CXorf61 is a target for T cell based immunotherapy of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is a high medical need disease with limited treatment options. CD8+ T cell-mediated immunotherapy may represent an attractive approach to address TNBC. The objectives of this study were to assess the expression of CXorf61 in TNBCs and healthy tissues and to evaluate its capability to induce T cell responses. We show by transcriptional profiling of a broad comprehensive set of normal human tissue that CXorf61 expression is strictly restricted to testis. 53% of TNBC patients express this antigen in at least 30% of their tumor cells. In CXorf61-negative breast cancer cell lines CXorf61 expression is activated by treatment with the hypomethylating agent 5-aza-2'-deoxycytidine. By vaccination of HLA-A*02-transgenic mice with CXorf61 encoding RNA we obtained high frequencies of CXorf61-specific T cells. Cloning and characterization of T cell receptors (TCRs) from responding T cells resulted in the identification of the two HLA-A*0201-restricted T cell epitopes CXorf6166-74 and CXorf6179-87. Furthermore, by in vitro priming of human CD8+ T cells derived from a healthy donor recognizing CXorf6166-74 we were able to induce a strong antigen-specific immune response and clone a human TCR recognizing this epitope. In summary, our data confirms this antigen as promising target for T cell based therapies.

Genes involved in the metastatic cascade of medullary thyroid tumours.

The process of how a benign tumour turns invasive and capable to survive in distant organs remains poorly understood, despite the evidence that metastasis formation is the primary cause of cancer patient mortality. This ignorance is partly due to the lack of appropriate animal models from which to investigate this complex process. The retinoblastoma (Rb) tumour suppressor pathway (pRb/E2F) is mutated in almost all human tumours, and a number of laboratories have now established pRb- or E2F-deficient mouse models. Consistent with the role of mutation in retinoblastoma in cancer biology, Rb heterozygous mice are prone to develop tumours. Among the ensuing tumours, the medullary thyroid carcinomas (MTCs) have a lessened tendency to form secondary cancers and metastases. Intriguingly, if an E2f3 mutation is introduced in this genetic background, more aggressive MTCs develop, which metastasize more frequently. Gene chip microarrays, however, provide an unbiased approach for examining the genome-wide expression levels and enable identification of a large set of metastasis-enriched gene sets. The identified genes may simply represent putative markers of the disease stage. Alternatively, genes may be identified that causally determine a link to the onset of metastasis. We describe the use of gene chip microarrays for identification of putative markers enriched in metastatic mouse MTCs. The chapter details how the most promising candidates are verified using additional methods, such as quantitative real-time PCR. In this case, co-transfection of the E2F-transcription factor using a heterologous reporter gene system is suggestive of E2Fs directly regulating putative metastasis markers.

Peripheral benzodiazepine receptor ligands induce apoptosis and cell cycle arrest in human hepatocellular carcinoma cells and enhance chemosensitivity to paclitaxel, docetaxel, doxorubicin and the Bcl-2 inhibitor HA14-1.

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is one of the most common causes of cancer deaths worldwide. Thus, novel therapies are urgently needed. A promising approach is the use of peripheral benzodiazepine receptor (PBR) ligands which inhibit the proliferation of various tumors. METHODS: PBR expression both in human HCC cell lines and in tumor specimens of HCC patients was analyzed by RT-PCR and immunostaining. To evaluate PBR ligands for the treatment of HCC, we tested their effects on human HCC cells. RESULTS: PBR was localized to the mitochondria both of HCC cell lines and tumor tissues of HCC patients. In contrast, normal liver did not express PBR. PBR ligands inhibited the proliferation of HCC cell lines by inducing apoptosis and cell cycle arrest. Apoptosis was characterized by a breakdown of the mitochondrial membrane potential, caspase-3 activation and nuclear degradation. Furthermore, pro-apoptotic Bax was overexpressed while anti-apoptotic Bcl-2 and Bcl-X(L) were suppressed. Cell cycle was arrested both at the G1/S- and G2/M-checkpoints. Synergistic anti-neoplastic effects were obtained by a combination of PBR ligands with cytostatic drugs (paclitaxel, docetaxel, doxorubicin), or with an experimental Bcl-2 inhibitor. CONCLUSIONS: This is the first report on the induction of apoptosis and cell cycle arrest by PBR ligands in HCC cells. Moreover, PBR ligands sensitized HCC cells to taxans and doxorubicin.