Perigestational dietary folic acid deficiency protects against medulloblastoma formation in a mouse model of nevoid basal cell carcinoma syndrome

Abstract

Hereditary nevoid basal cell carcinoma syndrome (NBCCS) is caused by PTCH1 gene mutations that result in diverse neoplasms including medulloblastoma (MB). Epidemiological studies report reduced pediatric brain tumor risks associated with maternal intake of prenatal vitamins containing folic acid (FA) and FA supplements specifically. We hypothesized that low maternal FA intake during the perigestational period would increase MB incidence in a transgenic NBCCS mouse model, which carries an autosomal dominant mutation in the Ptch1 gene. Female wild-type C57BL/6 mice (n = 126) were randomized to 1 of 3 diets with differing FA amounts: 0.3 mg/kg (low), 2.0 mg/kg (control), and 8.0 mg/kg (high) 1 mo prior to mating with Ptch1 (+/-) C57BL/6 males. Females were maintained on the diet until pup weaning; the pups were then aged for tumor development. Compared to the control group, offspring MB incidence was significantly lower in the low FA group (Hazard Ratio = 0.47; 95% confidence interval 0.27-0.80) at 1 yr. No significant difference in incidence was observed between the control and high FA groups. Low maternal perigestational FA levels may decrease MB incidence in mice genetically predisposed to tumor development. Our results could have implications for prenatal FA intake recommendations in the presence of cancer syndromes.

Figure 1. Experimental design

One hundred and twenty-six female mice were assigned in equal numbers to one of three maternal dietary FA groups (0.3 mg/kg (low), 2.0 mg/kg (control), or 8 mg/kg FA) one month prior to mating with Ptch1^+/− heterozygote male mice. Dams were kept on their respective diets until weaning of their litters at three weeks of age. Offspring were genotyped at 2 weeks of age and RBC folate measurements were performed on blood samples from dams taken at the time of weaning of their offspring. Upon weaning, offspring were switched to the standard mouse chow and followed for tumor development for 12 months.

Figure 2. Mean RBC folate measurements in dams at weaning

RBC folate was measured in dams at the weaning of their pups. RBC folate concentration increased significantly with increasing dietary FA dose (p<0.0001). The number of mice included in the data for each bar was 15 (0.3), 21 (2), and 17 (8).

Figure 3. Kaplan-Meier Survival curves comparing survival of Ptch1^+/− offspring of mothers assigned to three different doses of folic acid prior to conception through weaning

A. Overall survival. The difference in survival between offspring of different diet groups is not statistically significant (p=0.63). B. MB survival. The difference in MB survival between offspring of the low and control FA groups was statistically significant (p=0.02).

Figure 4. Pathologic analyses of tumor tissue

Photomicrographs of hematoxylin and eosin stained representative tissue showing Ptch1+/− brain sectioned in half (mid- or parasagittal sections). A) Representative section of brain with dilated lateral ventricle (highlighted by double-headed arrow). H – adjacent hippocampus. Image captured using 2X objective. B) Representative section of brain with medulloblastoma (denoted by asterisk) adjacent to uninvolved cerebellum (CB). Image captured using 2X objective. C) Higher magnification of medulloblastoma represented in B. The neoplasm is characterized by relatively monomorphous sheets of small undifferentiated cells. Image captured using 20X objective.