lds103

Linda D. Siracusa, PhD

Contact Dr. Siracusa

233 South 10th Street
BLSB 719
Philadelphia, PA 19107-5541

(215) 503-4536
(215) 923-4153 fax

Expertise and Research Interests

The study of genes that influence complex traits is a rapidly evolving field. The power of mammalian genetics coupled with the ability to scan entire genomes of individual animals has led to the discovery of chromosomal regions that harbor genes conferring susceptibility or resistance to different cancers. My laboratory focuses on identifying and characterizing genes that influence the development of cancer along the gastrointestinal tract. Our research combines molecular biology and classical genetics to investigate cancer susceptibility genes. Specifically, we focus on selected crosses between mouse mutations and inbred strains to determine the chromosomal location of modifier loci. We then screen candidate genes in each region and use transgenesis to prove the identity of novel modifier genes. Our goal is to understand the mechanisms of tumor suppression.

The system we chose involves the tumor suppressor gene Adenomatous Polyposis Coli (APC). Mutations in the human APC gene are responsible for the majority of inherited and sporadic colorectal cancers. Min mice have a mutation in the murine homologue of the human APC gene and develop multiple adenomas along their intestinal tract. Quantitative trait locus (QTL) studies identified a locus, Modifier of Min 1 (Mom1), which maps to the distal region of mouse chromosome 4. The Mom1 locus dramatically affects ApcMin-induced tumor number. We were the first to report that the secretory type II Phospholipase A2 (Pla2g2a) gene is a strong candidate for the Mom1 locus. Inbred mouse strains display 100% concordance between Pla2g2a allele type and tumor susceptibility. Expression and sequence analysis showed that Mom1 susceptible strains carry null alleles of the Pla2g2a gene. We developed a fluorescent enzyme activity assay for Pla2g2a which confirms and extends our molecular and immunohistochemical findings. Studies are in progress to establish the mechanism by which Pla2g2a prevents intestinal polyp formation as well as to investigate the role of PLA2G2A in human neoplasia and intestinal homeostasis.

We recently discovered a spontaneous dominant mutation, Modifier of Min 2 (Mom2), which is a potent suppressor of intestinal and colorectal tumorigenesis in ApcMin mice. One copy of the resistant Mom2R allele reduces small intestine adenoma numbers and colon adenoma incidence by ~90% in ApcMin mice. We localized Mom2 to the distal end of mouse chromosome 18 using a unique exclusion mapping strategy. Recent work combining quantitative RT-PCR and sequencing revealed a 4 bp duplication within exon 3 of the ATP synthase (Atp5a1) gene that is responsible for the Mom2 phenotype. Studies of tumor suppression are suggesting a novel mechanism of cellular lethality that is specific to the tumor lineage, which results in the suppression of polyp development. Further work is ongoing to understand the full effects of Mom2 on adenoma initiation, growth, and progression. Our work on this system is performed in collaboration with the laboratory of Dr. Arthur Buchberg at the Kimmel Cancer Center.

We have also established several reciprocal congenic lines of mice between the C3H/HeJ and C57BL/6J inbred strains that have a resistant Mom1 locus on an otherwise susceptible background and a susceptible Mom1 locus on an otherwise resistant background. Testing of these congenic lines has demonstrated significant differences in polyp formation in both the small and large intestines. Using these new congenic strains in designer crosses, offspring are aged and analyzed for tumor number, size and position and QTL analyses is performed to identify chromosomal regions that contain genes which act to significantly decrease polyp formation. We completed the first large-scale backcross to identify new modifier loci. Microarray analyses will determine gene expression profiles of existing and future lines to determine pathways that are prime candidates for conferring resistance phenotypes. We are using a similar strategy to assess the impact of the evolutionarily divergent Mus castaneus (CAST) genetic background on intestinal and colorectal tumorigenesis.

The comprehensive examination of novel modifier loci in humans will facilitate an understanding of the relationship between the effects of modifier genes on tumor initiation, growth, and progression. Further investigations will lead to insights about the role of modifier genes in human cancer risk assessment, tumor prevention, diagnosis, and their predictive value for response to treatment.

We recently predicted that microRNAs are candidates for tumor susceptibility genes. To this end, we established a database comparing the positions of mouse microRNAs across the genome with the positions of loci conferring susceptibility to eight different solid tumors. We established the MUSMIRSUS database (http://www.kimmelcancercenter.org/siracusa/musmirsus.htm) which compares the positions of tumor susceptibility loci with the positions of microRNAs in the mouse genome. Statistical analyses revealed a highly significant (p<0.001) association between the presence of microRNA genes and the locations of tumor susceptibility loci. Comparisons of the sequence of microRNAs between inbred strains that are known to differ in their resistance or susceptibility to specific types of tumors revealed polymorphisms, primarily in the regions upstream of the microRNA stem-loop structure. Studies are continuing to assess the level of expression of microRNAs in tissues of the gastrointestinal tract, in both normal and disease states. This research is performed in collaboration with Dr. Carlo Croce and Dr. George Calin at Ohio State University Medical Center.

Keywords

Cancer Risk; Cancer Genetics; Colorectal Cancer; Digestive Diseases and Disorders; Gastrointestinal System; Gene Expression; Gene Mutation; Genetic Library; Genetic Model; Immunohistochemistry; Laboratory Mouse; Mammalian Genetics; Model Design Development; Molecular Cloning; Molecular Oncology; Molecular Pathology; Nucleic Acid Sequence; Phenotype; Pleiotropism; Polymerase Chain Reaction; Protein Structure Function; Pulsed Field Gel Electrophoresis; Small Intestine; Southern and Northern Blotting; Tissue Cell Culture; Transcription; Transgenic Animals; Tumor Progression; Western Blotting