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Project Title:  A Novel Biodosimetry Method Reduce
Fiscal Year: FY 2011 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 09/01/2007  
End Date: 09/14/2011  
Task Last Updated: 12/19/2011 
Download report in PDF pdf
Principal Investigator/Affiliation:   Bacher, Jeff  Ph.D. / Promega Corporation 
Address:  Genetic Analysis 
2800 Woods Hollow Road 
Madison , WI 53711-5399 
Email: jeff.bacher@promega.com 
Phone: 608-277-2608  
Congressional District:
Web:  
Organization Type: INDUSTRY 
Organization Name: Promega Corporation 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Bailey, Susan  Colorado State University 
Halberg, Richard  University of Wisconsin 
Project Information: Grant/Contract No. NNX07AQ02G 
Responsible Center: NASA JSC 
Grant Monitor: Cucinott1a, Francis  
Center Contact: 281-483-0968 
noaccess@nasa.gov 
Solicitation / Funding Source: 2007 Space Radiation NNJ07ZSA001N 
Grant/Contract No.: NNX07AQ02G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) Cancer:Risk of Radiation Carcinogenesis
Human Research Program Gaps: (1) Cancer03:How can experimental models of carcinogenesis be applied to reduce the uncertainties in radiation quality effects from SPEs and GCR, including effects on tumor spectrum, burden, latency and progression (e.g., tumor aggression and metastatic potential)? (IRP Rev F)
(2) Cancer04:How can models of cancer risk be applied to reduce the uncertainties in dose-rate dependence of risks from SPEs and GCR?
(3) Cancer14:What biodosimetry methods are required for exploration missions and how can biomarker approaches be used for outcome prediction and surveillance? (IRP Rev F)
Flight Assignment/Project Notes: NOTE: End date is now 9/14/2011, per NSSC information (Ed., 9/23/2011)

NOTE: Received NCE through 8/31/2011, per C. Guidry/JSC (08/2010)

Task Description: Exposure of astronauts to space radiation during extended space missions may cause serious health problems. Accurate methods for measuring the biological effects of radiation exposure are, therefore, critical for estimating an individual's health risks. Biodosimetry measurements reflect variation in radiation sensitivity and consequently result in highly individualized estimates of dose and risk. Our novel biodosimetry approach is based on the hypothesis that non-coding repetitive DNA sequences are sensitive to radiation-induced mutations and that these mutations are not harmful to a cell. Therefore, mutations in non-coding repetitive DNA sequences can accumulate and provide a stable molecular record of genetic damage that can be used to determine cumulative radiation exposure and health risk. In our previous NASA grant, we demonstrated the feasibility of using radiation-induced mutations in non-coding repetitive DNA sequences to estimate radiation dose. Our initial data indicate that radiation-induced mutations in non-coding repetitive DNA markers are stable over time and additive over multiple exposures. In this successor proposal, we plan to extend our previous work by developing optimized multiplex marker panels for human and mouse biodosimetry, validate our approach by comparing our assay to current gold standard cytological methods and then utilize the novel system to assess risks from space radiation and improve our understanding of how these risks are affected by variations in dose rate, dose fractionation and genome stability. The main contribution of the proposed research to manned space exploration is the validation of a novel biodosimetry method for estimating dose and risks from exposure to space radiation. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Research Impact/Earth Benefits: Mutational load profiling, through analysis of mutations in tandem DNA repeat sequences, is a simple, non-invasive and generalized approach for monitoring an individual's cumulative record of mutations that may be useful for determining health risks and effectiveness of countermeasures for astronauts or other individuals exposed to ionizing radiation or chemical mutagens. Biomarkers identified in this study are also sensitive to free radical DNA damage and therefore may be useful markers for detection of cancer and other degenerative diseases in which oxidative stress is involved. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research. Biomarkers developed for this NASA project are currently being evaluated in clinical studies for use in the early detection of colon cancer.

Task Progress & Bibliography Information FY2011 
Task Progress: This research has led to the discovery and development of novel biomarkers and methodologies for monitoring radiation-induced mutations in humans and in mouse models. We demonstrated that under some conditions and in certain tissues (e.g., blood) our biodosimetry method can be used to assess dose. However, we have found that the dose response was influenced by numerous factors, including; radiation quality, dose rate, LET, time and tissue type. The sensitivity of our assay to a broad range of factors was surprising, but suggests a much broader utility of this approach for estimating an individual’s risk from radiation exposure. Biodosimetry measurements reflect the combined effects and interactions of all factors that influence mutation induction in an individual. Thus, biodosimetry is generally a rather poor estimator of actual dose, but can be an important estimator of an individual’s health risk from radiation exposure.

Our major findings include:

(1) Some non-coding microsatellite repeats are sensitive to radiation-induced mutations in a dose dependent manner (in some tissues) and therefore, monitoring changes in mutational load may is a viable biodosimetry method,

(2) Spontaneous mutations in microsatellite repeats accumulated linearly over time, indicating these mutations are stable,

(3) Fractionated exposures to iron ions, protons and gamma rays given in 24 hour intervals were additive,

(4) The relative biological effectiveness for induction of microsatellite mutations of 1 GeV/n iron ions was <1 and for 1 GeV/n protons <2,

(5) Microsatellite mutation induction was influenced by dose, dose rate, radiation quality, dose fractionation, LET, time, tissue type and DNA repair status,

(6) Microsatellite repeats containing long polyA runs typically occur within highly repetitive SINE and LINE elements and DNA damage induced recombination between these elements may contribute to the observed mutagenesis in polyA microsatellites,

(7) Microsatellite repeats containing long polyA runs are predicted to contain sequences associated with matrix attachment regions and this may be related to the observed differences in tissue specific radio-sensitivity,

(8) Radiation-induced microsatellite mutations appears to require a functional mismatch repair system in most tissues, suggesting error prone repair of radiation-induced lesions in repeat sequences,

(9) Split dose, dual ion experiments indicate a potential in vivo adaptive response to mixed beams of HZE iron ions and protons.

Knowledge gained from this research project will provide new insights into the effects of radiation on DNA mutagenesis and establishes novel biomarkers and methods with broad utility not only for the benefit of NASA, but also for future studies in radiation biology, toxicology, and cancer research. Biomarkers developed for this NASA project are currently being evaluated in clinical studies for use in the early detection of colon cancer.

Bibliography Type: Description: (Last Updated: 04/16/2019)  Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Steffen L, Weil M, Ray A, Genik P, Ullrich R, Fallgren C, Stroy M, Bouffler S, Gillan J, Bourdeau-Heller J, Leismann R, Bacher J. "Microsatellite Instability in Radiation-Induced Acute Myeloid Leukemia." Presented at the 55th Annual Meeting of the Radiation Research Society, Savannah, Georgia, October 3-7, 2009.

55th Annual Meeting of the Radiation Research Society, Savannah, Georgia, October 3-7, 2009. p. 32. http://c.ymcdn.com/sites/www.radres.org/resource/collection/0EA821A7-E616-428F-A197-38BFBD1E38D5/RRS_2009_Abstracts.pdf , Oct-2009

Abstracts for Journals and Proceedings Bourdeau-Heller J, Leisemann R, Halberg R, Steffen L, Storts D, Bacher J. "Microsatellite–Based Biodosimetry." 21st Annual NASA Space Radiation Investigators' Workshop, Port Jefferson, NY, May 16-19, 2010.

Program and abstracts. 21st Annual NASA Space Radiation Investigators' Workshop, Port Jefferson, NY, May 16-19, 2010. p. 101. http://www.abstractsonline.com/Plan/ViewAbstract.aspx?mID=2569&sKey=9e011f35-da29-44e9-81ca-4b52cf617fb4&cKey=333dabe7-4431-4dad-8578-0d77a4afc44c&mKey=5fb93787-2553-4b3b-a915-8c1ae71f127a , May-2010

Abstracts for Journals and Proceedings Bacher J, Bourdeau-Heller J, Leisemann R, Halberg R, Steffen L, Storts D. "Microsatellite Mutations in Mouse Tissues Induced by Low and High LET Radiation." 21st Annual NASA Space Radiation Investigators' Workshop, Port Jefferson, NY, May 16-19, 2010.

Program and abstracts. 21st Annual NASA Space Radiation Investigators' Workshop, Port Jefferson, NY, May 16-19, 2010. p. 37. , May-2010

Abstracts for Journals and Proceedings Bourdeau-Heller J, Leisemann R, Halberg R, Steffen L, Storts D, Bacher J. "Microsatellite mutations in mouse tissues induced by low and high LET radiation." Presented at the 56th Annual Meeting of Radiation Research Society, Maui, HI, September 25-29, 2010.

Proceedings of the 56th Annual Meeting of Radiation Research Society, Maui, HI, September 25-29, 2010. http://www.abstractsonline.com/Plan/ViewAbstract.aspx?mID=2569&sKey=9e011f35-da29-44e9-81ca-4b52cf617fb4&cKey=333dabe7-4431-4dad-8578-0d77a4afc44c&mKey=5fb93787-2553-4b3b-a915-8c1ae71f127a , Sep-2010

Abstracts for Journals and Proceedings Bourdeau-Heller J, Leisemann-Immel R, Steffen L, Halberg R, Betlach M, Storts D, Bacher J. "Microsatellites in Space." Presented at the 22nd Annual Space Radiation Investigators' Workshop, League City, TX, September 18-21, 2011.

22nd Annual NASA Space Radiation Investigators Workshop, League City, Texas, September 18-21, 2011. , Sep-2011

Articles in Peer-reviewed Journals Bacher JW, Sievers CK, Albrecht DM, Grimes IC, Weiss JM, Matkowskyj KA, Agni RM, Vyazunova I, Clipson L, Storts DR, Thliveris AT, Halberg RB. "Improved detection of microsatellite instability in early colorectal lesions." PLoS One. 2015 Aug 7;10(8):e0132727. eCollection 2015. https://doi.org/10.1371/journal.pone.0132727 ; PubMed PMID: 26252492; PubMed Central PMCID: PMC4529134 , Aug-2015
Patents US20090068646. Pending. Published March 2009. Mar-2009 Bacher J, Halberg R, Kent M. "Methods and Kits for Detecting Mutations."
Patents US 20080311565. Pending. Published December 2008. Dec-2008 Kent M, Bacher J, Megid W. "Methods and Kits for Detecting Germ Cell Genomic Instability."
Project Title:  A Novel Biodosimetry Method Reduce
Fiscal Year: FY 2010 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 09/01/2007  
End Date: 09/14/2011  
Task Last Updated: 10/29/2010 
Download report in PDF pdf
Principal Investigator/Affiliation:   Bacher, Jeff  Ph.D. / Promega Corporation 
Address:  Genetic Analysis 
2800 Woods Hollow Road 
Madison , WI 53711-5399 
Email: jeff.bacher@promega.com 
Phone: 608-277-2608  
Congressional District:
Web:  
Organization Type: INDUSTRY 
Organization Name: Promega Corporation 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Bailey, Susan  Colorado State University 
Halberg, Richard  University of Wisconsin 
Project Information: Grant/Contract No. NNX07AQ02G 
Responsible Center: NASA JSC 
Grant Monitor: Cucinott1a, Francis  
Center Contact: 281-483-0968 
noaccess@nasa.gov 
Solicitation / Funding Source: 2007 Space Radiation NNJ07ZSA001N 
Grant/Contract No.: NNX07AQ02G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) Cancer:Risk of Radiation Carcinogenesis
Human Research Program Gaps: (1) Cancer03:How can experimental models of carcinogenesis be applied to reduce the uncertainties in radiation quality effects from SPEs and GCR, including effects on tumor spectrum, burden, latency and progression (e.g., tumor aggression and metastatic potential)? (IRP Rev F)
(2) Cancer04:How can models of cancer risk be applied to reduce the uncertainties in dose-rate dependence of risks from SPEs and GCR?
(3) Cancer14:What biodosimetry methods are required for exploration missions and how can biomarker approaches be used for outcome prediction and surveillance? (IRP Rev F)
Flight Assignment/Project Notes: NOTE: End date is now 9/14/2011, per NSSC information. New Gaps added per HRR information (Ed., 9/23/2011)

NOTE: Received NCE through 8/31/2011, per C. Guidry/JSC (08/2010)

Task Description: Exposure of astronauts to space radiation during extended space missions may cause serious health problems. Accurate methods for measuring the biological effects of radiation exposure are, therefore, critical for estimating an individual's health risks. Biodosimetry measurements reflect variation in radiation sensitivity and consequently result in highly individualized estimates of dose and risk. Our novel biodosimetry approach is based on the hypothesis that non-coding repetitive DNA sequences are sensitive to radiation-induced mutations and that these mutations are not harmful to a cell. Therefore, mutations in non-coding repetitive DNA sequences can accumulate and provide a stable molecular record of genetic damage that can be used to determine cumulative radiation exposure and health risk. In our previous NASA grant, we demonstrated the feasibility of using radiation-induced mutations in non-coding repetitive DNA sequences to estimate radiation dose. Our initial data indicate that radiation-induced mutations in non-coding repetitive DNA markers are stable over time and additive over multiple exposures. In this successor proposal, we plan to extend our previous work by developing optimized multiplex marker panels for human and mouse biodosimetry, validate our approach by comparing our assay to current gold standard cytological methods and then utilize the novel system to assess risks from space radiation and improve our understanding of how these risks are affected by variations in dose rate, dose fractionation and genome stability. The main contribution of the proposed research to manned space exploration is the validation of a novel biodosimetry method for estimating dose and risks from exposure to space radiation. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Research Impact/Earth Benefits: Mutational load profiling, through analysis of mutations in tandem DNA repeat sequences, is a simple, non-invasive and generalized approach for monitoring an individual's cumulative record of mutations that may be useful for determining health risks and effectiveness of countermeasures for astronauts or other individuals exposed to ionizing radiation or chemical mutagens. Biomarkers identified in this study are also sensitive to free radical DNA damage and therefore may be useful markers for detection of cancer and other degenerative diseases in which oxidative stress is involved. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Task Progress & Bibliography Information FY2010 
Task Progress: This research has led to the discovery and development of novel biomarkers and methodologies for monitoring radiation-induced mutations in humans and in mouse models. We have demonstrated that under some conditions and in certain tissues (e.g., blood) our biodosimetry method can be used to assess dose. However, we have found that the dose response was influenced by numerous factors, including; radiation quality, dose rate, LET, time and tissue type. Biodosimetry measurements reflect the combined affects and interactions of all factors that influence mutation induction in an individual. Thus, biodosimetry is generally a rather poor estimator of actual dose, but can be an important estimator of an individuals health risk from radiation exposure.

Our major findings include:

• Mutation induction in certain types of coding and non-coding microsatellite repeats is dose dependent in most tissues

• Microsatellite markers sensitive to radiation-induced mutations are found almost exclusively within highly repetitive SINE and LINE elements in human and mouse genomes and recombination repair between these elements may contribute to mutagenesis

• Microsatellite mutation induction is influenced by radiation quality, dose rate, LET, time and tissue type

• The relative biological effectiveness for induction of microsatellite mutations of 1 GeV/n iron ions was less than that of gamma rays. The RBE of 1 GeV/n protons was <2

• Split dose, dual ions experiments indicate a potential adaptive response to sequential exposures of iron ions followed by protons, but not vice versa

• There was evidence of delayed onset genomic instability in some tissue types (e.g., buccal cells) following exposure to 1 GeV/n iron ions and a high incidence of liver tumors within 2 years of exposure

• Fractionated exposure to iron ions was additive (24 hours between doses)

• Mismatch repair deficient mice exhibit a much higher level of spontaneous mutations, but only show radiation-induced mutations in colon

Bibliography Type: Description: (Last Updated: 04/16/2019)  Show Cumulative Bibliography Listing
 
Articles in Peer-reviewed Journals Haines J, Bacher J, Coster M, Huiskamp R, Meijne E, Mancuso M, Pazzaglia S, Bouffler S. "Microsatellite instability in radiation-induced murine tumours; influence of tumour type and radiation quality." International Journal of Radiation Biology 2010 Jul;86(7):555-68. PMID: 20545567 , Jul-2010
Articles in Peer-reviewed Journals Halberg RB, Waggoner J, Rasmussen K, White A, Clipson L, Prunuske AJ, Bacher JW, Sullivan R, Washington MK, Pitot HC, Petrini JH, Albertson DG, Dove WF. "Long-lived Min mice develop advanced intestinal cancers through a genetically conservative pathway." Cancer Research 2009 Jul 15;69(14):5768-75. PMID: 19584276 , Jul-2009
Project Title:  A Novel Biodosimetry Method Reduce
Fiscal Year: FY 2009 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 09/01/2007  
End Date: 08/31/2011  
Task Last Updated: 07/03/2009 
Download report in PDF pdf
Principal Investigator/Affiliation:   Bacher, Jeff  Ph.D. / Promega Corporation 
Address:  Genetic Analysis 
2800 Woods Hollow Road 
Madison , WI 53711-5399 
Email: jeff.bacher@promega.com 
Phone: 608-277-2608  
Congressional District:
Web:  
Organization Type: INDUSTRY 
Organization Name: Promega Corporation 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Bailey, Susan  Colorado State University 
Halberg, Richard  University of Wisconsin 
Project Information: Grant/Contract No. NNX07AQ02G 
Responsible Center: NASA JSC 
Grant Monitor: Cucinott1a, Francis  
Center Contact: 281-483-0968 
noaccess@nasa.gov 
Solicitation / Funding Source: 2007 Space Radiation NNJ07ZSA001N 
Grant/Contract No.: NNX07AQ02G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) Cancer:Risk of Radiation Carcinogenesis
Human Research Program Gaps: (1) Cancer03:How can experimental models of carcinogenesis be applied to reduce the uncertainties in radiation quality effects from SPEs and GCR, including effects on tumor spectrum, burden, latency and progression (e.g., tumor aggression and metastatic potential)? (IRP Rev F)
(2) Cancer04:How can models of cancer risk be applied to reduce the uncertainties in dose-rate dependence of risks from SPEs and GCR?
(3) Cancer14:What biodosimetry methods are required for exploration missions and how can biomarker approaches be used for outcome prediction and surveillance? (IRP Rev F)
Flight Assignment/Project Notes: NOTE: Received NCE through 8/31/2011, per C. Guidry/JSC (08/2010)

Task Description: Exposure of astronauts to space radiation during extended space missions may cause serious health problems. Accurate methods for measuring the biological effects of radiation exposure are, therefore, critical for estimating an individual's health risks. Biodosimetry measurements reflect variation in radiation sensitivity and consequently result in highly individualized estimates of dose and risk. Our novel biodosimetry approach is based on the hypothesis that non-coding repetitive DNA sequences are sensitive to radiation-induced mutations and that these mutations are not harmful to a cell. Therefore, mutations in non-coding repetitive DNA sequences can accumulate and provide a stable molecular record of genetic damage that can be used to determine cumulative radiation exposure and health risk. In our previous NASA grant, we demonstrated the feasibility of using radiation-induced mutations in non-coding repetitive DNA sequences to estimate radiation dose. Our initial data indicate that radiation-induced mutations in non-coding repetitive DNA markers are stable over time and additive over multiple exposures. In this successor proposal, we plan to extend our previous work by developing optimized multiplex marker panels for human and mouse biodosimetry, validate our approach by comparing our assay to current gold standard cytological methods and then utilize the novel system to assess risks from space radiation and improve our understanding of how these risks are affected by variations in dose rate, dose fractionation and genome stability. The main contribution of the proposed research to manned space exploration is the validation of a novel biodosimetry method for estimating dose and risks from exposure to space radiation. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Research Impact/Earth Benefits: Mutational load profiling, through analysis of mutations in tandem DNA repeat sequences, is a simple, non-invasive and generalized approach for monitoring an individual's cumulative record of mutations that may be useful for determining health risks and effectiveness of countermeasures for astronauts or other individuals exposed to ionizing radiation or chemical mutagens. Biomarkers identified in this study are also sensitive to free radical DNA damage and therefore may be useful markers for detection of cancer and other degenerative diseases in which oxidative stress is involved. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Task Progress & Bibliography Information FY2009 
Task Progress: Irradiation and sample collection began in the first year and has been successfully completed for all but two of our experiments, which are scheduled for fall 2009. In the 1st year we focused on identifying new mouse and human microsatellite markers and constructed mouse and human multiplexes assay for small-pool PCR (SP-PCR). In the 2nd year we have improved our SP-PCR assay to dramatically increase throughput and accuracy while reducing costs. Mutational analysis of samples began in the later part of the first year and became the primary focus in the later part of the 2nd year. We have been able to successfully culture primary buccal cells and therefore will be using primary human blood and buccal cells for dose response studies instead of transformed cell lines. Human research protocols these new experiments have been approved by our institute and are under review by Brookhaven National Laboratory. A brief description of each planned experiment and current status is given below.

Experiment 1. Biomarker discovery: Suitable microsatellite repeats in mouse and human genomes were identified and screened for sensitivity to radiation-induced mutations. We identified over 300 new microsatellite markers and constructed mouse and human multiplexes assay for small-pool PCR (SP-PCR). Status: completed.

Experiment 2. Dose Response in mice: Mice were exposed to various doses of HZE iron particles, protons or gamma rays, allowed to recover for 3 days, then samples were collected for later mutation testing. Status: irradiations and sample collection completed.

Experiment 3. Dose Response in human cells: Cultured primary human buccal and blood cells will be irradiated at NSRL-09C and analyzed for mutations using SP-PCR. IRB protocols are under review.

Experiment 4. Stability: Blood and buccal samples from B6 mice were be exposed to iron ions, protons or gamma rays and analyzed for mutations after 3 days, 3 mo, 12 mo and 18 mo. Irradiations have been completed on all mice and samples collected for all time points except 18 months. Analysis of samples is ongoing.

Experiment 5. Mutations in coding repeats: Irradiated mismatch repair deficient mice containing the SupFG1 mutation reporter were screened for mutations in (C)8 and (G)7 coding repeats of the SupFG1 gene. DNA sequencing revealed that 97% of the SupFG1 mutations were insertions or deletions in the (G)7 or the (C)8 mononucleotide repeats. Mutations in these short repeats exited a dose response that was highly correlated (but about 10-fold lower) with mutations in longer non-coding repeats. Status: completed.

Experiment 6. Dose rate effects: B6 mice were irradiated with 1 Gy iron, protons and gamma rays at dose rates of 0, 10, 20, 50, 100 and 150 cGy/minute, then analyzed for mutations by SP-PCR. Preliminary results indicate that mutation frequency increases linearly in mouse blood cells with increasing dose rates up to 50 cGy/min iron ions. This trend did not occur with protons or gamma rays, but we will need to analyze more samples to confirm this. Status: analysis ongoing.

Experiment 7. Dose- fractionation: B6 mice were exposed to 0.5 Gy of iron, protons or gamma rays at 0.5 Gy/m followed by another 0.5 Gy dose 3 days later. Mutation frequency after a single 1 Gy dose will be compared to 2x 0.5 Gy dose. Status: irradiations and sample collection complete.

Experiment 8. Adaptive response: B6 mice were exposed to different combinations of a 0.1 Gy priming dose followed 24 hr later by a 0.9 Gy challenge dose. Samples were collected 3 days later for mutation analysis by SP-PCR to determine if mutation frequencies are additive, lower than expected or higher than expected. Status: irradiations and sample collection complete.

Experiment 9. Aging effects: Young (6 wk) and old (18 mo) B6, CBA/Ca and Balb/c mice will be irradiated and screened for mutations by SP-PCR and chromosomal and telomere aberrations using Giemsa staining and telomere FISH. Status: scheduled for NSRL-09C.

Experiment 10. DNA mismatch repair expression: qPCR, IHC, Western Blot and methylation specific PCR assays will be performed on tissues from irradiated mice to determine whether mismatch repair gene expression or methylation patterns are altered following radiation exposure. Status: irradiations and sample collection complete. qPCR and IHC assays are in development.

Experiment 11. Tissue Effects: Msh2-/- deficient mice were irradiated with 1 Gy iron ions and are being screened for microsatellite mutations in blood, buccal cells, spleen, colon, liver and brain tissues. Mutation frequency differences were observed in different tissues. Msh2+/+ and +/- tissues exhibit similar mutation frequencies. Msh2 deficient tissues had approximately 10-fold higher rates of mutation rates compared to wild type. Status: irradiations and sample collection complete. Analysis ongoing.

Experiment 12. LET Effects: Mice were irradiated with iron ions of different LET (300, 600 and 1000 MeV/n) and blood samples will be tested for mutations by SP-PCR. Status: Irradiations and sample collection complete.

Bibliography Type: Description: (Last Updated: 04/16/2019)  Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Bourdeau-Heller J, Steffen L, Leisemann R, Bailey S, Halberg R, Stanhope S, Storts D, Bacher J. "Microsatellite Mutations Induced by High and Low-LET Radiation." Radiation mutagenesis session, Heavy Ions in Therapy and Space Symposium, Cologne, Germany, July 6-10, 2009.

Heavy Ions in Therapy and Space Symposium, Cologne, Germany, July 2009. p. 230. http://www.heavyions2009.de/images/pdf/HeavyIons_AbstractBook_B5.pdf , Jul-2009

Abstracts for Journals and Proceedings Steffen L, Weil M, Ray F, Genik P, Ullrich R, Fallgren C, Story MD, Gillan J, Bouffler S, Bourdeau-Heller J, Leisemann R, Bacher J. "Microsatellite Instability in Radiation-Induced Acute Myeloid Leukemia." Radiation carcinogenesis session, Heavy Ions in Therapy and Space Symposium, Cologne, Germany, July 6-10, 2009.

Heavy Ions in Therapy and Space Symposium, Cologne, Germany, July 2009. p. 223. http://www.heavyions2009.de/images/pdf/HeavyIons_AbstractBook_B5.pdf , Jul-2009

Articles in Peer-reviewed Journals Haines J, Bacher J, Huiskamp R, Meijne E, Mancuso M, Pazzaglia S, Bouffler S. "Microsatellite Instability in Radiation-induced Murine tumours; Influence of Tumour Type and Radiation Quality." International Journal of Radiation Research (in press). Expected publication December 2009. , Jul-2009
Articles in Peer-reviewed Journals Halberg R, Jesse Waggoner J, Rasmussen K, White A, Clipson L, Bacher J, Sullivan R, Washington M, Pitot H, Petrini J, Albertson D, Dove W. "Long-lived Min Mice Develop Advanced Intestinal Cancers Through A Genetically Conservative Pathway." Cancer Research (in press). Expected publication December 2009. , Jul-2009
Project Title:  A Novel Biodosimetry Method Reduce
Fiscal Year: FY 2008 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 09/01/2007  
End Date: 08/31/2010  
Task Last Updated: 07/11/2008 
Download report in PDF pdf
Principal Investigator/Affiliation:   Bacher, Jeff  Ph.D. / Promega Corporation 
Address:  Genetic Analysis 
2800 Woods Hollow Road 
Madison , WI 53711-5399 
Email: jeff.bacher@promega.com 
Phone: 608-277-2608  
Congressional District:
Web:  
Organization Type: INDUSTRY 
Organization Name: Promega Corporation 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Bailey, Susan  Colorado State University 
Halberg, Richard  University of Wisconsin 
Project Information: Grant/Contract No. NNX07AQ02G 
Responsible Center: NASA JSC 
Grant Monitor: Cucinott1a, Francis  
Center Contact: 281-483-0968 
noaccess@nasa.gov 
Solicitation / Funding Source: 2007 Space Radiation NNJ07ZSA001N 
Grant/Contract No.: NNX07AQ02G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) Cancer:Risk of Radiation Carcinogenesis
Human Research Program Gaps: (1) Cancer03:How can experimental models of carcinogenesis be applied to reduce the uncertainties in radiation quality effects from SPEs and GCR, including effects on tumor spectrum, burden, latency and progression (e.g., tumor aggression and metastatic potential)? (IRP Rev F)
(2) Cancer04:How can models of cancer risk be applied to reduce the uncertainties in dose-rate dependence of risks from SPEs and GCR?
(3) Cancer14:What biodosimetry methods are required for exploration missions and how can biomarker approaches be used for outcome prediction and surveillance? (IRP Rev F)
Task Description: Exposure of astronauts to space radiation during extended space missions may cause serious health problems. Accurate methods for measuring the biological effects of radiation exposure are, therefore, critical for estimating an individual¿s health risks. Biodosimetry measurements reflect variation in radiation sensitivity and consequently result in highly individualized estimates of dose and risk. Our novel biodosimetry approach is based on the hypothesis that non-coding repetitive DNA sequences are sensitive to radiation-induced mutations and that these mutations are not harmful to a cell. Therefore, mutations in non-coding repetitive DNA sequences can accumulate and provide a stable molecular record of genetic damage that can be used to determine cumulative radiation exposure and health risk. In our previous NASA grant, we demonstrated the feasibility of using radiation-induced mutations in non-coding repetitive DNA sequences to estimate radiation dose. Our initial data indicate that radiation-induced mutations in non-coding repetitive DNA markers are stable over time and additive over multiple exposures. In this successor proposal, we plan to extend our previous work by developing optimized multiplex marker panels for human and mouse biodosimetry, validate our approach by comparing our assay to current gold standard cytological methods and then utilize the novel system to assess risks from space radiation and improve our understanding of how these risks are affected by variations in dose rate, dose fractionation and genome stability. The main contribution of the proposed research to manned space exploration is the validation of a novel biodosimetry method for estimating dose and risks from exposure to space radiation. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Research Impact/Earth Benefits: Mutational load profiling, through analysis of mutations in tandem DNA repeat sequences, is a simple, non-invasive and generalized approach for monitoring an individual's cumulative record of mutations that may be useful for determining health risks and effectiveness of countermeasures for astronauts or other individuals exposed to ionizing radiation or chemical mutagens. Biomarkers identified in this study are also sensitive to free radical DNA damage and therefore may be useful markers for detection of cancer and other degenerative diseases in which oxidative stress is involved. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Task Progress & Bibliography Information FY2008 
Task Progress: We have previously demonstrated that mutations in selected biomarkers exhibit a dose response to radiation exposure in normal AG01522 human fibroblasts cell cultures. A dose-dependent response was observed in vivo in mouse blood, cheek and brain cells in tissue samples collected 10 weeks after exposure. A significant increase in radiation-induced mutations in mononucleotide repeats was detectible in mouse blood and cheek samples up to 26 weeks after radiation exposure and these mutations were additive over multiple exposures. In this successor proposal we plan to build on our previous work and ultimately plan to validate our biodosimetry method for assessing radiation exposure in human lymphoblast cells and in vivo in mouse blood and buccal cells. We will confirm that mutation frequency from fractionated exposures is additive and extend stability studies out to about 2 years post irradiation with gamma, protons and iron ions.

The usefulness of our biodosimetry assay as a surrogate biomarker for estimating radiation-induced cancer risk is being investigated by looking for correlation with other known cancer risk factors, such as chromosomal aberrations and mutations in coding repeats. Mismatch repair deficient SupFG1 mutation reporter mice were irradiated and screened for mutations in (C)8 and (G)7 coding repeats of the SupFG1 transgene and compared to mutations observed in tandem DNA repeats. The sensitivity of our PCR-based mutation assay was 10-fold greater than that observed for the SupFG1 transgenic mouse mutation assay. DNA sequencing revealed that almost all (97%) of the SupFG1 mutations were insertions or deletions in the (G)7 or the (C)8 mononucleotide repeats. The susceptibility of short coding repeats to radiation-induced mutations is troubling since there are thousands of similar coding repeats within the human genome that may be vulnerable. Radiation-induced mutations in non-coding repeats were highly correlated with mutations in short coding repeats (which are considered biomarkers for cancer risk) and therefore may be useful as surrogate markers for cancer “risk” as well as “dose”. Correlation with chromosomal aberrations is currently being investigated.

We have completed two NSRL runs thus far. During the NSRL-08A run we started experiments designed to test the effects of dose rate on mutation induction in tandem repeats and also experiments to determine the stability of mutations in repeats over time. To do this human AG9389 lymphoblast cells were exposed to 1 Gy of iron ions, protons and gamma rays at various dose rates (0.1, 0.2, 0.5 and 1 Gy/min), allowed to recover for 3 days, then snap frozen for later mutational analysis. We also irradiated C57BL/6 mice with 1 Gy of iron ions, protons or gamma rays and collected blood and tissue samples (cheek swabs, spleen, liver, brain and colon) after 3 days. Additional groups of mice from NSRL-08A run will be tested after 4 months, 1 year and 21 months to determine the stability of mutations in DNA repeat markers and chromosomal aberrations.

During the NSRL-08B run we started experiments designed to validate our biodosimetry method by comparing results of obtained using tandem repeat markers to the gold standard, chromosomal aberration analysis. Human AG9389 lymphoblast cells were exposed to doses of 0.1, 0.2, 0.5, 1 and 2 Gy of iron ions, protons or gamma rays. The cells were allowed to recover for 3 days then either snap frozen for mutational analysis or shipped to co-investigator Susan Bailey for chromosomal analysis. The cells will be tested for DNA repeat mutations using small-pool PCR and for chromosomal aberrations using Giemsa staining and whole chromosome painting for translocations. This validation experiment is will be repeated at NSRL-08C so we can compare results from two independent experiments.

Other confounding effects that may affect biodosimetry measurements are being investigated. For example, the deleterious effects of radiation exposure may increase with age. To test this, 20-month-old C57BL/6, CBA/Ca and Balb/c mice will be irradiated and results for tandem repeat mutations, mismatch repair gene expression, mismatch repair gene methylation status and telomere stability will be compared to 2-month-old mice. Mice are currently being aged and irradiation experiments are planned in 2009 at NSRL-09C. We are also investigating the effects of dose rate, dose fractionation, dual ions and the effect of radiation on DNA mismatch repair gene expression and how these may alter the frequency of radiation-induced mutations in tandem repeats, and thus the accuracy of biodosimetry measurements.

Bibliography Type: Description: (Last Updated: 04/16/2019)  Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Ensenberger MG, Megid WA, Halberg RB, Steffen LS, Bourdeau-Heller JM, Stanhope SA, Kent-First MG, Prolla TA, Storts DR, Bacher JW. "A Novel Biodosimetry Method." Presented at the NASA Human Research Program Investigators' Workshop, League City, Texas, January 2008.

2008 NASA Human Research Program Investigators' Workshop, January 2008. , Jan-2008

Abstracts for Journals and Proceedings Bacher JW, Ensenberger MG, Megid WA, Halberg RB, Steffen LS, Bourdeau-Heller, JM, Stanhope SA, Grochowski E, Storts DR. "A Novel Biodosimetry Method." 19th Annual NASA Space Radiation Investigators Workshop, Philadelphia, PA, June 30-July 2, 2008.

Proceedings from the 19th Annual NASA Space Radiation Investigators Workshop, July 2008. , Jul-2008

Project Title:  A Novel Biodosimetry Method Reduce
Fiscal Year: FY 2007 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 09/01/2007  
End Date: 08/31/2010  
Task Last Updated: 10/11/2007 
Download report in PDF pdf
Principal Investigator/Affiliation:   Bacher, Jeff  Ph.D. / Promega Corporation 
Address:  Genetic Analysis 
2800 Woods Hollow Road 
Madison , WI 53711-5399 
Email: jeff.bacher@promega.com 
Phone: 608-277-2608  
Congressional District:
Web:  
Organization Type: INDUSTRY 
Organization Name: Promega Corporation 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Bailey, Susan  Colorado State University 
Halberg, Richard  University of Wisconsin 
Project Information: Grant/Contract No. NNX07AQ02G 
Responsible Center: NASA JSC 
Grant Monitor:  
Center Contact:   
Solicitation / Funding Source: 2007 Space Radiation NNJ07ZSA001N 
Grant/Contract No.: NNX07AQ02G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:  
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:  
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) Cancer:Risk of Radiation Carcinogenesis
Human Research Program Gaps: (1) Cancer03:How can experimental models of carcinogenesis be applied to reduce the uncertainties in radiation quality effects from SPEs and GCR, including effects on tumor spectrum, burden, latency and progression (e.g., tumor aggression and metastatic potential)? (IRP Rev F)
(2) Cancer04:How can models of cancer risk be applied to reduce the uncertainties in dose-rate dependence of risks from SPEs and GCR?
(3) Cancer14:What biodosimetry methods are required for exploration missions and how can biomarker approaches be used for outcome prediction and surveillance? (IRP Rev F)
Task Description: Exposure of astronauts to space radiation during extended space missions may cause serious health problems. Accurate methods for measuring the biological effects of radiation exposure are, therefore, critical for estimating an individual¿s health risks. Biodosimetry measurements reflect variation in radiation sensitivity and consequently result in highly individualized estimates of dose and risk. Our novel biodosimetry approach is based on the hypothesis that non-coding repetitive DNA sequences are sensitive to radiation-induced mutations and that these mutations are not harmful to a cell. Therefore, mutations in non-coding repetitive DNA sequences can accumulate and provide a stable molecular record of genetic damage that can be used to determine cumulative radiation exposure and health risk. In our previous NASA grant, we demonstrated the feasibility of using radiation-induced mutations in non-coding repetitive DNA sequences to estimate radiation dose. Our initial data indicate that radiation-induced mutations in non-coding repetitive DNA markers are stable over time and additive over multiple exposures. In this successor proposal, we plan to extend our previous work by developing optimized multiplex marker panels for human and mouse biodosimetry, validate our approach by comparing our assay to current gold standard cytological methods and then utilize the novel system to assess risks from space radiation and improve our understanding of how these risks are affected by variations in dose rate, dose fractionation and genome stability. The main contribution of the proposed research to manned space exploration is the validation of a novel biodosimetry method for estimating dose and risks from exposure to space radiation. Completion of this research should provide new insights into the effects of space radiation on DNA mutagenesis and establishes panels of human and mouse biomarkers with broad utility for future studies in radiation biology, toxicology and cancer research.

Research Impact/Earth Benefits: 0

Task Progress & Bibliography Information FY2007 
Task Progress: New project for FY2007.

Bibliography Type: Description: (Last Updated: 04/16/2019)  Show Cumulative Bibliography Listing
 
 None in FY 2007