10 normal breast tissue samples were studied with RT-PCR. Expression of PRAME was studied with IHC in 37 benign breast lesions, in 54 breast cancer patients from both tumor and normal breast tissue. RT-PCR and IHC results for PRAME were compared in this study. Results: PRAME was found to be expressed in 50 % of the breast cancer and 25 % of the benign breast lesions. Using IHC method, (+), (++) and (+++) staining for PRAME expression were found in 29,6%, 31,5% and 3,7% of the cases, respectively in invasive component of the breast cancer. PRAME expression detected by both IHC and RT-PCR was compared with prognostic parameters. PRAME expression in breast cancer was found to be associated with high tumor grade and negative hormone receptor. We found an important association between PRAME RT-PCR and of PRAME IHC. Conclusion: Both RT-PCR in fresh tissues and IHC method in paraffin embedded tissues can be used to identify PRAME expression and the predictive role of PRAME expression.

Both Methylation and Copy Number Variation Participated in the Varied Expression of PRAME in Multiple Myeloma.

PurposeThe cancer-testis antigen, which is a preferentially expressed antigen of melanoma (PRAME), is an ideal target for immunotherapy and cancer vaccines. Since the expression of this antigen is relevant to therapy responses, the heterogeneity in its expression and the underlying mechanism need to be investigated.Patients and MethodsPlasma cell sorting was performed in 48 newly diagnosed multiple myeloma (MM) patients. Real-time quantitative PCR was performed to examine the PRAME transcript levels and gene copy numbers. Bisulfate clone sequencing of the PRAME promoter and exon 1b regions was performed in 4 patients. Quantitative methylation-specific PCR of the +287 CpG site was performed for all patients. The human MM cell lines RPMI8226, LP-1 and MOLP-2 were treated with 5-azacytidine.ResultsThe median PRAME transcript level was 3.1% (range: 0–298.3%) in the plasma cells sorted from the 48 MM patients. Eleven (22.9%) and 37 (77.1%) patients were individually categorized into the PRAME low- and high-expression groups according to the cut-off value of 0.05%. The methylation ratios of the promoter and the 3ʹ region of exon 1b region were both negatively related to the transcript levels. The degrees of methylation at the +287 CpG site were significantly negatively related to the transcript levels in all 48 patients (r=−0.44, P=0.0018), and those in the high-expression group (r=−0.69, P<0.0001) but not those in the low-expression group (r=−0.27, P=0.43). All 5 patients with homozygous deletions were categorized into the low-expression group. There were no significant differences in the PRAME transcript levels between the hemizygous deletion (n=8) and no deletion (n=35) groups (P=0.40). Furthermore, the PRAME transcript levels significantly increased in the MM cell lines after treatment with 5-azacytidine.ConclusionBoth methylation and copy number variation may participate in the regulation of PRAME expression in MM; in patients with no homozygous deletion, PRAME expression is mainly controlled by methylation, and a proportion of fairly low expression is caused by homozygous deletion.

WT1, PRAME, and PR3 mRNA Expression in Acute Myeloid Leukemia (AML).

Several tumor-associated antigens (TAAs) were recently identified, that could qualify as targets for immunotherapy, they could qualify (on RNA-level) for monitoring of tumor load. Here, we studied the expression levels of the immunogenic antigens PRAME (preferentially expressed antigen of melanoma), WT1 (Wilms' tumor gene), and PR3 (proteinase 3) on myeloid blasts by real-time quantitative polymerase chain reaction and correlated these data to the state and course of disease and to the defined subgroups of acute myeloid leukemia (AML). At first diagnoses, 41 of 47 patients tested showed overexpression of PRAME (87%), 38 of WT1 (81%), and 26 of PR3 (55%), with the highest expression levels for PRAME (2048-fold), followed by WT1 (486-fold) and PR3 (196-fold). Thereby, with 70%, the most frequent combination at first diagnoses was detected to be PRAME and WT1 (33/47 patients). Overall, 21 patients (45%) revealed overexpression for all 3 TAAs. Moreover, the highest expression levels of PRAME were found to be correlated with the FAB subtype M5, cytogenetic unfavorable risk groups, and AMLs arising from myelodysplasia (secondary AML; P=0.02). To compare TAA expression levels in the course of disease, expression data were calculatory adjusted to 100% blasts, revealing a relative increase in the PRAME expression levels during the course of persistent disease (3/4 cases). Independent of stage of disease, by trend, higher TAA expression levels were found on blasts derived from peripheral blood than those derived from the bone marrow. In conclusion, it is suggested that vaccine strategies for cancer immunotherapy should comprise different TAA peptides anticipating the diverse TAA expression levels on blasts evolving during the course of disease or treatment.

DC Vaccination Induces Antigen Specific Immune Responses in AML Patients: A 1-Year Interim Assessment

Acute myeloid leukemia (AML) is a lethal hematological malignancy characterized by high rates of relapse (50%). Studies of immune responses in AML patients led to identification of two leukemia-associated antigens (Wilms' tumor protein 1 (WT1) and preferentially expressed antigen of melanoma (PRAME) for which cellular and humoral immune responses were detected. Based on expression profiles in AML and normal tissues, WT1 and PRAME were selected as target antigens for a DC vaccine, aiming to elicit adaptive and innate immune responses in AML patients. The primary objective of the trial (EudraCT No.: 2014-003520-44) is to evaluate safety and feasibility of active immunotherapy with antigen-loaded autologous DCs. The secondary objectives are to assess DC vaccination-induced T cell responses against WT1 and PRAME and to evaluate the clinical outcome over a period of 2 years or until disease progression/relapse. WT1-positive AML patients, with or without PRAME positivity were included, if they had a morphological remission with or without hematological recovery (CRi) after induction chemotherapy and were not eligible for allogeneic hematopoietic stem cell transplantation. DCs were administered intradermally (2.5x106 WT1 plus 2.5x106 PRAME DCs) on a weekly basis for 4 consecutive weeks, followed by administration on week 6 and then every 4 weeks until 2 years, tumor progression or patient withdrawal. This report focusses on the immune-monitoring data collected during the first year of treatment. Multiple parameters were evaluated, including WT1- and PRAME specific T cell responses in peripheral blood (IFN-g ELISPOT), the recurrence of WT1- and/or PRAME-positive AML blasts in bone marrow (qRT-PCR) and peripheral blood (flow cytometry), and occurrence of AML-specific mutations in bone marrow (NGS). DCs were assessed for expression of WT1 and PRAME and costimulatory molecules by flow cytometry, while secretion of IL10 and IL12 was evaluated by double-color ELISPOT. Twenty CR/CRi AML patients were vaccinated, and vaccination was well tolerated, and no signs of autoimmunity were observed. 12/20 patients remained stable throughout the first year of treatment, while 8/20 relapsed. 2/8 relapsed patients died due to progression of disease during the first year. In patients in stable remission, 7/12 patients had no detectable IFN-γ ELISPOT response in peripheral blood, 3/12 mounted a response and 2/12 were not evaluable due to high background production of IFN-γ. Positive IFN-γ ELISPOT responses were observed within 1-30 weeks of treatment and associated with a decrease over time in levels of WT1 (3/3) and/or PRAME mRNA (1/3) expression. Except for 2 patients who remained MRD-negative throughout the first year of treatment, ELISPOT-negative patients in remission (5/7) exhibited low, relatively stable levels of target antigens. Stable remissions were associated with one (3/12) or no (8/12) AML-related mutations while one patient was not evaluable. AML relapses were observed within 50 days in 4 patients, within 51-100 days in 1 patient, and after more than 150 days in 3 patients. Overall, 6/8 relapsing patients exhibited positive IFN-γ ELISPOT responses, while 2 patients relapsing in &lt;50 days failed to respond and one patient relapsing on day 77 exhibited an IFN-γ response at baseline. In all other relapsing patients, IFN-γ ELISPOT responses were detected within 6-30 weeks after vaccination, in association with rapid and progressive increases in levels of WT1 and/or PRAME. AML-related gene mutations were detected in 6/8 relapses, while one patient was not evaluable, and one had no mutation; In 4 of the relapsing patients, 3 to 6 mutations were detected, while in 2/8 a single mutation was observed. In conclusion, IFN-γ responses to WT1 and/or PRAME were detected in peripheral blood of 75% (6/8) of relapsing patients, but only 25% (3/12) of the patients in remission. It is tempting to speculate that detection of IFN-γ response in peripheral blood is linked to the concomitant presence of AML blasts in the periphery of relapsing patients. Stable or decreasing levels of WT1 and/or PRAME mRNA in bone marrow of most patients in remission is compatible with the hypothesis that a local response against AML antigens may be ongoing, despite a negative IFN-γ response in peripheral blood. Monitoring of the patients currently in remission may shed some light on the role of DC vaccination on the prevention of AML recurrence. Disclosures Eckl: Medigene Immunotherapies GmbH: Employment. Raffegerst:Medigene Immunotherapies GmbH: Employment. Schnorfeil:Medigene Immunotherapies GmbH: Employment. Prinz-Schulz:Medigene Immunotherapies GmbH: Employment. Fingerhut:Medigene Immunotherapies GmbH: Employment. Bigalke:BioNTech Innovative Manufacturing Services GmbH: Employment. Pinkernell:Medigene AG: Employment. Schendel:Medigene AG: Employment, Equity Ownership, Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents &amp; Royalties. Tafuri:Medigene Immunotherapies GmbH: Employment.

PRAME Expression as a Potential Biomarker for Hematogenous Recurrence of Esophageal Squamous Cell Carcinoma.

To investigate the function of preferentially expressed antigen of melanoma (PRAME) in esophageal squamous cell carcinoma (ESCC). mRNA expression levels of PRAME were analyzed in resected esophageal tissues of 150 ESCC patients and correlated with clinicopathological parameters. We also investigated the potential function of PRAME by analyzing coordinately expressed genes in 13 ESCC cell lines. RT-qPCR analysis of clinical samples revealed aberrantly high PRAME expression in tumors compared with normal esophageal tissues. High PRAME expression was significantly associated with shorter disease-specific survival and hematogenous recurrence, but not with overall recurrence. The cumulative incidence of hematogenous recurrence was significantly greater for patients with high compared to those with low PRAME expression. In vitro, PCR array analysis revealed that PRAME was coordinately expressed with EGFR, ITGB, and TCF3. PRAME is overexpressed in ESCC tissues and may serve as a novel biomarker for predicting hematogenous recurrence.

PRAME as a Potential Biomarker for Liver Metastasis of Gastric Cancer.

Liver metastasis of gastric cancer (GC) is highly associated with poor prognosis. The development of sensitive biomarkers for detecting and predicting liver metastasis is required for better clinical outcome. In this study, we aimed to identify novel genes associated with liver metastasis of GC. Global expression profiling of 57,749 genes was performed using surgically resected gastric tissues from four patients with liver metastasis to identify candidate genes. The mRNA expression levels of the selected candidate gene were analyzed in the resected gastric tissues of 300 GC patients and correlated with clinicopathological parameters. Fourteen GC cell lines were subjected to mRNA expression and polymerase chain reaction (PCR) array analysis. Among 25 candidate genes identified by transcriptome analysis, preferentially expressed antigen of melanoma (PRAME) was selected for subsequent analyses. mRNA expression analysis of clinical samples revealed the aberrant expression of PRAME in GC tissues, and its high expression was significantly related to differentiated phenotype and vessel invasion, as well as liver metastasis. High PRAME expression was significantly associated with hepatic recurrence after curative surgery, and cumulative incidences of hepatic recurrence were significantly greater in patients with high PRAME expression compared with patients with low PRAME expression. In an in vitro analysis, overexpression was observed in all GC cell lines compared with a normal epithelial cell line. PCR array analysis revealed the coordinate expression of MMP9, OCLN, IL1RN, and MST1R. PRAME is related to the malignant potential of GC and could serve as a novel biomarker for the detection and prediction of liver metastasis.

Growth inhibitor of human hepatic carcinoma HepG2 cells by evodiamine is associated with downregulation of PRAME.

Hepatocellular carcinoma (HCC) patients have low 5-year survival due to the delayed diagnosis, so it is necessary to develop an alternative treatment. Preferentially expressed antigen of melanoma (PRAME) has a high expression in HCC patients, and the effects of evodiamine on HCC are less characterized, although evodiamine has anti-tumor activities in several tumor types. To investigate the effects of evodiamine on PRAME expression, the in vitro PRAME expression in HepG2 cells after incubated with evodiamine was determined by RT-PCR and western blot. Cell viability, migration, invasion, and apoptosis of evodiamine-incubated HepG2 cells were evaluated by Cell Counting Kit-8, wound healing, transwell assay, and Annexin V-FITC/PI double-staining assay, respectively. To evaluate the mechanism of the regulation of evodiamine on the PRAME expression, chromatin immunoprecipitation coupled with quantitative PCR was employed. Xenograft model was used to evaluate the effects of evodiamine on tumor growth, survival rate, and the PRAME expression. The PRAME expression was inhibited in evodiamine-treated HepG2 cells in vitro and in vivo. The tumor metastasis and growth were inhibited resulting from evodiamine incubation. The evodiamine inhibited the PRAME expression through trimethylation of H3K27. In this study, evodiamine contributes to in vitro and in vivo tumor cell growth inhibition. To achieve this inhibition, the PRAME expression may be repressed through trimethylation resulting from epigenetic regulation of evodiamine.

PRAME Gene Copy Number Variation Is Related to Its Expression in Multiple Myeloma.

Multiple myeloma (MM) patients commonly present abnormal expression of cancer-testis antigens, which may serve as immunotherapeutic targets and prognostic factors. We previously reported that preferentially expressed antigen of melanoma (PRAME) overexpression in bone marrow mononuclear cells is related to progression in MM patients treated with non-bortezomib-containing regimens. The mechanism underlying variations in PRAME expression remains unknown. To investigate the impact of gene copy number variation (CNV) on PRAME expression, plasma cells were sorted from 50 newly diagnosed patients and 8 healthy volunteers to measure PRAME transcript levels and gene copy numbers by real-time quantitative polymerase chain reaction. A total of 14 (28.0%), 7 (14.0%), and 29 (58.0%) patients exhibited overexpression, expression within the normal range, and low expression, respectively. PRAME overexpression was significantly related to a lower 1-year progression-free survival rate compared with PRAME low expression (20.0% vs. 88.9%, p = 0.043). The mean PRAME gene copy number relative to albumin (ALB) in normal samples was ∼1.0, whereas 4.0%, 24.0%, 70.0%, and 2.0% of patients had PRAME gene relative copy numbers of approximately 0, 0.5, 1.0, and 2.0, respectively. Patients with PRAME gene deletion (relative copy number of 0 or 0.5) had significantly higher frequency of PRAME nonoverexpression and lambda light chain expression than those with no deletion (p = 0.011 and 0.003). Thus, PRAME gene CNV occurs in MM. Gene deletion may be one mechanism leading to PRAME nonoverexpression and related to immunoglobulin lambda light chain locus rearrangement. PRAME overexpression in plasma cells might be an adverse prognostic factor for progression in MM.

Methylation pattern of preferentially expressed antigen of melanoma in acute myeloid leukemia and myelodysplastic syndromes.

Preferentially expressed antigen of melanoma (PRAME), a tumor-associated antigen, is overexpressed in a variety of hematologic malignancies with a great variation in expression. The majority of patients with acute myeloid leukemia (AML) 1-eight-twenty one (ETO)+ AML and a certain number of myelodysplastic syndromes (MDS) have an abnormally high increase in PRAME expression level. The landscape of PRAME methylation requires evaluation in order to determine the most relevant sites and the exact association of its methylation with expression level and type of disease. In the present study, bone marrow samples collected from 8 AML1-ETO+ AML, 4 MDS, 3 AML1-ETO- AML and 2 normal volunteers underwent bisulfate sequencing to analyze the methylation status of all four 5'-C-phosphate-G-3' (CpG) regions within the entire PRAME gene. The median PRAME transcript level of 15 patients was 204.5% (range, 0.02-710.3%). PRAME transcript levels were inversely associated with the degree of methylation of the -389 to -146 CpG sites (r=-0.69; P=0.002) in the 3' part of the promoter region and the +132 to +363 CpG sites (r=-0.69; P=0.006) in the exon 1b region. However, not every sample strictly followed this correlation: Certain samples with high degrees of methylation demonstrated abnormally high expression levels, and vice versa. The methylation ratios of CpG sites in exon 1a were low for all samples (range, 0.0-13.8%), and those in exon 2 were similar in 16 samples (range, 72.4-93.4%), with the exception of one patient with high expression (425.2%) and significantly low degree of methylation in the PRAME gene (22.2%). MDS patients revealed similar methylation ratios in the 3' section of the promoter region, but tended to have lower methylation ratios in the exon 1b region (P=0.62 and P=0.09, respectively) compared with those observed in AML1-ETO+ patients with AML and similar degree of PRAME overexpression. Therefore, the hypomethylation of CpG sites in the 3' part of the promoter region and in exon 1b was typically found with PRAME overexpression in AML and MDS. Methylation of other CpG islands, epigenetic and genetic mechanisms, and type of disease may also be involved.

Gene expression of MAGE-A3 and PRAME tumor antigens and EGFR mutational status in Taiwanese non-small cell lung cancer patients.

To determine the frequency of expression of the tumor-associated antigens (TAAs) melanoma-associated antigen A3 (MAGE-A3) and preferentially expressed antigen of melanoma (PRAME) and the rate of EGFR mutations in a Taiwanese non-small cell lung cancer (NSCLC) population including only adenocarcinomas and squamous cell carcinomas. Furthermore, to investigate associations between TAA expression and EGFR mutations and to evaluate these TAAs as prognostic markers for overall survival. The occurrence of single nucleotide polymorphisms in MAGEA3 and PRAME was also assessed. Archival fresh-frozen tumor tissue specimens were tested by quantitative reverse transcription polymerase chain reaction assays to detect MAGE-A3 and PRAME expression. EGFR mutations were detected by mass spectroscopy and single nucleotide polymorphisms by gene sequencing. Of the 156 adenocarcinomas examined, 3.3% expressed MAGE-A3, 32.2% expressed PRAME and 62.8% had EGFR mutations. Of the 128 squamous cell carcinomas, 29.8% expressed MAGE-A3, 59.2% expressed PRAME and 20.5% harbored EGFR mutations. TAA expression was similar across subgroups determined by patient or tumor characteristics. There was no association between TAA expression and EGFR mutation status and TAA expression was found not to be a prognostic marker for survival. Single nucleotide polymorphisms were identified, one of which with a possible impact on MAGE-A3 expression. In this NSCLC population, expression of MAGE-A3 and PRAME was more frequent in squamous cell carcinomas than in adenocarcinomas tumors. EGFR mutations were not associated with TAA expression for either histology and were three times more frequent in adenocarcinomas than in squamous cell carcinomas tumors.

Identification of Preferentially Expressed Antigen of Melanoma as a Potential Tumor Suppressor in Lung Adenocarcinoma.

BackgroundPreferentially expressed antigen of melanoma (PRAME) is known as a tumor-associated antigen that is altered in a variety of malignancies, including lung cancer. However, the role of PRAME in lung cancer remains unclear.Material/MethodsWe analyzed the expression of PRAME in human lung adenocarcinomas and studied the function of PRAME using small interfering RNA (siRNA)-induced gene knockdown in lung cancer cell lines PC9 and A549.ResultsWe found that PRAME expression is down-regulated in lung adenocarcinomas. Knockdown of PRAME promoted proliferation and suppressed apoptosis of PC9 and A549 cells.ConclusionsIn line with its roles in controlling cell growth, RPAME regulates multiple critical cell-growth related genes, including IGF1R oncogene. IGF1R up-regulation contributes to increase of cell growth upon the knockdown of PRAME. Taken together, our results suggest that PRAME has inhibitory roles in lung cancer.

Impact of Preferentially Expressed Antigen of Melanoma on the Prognosis of Hepatocellular Carcinoma

Background: Retinoids, vitamin A and its derivatives, have an antitumor effect on hepatocellular carcinoma (HCC). The function of retinoids is exerted by the complex of retinoic acid (RA) with the heterodimer of retinoid X receptor and the RA receptor. The preferentially expressed antigen of melanoma (PRAME) acts as a dominant repressor of RA signaling by binding to the complex. The significance of PRAME on the prognosis of HCC remains to be clarified. Methods: PRAME mRNA expression was examined by quantitative real-time polymerase chain reaction in both tumor and non-tumor tissues of 100 HCC patients who received surgical resection. The effect of PRAME knockdown on DR5-mediated RA transcriptional activity was examined. Results: In tumor tissues, there were significant associations among PRAME expression, clinical stage, tumor markers, and tumor numbers. In non-tumor tissues, there were significant associations among PRAME expression, overall survival, and disease-free survival. The knockdown of PRAME caused no reduction in DR5-mediated transcriptional activity of RA, suggesting that PRAME acts via other mechanisms than the DR5 RA-responsive elements. Conclusion: Our findings indicate that PRAME expression is a novel prognostic marker in HCC patients.

PRAME Expression and Its Clinical Relevance in Hodgkin's Lymphoma

Objectives: Although Hodgkin's lymphoma (HL) is one of the most curable cancers in adult patients, new targets have to be defined in cases resistant to traditional chemotherapy. The preferentially expressed antigen of melanoma (PRAME) is a cancer testis antigen and its expression is very scarce or absent in normal tissues. For this reason PRAME is a promising candidate for tumor immunotherapy. The aim of this study is to understand the correlation of PRAME expression with prognostic factors in HL, to determine the utility of PRAME as a targeted molecule for immunotherapy and to compare real-time polymerase chain reaction (real-time PCR) and immunohistochemistry (IHC) for the detection of PRAME. Methods: In 82 patients, PRAME was studied using real-time PCR and IHC. Data analyses were performed using statistical methods such as t test, Mann-Whitney U test, χ² test, Kaplan-Meier method, log-rank test and Cox regression analysis. Results: PRAME was detected in 15 (18.3%) patients using IHC and in 8 (9.8%) patients using real-time PCR. A correlation was found between PRAME positivity and higher International Prognostic Score (p = 0.039). PRAME positivity detected using real-time PCR was found to be correlated with shorter disease-free survival (DFS) and overall survival (OS, p = 0.0005). Discussion: The demonstration of PRAME especially in histiocytes and Reed-Sternberg cells may provide guidance for immunotherapy. Although PRAME positivity increases the risk for death (3.56), independent risk factors that affected DFS and OS occurred in advanced age and high-risk groups. Conclusion: Although real-time PCR is sensitive in the detection of PRAME, IHC can be another useful method. Despite the need for studies conducted on larger patient samples, PRAME expression is considered as a poor prognostic parameter in HL.

Preferentially Expressed Antigen of Melanoma (PRAME) and Wilms' Tumor 1 (WT 1) Genes Expression in Childhood Acute Lymphoblastic Leukemia, Prognostic Role and Correlation with Survival.

BACKGROUND:Acute lymphocytic leukemia (ALL) is the most common hematologic malignancy in children. In young children it is also largely curable, with more than 90% of afflicted children achieving long-term remission. PRAME (Preferentially expressed antigen of melanoma) gene belongs to Group 3 class I HLA-restricted widely expressed antigens in which genes encoding widely expressed tumor antigens have been detected in many normal tissues as well as in histologically different types of tumors with no preferential expression on a certain type of cancer. It has been found to be expressed in a variety of cancer cells as leukemia & lymphoma. PRAME monitoring can be useful for detection of minimal residual disease and subsequent relapses particularly those leukemias in which specific tumor markers are unavailable. Wilms’ tumor1 (WT1) gene was identified as a gene that plays an important role in normal kidney development and inactivation of its function was shown to result in the development of Wilms’ tumors in paediatric patients. Disruption of WT1 function has been implicated in the formation of many different tumor types.AIM:to study how PRAME & WT 1 genes expression patterns influence cancer susceptibility & prognosis.PATIENTS & METHODS:50 patients with denovo childhood acute lymphoblastic leukemia, as well as 50 age and sex matched apparently healthy volunteers were genotyped for PRAME and WT1 genes expression by reverse transcription polymerase chain reaction (RT-PCR).RESULTS:PRAME gene was expressed in 34 of the patients (68%) and WT1 gene was expressed in 26 of the patients (52%). Expression of both genes was significantly higher compared to controls (P < 0.0001). Analysis of relapse free survival among our patients revealed that patients expressing PRAME gene or WT1 gene had better relapse free survival (p value=0.02 and 0.01 respectively). Relapse free survival increased significantly among patients coexpressing PRAME and WT 1(p value =0.001).CONCLUSION:It is concluded that the expression of PRAME and WT1 genes are indicators of favorable prognosis and can be useful tools for monitoring minimal residual disease (MRD) in acute leukemia especially in patients without known genetic markers. Differential expression between acute leukemia patients and healthy volunteers suggests that the immunogenic antigens (PRAME and WT1) are potential candidates for immunotherapy in childhood acute leukemia.

Prognostic Significance of WT1 and PRAME Gene Expression in Bone Marrow Samples of MDS and AML Patients Treated with Azacytidine

Introduction: The Wilms’ tumor gene (WT1) and preferentially expressed antigen of melanoma (PRAME) gene are frequently overexpressed in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) Combined quantitation of PRAME and WT1 transcript levels could provide a powerful molecular marker in newly diagnosed MDS patients, enabling the early identification of relapse through minimal residual disease monitoring and also to enable alternate treatment strategies (Qin et al, 2012).In this study, we investigated the prognostic impact of WT1 and PRAME gene expression on MDS and AML patients treated with azacytidine.Methods:In this ethically approved, retrospective study, we studied WT1 and PRAME gene expression in the bone marrow of 20 patients with MDS and AML (unsuitable for more intensive chemotherapy) prior to starting epigenetic therapy with azacytidine (100mg/M2 sc for 5 days every 28 days). 20 patients with lymphoma not involving the bone marrow were used as a control group. Quantitative assessment of WT1 and PRAME transcript levels was performed on formalin fixed paraffin-embedded trephine biopsies, using the WT1 Ipsogen ProfileQuant and a ‘homebrew’ kit respectively. A control GADPH was used. Overall survival was extimated using the Kaplan-Meier method and compared using the log-rank test. The logistic regression model was used to correlate IPSS-R with WT-1 and PRAME gene expression levels.Results: The median age of the patient group was 71. 16 patients had MDS (RCMD: 3; RARS: 2; RAEB1: 2; RAEB2: 5; CMML: 4) and 4 had AML. Median age of the control group was 61. Median number of cycles of azacytidine was 4 (range 1-21). In 11 of the patient group and in all of the controls, neither PRAME nor WT1 gene expression was detected.. PRAME gene expression without WT1 gene expression was detected in 5 patients, whilst, in the remaining 4 patients, both PRAME and WT1 gene expression was detected. There was no significant difference in mean percentage of bone marrow blasts between patients with detectable PRAME +/- WT1 gene expression (16.67%) and those without (12.64%).Median gene expression levels of PRAME +/- WT1 significantly correlated with IPSS-R (p=0.003).Overall survival (OS) was significantly inferior in patients who were positive for PRAME +/- WT1 (p=0.0103). Median OS for patients positive for both PRAME and WT1 was 270 days and for patients who were PRAME positive and WT1 negative was 300 days, whilst patients negative for both had a median OS of 630 days (figure 1).Conclusions: Our study results show that patients who have detectable WT1 and PRAME expression in their bone marrow prior to azacytidine therapy have more advanced disease in terms of IPSS-R score and have a much inferior OS compared to patients without detectable WT1 and PRAME. Such patients should be considered for either additional or alternate therapy to single agent azacytidine. DisclosuresNo relevant conflicts of interest to declare.

Prognostic significance of PRAME expression based on immunohistochemistry for diffuse large B-cell lymphoma patients treated with R-CHOP therapy.

The preferentially expressed antigen of melanoma (PRAME), a tumor-associated antigen, is considered a prognostic marker for various human malignancies. The prognostic significance of PRAME expression for diffuse large B-cell lymphoma (DLBCL) patients treated with rituximab-containing chemotherapy has not been evaluated to date, and the ability of immunohistochemistry (IHC) to detect PRAME expression in these patients has not yet been studied, although IHC is simple to perform in clinical practice. We evaluated the prognostic significance of PRAME expression based on IHC analysis in 160 DLBCL patients treated with R-CHOP therapy. There was a significant association between higher PRAME expression and shorter progression-free survival (PFS), and a trend toward shorter overall survival (OS) in patients with higher PRAME expression than that in patients with lower PRAME expression (5-year PFS, 48.1 vs. 61.1 %; 5-year OS, 65.6 vs. 79.1 %). Patients with high PRAME expression tended to have lower chemotherapeutic responses. Thus, IHC is useful for detecting and assessing PRAME expression in DLBCL. Further, we found a positive correlation between IHC and quantitative real-time RT-PCR measurements of PRAME expression. Our findings indicate that IHC results of PRAME expression can be a novel prognostic maker in DLBCL patients treated with R-CHOP therapy.

Bortezomib improves progression-free survival in multiple myeloma patients overexpressing preferentially expressed antigen of melanoma

Background Significant efforts have been made to identify factors that differentiate patients treated with novel therapies, such as bortezomib in multiple myeloma (MM). The exact expression pattern and prognostic value of the cancer/testis antigen preferentially expressed antigen of melanoma (PRAME) in MM are unknown and were explored in this study. Methods The transcript level of PRAME was detected in bone marrow specimens from 100 newly diagnosed MM patients using real-time quantitative polymerase chain reaction, and the prognostic value of PRAME was determined through retrospective survival analysis. PRAME expression higher than the upper limit of normal bone marrow was defined as PRAME overexpression or PRAME (+). Results Sixty-two patients (62.0%) overexpressed PRAME. PRAME overexpression showed no prognostic significance to either overall survival (n=100) or progression-free survival (PFS, n=96, all P &gt;0.05) of patients. The patients were also categorized according to regimens with or without bortezomib. PRAME overexpression tended to be associated with a lower two-year PFS rate in patients treated with non-bortezomib-containing regimens (53.5% vs. 76.9%, P=0.071). By contrast, it was not associated with the two-year PFS rate in patients with bortezomib-containing regimens (77.5% vs. 63.9%, P &gt;0.05). When the patients were categorized into PRAME (+) and PRAME (-) groups, treatment with bortezomib-containing regimens predicted a higher two-year PFS rate in PRAME (+) patients (77.5% vs. 53.5%, P=0.027) but showed no significant effect on two-year PFS rate in PRAME (-) patients (63.9% vs. 76.9%, P &gt;0.05). Conclusion PRAME overexpression might be an adverse prognostic factor of PFS in MM patients treated with non-bortezomib-containing regimens. Bortezomib improves PFS in patients overexpressing PRAME.