Ketoprofen Combined with UVA Irradiation Exerts Higher Selectivity in the Mode of Action against Melanotic Melanoma Cells than against Normal Human Melanocytes

Malignant melanoma is responsible for the majority of skin cancer-related deaths. The methods of cancer treatment include surgical removal, chemotherapy, immunotherapy, and targeted therapy. However, neither of these methods gives satisfactory results. Therefore, the development of new anticancer therapeutic strategies is very important and may extend the life span of people suffering from melanoma. The aim of this study was to examine the effect of ketoprofen (KTP) and UVA radiation (UVAR) therapy on cell proliferation, apoptosis, and cell cycle distribution in both melanotic melanoma cells (COLO829) and human melanocytes (HEMn-DP) in relation to its supportive effect in the treatment of melanoma. The therapy combining the use of pre-incubation with KTP and UVAR causes a significant increase in the anti-proliferative properties of ketoprofen towards melanoma cells and the co-exposure of melanotic melanoma cells induced apoptosis shown as the mitochondrial membrane breakdown, cell-cycle deregulation, and DNA fragmentation. Moreover, co-treatment led to GSH depletion showing its pro-apoptotic effect dependent on ROS overproduction. The treatment did not show a significant effect on normal cells—melanocytes—which indicates its high selectivity. The results suggest a possible benefit from the use of the ketoprofen and ultraviolet A irradiation as a new concept of melanotic melanoma therapy.

Advances in Pyrazole Based Scaffold as Cyclin-Dependent Kinase 2 Inhibitors for the Treatment of Cancer

: The transformation of a normal cell into a tumor cell is one of the initial steps in cell cycle deregulation. The cell cycle is regulated by cyclin-dependent kinases (CDKs) that belong to the protein kinase family. CDK2 is an enchanting target for specific genotypes tumors since cyclin E is selective for CDK2 and the deregulation of specific cancer forms. Thus, CDKs inhibitor specifically CDK2/cyclin A-E has the potential to be a valid cancer target as per the currently undergoing clinical trials. Mostly pyrazole scaffolds have shown selectivity and potency for CDK2 inhibitors. This review demonstrates pyrazole and pyrazole fused with other heterocyclic rings for anti-proliferative activity. Based on the in vitro and molecular docking studies, the IC50 value of various hybrids is revealed to display the most potent analogs for CDK2 inhibition. Thus, the review emphasizes various lead analogs of pyrazole hybrids which can be found to be very potent and selective for anti-cancer drugs.

Transcription/Replication Conflicts in Tumorigenesis and Their Potential Role as Novel Therapeutic Targets in Multiple Myeloma

Plasma cells (PCs) have an essential role in humoral immune response by secretion of antibodies, and represent the final stage of B lymphocytes differentiation. During this differentiation, the pre-plasmablastic stage is characterized by highly proliferative cells that start to secrete immunoglobulins (Igs). Thus, replication and transcription must be tightly regulated in these cells to avoid transcription/replication conflicts (TRCs), which could increase replication stress and lead to genomic instability. In this review, we analyzed expression of genes involved in TRCs resolution during B to PC differentiation and identified 41 genes significantly overexpressed in the pre-plasmablastic stage. This illustrates the importance of mechanisms required for adequate processing of TRCs during PCs differentiation. Furthermore, we identified that several of these factors were also found overexpressed in purified PCs from patients with multiple myeloma (MM) compared to normal PCs. Malignant PCs produce high levels of Igs concomitantly with cell cycle deregulation. Therefore, increasing the TRCs occurring in MM cells could represent a potent therapeutic strategy for MM patients. Here, we describe the potential roles of TRCs resolution factors in myelomagenesis and discuss the therapeutic interest of targeting the TRCs resolution machinery in MM.

OTEH-10. Evolutionary trajectory of epigenomic of gliomas

Gliomas are the most common malignant brain tumor, have an aggressive behavior, and invariably relapse and progress. Despite the recent advancements, little is known about the role of the epigenome in glioma disease progression and recurrence. To investigate the molecular dynamics over time and in response to therapeutic pressures, the Glioma Longitudinal AnalySiS (GLASS) Consortium, a multinational collaboration, is investigating epigenome-wide molecular data from primary and recurrent matched pairs, including IDH mutant (IDHmut) and IDH wildtype (IDHwt) gliomas. We have compiled a total of 357 samples comprising 143 primary-recurrent pairs profiled by DNA methylation, of which 157 samples have genomic data (WXS/WGS) and 120 have transcriptomic data (RNAseq). IDHwt gliomas have a distinct epigenetic evolution compared to IDHmut after treatment. IDHwt gliomas are more epigenetically stable over time, while IDHmut gliomas display a loss of DNA methylation throughout disease progression. Next, we investigated the molecular drivers of longitudinal gliomas by integration of DNA methylation and gene expression data. We identified epigenetic activation of cell cycle pathways in recurrent IDHmut compared to initial tumors. Transcription factors musculin, ZNF367, and ZNF682 are enriched among recurrent IDHmut gliomas and potentially regulate IDHmut recurrence and/or progression. We next used a DNA methylation-based deconvolution approach to estimate the tumor microenvironment (TME) composition. We found that the TME among IDHmut subtypes (Codel, GCIMP-high, and GCIMP-low) presented less immune infiltration than IDHwt (Classic-like, Mesenchymal-like, and PA-like). Post-treatment, we found a decrease of CD4+T and an increase of CD8+T cells in IDHmut. In conclusion, IDHmut gliomas present a more unstable epigenome, while the epigenome of IDHwt gliomas seems relatively preserved after treatment. We identified potential master regulators of cell cycle deregulation of IDHmut recurrence. Finally, the TME differs across IDHmut and IDHwt gliomas and the cell composition changes over time.

Epigenetic mechanisms underlying prostate cancer radioresistance

Radiotherapy (RT) is one of the mainstay treatments for prostate cancer (PCa), a highly prevalent neoplasm among males worldwide. About 30% of newly diagnosed PCa patients receive RT with a curative intent. However, biochemical relapse occurs in 20–40% of advanced PCa treated with RT either alone or in combination with adjuvant-hormonal therapy. Epigenetic alterations, frequently associated with molecular variations in PCa, contribute to the acquisition of a radioresistant phenotype. Increased DNA damage repair and cell cycle deregulation decreases radio-response in PCa patients. Moreover, the interplay between epigenome and cell growth pathways is extensively described in published literature. Importantly, as the clinical pattern of PCa ranges from an indolent tumor to an aggressive disease, discovering specific targetable epigenetic molecules able to overcome and predict PCa radioresistance is urgently needed. Currently, histone-deacetylase and DNA-methyltransferase inhibitors are the most studied classes of chromatin-modifying drugs (so-called ‘epidrugs’) within cancer radiosensitization context. Nonetheless, the lack of reliable validation trials is a foremost drawback. This review summarizes the major epigenetically induced changes in radioresistant-like PCa cells and describes recently reported targeted epigenetic therapies in pre-clinical and clinical settings.

Deregulation of cell cycle and related microRNA expression induced by vinyl chloride monomer in hepatocytes of rats

Vinyl chloride (VC) is a confirmed human carcinogen associated with hepatocellular carcinoma and angiosarcoma. However, the role of microRNAs (miRNAs) in liver cell cycle changes under VC exposure remains unclear, which prevents research on the mechanism of VC-induced carcinogenesis. In this study, male rats were injected intraperitoneally with VC (0, 5, 25, and 125 mg/kg body weight) for 6, 8, and 12 weeks. Cell cycle analysis of liver cells, miRNA-222, miRNA-199a, miRNA-195, and miRNA-125b expression in the liver and serum, and target protein expression were performed at different time points. The results showed a higher percentage of hepatocytes in the G1/G0 and S phases at the end of 6 and 12 weeks of VC exposure, respectively. MiRNA-222 expression decreased initially and then increased, whereas miRNA-199a, miRNA-195, and miRNA-125b expression increased initially and then decreased, which corresponded with changes in cell cycle distribution and related target proteins expression (p27, cyclinA, cyclinD1, and CDK6). The corresponding expression levels of miRNAs in serum did not change. Dynamic changes in miR-222, miR-199a, miR-195, and miR-125b induced by VC can lead to cell cycle deregulation by affecting cell cycle-related proteins, and these miRNAs can serve as early biomarkers for malignant transformation caused by VC.

Zrsr2 Deficient Zebrafish Display Hematopoietic Defects and U12-Type Intron Retention in mRNA Processing Genes

ZRSR2 is a small nuclear riboprotein necessary for assembly of the minor spliceosome and subsequent splicing of U12-type introns. It is involved in the recognition of 3' splice sites during the assembly of the spliceosome. Somatic mutations in ZRSR2 gene are present in hematopoietic malignancies, most notably myelodysplastic syndrome (MDS). These mutations are spread throughout the gene, indicating that its function is critical for hematopoiesis. In spite of these clinical associations, the role of ZRSR2 in hematopoiesis has not been well studied. Zebrafish make an excellent animal model for investigating this gene as all of the functional domains in the human protein are conserved with a high level of protein similarity in zebrafish. Additionally, hematopoietic development is well understood in this species. We used CRISPR/Cas9 to generate a zebrafish zrsr2 knockout model with an 11 base pair deletion (zrsr2Δ11) that results in frameshift with premature truncation and loss of all functional domains (p.W167fs*175). The zrsr2Δ11/Δ11 mutants began to appear morphologically different from their siblings by 4 days post fertilization (dpf), showing aberrant development in the mandible and pharyngeal arches. The zrsr2Δ11/Δ11 mutants developed mild to severe edema by 6 dpf and died by 8 dpf. To understand its role in hematopoiesis, we performed o-dianisidine staining and whole mount in situ hybridization (WISH) with lineage-specific markers during embryonic development. Our data showed that zrsr2Δ11/Δ11 embryos exhibit mild anemia by 2 dpf, suggesting that primitive hematopoiesis is defective. WISH showed that zrsr2Δ11/Δ11 mutants have normal early hematopoietic stem cell (HSC) emergence (c-myb) at 36 hours post fertilization. However, these cells are absent through 3 and 5 dpf. At 5 dpf zrsr2Δ11/Δ11 mutants had severely reduced expression of hematopoietic markers for erythroid (hbae1), lymphoid (rag1), and myeloid (mpo) lineages. This verifies that knockout of zrsr2 in zebrafish disrupted normal hematopoiesis and leads to cytopenias - emulating the symptoms of MDS. We then investigated the transcriptional and splicing changes underlying these phenotypes using RNA sequencing on zrsr2Δ11/Δ11 mutant embryos at 3dpf. We found a total of 285 intron retention events in zrsr2Δ11/Δ11 mutants as compared to the control embryos, with about 76% of those events occurring within U12-type introns. Genes with retained introns were associated with mRNA decay and destabilization. We also found that genes related to cell cycle arrest were upregulated in zrsr2Δ11/Δ11 mutants . These results indicate that aberrant mRNA splicing and cell cycle deregulation contribute to arrested hematopoiesis when zrsr2 is knocked out in zebrafish. Disclosures No relevant conflicts of interest to declare.

PARP1 Deficiency Reduces Tumour Growth by Decreasing E2F1 Hyperactivation: A Novel Mechanism in the Treatment of Cancer

In recent years, poly (ADP-ribose) polymerase (PARP) inhibitors have been evaluated for treating homologous recombination-deficient tumours, taking advantage of synthetic lethality. However, increasing evidence indicates that PARP1 exert several cellular functions unrelated with their role on DNA repair, including function as a co-activator of transcription through protein-protein interaction with E2F1. Since the RB/E2F1 pathway is among the most frequently mutated in many tumour types, we investigated whether the absence of PARP activity could counteract the consequences of E2F1 hyperactivation. Our results demonstrate that genetic ablation of Parp1 extends the survival of Rb-null embryos, while genetic inactivation of Parp1 results in reduced development of pRb-dependent tumours. Our results demonstrate that PARP1 plays a key role as a transcriptional co-activator of the transcription factor E2F1, an important component of the cell cycle regulation. Considering that most oncogenic processes are associated with cell cycle deregulation, the disruption of this PARP1-E2F1 interaction could provide a new therapeutic target of great interest and a wide spectrum of indications.