Thyroid diseases have a complex and multifactorial aetiology. Despite the numerous studies on the signals referable to the malignant transition, the molecular mechanisms concerning the role of oxidative stress remain elusive. Based on its strong oxidative power, H2O2 could be responsible for the high level of oxidative DNA damage observed in cancerous thyroid tissue and hyperactivation of mitogen-activated protein kinase (MAPK) and PI3K/Akt, which mediate ErbB signaling. Increased levels of 8-oxoG DNA adducts have been detected in the early stages of thyroid cancer. These DNA lesions are efficiently recognized and removed by the base excision repair (BER) pathway initiated by 8-oxoG glycosylase1 (OGG1). This study investigated the relationships between the EGFR and OGG1-BER pathways and their mutual regulation following oxidative stress stimulus by H2O2 in human thyrocytes. We clarified the modulation of ErbB receptors and their downstream pathways (PI3K/Akt and MAPK/ERK) under oxidative stress (from H2O2) at the level of gene and protein expression, according to the mechanism defined in a human non-pathological cell system, Nthy-ori 3-1. Later, on the basis of the results obtained by gene expression cluster analysis in normal cells, we assessed the dysregulation of the relationships in a model of papillary thyroid cancer with RET/PTC rearrangement (TPC-1). Our observations demonstrated that a H2O2 stress may induce a physiological cross-regulation between ErbB and OGG1-BER pathways in normal thyroid cells (while this is dysregulated in the TPC-1 cells). Gene expression data also delineated that MUTYH gene could play a physiological role in crosstalk between ErbB and BER pathways and this function is instead lost in cancer cells. Overall, our data on OGG1 protein expression suggest that it was physiologically regulated in response to oxidative modulation of ErbB, and that these might be dysregulated in the signaling pathway involving AKT in the progression of thyroid malignancies with RET/PTC rearrangements.

Colorectal cancer (CRC) is a multistep process that arises in the colic tissue microenvironment. Oxidative stress plays a role in mediating CRC cell survival and progression, as well as promoting resistance to therapies. CRC progression is associated with Wnt/β-Catenin signaling dysregulation and loss of proper APC functions. Cancer recurrence/relapse has been attributed to altered ROS levels, produced in a cancerous microenvironment. The effect of oxidative distress on Wnt/β-Catenin signaling in the light of APC functions is unclear. This study evaluated the effect of H2O2-induced short-term oxidative stress in HCT116, SW480 and SW620 cells with different phenotypes of APC and β-Catenin. The modulation and relationship of APC with characteristic molecules of Wnt/β-Catenin were assessed in gene and protein expression. Results indicated that CRC cells, even when deprived of growth factors, under acute oxidative distress conditions by H2O2 promote β-Catenin expression and modulate cytoplasmic APC protein. Furthermore, H2O2 induces differential gene expression depending on the cellular phenotype and leading to favor both Wnt/Catenin-dependent and -independent signaling. The exact mechanism by which oxidative distress can affect Wnt signaling functions will require further investigation to reveal new scenarios for the development of therapeutic approaches for CRC, in the light of the conserved functions of APC.

Background: Bromodomain and Extra-Terminal domain (BET) proteins are epigenetic readers that play a fundamental role in transcription regulation. Preclinical and early clinical evidences sustain BET targeting as a promising novel approach for cancer treatment. BET degraders are chimeric compounds merging a BET inhibitor, which allows the binding to BET bromodomains, linked to an additional small molecule, which mediates the binding to an E3 ubiquitin ligase complex, triggering BET proteins degradation via the proteasome. These compounds are based on the concept of the PROTAC (Proteolysis targeting Chimera) (Sakamoto et al, PNAS 2001). MZ1 is a potent BRD4 degrader that exploits JQ1 for BET bromodomain binding and the von Hippel Lindau-containing complex for protein degradation (Zengerle et al, ACS Chem Biol 2015; Gadd et al., Nat Chem Biol 2017). MZ1 showed potent antiproliferative activity in AML cell lines (Chan et al. J Med Chem 2017). Here, we present data on the antitumor activity of MZ1 in diffuse large B cell lymphoma of the activated B cell like (ABC DLBCL). Materials and Methods: Established lymphoma cell lines were exposed for 72 hours to increasing doses of the compounds. Cell proliferation was evaluated by using MTT assay. FACS analysis was performed to measure apoptotic activation (Annexin V, threshold of 1.5 fold) after 72 hours of treatment. Results: The antitumor activity of the BET degrader MZ1, of its negative control epimer cisMZ1 and, as comparison, of the BET inhibitor OTX015/MK-8628 was assessed in seven cell lines derived from diffuse large B cell lymphoma of the activated B cell like (ABC DLBCL). MZ1 was very active with a median IC50 of 50nM (range, 5-150 nM). No activity was seen with cisMZ1. BET degrader was more active in vitro than the BET inhibitor, which presented a median IC50 of 125 nM (range, 80-400 nM) (P=0.024). Importantly, MZ1 (500nM, 72 h) induced apoptosis in all the seven ABC DLBCL cell lines, while, in accordance with previous data (Boi et al, Clin Cancer Res 2015), OTX015 (500nM, 72 h) was cytotoxic only in 2/7 cell lines (P=0.021). No apoptosis was observed with cisMZ1 (500nM, 72 h). Conclusion: The BRD4 degrader MZ1 had a strong cytotoxic activity in all the ABC-DLBCL cell lines we tested, and, at least in vitro, it elicited more profound effects than BET inhibitors, and encourages further investigations. Citation Format: Chiara Tarantelli, Hillarie Ekeh, Carmelo Moscatello, Eugenio Gaudio, Andrea Testa, Emanuele Zucca, Anastasios Stathis, Alessio Ciulli, Francesco Bertoni. The BRD4 degrader MZ1 exhibits potent antitumoral activity in diffuse large B cell lymphoma of the activated B cell-like type [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A179.

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