Multi-omics analysis of overexpressed tumor-associated proteins: gene expression, immunopeptide presentation, and antibody response in oropharyngeal squamous cell carcinoma, with a focus on cancer-testis antigens.

Introduction: The human leukocyte antigen complex (HLA) is essential for inducing specific immune responses to cancer by presenting tumor-associated peptides (TAP) to T cells. Overexpressed tumor associated antigens, mainly cancer-testis antigens (CTA), are outlined as essential targets for immunotherapy in oropharyngeal squamous cell carcinoma (OPSCC). This study assessed the degree to which presentation, gene expression, and antibody response (AR) of TAP, mainly CTA, are correlated in OPSCC patients to evaluate their potential as immunotherapy targets. Materials and methods: Snap-frozen tumor (NLigand/RNA=40), healthy mucosa (NRNA=6), and healthy tonsils (NLigand=5) samples were obtained. RNA-Seq was performed using Illumina HiSeq 2500/NovaSeq 6000 and whole exome sequencing (WES) utilizing NextSeq500. HLA ligands were isolated from tumor tissue using immunoaffinity purification, UHPLC, and analyzed by tandem MS. Antibodies were measured in serum (NAb=27) utilizing the KREX™ CT262 protein array. Data analysis focused on 312 proteins (KREX™ CT262 panel + overexpressed self-proteins). Results: 183 and 94 of HLA class I and II TAP were identified by comparative profiling with healthy tonsils. Genes from 26 TAP were overexpressed in tumors compared to healthy mucosa (LFC>1; FDR<0.05). Low concordance (r=0.25; p<0.0001) was found between upregulated mRNA and class I TAP. The specific mode of correlation of TAP was found to be dependent on clinical parameters. A lack of correlation was observed both between mRNA and class II TAP, as well as between class II tumor-unique TAP (TAP-U) presentation and antibody response (AR) levels. Discussion: This study demonstrates that focusing exclusively on gene transcript levels fails to capture the full extent of TAP presentation in OPSCC. Furthermore, our findings reveal that although CTA are presented at relatively low levels, a few CTA TAP-U show potential as targets for immunotherapy.

PRAMEF12, a novel cancer/testis gene, regulates proliferation and apoptosis to promote progression of glioma.

Aim: To evaluate whether PRAMEF12 can serve as a diagnostic biomarker for glioma. Methods: We examined PRAMEF12 expression in multiple normal and glioma tissues. The diagnostic value of PRAMEF12 was evaluated using receiver operating characteristic curve analysis. The effect of PRAMEF12 ablation on proliferation, cell cycle and apoptosis was investigated. Database analyses were utilized for functional enrichment analysis. Results: PRAMEF12 expression in normal tissue was restricted to the human testis. PRAMEF12 displayed significant diagnostic value in glioma. PRAMEF12 knockdown inhibited cell proliferation, induced apoptosis and resulted in induction of S-phase cell cycle arrest. Pathway enrichment analysis indicated that PRAMEF12 may participate in cancer. Conclusion: PRAMEF12, a novel cancer/testis gene, may be a potential new diagnostic biomarker for glioma.

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How the Intrinsically Disordered N-Terminus of Cancer/Testis Antigen MAGEA10 Is Responsible for Its Expression, Nuclear Localisation and Aberrant Migration.

Melanoma-associated antigen A (MAGEA) subfamily proteins are normally expressed in testis and/or placenta. However, aberrant expression is detected in the tumour cells of multiple types of human cancer. MAGEA expression is mainly observed in cancers that have acquired malignant phenotypes, invasiveness and metastasis, and the expression of MAGEA family proteins has been linked to poor prognosis in cancer patients. All MAGE proteins share the common MAGE homology domain (MHD) which encompasses up to 70% of the protein; however, the areas flanking the MHD region vary between family members and are poorly conserved. To investigate the molecular basis of MAGEA10 expression and anomalous mobility in gel, deletion and point-mutation, analyses of the MAGEA10 protein were performed. Our data show that the intrinsically disordered N-terminal domain and, specifically, the first seven amino acids containing a unique linear motif, PRAPKR, are responsible for its expression, aberrant migration in SDS-PAGE and nuclear localisation. The aberrant migration in gel and nuclear localisation are not related to each other. Hiding the N-terminus with an epitope tag strongly affected its mobility in gel and expression in cells. Our results suggest that the intrinsically disordered domains flanking the MHD determine the unique properties of individual MAGEA proteins.

An immunoinformatics assessment of the cancer testis antigen, DDX53, as a potential early esophageal cancer antigen.

T-lymphocytes have been implicated in facilitating a pro-inflammatory, pro-tumorigenic microenvironment that worsens prognosis for esophageal carcinoma (ESCA). In this study, we identified tumor resident, T-cell receptor (TCR) complementarity determining region-3 (CDR3) amino acid sequences and employed an algorithm particularly suited to the big data setting to evaluate TCR CDR3-cancer testis antigen (CTA) chemical complementarities. Chemical complementarity of the ESCA TCR CDR3s and the cancer testis antigen DDX53 represented a disease-free survival (DFS) distinction, whereby the upper fiftieth percentile complementarity group correlated with worse DFS. The high TCR CDR3-DDX53 complementarity group also represented a greater proportion of tumor samples lacking DDX53 expression. These data and analyses raise the question of whether the TCR CDR3-DDX53 chemical complementarity assessment detected an ESCA immune response that selected for DDX53-negative cells?

Cancer/Testis Antigens as Targets for RNA-Based Anticancer Therapy.

In the last few decades, RNA-based drugs have emerged as a promising candidate in the treatment of various diseases. The introduction of messenger RNA (mRNA) as a vaccine or therapeutic agent enables the production of almost any functional protein/peptide. The key to applying RNA therapy in clinical trials is developing safe and effective delivery systems. Exosomes and lipid nanoparticles (LNPs) have been exploited as promising vehicles for drug delivery. This review discusses the feasibility of exosomes and LNPs as vehicles for mRNA delivery. Cancer/testis antigens (CTAs) show restricted expression in normal tissues and widespread expression in cancer tissues. Many of these CTAs show expression in the sera of patients with cancers. These characteristics of CTAs make them excellent targets for cancer immunotherapy. This review summarizes the roles of CTAs in various life processes and current studies on mRNAs encoding CTAs. Clinical studies present the beneficial effects of mRNAs encoding CTAs in patients with cancers. This review highlight clinical studies employing mRNA-LNPs encoding CTAs.

Characterization of Cancer/Testis Antigens as Prognostic Markers of Ovarian Cancer.

The main goal of this study was to characterize cancer/testis antigens (CTAs) as potential molecular markers of ovarian cancer. First, we gathered and analyzed a significantly large dataset of 21 selected CTAs that are encoded by 32 genes; the dataset consisted of the mutation data, expression data, and survival data of patients with ovarian cancer (n = 15,665). The 19 functionally significant missense mutations were identified in 9 CTA genes: ACRBP, CCT4, KDM5B, MAGEA1, MAGEA4, PIWIL1, PIWIL2, PRAME, and SPA17. The analysis of the mRNA expression levels of 21 CTAs in healthy and tumor ovarian tissue showed an up-regulation in the expression level of AKAP3, MAGEA4, PIWIL1, and PRAME in tumor samples and a down-regulation in the expression level of CTAG1A, CTAG1B, MAGEC1, and PIWIL2. The CCT4 up-regulation and PRAME mutations were correlated with a good prognosis for ovarian cancer, while higher levels of GAGE2A and CT45A1 mRNAs were correlated with a poor prognosis for ovarian cancer patients. Thus, GAGE2, CT45, CCT4, and PRAME cancer/testis antigens can be considered as potential prognostic markers for ovarian tumors, and GAGE2, CCT4, and PRAME were revealed to be correlated with the prognosis for ovarian cancer patients for the first time.

Aberrant activation of five embryonic stem cell-specific genes robustly predicts a high risk of relapse in breast cancers.

BACKGROUND: In breast cancer, as in all cancers, genetic and epigenetic deregulations can result in out-of-context expressions of a set of normally silent tissue-specific genes. The activation of some of these genes in various cancers empowers tumours cells with new properties and drives enhanced proliferation and metastatic activity, leading to a poor survival prognosis. RESULTS: In this work, we undertook an unprecedented systematic and unbiased analysis of out-of-context activations of a specific set of tissue-specific genes from testis, placenta and embryonic stem cells, not expressed in normal breast tissue as a source of novel prognostic biomarkers. To this end, we combined a strict machine learning framework of transcriptomic data analysis, and successfully created a new robust tool, validated in several independent datasets, which is able to identify patients with a high risk of relapse. This unbiased approach allowed us to identify a panel of five biomarkers, DNMT3B, EXO1, MCM10, CENPF and CENPE, that are robustly and significantly associated with disease-free survival prognosis in breast cancer. Based on these findings, we created a new Gene Expression Classifier (GEC) that stratifies patients. Additionally, thanks to the identified GEC, we were able to paint the specific molecular portraits of the particularly aggressive tumours, which show characteristics of male germ cells, with a particular metabolic gene signature, associated with an enrichment in pro-metastatic and pro-proliferation gene expression. CONCLUSIONS: The GEC classifier is able to reliably identify patients with a high risk of relapse at early stages of the disease. We especially recommend to use the GEC tool for patients with the luminal-A molecular subtype of breast cancer, generally considered of a favourable disease-free survival prognosis, to detect the fraction of patients undergoing a high risk of relapse.

Expression of four cancer-testis antigens in TNBC indicating potential universal immunotherapeutic targets.

OBJECTIVE: Immunotherapy is an attractive treatment for breast cancer. Cancer-testis antigens (CTAs) are potential targets for immunotherapy for their restricted expression. Here, we investigate the expression of CTAs in breast cancer and their value for prognosis. So as to hunt for a potential panel of CTAs for universal immunotherapeutic targets. MATERIAL AND METHODS: A total of 137 breast cancer tissue specimens including 51 triple-negative breast cancer (TNBC) were assessed for MAGE-A4, MAGEA1, NY-ESO-1, KK-LC-1 and PRAME expression by immunohistochemistry. The expression of PD-L1 and TILs was also calculated and correlated with the five CTAs. Clinical data were collected to evaluate the CTA's value for prognosis. Data from the K-M plotter were used as a validation cohort. RESULTS: The expression of MAGE-A4, NY-ESO-1 and KK-LC-1 in TNBC was significantly higher than in non-TNBC (P = 0.012, P = 0.005, P < 0.001 respectively). 76.47% of TNBC expressed at least one of the five CTAs. Patients with positive expression of either MAGE-A4 or PRAME had a significantly extended disease-free survival (DFS). Data from the Kaplan-Meier plotter confirm our findings. CONCLUSIONS: MAGE-A4, NY-ESO-1, PRAME and KK-LC-1 are overexpressed in breast cancer, especially in TNBC. Positive expression of MAGE-A4 or PARME may be associated with prolonged DFS. A panel of CTAs is attractive universal targets for immunotherapy.

TRB CDR3-cancer testis antigen chemical complementarity scoring for identifying productive immune responses in renal cell carcinoma.

BACKGROUND: Immunogenomics approaches to the characterization of renal cell carcinoma (RCC) have helped to better our understanding of the features of RCC immune dysfunction. However, much is still unknown with regard to specific immune interactions and their impact in the tumor microenvironment. OBJECTIVE: This study applied chemical complementarity scoring for the TRB complementarity determining region-3 (CDR3) amino acid sequences and cancer testis antigens (CTAs) to determine whether such complementarity correlated with survival and the expression of immune marker genes. METHODS: TRB recombination reads from RCC tumor samples from RNAseq files obtained from two separate databases, Moffitt Cancer Center and The Cancer Genome Atlas (TCGA), were evaluated. Chemical complementarity scores (CSs) were calculated for TRB CDR3-CTA pairs and survival assessments based on those CSs were performed. RESULTS: Moffitt Cancer Center and TCGA cases representing the upper 50th percentile of chemical CSs for TRB CDR3 amino acid sequences and the CTA POTEA were found to be associated with a better overall survival (OS) Also, greater tumor RNA expression of multiple immune signature genes, including granzyme A, granzyme B, and interferon-gamma were correlated with the higher chemical CSs. CONCLUSIONS: These results indicate that TRB CDR3-CTA chemical complementarity scoring may be useful in distinguishing RCC cases with a productive, anti-tumor immune response from cases where basic immune parameter assessments are inconsistent with a productive immune response.

The Price of Human Evolution: Cancer-Testis Antigens, the Decline in Male Fertility and the Increase in Cancer.

The increasing frequency of general and particularly male cancer coupled with the reduction in male fertility seen worldwide motivated us to seek a potential evolutionary link between these two phenomena, concerning the reproductive transcriptional modules observed in cancer and the expression of cancer-testis antigens (CTA). The phylostratigraphy analysis of the human genome allowed us to link the early evolutionary origin of cancer via the reproductive life cycles of the unicellulars and early multicellulars, potentially driving soma-germ transition, female meiosis, and the parthenogenesis of polyploid giant cancer cells (PGCCs), with the expansion of the CTA multi-families, very late during their evolution. CTA adaptation was aided by retrovirus domestication in the unstable genomes of mammals, for protecting male fertility in stress conditions, particularly that of humans, as compensation for the energy consumption of a large complex brain which also exploited retrotransposition. We found that the early and late evolutionary branches of human cancer are united by the immunity-proto-placental network, which evolved in the Cambrian and shares stress regulators with the finely-tuned sex determination system. We further propose that social stress and endocrine disruption caused by environmental pollution with organic materials, which alter sex determination in male foetuses and further spermatogenesis in adults, bias the development of PGCC-parthenogenetic cancer by default.

Cancer/testis antigens: promising immunotherapy targets for digestive tract cancers.

Digestive tract cancers, including esophageal, gastric, and colorectal cancers, are the major cause of death among cancer patients worldwide due to the heterogeneity of cancer cells, which limits the effectiveness of traditional treatment methods. Immunotherapy represents a promising treatment strategy for improving the prognosis of patients with digestive tract cancers. However, the clinical application of this approach is limited by the absence of optimal targets. Cancer/testis antigens are characterized by low or absent expression in normal tissues, but high expression in tumor tissues, making them an attractive target for antitumor immunotherapy. Recent preclinical trials have shown promising results for cancer/testis antigen-targeted immunotherapy in digestive cancer. However, practical problems and difficulties in clinical application remain. This review presents a comprehensive analysis of cancer/testis antigens in digestive tract cancers, covering their expression, function, and potential as an immunotherapy target. Additionally, the current state of cancer/testis antigens in digestive tract cancer immunotherapy is discussed, and we predict that these antigens hold great promise as an avenue for breakthroughs in the treatment of digestive tract cancers.

Expression of cancer-testis antigens in the immune microenvironment of non-small cell lung cancer.

The antigenic repertoire of tumors is critical for successful anti-cancer immune response and the efficacy of immunotherapy. Cancer-testis antigens (CTAs) are targets of humoral and cellular immune reactions. We aimed to characterize CTA expression in non-small cell lung cancer (NSCLC) in the context of the immune microenvironment. Of 90 CTAs validated by RNA sequencing, eight CTAs (DPEP3, EZHIP, MAGEA4, MAGEB2, MAGEC2, PAGE1, PRAME, and TKTL1) were selected for immunohistochemical profiling in cancer tissues from 328 NSCLC patients. CTA expression was compared with immune cell densities in the tumor environment and with genomic, transcriptomic, and clinical data. Most NSCLC cases (79%) expressed at least one of the analyzed CTAs, and CTA protein expression correlated generally with RNA expression. CTA profiles were associated with immune profiles: high MAGEA4 expression was related to M2 macrophages (CD163) and regulatory T cells (FOXP3), low MAGEA4 was associated with T cells (CD3), and high EZHIP was associated with plasma cell infiltration (adj. P-value < 0.05). None of the CTAs correlated with clinical outcomes. The current study provides a comprehensive evaluation of CTAs and suggests that their association with immune cells may indicate in situ immunogenic effects. The findings support the rationale to harness CTAs as targets for immunotherapy.

Sperm protein antigen 17 and Sperm flagellar 1 cancer testis antigens are expressed in a rare case of ciliated foregut cyst of the common hepatic duct.

INTRODUCTION: Ciliated foregut cysts (CFCs) are frequently described in liver, pancreas and gallbladder and generally considered benign although one case of squamous cell metaplasia and five cases of squamous cell carcinoma arising from a ciliated hepatic foregut cyst have been reported. Here we explore two cancer-testis antigens (CTAs), Sperm protein antigen 17 (SPA17) and Sperm flagellar 1 (SPEF1) expression in a rare case of CFC of the common hepatic duct MATERIALS AND METHODS: 3 µm-thick CFC sections were immunohistochemically treated with antibodies raised against human SPA17 or SPEF1. In silico Protein-Protein Interaction (PPI) network and differential protein expression were also investigated RESULTS: Immunohistochemistry revealed SPA17 and SPEF1 in the cytoplasm of ciliated epithelium. SPA17, but not SPEF1, was also detected in cilia. The PPI networks demonstrated that other CTAs are significantly predicted functional partners with SPA17 and SPEF1. The differential protein expression demonstrated that SPA17 was higher in breast cancer, cholangiocarcinoma, liver hepatocellular carcinoma, uterine corpus endometrial carcinoma, gastric adenocarcinoma, cervical squamous cell carcinoma, bladder urothelial carcinoma. SPEF1 expression was higher in breast cancer, cholangiocarcinoma, uterine corpus endometrial carcinoma and kidney renal papillary cell carcinoma CONCLUSIONS: Our study suggests that further characterization of SPA17 and SPEF1 in patients with CFCs might provide significant insights to understand the mechanisms underlying their potential to malignant transformation.

Electrostatic Complementarities of Glioblastoma-Resident T-Cell Receptors and Cancer Testis Antigens Linked to Poor Outcomes and High Levels of Sphingosine Kinase-2 Expression.

INTRODUCTION: Glioblastoma (GBM) is the most aggressive primary brain tumor in adults. Despite a growing understanding of glioblastoma pathology, the prognosis remains poor. METHODS: In this study, we used a previously extensively benchmarked algorithm to retrieve immune receptor (IR) recombination reads from GBM exome files available from the cancer genome atlas. The T-cell receptor complementarity determining region-3 (CDR3) amino acid sequences that represent the IR recombination reads were assessed and used for the generation of chemical complementarity scores (CSs) that represent potential binding interactions with cancer testis antigens (CTAs), which is an approach particularly suited to a big data setting. RESULTS: The electrostatic CSs representing the TRA and TRB CDR3s and the CTAs, SPAG9, GAGE12E, and GAGE12F, indicated that an increased electrostatic CS was associated with worse disease-free survival (DFS). We also assessed the RNA expression of immune marker genes, which indicated that a high-level expression of SPHK2 and CIITA genes also correlated with high CSs and worse DFS. Furthermore, apoptosis-related gene expression was revealed to be lower when the TCR CDR3-CTA electrostatic CSs were high. CONCLUSION: Adaptive IR recombination reads from exome files have the potential to aid in GBM prognoses and may provide opportunities to detect unproductive immune responses.

Identification of Cancer/Testis Antigens Related to Gastric Cancer Prognosis Based on Co-Expression Network and Integrated Transcriptome Analyses.

Background: Gastric cancer is a worldwide life-threatening cancer. The underlying cause of it is still unknown. We have noticed that some cancer/testis antigens (CTAs) are up-regulated in gastric cancer. The role of these genes in gastric cancer development is not fully understood. The main aim of the current study was to comprehensively investigate CTAs' expression and function in stomach adenocarcinoma (STAD). Materials and Methods: A comprehensive list of CTA genes was compiled from different databases. Transcriptome profiles of STAD were downloaded from the cancer genome atlas (TCGA) database and analyzed. Differentially-expressed CTAs were identified. Pathway enrichment analysis, weighted gene correlation network analysis (WGCNA), and overall survival (OS) analysis were performed on differentially-expressed CTA genes. Results: Pathway enrichment analysis indicates that CTA genes are involved in protein binding, ribonucleic acid processing, and reproductive tissues. WGCNA showed that six differentially-expressed CTA genes, namely Melanoma antigen gene (MAGE) family member A3, A6, A12 and chondrosarcoma associated gene (CSAG) 1, 2, and 3, were correlated. Up-regulation of MAGEA11, MAGEC3, Per ARNT SIM domain containing 1 (PASD1), placenta-specific protein 1 (PLAC1) and sperm protein associated with the nucleus X-linked family member (SPANXB1) were significantly associated with lower OS of patients. Conclusion: MAGEA11, MAGEC3, PASD1, PLAC1, and SPANXB1 can be investigated as prognostic biomarkers in basic and clinical studies. Further functional experiments are needed to understand the exact interaction mechanisms of these genes.

Gene-based cancer-testis antigens as prognostic indicators in hepatocellular carcinoma.

Cancer/testis antigens (CTAs) are reproductive tissue-restricted genes, frequently ectopic expressed in tumors. CTA genes associate with a poor prognosis in some solid tumors, due to their potential roles in the tumorigenesis and progression. However, whether CTAs relate with hepatocellular carcinoma (HCC) remains unclear. In this study, the prognostic signatures based on CTA genes were investigated and validated in three cohorts including Chinese HCC patients with hepatitis B virus infection (CHCC-HBV), International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA) cohorts. Univariate, LASSO, and multivariate Cox regression analyses were used to screen prognostic genes and develop the prognostic gene signature. A prognosis model was established with six CTA genes (SSX1, CTCFL, OIP5, CEP55, NOL4, and TPPP2) in CHCC-HBV cohort, and further validated in the ICGC and TCGA cohorts. The CTA signature was an essential prognostic predictor independent of other clinical pathological factors. High-risk group exhibited up-regulated cell cycle-related and tumor-related pathways and more M0 macrophage, activated mast cell, activated memory CD4+ T cell, and memory B cell infiltration. Furthermore, CTA signature correlated with the sensitivity to multiple chemotherapy drugs. Our results highlighted that the CTA gene profiling was a prognostic assessment tool for HCC patients.

Identification of TSGA10 and GGNBP2 splicing variants in 5' untranslated region with distinct expression profiles in brain tumor samples.

Introduction: Brain tumors (BTs) are perceived as one of the most common malignancies among children. The specific regulation of each gene can play a critical role in cancer progression. The present study aimed to determine the transcripts of the TSGA10 and GGNBP2 genes, considering the alternative 5'UTR region, and investigating the expression of these different transcripts in BTs. Material and methods: Public data on brain tumor microarray datasets in GEO were analyzed with R software to evaluate the expression levels of TSGA10 and GGNBP2 genes (the Pheatmap package in R was also used to plot DEGs in a heat map). In addition, to validate our in-silico data analysis, RT-PCR was performed to determine the splicing variants of TSGA10 and GGNBP2 genes in testis and brain tumor samples. The expression levels of splice variants of these genes were analyzed in 30 brain tumor samples and two testicular tissue samples as a positive control. Results: In silico results show that the differential expression levels of TSGA10 and GGNBP2 were significant in the GEO datasets of BTs compared to normal samples (with adjusted p-value<0.05 and log fold change > 1). This study's experimental results showed that the TSGA10 gene produces four different transcripts with two distinct promoter regions and splicing exon 4. The relative mRNA expression of transcripts without exon 4 was higher than transcripts with exon 4 in BT samples (p-value<001). In GGNBP2, exon 2 in the 5'UTR region and exon 6 in the coding sequence were spliced. The expression analysis results showed that the relative mRNA expression of transcript variants without exon 2 was higher than other transcript variants with exon 2 in BT samples (p-value<001). Conclusion: The decreased expression levels of transcripts with longer 5'UTR in BT samples than in testicular or low-grade brain tumor samples may decrease their translation efficiency. Therefore, decreased amounts of TSGA10 and GGNBP2 as potential tumor suppressor proteins, especially in high-grade brain tumors, may cause cancer development by angiogenesis and metastasis.

Biology of Cancer-Testis Antigens and Their Therapeutic Implications in Cancer.

Tumour-specific antigens have been an area of interest in cancer therapy since their discovery in the middle of the 20th century. In the era of immune-based cancer therapeutics, redirecting our immune cells to target these tumour-specific antigens has become even more relevant. Cancer-testis antigens (CTAs) are a class of antigens with an expression specific to the testis and cancer cells. CTAs have also been demonstrated to be expressed in a wide variety of cancers. Due to their frequency and specificity of expression in a multitude of cancers, CTAs have been particularly attractive as cancer-specific therapeutic targets. There is now a rapid expansion of CTAs being identified and many studies have been conducted to correlate CTA expression with cancer and therapy-resistant phenotypes. Furthermore, there is an increasing number of clinical trials involving using some of these CTAs as molecular targets in pharmacological and immune-targeted therapeutics for various cancers. This review will summarise the current knowledge of the biology of known CTAs in tumorigenesis and the regulation of CTA genes. CTAs as molecular targets and the therapeutic implications of these CTA-targeted anticancer strategies will also be discussed.

A Highly Sensitive Flow Cytometric Approach to Detect Rare Antigen-Specific T Cells: Development and Comparison to Standard Monitoring Tools.

Personalized vaccines against patient-unique tumor-associated antigens represent a promising new approach for cancer immunotherapy. Vaccine efficacy is assessed by quantification of changes in the frequency and/or the activity of antigen-specific T cells. Enzyme-linked immunosorbent spot (ELISpot) and flow cytometry (FCM) are methodologies frequently used for assessing vaccine efficacy. We tested these methodologies and found that both ELISpot and standard FCM [monitoring CD3/CD4/CD8/IFNγ/Viability+CD14+CD19 (dump)] demonstrate background IFNγ secretion, which, in many cases, was higher than the antigen-specific signal measured by the respective methodology (frequently ranging around 0.05-0.2%). To detect such weak T-cell responses, we developed an FCM panel that included two early activation markers, 4-1BB (CD137) and CD40L (CD154), in addition to the above-cited markers. These two activation markers have a close to zero background expression and are rapidly upregulated following antigen-specific activation. They enabled the quantification of rare T cells responding to antigens within the assay well. Background IFNγ-positive CD4 T cell frequencies decreased to 0.019% ± 0.028% and CD8 T cells to 0.009% ± 0.013%, which are 19 and 13 times lower, respectively, than without the use of these markers. The presented methodology enables highly sensitive monitoring of T-cell responses to tumor-associated antigens in the very low, but clinically relevant, frequencies.

Myxoid Liposarcomas: Systemic Treatment Options.

OPINION STATEMENT: Myxoid/round-cell liposarcoma (MRCL) account for 30% of liposarcomas and are the most chemo-sensitive subtype of liposarcoma. The 5-year local relapse and distant metastasis rates are 10% and 20%, respectively. In the advanced setting, the first-line median progression-free survival and overall survival is 9 and 30 months, respectively. The overall response rate (ORR) by RECIST with anthracycline-based chemotherapy is around 40% and with trabectedin is 20%, although response is higher when captured by CHOI criteria. Anthracycline-based combination chemotherapy regimens remain the standard of care first-line treatment option. However, trabectedin is also effective and may be considered in the first-line setting when anthracyclines cannot be prescribed. Beyond chemotherapy, new therapeutic classes are being developed, including autologous adoptive modified T cell receptor cellular therapies which have shown promising results thus far. These new therapies utilize the immunogenic potential of cancer testis antigens, NY-ESO-1 and MAGE-A4, which are expressed in the vast majority of MRCL. Early phase trials have shown encouraging results with up to 40% ORR and a median progression-free survival up to 8.7 months. Other innovative strategies are being developed, tailored to the molecular biology of MRCL. This review summarizes current evidence for the use of standard chemotherapy and the new biomarker-selected treatments under development.