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The II melanoma translational meeting of the Spanish Melanoma Group (GEM) was held in Barcelona, Spain, at the Hospital Universitari Quiron-Dexeus on November 3 rd , 2016. The conference featured leaders in the fields of oncology, immunology and dermatology, all working at both national and international levels on basic research with direct applications to the clinical setting. The objective was to present cutting edge research on melanoma, mainly from Spanish research groups, but also from some international groups, with the aim of generating a network for future national and international collaborations. During a single day, fifteen speakers, seven biologists and eight clinicians, with a special focus on translational research, outlined the main findings of their ongoing studies. The moderators in every session were recognized GEM oncologists who guided discussion from the clinical point of view regarding the preclinical data presented. Herein, we summarize the main topics discussed during the meeting.

LVs have proven to be efficient vehicles to deliver one or more tgs to any cell type of choice, which has led to a promising list of therapeutic applications. As the demand for experimentation in gene delivery to specific cell types increases, technologies that precisely target LVbased gene expression will become more important for research and clinical applications. Four main groups of strategies with their own possibilities as well as difficulties have been developed so far. Self-evidently, further optimization and fine-tuning of these strategies is a necessity to fulfill the expectations for targeted LV delivery in vivo. In addition to extra optimization steps, combinations of two or more of these strategies can also lead to an overall more selective, efficient and most importantly, safe LV system. Several attempts to combine the different strategies have been reported (Brown, Cantore et al. 2007; Pariente, Morizono et al. 2007; Escors and Breckpot 2011). Pariente et al. for example, reported on a LV that was transductionally targeted to prostate cancer bone metastases by a modified sindbis virus envelope that interacts with PSCA and transcriptionally targeted with a prostate cell specific promoter. This dual-targeted LV enhanced specificity to prostate cancer bone metastases after systemic delivery with respect to individual transcriptional or transduction targeting. As the developed targeting strategies already resulted in a major step forward for LV-based gene therapy, their potential will most likely be more exploited in the future, paving the way towards an all-embracing LV-based tg vehicle for the gene therapeutic field.

Historically, protection against virus infections has relied on the use of vaccines, but the induction of an immune response requires several days and in certain situations, like in newborn animals that may be infected at birth and die in a few days, there is not sufficient time to elicit a protective immune response. Immediate protection in new born could be provided either by vectors that express virus-interfering molecules in a tissue specific form, or by the production of animals expressing resistance to virus replication. The mucosal surface is the largest body surface susceptible to virus infection that can serve for virus entry. Then, it is of high interest to develop strategies to prevent infections of these areas. Virus growth can be interfered intracellularly, extracellularly or both. The antibodies neutralize virus intra- and extracellularly and their molecular biology is well known. In addition, antibodies efficiently neutralize viruses in the mucosal areas. The autonomy of antibody molecules in virus neutralization makes them functional in cells different from those that produce the antibodies and in the extracellular medium. These properties have identified antibodies as very useful molecules to be expressed by vectors or in transgenic animals to provide resistance to virus infection. A similar role could be played by antimicrobial peptides in the case of bacteria. Intracellular interference with virus growth (intracellular immunity) can be mediated by molecules of very different nature: (i) full length or single chain antibodies; (ii) mutant viral proteins that strongly interfere with the replication of the wild type virus (dominant-negative mutants); (iii) antisense RNA and ribozyme sequences; and (iv) the product of antiviral genes such as the Mx proteins. All these molecules inhibiting virus replication may be used to obtain transgenic animals with resistance to viral infection built in their genomes. We have developed two strategies to target into mucosal areas either antibodies to provide immediate protection, or antigens to elicit immune responses in the enteric or respiratory surfaces in order to prevent virus infection. One strategy is based on the development of expression vectors using coronavirus derived defective RNA minigenomes, and the other relies on the development of transgenic animals providing virus neutralizing antibodies in the milk during lactation. Two types of expression vectors are being engineered based on transmissible gastroenteritis coronavirus (TGEV) defective minigenomes. The first one is a helper virus dependent expression system and the second is based on self-replicating RNAs including the information required to encode the TGEV replicase. The minigenomes expressing the heterologous gene have been improved by using a two-step amplification system based on cytomegalovirus (CMV) and viral promoters. Expression levels around 5 micrograms per 10(6) cells were obtained. The engineered minigenomes will be useful to understand the mechanism of coronavirus replication and for the tissue specific expression of antigen, antibody or virus interfering molecules. To protect from viral infections of the enteric tract, transgenic animals secreting virus neutralizing recombinant antibodies in the milk during lactation have been developed. Neutralizing antibodies with isotypes IgG1 or IgA were produced in the milk with titers of 10(6) in RIA that reduced virus infectivity by one million-fold. The recombinant antibodies recognized a conserved epitope apparently essential for virus replication. Antibody expression levels were transgene transgene copy number independent and were related to the transgene integration site. This strategy may be of general use since it could be applied to protect newborn animals against infections of the enteric tract by viruses or bacteria for which a protective MAb has been identified. Alternatively, the same strategy could be used to target the expression of antibio