DOE Low Dose Radiation Research Program Workshop I Abstracts November 10-12, 1999, Washington, D.C.

41. DNA Damage vs. Cell Killing by Low-Dose-Rate Gamma Radiation Ultrasensitive Measures, and Implications for Mechanistically Modeled Cancer Risk

K.T. Bogen, M. Weinfeld, X.C. Le, A.D. Murtha, R. Langlois, and G. Keating
HEA L-396, Lawrence Livermore National Laboratory, University of California, 7000 East Avenue, Livermore, CA 94550-9900
bogen@LLNL.gov

Summary: To obtain information critical to improved radiation-risk extrapolation, this project will develop and apply new, ultra-efficient methods allowing direct comparison of low-dose dose-response (LDDR) relations for DNA damage, mutations, and cell killing in different types of human cells, exposed in vitro to low-level gamma radiation.

Abstract: To obtain information critical to improved radiation-risk extrapolation, we will develop and apply new, ultra-efficient methods allowing direct comparison of low-dose dose-response (LDDR) relations for DNA damage, mutations, and cell killing in different types of human cells, exposed in vitro to low-level gamma radiation. Using these methods, we will examine potential LDDR nonlinearities in cell killing (i.e., "hypersensitivity") and in DNA damage/mutation (e.g., due to induced DNA repair). We will also explore how induced DNA damage, mutations and cell killing are each affected by cell-division rate during low-level exposure.

Our methods will include parallel applications of: (1) a new assay for thymine-glycol DNA damage, shown recently to reach zeptomolar (10^-21 M) levels of sensitivity; and (2) unprecedented simultaneous/rapid detection of clonogenic death vs. somatic mutations in ~1 million microcolonies/assay, by means of a novel application of current "gel-microdrop" (GMD) and flow-cytometry (FC) technology. Data involving assay #1 will be obtained in collaboration with ongoing work at the National Cancer Institute of Canada (NCIC). In collaboration with LLNL investigators, data obtained using assay #1 will be supplemented by ultrahigh-sensitivity measures of reduced clonogenic survival based on GMD-FC analysis of the same gamma-exposed cells. To develop/demonstrate assay #2, LLNL investigators will (in Year 1) first modify a line of mutagen-sensitive Chinese hamster ovary (CHO) cells (previously developed at LLNL) to express a FC-detectable fluorescent protein, and then apply existing GMD-encapsulation technology and FC to these CHO cells after low-level mutagenic exposures to demonstrate rapid, ultrasensitive dual detection of reduced clonogenic survival and gene inactivation. Similarly modified lines of human cells will be created at LLNL in Years 2 and 3, to which assays #1 and #2 will be applied in parallel after low-level gamma exposure.

By providing direct comparisons of thymine-glycol damage, mutations, and reduced clonogenic survival in human cells caused by very low-levels of continuous gamma-ray exposure, these studies will reveal LDDR relations critical to improving biologically based predictions of cancer risk posed by low-level exposures to ionizing radiation. Specifically, data obtained will be incorporated into a biologically based cancer-risk model that accounts for competing effects of gamma-induced production vs. killing of premalignant cells. Model predictions will explore how these competing endpoints are likely to shape LDDR for gamma-induced cancer risk. [This work will be performed under the auspices of the U.S. Department of Energy at Lawrence Livermore National Laboratory under contract W-7405-ENG-48.]