Epigenetic treatment combinations to effectively target cisplatin-resistant germ cell tumors: past, present, and future considerations.

BACKGROUND: Type II germ cell tumors represent the most common solid malignancy in men aged 15-45 years. Despite high cure rates of >90% over all stages, 10-15% of advanced patients develop treatment resistance and potentially succumb to their disease. Treatment of refractory germ cell tumors remains unsatisfactory, and new approaches are needed to further improve outcomes. OBJECTIVES: With this narrative review, we highlight epigenetic mechanisms related to resistance to standard systemic treatment, which may act as promising targets for novel combined epigenetic treatment approaches. MATERIALS AND METHODS: A comprehensive literature search of PubMed and MEDLINE was conducted to identify original and review articles on resistance mechanisms and/or epigenetic treatment of germ cell tumors in vitro and in vivo. Review articles were hand-searched to identify additional articles. RESULTS: Distinct epigenetic phenomena have been linked to chemotherapy resistance in germ cell tumors, among which DNA hypermethylation, histone acetylation, and bromodomain proteins appear as promising targets for therapeutic exploitation. Inhibitors of key regulators, for example DNA methyltransferases (e.g. decitabine, guadecitabine), histone deacetylases (e.g. romidepsin), and bromodomain proteins (e.g. JQ1) decreased cell viability, triggered apoptosis, and growth arrest. Additionally, these epigenetic drugs induced differentiation and led to loss of pluripotency and re-sensitization towards cisplatin in cell lines and animal models. DISCUSSION: Epigenetic treatments hold promise to (i) reduce the treatment burden of and (ii) overcome resistance to standard cisplatin-based chemotherapy. Combined approaches may enhance activity, while the ideal target and treatment combination of epigenetic drugs, either with another epigenetic agent or conventional cytotoxic agents need to be defined. CONCLUSION: Epigenetic (combination) treatment for germ cell tumors should be further explored in pre-clinical and clinical research for its potential to further improve germ cell tumor treatment.

Breath analysis as promising indicator of hemodialysis efficiency.

BACKGROUND: The measurement of trimethylamine and isoprene in exhaled breath collected from dialysed patients indicates the changes in concentration of both compounds during dialysis. The aim of the presented study was to confirm diagnostic usefulness of TMA and isoprene detected in breath, as potential biomarkers of hemodialysis efficiency. METHODS: The samples of exhaled breath were collected from 22 dialyzed patients (9 women, 13 men) before and after hemodialysis (HD). All analyses were carried out using a gas chromatograph equipped with a mass spectrometer. Thermal desorption was used as breath sample enrichment method. RESULTS: Chromatographic analysis of breath samples indicated statistically significant differences in trimethylamine (TMA) and 2-methyl-1,3-butadiene (isoprene) concentrations in patients' breath collected before and after HD. TMA concentrations measured in breath samples, before dialysis, ranged from 0.024 to 0.461 nmol/L. After dialysis, the values of detected TMA were lower versus output values and ranged from 0.008 to 0.050 nmol/L. Isoprene concentrations before dialysis were present in the range from 0.236 to 9.718 nmol/L, after dialysis in the range from 0.478 to 26.182 nmol/L. Additionally, the dependences of TMA and isoprene concentrations, detected in breath with renal efficiency parameters detected in blood, were studied. The relationships between TMA and urea (r = 0.67; p < 0.00001) and creatinine (r = 0.61; p = 0.00002) were checked. In case of isoprene considerably higher concentrations were observed after dialysis, but no statistically significant correlation of isoprene with blood parameters was noticed. CONCLUSION: The observed decrease of TMA concentrations during dialysis could be useful as a measure of dialysis efficiency. The explanation of isoprene increase in breath during dialysis requires further investigation.

The application of chromatographic breath analysis in the search of volatile biomarkers of chronic kidney disease and coexisting type 2 diabetes mellitus.

Chromatographic studies on breath composition are aimed at finding volatile markers useful for medical diagnostics or in screening investigations. Studies leading to the development of screening breath tests are especially important for the diagnostics of chronic kidney disease (CKD) and type 2 diabetes mellitus (T2DM). The aim of the presented study was to confirm diagnostic usefulness of chosen volatile compounds detected in breath, which are suggested as potential biomarkers of renal dysfunction and diabetes. Breath analysis were carried out in three groups: 10 healthy volunteers, 10 patients with CKD and 10 patients with CKD and T2DM. All exhaled air samples were analyzed using gas chromatograph (Agilent 6890GC) coupled with mass spectrometer (5975MSD). Thermal desorption was applied as the enrichment method. TMA was detected only in CKD patients. Higher breath concentrations of methanethiol (MeSH) were observed in CKD patients with coexisting diabetes than in patients with renal dysfunction only or in the healthy group. There was a tendency of increasing MeSH concentration in breath with increasing total glutathione in plasma (r=0.53, p=0.0026). Also, a trend of increasing dimethylsulfide (DMS) levels detected in breath was noticed with an increase of hydrogen sulfide concentration in plasma (r=0.74; p=0.00001) as well as with aspartate aminotransferase (AST), (r=0.61; p=0.001). The presented results suggest the possibility of applying TMA, MeSH, and DMS detection in breath as diagnostic methods.

Stem cell-like transcriptional reprogramming mediates metastatic resistance to mTOR inhibition.

Inhibitors of the mechanistic target of rapamycin (mTOR) are currently used to treat advanced metastatic breast cancer. However, whether an aggressive phenotype is sustained through adaptation or resistance to mTOR inhibition remains unknown. Here, complementary studies in human tumors, cancer models and cell lines reveal transcriptional reprogramming that supports metastasis in response to mTOR inhibition. This cancer feature is driven by EVI1 and SOX9. EVI1 functionally cooperates with and positively regulates SOX9, and promotes the transcriptional upregulation of key mTOR pathway components (REHB and RAPTOR) and of lung metastasis mediators (FSCN1 and SPARC). The expression of EVI1 and SOX9 is associated with stem cell-like and metastasis signatures, and their depletion impairs the metastatic potential of breast cancer cells. These results establish the mechanistic link between resistance to mTOR inhibition and cancer metastatic potential, thus enhancing our understanding of mTOR targeting failure.

Expression of the tumour suppressor gene CADM1 is associated with favourable outcome and inhibits cell survival in neuroblastoma.

Cell adhesion molecule 1 (CADM1) is a putative tumour suppressor gene, which is downregulated in many solid tumours. In neuroblastoma, loss of CADM1 expression has recently been found in disseminated tumours with adverse outcome, prompting us to investigate its role in neuroblastoma tumour progression. Oligonucleotide-microarray analysis of 251 neuroblastoma specimens demonstrated that CADM1 downregulation is associated with unfavourable prognostic markers like disseminated stage 4, age >18 months, MYCN amplification and chromosome 11q alterations (P<0.001 each). Furthermore, low CADM1 expression was significantly correlated with unfavourable gene expression-based classification (P<0.001) and adverse patient outcome (P<0.001). Bisulphite sequencing and genetic analysis of 18 primary neuroblastomas suggested that neither haploinsufficiency nor hypermethylation is regularly involved in CADM1 gene silencing in neuroblastoma, which is in contrast to results obtained in other malignancies. In addition, no mutations disrupting the CADM1 reading frame were found in 25 primary neuroblastomas. Over-expression of CADM1 in neuroblastoma cells resulted in significant reduction of proliferation, viability and colony formation in soft agar. Collectively, our results suggest that downregulation of CADM1 tumour suppressor gene expression is a critical event in neuroblastoma pathogenesis resulting in tumour progression and unfavourable patient outcome.

MYCN-status in neuroblastoma: characteristics of tumours showing amplification, gain, and non-amplification.

While the role of MYCN-amplification (MNA) for risk assessment in neuroblastoma is undisputed, the phenomenon of gene copy excess below the amplification threshold is rarely described. To discuss biological characteristics and the clinical impact of the so-called MYCN-gain versus amplified or non-amplified cases, we investigated the MYCN status of 659 patients uniformly analysed by fluorescence in situ hybridisation. The number of MYCN-amplified tumours in our cohort was 18% (116/659); an additional 38 tumours (6%) displayed MYCN-gain. Both alterations were associated with an advanced stage disease, an increased patient age and further chromosomal alterations. Most of the amplified neuroblastomas displayed 1p aberrations, whereas MYCN-gain tumours correlated with 11q alterations. In contrast to the amplified cases, tumours with gain displayed no increased MYCN RNA levels. MNA versus non-amplification discriminated between good and poor outcomes, independent of stage, age and the degree of amplification. However, patients with amplified tumours showed a significantly better outcome when this was combined with non-stage 4 disease and age <1 year versus stage 4 and age < 1 year. Although MYCN-gain was associated with poor event-free-survival (EFS) in stages 1-3, 4S (P=0.005), this might be related to associated genetic aberrations and not to the MYCN-gain itself. A survival difference between neuroblastomas with gain and single copy MYCN could not be delineated. In conclusion, MNA predicts a poor outcome for neuroblastoma patients of all stages and age. MYCN-gain is also a characteristic feature of advanced stage tumours and older patients, but is not associated with higher MYCN expression and appears not to be discriminative in predicting patient outcome.