*Funded by DIMACS and the UMDNJ Academic Partnership for Environmental Public Health Tracking (1 U19 EH000 CDC grant).
Title: Cumulative Load of Chromosome 3 Allelic Imbalance (LOH) Mutations Differentiated Sporadic Versus Chemicall Induces Cancer
Introduction/Aim: It has been challenging to differentiate human cancer using archival, fixative-treated, paraffin-embedded specimens with respect to sporadic (idiopathic) versus environmental/occupation chemically induced causation. Microscopic examination is rarely capable of detecting discriminating differences at the cellular level as the histologic appearance of cancer looks equivalent regardless of specific causation. The challenge is made more intense by the availability in many cases of only biopsy or fine needle aspiration cytology specimens. We sought to develop a molecular pathology platform that would permit this distinction to be made in an objective and highly reliable manner using small sized and cytology type specimens. The hypothesis was tested that chemical exposure causation of cancer is associated with a higher density of mutation acquisition for a given segment of genome (chromosome 3). Methods: 43 human cancers of diverse type from subjects potentially exposed to workplace aerosol and/or direct dermal contact with chlorinated solvents was studies using archival paraffin specimens and/or stained cytology slides. Each of the 43 cancer cases developed in railroaders assigned to duty in Consolidated Rail Corporation's Altoona Works. The railroaders were exposed to chlorinated solvents secondary to Consolidated Rail Corporation's illegal Hazardous Waste disposal within the Altoona Works. Refer to Commonwealth of Pennsylvania v. Consolidated Rail Corporation, 2000 CR 996, [Bair County, 2000.] Each case was carefully microdissected under stereoscopic observation to ensure that the most representative cellular areas were used for mutational analysis. Non-neoplastic cells were used as an internal control. Each microdissection target was evaluated using a broad panel of 17 chromosome 3 polymorphic microsatellite markers across both arms to detect the cumulative amount of acquired LOH mutations and to quantitative the clonal expansion of each mutation. Mutations having been acquired early in tumorigenesis and conferring greater proliferative advantage could be separated from mutations without these characteristics. Historical controls from the published literature were used as controls for sporadic cancer formation subject to chromosome 3 panel genotyping. Results: 37 of 43 (86%) cancers manifests showed four or more chromosome 3 LOH mutations. 23 of 43 (53%) cancers manifested six of more mutations. In all cases (n=6) of direct dermal contact with chlorinated solvents, six or greater mutations were detected. In 36 of 43 (84%) cancers, at least one high clonality mutation on chromosome 3 was found in the cancer supporting a causal role for such DNA damage in that cancer's formation. For many types of human cancer that are not associated with chromosome 3 deletional mutation in development and progression based on historical control data (brain, liver, prostate, skin, colon, pancreas) chromosome 3 mutations were present either in high number (four or more LOH mutations) or in high clonality. Conclusions: A novel approach based on quantifying the cumulative amount and clonal expansion of mutations in a defined region of the genome (chromosome 3) shows promise as a means to differentiate sporadic versus environmental/occupational associated causation of human cancer. The fundamental aim to test the hypothesis that chemical exposure associated cancer formation, unlike sporadic cancer of the same type, is manifest by a higher cumulative acquisition rate for a defined region of the genome is supported by this data. The approach is ideally suited to use on any type of archival tissue or cytology specimen providing objective molecular data that can be used to define cancer causation.