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41 p.-7 fig., ICAP-1 regulates β1 integrin activation and cell adhesion. Here we used ICAP-1-null mice to study ICAP-1 potential involvement during immune cell development and function. Integrin α4β1-dependent adhesion was comparable between ICAP-1-null and control thymocytes, but lack of ICAP-1 caused a defective single positive (SP) CD8+ cell generation, thus unveiling an ICAP-1 involvement in SP thymocyte development. ICAP-1 bears a nuclear localization signal and we found it displayed a strong nuclear distribution in thymocytes. Interestingly, there was a direct correlation between the lack of ICAP-1 and reduced levels in SP CD8+ thymocytes of Runx3, a transcription factor required for CD8+ thymocyte generation. In the spleen, ICAP-1 was found evenly distributed between cytoplasm and nuclear fractions, and ICAP-1-/- spleen T and B cells displayed upregulation of α4β1-mediated adhesion, indicating that ICAP-1 negatively controls their attachment. Furthermore, CD3+ - and CD19+ -selected spleen cells from ICAP-1-null mice showed reduced proliferation in response to T and B cell stimuli, respectively. Finally, loss of ICAP-1 caused a remarkable decrease in marginal zone B cell frequencies and a moderate increase in follicular B cells. Together, these data unravel an ICAP-1 involvement in the generation of SP CD8+ thymocytes and in the control of marginal zone B cell numbers., This work was supported by grants SAF2017-85146-R and PID2020-116291RB-I00 from the Ministerio de Ciencia e Innovación (MICINN) to J.T, PID2019-105623RB-I00 from MICINN to M.L.T,BFU2013-48828-P from MICINN to Y.R.C., ERC Synergy Grant (2018) to R.F., RTI2018-095497-B-I00 from MICINN to A.H, and RTI2018-093938-B-I100 from MICINN and (RD16/0011/0002, TERCEL) from Instituto de Salud Carlos III to AGZ.

39 p.-7 fig., Combined treatment of metastatic melanoma with BRAF and MEK inhibitors has improved survival, but the emergence of resistance represents an important clinical challenge. Targeting ERK is a suitable strategy currently being investigated in melanoma and other cancers. To anticipate possible resistance to ERK inhibitors (ERKi), we used SCH772984 (SCH) as a model ERKi to characterize resistance mechanisms in two BRAF V600E melanoma cell lines. The ERKi-resistant cells were also resistant to vemurafenib (VMF), trametinib (TMT), and to combined treatment with either VMF and SCH or TMT and SCH. Resistance to SCH involved stimulation of the IGF-1R-MEK5-Erk5 signaling pathway, which counteracted inhibition of Erk1/2 activation and cell growth. Inhibition of IGF-1R with linsitinib blocked Erk5 activation in SCH-resistant cells and decreased their growth in 3D spheroid growth assays as well as in NOD scid gamma (NSG) mice. Cells doubly resistant to VMF and TMT or to VMF and SCH also exhibited downregulated Erk1/2 activation linked to stimulation of the IGF-1R-MEK5-Erk5 pathway, which accounted for resistance. In addition, we found that the decreased Erk1/2 activation in SCH-resistant cells involved reduced expression and function of TGF-alpha. These data reveal an escape signaling route that melanoma cells use to bypass Erk1/2 blockade during targeted melanoma treatment and offer several possible targets whose disruption may circumvent resistance., This work was supported by grants SAF2014-53059-R, SAF2017-85146-R and RD12/0036/0061 to J.T., and PI15/01180 to A.E-O.

36 p.-6 fig., The interaction of multiple myeloma (MM) cells with the bone marrow (BM) microenvironment promotes MM cell retention, survival and resistance to different anti‐MM agents, including proteasome inhibitors (PIs) such as bortezomib (BTZ). The α4β1 integrin is a main adhesion receptor mediating MM cell‐stroma interactions and MM cell survival, and its expression and function are downregulated by BTZ, leading to inhibition of cell adhesion‐mediated drug resistance (CAM‐DR) and MM cell apoptosis. Whether decreased α4β1 expression and activity is maintained or recovered upon development of resistance to BTZ represents an important question, as a potential rescue of α4β1 function could boost MM cell survival and disease progression. Using BTZ‐resistant MM cells, we found that they not only rescue their α4β1 expression, but its levels were higher than in parental cells. Increased α4β1 expression in resistant cells correlated with enhanced α4β1‐mediated cell lodging in the BM, and with disease progression. BTZ‐resistant MM cells displayed enhanced NF‐κB pathway activation relative to parental counterparts, which contributed to upregulated α4 expression and to α4β1‐dependent MM cell adhesion. These data emphasize the upregulation of α4β1 expression and function as a key event during resistance to BTZ in MM, which might indirectly contribute to stabilize this resistance, as stronger MM cell attachment to BM stroma will regain CAM‐DR and MM cell growth and survival. Finally, we found a strong correlation between high ITGB1 (integrin β1) expression in MM and poor progression‐free survival (PFS) and overall survival (OS) during treatment of MM patients with BTZ and IMIDs, and combination of high ITGB1 levels and presence of the high‐risk genetic factor amp1q causes low PFS and OS. These results unravel a novel prognostic value for ITGB1 in myeloma., This work was supported by grants SAF2014-53059-R and SAF2017-85146-R from the Ministerio de Ciencia, Innovación y Universidades (MCIU) to JT; SAF2017-89672-R from MCIU to FM; by the Research Institute Hospital 12 de Octubre (i+12) and grants from Instituto de Salud Carlos III (ISCIII) and CIBERONC to JML; by grants from ISCIII PI16/02024,PI17/00701 and PI19/01352, TRASCAN (EPICA and Immunocell), Fundació La Marató de TV3, the Accelerator award CRUK/AIRC/AECC joint funder-partnership, CIBERONC(CB16/12/00489) and co-financed with FEDER funds MINECO Explora (RTHALMY),Gobierno de Navarra, Departamento de Salud 40/2016 and Departamento de Industria (Proyecto Estrategico, Reto Genomica, DIANA) and Fundación Ramón Areces (PREMAMM) to FP and XA The study was also supported by the Multiple Myeloma Research Foundation Networks of excellence, the International Myeloma Foundation (Brian van Novis), and the Qatar National Research Fund award 7-916-3-237.