Os objetivos deste trabalho foram os de avaliar a eficácia da cimetidina como única forma de tratamento para reduzir o tamanho de melanomas localizados em região ventral de cauda e períneo em equinos tordilhos, determinar a eficácia da droga no controle do aparecimento de novas formações nas mesmas regiões e avaliar a duração do tratamento com cimetidina após a sua suspensão com relação à manutenção da redução de tamanho e do aparecimento de novas formações. Para o desenvolvimento do trabalho foram utilizados 32 equinos tordilhos, sem restrições de raça ou sexo, com idade entre sete e 30 anos, com melanomas em região de cauda e períneo de até 5,0 cm de diâmetro e sem histórico anterior de tratamento com cimetidina ou procedimento cirúrgico para esta finalidade. Os animais foram divididos em dois grupos: 21 animais no grupo tratamento que receberam 18 mg/kg de cimetidina por via oral, BID durante noventa dias e 11 animais no grupo controle, que não receberam nenhuma medicação ou placebo. Os equinos foram avaliados quinzenalmente durante o período de tratamento e monitorados mensalmente durante 150 dias após o término da administração da droga. A cimetidina, na dose de 18 mg/kg VO BID, apresenta ação sobre os melanomas em equinos tordilhos, mas sua ação mostrou-se bastante variável e individual, além de não apresentar efetividade na manutenção da redução do volume após o término do tratamento. / The purpose of this study was to evaluate the efficacy of cimetidine as treatment for melanoma present in the perineal area and ventral tail of Grey horses. Reduction in tumor volume and the capability of the drug to control the appearance of new nodules were evaluated. Thirty two grey horses with ages ranging between sete and 30 years and no predilection of breed or gender were used in this study. All the animals selected had no attempted treatment prior to this study. The animals were divided in two groups: the treatment group, consisting of 21 horses and the control group consisting of 11 animals. Cimetidine was administered at a dosage of 18 mg/kg orally twice a day for a total of ninety days. During the lenght of treatment, the group was evaluated every two weeks, and all the animals were monitored once a month for another 150 days after the end of therapy. The efficacy of the treatment with cimetidine was observed in 11 animals of the treatment group which showed size reduction of the masses during the drug administration period followed by a slight increase and stabilization at lower volumes than the initial measures. In addition, the animals that responded to the treatment presented a very variable and individual response.

Os objetivos deste trabalho foram os de avaliar a eficácia da cimetidina como única forma de tratamento para reduzir o tamanho de melanomas localizados em região ventral de cauda e períneo em equinos tordilhos, determinar a eficácia da droga no controle do aparecimento de novas formações nas mesmas regiões e avaliar a duração do tratamento com cimetidina após a sua suspensão com relação à manutenção da redução de tamanho e do aparecimento de novas formações. Para o desenvolvimento do trabalho foram utilizados 32 equinos tordilhos, sem restrições de raça ou sexo, com idade entre sete e 30 anos, com melanomas em região de cauda e períneo de até 5,0 cm de diâmetro e sem histórico anterior de tratamento com cimetidina ou procedimento cirúrgico para esta finalidade. Os animais foram divididos em dois grupos: 21 animais no grupo tratamento que receberam 18 mg/kg de cimetidina por via oral, BID durante noventa dias e 11 animais no grupo controle, que não receberam nenhuma medicação ou placebo. Os equinos foram avaliados quinzenalmente durante o período de tratamento e monitorados mensalmente durante 150 dias após o término da administração da droga. A cimetidina, na dose de 18 mg/kg VO BID, apresenta ação sobre os melanomas em equinos tordilhos, mas sua ação mostrou-se bastante variável e individual, além de não apresentar efetividade na manutenção da redução do volume após o término do tratamento. / The purpose of this study was to evaluate the efficacy of cimetidine as treatment for melanoma present in the perineal area and ventral tail of Grey horses. Reduction in tumor volume and the capability of the drug to control the appearance of new nodules were evaluated. Thirty two grey horses with ages ranging between sete and 30 years and no predilection of breed or gender were used in this study. All the animals selected had no attempted treatment prior to this study. The animals were divided in two groups: the treatment group, consisting of 21 horses and the control group consisting of 11 animals. Cimetidine was administered at a dosage of 18 mg/kg orally twice a day for a total of ninety days. During the lenght of treatment, the group was evaluated every two weeks, and all the animals were monitored once a month for another 150 days after the end of therapy. The efficacy of the treatment with cimetidine was observed in 11 animals of the treatment group which showed size reduction of the masses during the drug administration period followed by a slight increase and stabilization at lower volumes than the initial measures. In addition, the animals that responded to the treatment presented a very variable and individual response.

Domestic animals have become very different from their wild ancestors during domestication and animal breeding. This provides a good model to unravel the molecular mechanisms underlying phenotypic variation. In my thesis I have studied genes affecting two important traits, leanness in pigs and hair greying-associated melanoma in horses. In the first part of the thesis, I focused on an intronic mutation leading to more muscle growth and less fat deposition in domestic pigs to identify a transcription factor (TF) that binds to the regulatory element overlapping with the mutation. The aim has been to further study the function of the previously unknown TF in mouse myoblast cells and in insulin-producing cells (Paper I-III). We discovered a new TF ZBED6 binding to intron 3 of the IGF2 gene, in which a single nucleotide substitution in pigs abrogates the binding and causes increased leanness in domestic pigs. Silencing of ZBED6 expression in mouse myoblasts increased Igf2 expression, cell proliferation and migration, and myotube formation. This result is in line with the increased leanness phenotype in mutant pigs. Chromatin Immunoprecipitation-sequencing (ChIP-seq) using an anti-ZBED6 antibody identified 1200 ZBED6 target genes besides IGF2 and many are TFs controlling fundamental biological processes. In the first follow-up study we found ZBED6 mainly affected the expression of muscle protein genes by directly regulating Igf2 and Twist2 expression, in agreement with our previous observation of faster myotube formation in ZBED6-silenced cells. ChIP-seq with antibodies against six different histone modifications revealed that ZBED6 preferentially binds to active promoters and modulates transcriptional activity by a novel mechanism rather than by recruiting repressive histone modifications. The second follow-up study revealed that ZBED6 affects the morphology and insulin content and release in pancreatic ß cells. In the second part (Paper IV), we investigate the functional significance of an intronic duplication in the Syntaxin 17 (STX17) gene causing hair greying and melanoma in horses. We found two Microphtalmia-associated transcription factor (MITF) binding sites within the duplication and showed that the duplicated sequence up-regulates reporter gene expression in a melanocyte-specific manner both by reporter assays in mouse melanocytes and in transgenic zebrafish. These results established that the intronic duplication acts as a melanocyte-specific enhancer that becomes much stronger when it is duplicated.