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Resumen del trabajo presentado al 63rd Annual Meeting Biophysical Society, celebrado en Baltimore, Maryland (USA) del 2 al 6 de marzo de 2019., Immune cells have an important role in the tumor-microenvironment. Macrophages may tune the immune response toward inflammatory or tolerance pathways. Tumor associated macrophages (TAM) have immunosuppressive functions and they are considered a therapeutic target in cancer. The aim of this study was to evaluate the effects of trabectedin, a new class of antitumor agent, on the tumor-microenvironment through the study of electrophysiological and molecular phenotype of macrophages. Experiments were performed using the whole-cell configuration of the patch-clamp technique in resident peritoneal mouse macrophages under different types of polarization. Trabectedin decreased macrophage viability and increased ROS production. Trabectedin does not directly interact with KV1.5 and KV1.3 channels, but treatment (16 h) of macrophages with sub-cytotoxic concentrations (0.1-5 nM) increased their KV current in a concentration-dependent manner due to an upregulation of KV1.3 channels. In vitro generated TAM (TAMiv), by a co-culture of ID8 cells and macrophages, exhibited a M2 phenotype. TAMiv generated a small KV current, similarly to M2 polarized macrophages, and expressed high levels of M2 markers. In this study, we demonstrated that TAMiv polarization could be re-educated by using sub-cytotoxic concentration of trabectedin. TAMiv treated with sub-cytotoxic concentration of trabectedin exhibited an upregulation of KV1.3 channels and their M2 phenotype changed towards M1 pro-inflammatory one., Funded by PharmaMar, CSIC 201820E104, CIBERCV and SAF2016-75021-R.

[Background]: Despite recent advances in the treatment of multiple myeloma (MM), the prognosis of most patients remains poor, and resistance to traditional and new drugs frequently occurs. EDO-S101 is a novel therapeutic agent conceived as the fusion of a histone deacetylase inhibitor radical to bendamustine, with the aim of potentiating its alkylating activity. [Methods]: The efficacy of EDO-S101 was evaluated in vitro, ex vivo and in vivo, alone, and in combination with standard anti-myeloma agents. The underlying mechanisms of action were also evaluated on MM cell lines, patient samples, and different murine models. [Results]: EDO-S101 displayed potent activity in vitro in MM cell lines (IC 1.6-4.8 μM) and ex vivo in cells isolated from MM patients, which was higher than that of bendamustine and independent of the p53 status and previous melphalan resistance. This activity was confirmed in vivo, in a CB17-SCID murine plasmacytoma model and in de novo Vk∗MYC mice, leading to a significant survival improvement in both models. In addition, EDO-S101 was the only drug with single-agent activity in the multidrug resistant Vk12653 murine model. Attending to its mechanism of action, the molecule showed both, a HDACi effect (demonstrated by α-tubulin and histone hyperacetylation) and a DNA-damaging effect (shown by an increase in γH2AX); the latter being again clearly more potent than that of bendamustine. Using a reporter plasmid integrated into the genome of some MM cell lines, we demonstrate that, apart from inducing a potent DNA damage, EDO-S101 specifically inhibited the double strand break repair by the homologous recombination pathway. Moreover, EDO-S101 treatment reduced the recruitment of repair proteins such as RAD51 to DNA-damage sites identified as γH2AX foci. Finally, EDO-S101 preclinically synergized with bortezomib, both in vitro and in vivo. [Conclusion]: These findings provide rationale for the clinical investigation of EDO-S101 in MM, either as a single agent or in combination with other anti-MM drugs, particularly proteasome inhibitors., This work was in part funded by Mundipharma-EDO GmbH, the Spanish Instituto de Salud Carlos III (ISCIII-FIS), (PI 15/0067 and PI 15/2156) and FEDER, the Spanish RTICC (RD12/0036/0058), Spanish Association Against Cancer (AECC, GCB120981SAN), and the Regional Council of Castilla y León (GRS 1175/A/15 and FIC335U14). All this funding was employed for the execution of experiments and the analysis of data. AALI was supported by a grant from the Spanish Society of Hematology and Hemotherapy. MMS is supported by the Network of Centers for Regenerative Medicine and Cellular Therapy from Castilla y León, Spain.

The benefit of intrathecal therapy and systemic rituximab on the outcome of diffuse large B-cell lymphoma at risk of central nervous system disease is controversial. Furthermore, the effect of intrathecal treatment and rituximab in diffuse large B-cell and Burkitt lymphoma with occult leptomeningeal disease detected by flow cytometry at diagnosis is unknown. Untreated diffuse large B-cell (n=246) and Burkitt (n=80) lymphoma at clinical risk of central nervous system disease and having had pre-treatment cerebrospinal fluid were analyzed by flow cytometry and cytology. Spinal fluid involvement was detected by flow cytometry alone (occult) in 33 (13%) diffuse large B-cell and 9 (11%) Burkitt lymphoma patients, and detected by cytology in 11 (4.5%) and 5 (6%) patients, respectively. Diffuse large B-cell lymphoma with occult spinal fluid involvement had poorer survival (P=0.0001) and freedom from central nervous system relapse (P, The work was partially supported by the following grants RD06⁄0020⁄0035 and RD12/0036/0048 from RETICS-FEDER (Instituto de Salud Carlos III, Ministerio de Economia y Competitividad, Madrid, Spain) and an unrestricted grant from Mundipharma Pharmaceuticals, SL (Madrid, Spain).