GENETIC AND MOLECULAR BASIS OF LEUKEMIA AND MAMMARY CANCER METASTASIS

KENT W. HUNTER, Ph.D., Associate Member

We are interested in identifying genes that are involved in the induction and progression of breast cancer. Identification and analysis of novel genes associated with cancer will provide valuable clues for the design of new anticancer therapeutic strategies and early intervention. Our laboratory uses a strain of mice that has a heritable form of breast cancer with a very high incidence of metastasis to the lung to identify and characterize these potential modifier/suppressor genes. The presence of metastatic modifier or suppressor genes has been demonstrated by a combination of somatic cell hybrid or transfection analyses. To date, only four potential metastatic modifier/suppressor genes have been identified, KAI1, KiSS-1, E-cadherin and NM23.

This laboratory strives to increase the understanding of the process of breast cancer metastasis by localizing, identifying, and characterizing genes that influence the risk for metastatic involvement in breast cancer, utilizing the mouse as a model genetic system. Identification of metastasis modifier or suppressor genes will have important prognostic value, permitting the identification of those patients that should be closely monitored for metastatic involvement, or those that should undergo early anti-metastatic therapy to eliminate metastases that are not clinically apparent at the time of primary tumor diagnosis/treatment. In addition, identification and characterization of modifier/suppressor genes may reveal novel approaches for the treatment of disseminated tumors and early stage tumors at risk for metastasis that are more effective than current therapeutic strategies. Mice with breast cancer have been bred to a variety of laboratory mouse strains and a number of inbred strains have been identified that harbor genes that alter various aspects of the oncogenic process. Using high-resolution mouse mapping techniques, we are attempting to identify these genes and subsequently analyze them for their potential role in human breast cancer.

Metastasis is one of the most important aspects of neoplastic disease and one of the most poorly understood. A significant fraction of patients diagnosed with solid tumors already have metastases at the time of primary tumor diagnosis. The dispersal of metastatic tumors throughout the body often precludes surgical removal, and the tumors often prove refractory to anticancer therapies. As a result, many cancer patients succumb either to metastatic burden, rather than the primary tumor, or to complications associated with antimetastatic therapy. Analysis of human tumors has demonstrated specific alterations, including gene amplification and loss of heterozygosity (LOH), at various stages of tumorigenesis and metastatic progression.

FVB/N-TgN(MMTVPyMT) is a transgenic mouse that carries the polyoma middle T antigen driven by the mouse mammary tumor virus (MMTV) enhancer/promoter, and develops synchronously appearing multifocal tumors involving all of the mammary glands with extensive pulmonary metastases. Females develop palpable tumors within 8 weeks of birth, independent of pregnancy. We are utilizing this animal model to search for dominant alleles that affect the initiation and/or progression of mammary carcinogenesis. The transgenic animal has been bred to 25 different inbred mice, the progeny aged to permit tumor induction and potential metastasis, and subsequently analyzed for metastatic progression. Significant reduction in the number of pulmonary metastases was observed for several strains, including the inbred strain DBA/2J. Using a mouse genetic mapping resource known as the AKXD recombinant inbred panel, we have preliminary mapping data for two loci that control the susceptibility of DBA/2J mice to pulmonary metastatic involvement. We are currently confirming and refining the mapping data by the generation and analysis of an FVB/N-TgN(MMTVPyMT) versus DBA/2J backcross. In addition, we are analyzing a number of potential genes as candidate metastasis susceptibility genes.

Over 180,000 new cases of breast cancer are diagnosed every year, but the high-risk genetic determinants BRCA1 and BRCA2 account for only approximately 10% of all cases. There is a growing body of literature to suggest the presence of a number of other breast cancer susceptibility genes in the general population. However, since these susceptibility genes are not as penetrant, the tumors appear sporadic rather than hereditary, as was observed for BRCA1 and BRCA2. Identification of these less penetrant genes will likely reveal important insights into the etiology of breast cancer. However, due to the large population sizes required, it is difficult to perform this research in humans.

To identify novel breast cancer sus-ceptibility genes, our laboratory is studying two different inbred strains of mice. The FVB/N-TgN(MMTVPyMT) mouse develops breast cancer spontaneously, approximately 60 days after birth. We observed an acceleration in the appearance of the disease when the FVB/N-TgN(MMTVPyMT) mice were mated to the inbred strain I/LnJ. In addition, animals derived from the mating showed a significant reduction in the growth rate of the tumors. These results strongly suggest the presence of susceptibility or modifier genes in the I/LnJ mice. Identification of these genetic determinants might enable identification of more aggressive tumors in human breast cancer, enabling physicians to tailor treatment to individual tumor types.

We have, therefore, generated a [FVB/N-TgN(MMTVPyMT) x I/LnJ] backcross panel to map the genetic location of the modifier genes. A total of 127 animals were generated and a genome screen performed with more than 60 genetic markers. We have identified three genes that control the acceleration of disease appearance. The acceleration appears not to be due to a simple dominance affect of the I/LnJ genes. Rather a complex interaction with I/LnJ and FVB/NJ genes appears to be responsible. In addition, at least two additional genes have been identified that control the tumor growth rate. Like the genes that accelerate the appearance of the tumor, the genes that control the tumor growth rate also appear to function by synergistic interaction. We are currently developing high resolution mapping reagents to further localize the genetic position of these modifier genes prior to initiating positional cloning projects. We are also analyzing interesting genes located in the modifier gene intervals to determine whether they might be good candidates for the tumor latency and progression modifier genes.

LE VOYER, T., LU, Z., HUNTER, K. Microsatellite DNA variants between the FVB/NJ, C58/J and I/LnJ mouse strains. Mammalian Genome (in press).

WILLIAMS, R.W., HOLDENER-KENNY, B.C., ANGEL, J.M., OAKEY, R., HUNTER, K.W. Chromosome 7. Mamm. Genome 8:S136-159, 1998 and online: http://www.informatics.jax.org/bin/ccr/contents

WILLIAMS, M., LYU, M.S., YANG, Y.L., LIN, E.P., DUNBRACK, R., BIRREN, B., CUNNINGHAM J., HUNTER, K.W. Ier5, a novel member of the slow-kinetics immediate-early genes. Genomics 55:327-334, 1999.

YANG, Y-L., LEI, G., HOLLAND, C., KITAMURA, T., HUNTER, K., CUNNINGHAM, J.M. Identification of the receptors for the polytropic and xenotropic murine leukemia viruses. Nature Genet. 21:216-219, 1999.

LIFSTED, T., LE VOYER, T., WILLIAMS, M., MULLER, W., KLEIN-SZANTO, A., BUETOW, K.H., HUNTER, K.W. Identification of inbred mouse strains harboring genetic modifiers of mammary tumor age of onset and metastatic progression. Intl. J. Cancer 77:640-644, 1998.

^§   Fox Chase researcher

^a   M.-S. Lyu: Present address-National Cancer Institute, Bethesda, MD 20892

^b   K.H. Buetow: NIH/DCEG/LPG, Bethesda, MD 20892

Illustrations or unpublished data in these reports should not be used without permission of the author.

Fox Chase Cancer Center

Scientific Report 1998