Abstract 5092: Establishment of an induction protocol for medulloblastoma during prenatal stages of a floxed-cre inducible model
Authors: Gallo-Oller Gabriel , Azatyan Ani , Baryawno Ninib , Kogner Per , Johnsen John Inge 
Published: August 2018

Abstract
Advances in mouse genetics in the last decades have improved the site-specific and conditional control of specific genes in vivo. Despite the availability of different recombinases, the most popular and best-characterized model is the Cre-loxP system. To achieve the recombination in specific tissues or cell types, Cre expression is regulated by a cell-specific promoter and it is also possible to use a tamoxifen-inducible form of Cre for temporal control of the recombination process. Different models have been described using this technology. However, the recombination in the central nervous system tissues is still problematic. Here, we describe a pilot study to induce the recombination process in the brain at prenatal stages in a Cre-loxP system.
To test the temporal control of the recombination in the brain, we used a Ptch1fl/fl mouse line in combination with the previously described Math1-creERT2 mouse line that can be induced by tamoxifen administration. All experimental procedures on animals, together with mouse breeding, were performed in accordance with the guidelines of the animal ethics committee of the Karolinska Institutet. The detection of the floxed allele and the cre gene was carried out by PCR at each breeding step.
In order to control the specific time point of the induction, the presence of the males for mating was controlled for two days. After this, females were monitored and the induction of the model was carried out at E14.5 (embryonic days) for two days by oral administration of tamoxifen. Issues due to tamoxifen administrations as problems in delivery and mortality/toxicity in pups were not detected. The litters obtained were genotyped to detect the correct combination, Ptch1fl/fl; Math1creERT2 +/-. Mice with the correct genotype developed neurological symptoms between 7 and 9 weeks old, and once the brain was dissected, an abnormal size of the cerebellum was detected. Cerebellum was dissected and the cells obtained were grown and maintained in different specific culture media. The deleted allele of Ptch1 was detected by PCR in the tumoral tissue and in the cell cultures.
Here, we described a protocol to induce a Cre-loxP system during embryonic stages. The present protocol avoids the problems described for extensive tamoxifen administration. We also described new culture media to grow and expand the tumor cells obtained. Our findings could help to improve the induction of this kind of systems and study mutations during embryonic stages related with the development and progression of medulloblastoma. The culture of cells derived from these systems, can be used to establish therapeutic mouse models and to evaluate novel treatment strategies for this tumor.
Citation Format: Gabriel Gallo-Oller, Ani Azatyan, Ninib Baryawno, Per Kogner, John Inge Johnsen. Establishment of an induction protocol for medulloblastoma during prenatal stages of a floxed-cre inducible model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5092.