Radiation Protection Dosimetry (2016), Vol. 169, No. 1–4, pp. 158–164 doi:10.1093/rpd/ncw107 14C BOMB-PULSE DATING AND STABLE ISOTOPE ANALYSIS FOR GROWTH RATE AND DIETARY INFORMATION IN BREAST CANCER? K. Lång1,*,†, K. Eriksson Stenström2,†, A. Rosso3, M. Bech4, S. Zackrisson1, D. Graubau5 and S. Mattsson6 1Department of Translational Medicine, Division of Diagnostic Radiology, Lund University, Malmö, Sweden 2Department of Physics, Division of Nuclear Physics, Lund University, Lund, Sweden 3Epidemiology and Register Centre South, Skåne University Hospital, Lund, Sweden 4Department of Clinical Sciences, Division of Medical Radiation Physics, Lund University, Lund, Sweden 5Department of Clinical Sciences, Division of Pathology, Lund University, Lund, Sweden 6Department of Translational Medicine, Division of Medical Radiation Physics, Lund University, Malmö, Sweden *Corresponding author: [email protected] The purpose of this study was to perform an initial investigation of the possibility to determine breast cancer growth rate with 14C bomb-pulse dating. Tissues from 11 breast cancers, diagnosed in 1983, were retrieved from a regional biobank. The esti- mated average age of the majority of the samples overlapped the year of collection (1983) within 3σ. Thus, this first study of tumour tissue has not yet demonstrated that 14C bomb-pulse dating can obtain information on the growth of breast cancer. However, with further refinement, involving extraction of cell types and components, there is a possibility that fundamental knowledge of tumour biology might still be gained by the bomb-pulse technique. Additionally, δ13C and δ15N analyses were performed to obtain dietary and metabolic information, and to serve as a base for improvement of the age determination. INTRODUCTION using the 14C bomb-pulse technique, with 14C mea- surements on e.g. gallstones(14) and Alzheimer pla- Large amounts of the radioactive carbon isotope 14C ques(15). A common denominator for these studies is were produced during atmospheric testing of nuclear slow turnover (years). Another example is Gonçalves weapons in the late 1950s and early 1960s. As a con- et al.(16), who demonstrated that human atheroscler- sequence, the concentration of 14C in air was almost otic plaques, known to cause e.g. heart attacks and doubled by 1963. Since the 14C produced was incor- strokes, develop slowly (mean biological ages 5–15 porated in atmospheric CO and introduced into the 2 years). Quantification of the age of various struc- global carbon cycle, all organisms living during the tures in the plaque has gained a better understanding bomb-pulse era, including humans, have been – of the development of the plaque and opened for labelled with bomb-14C(1 3). When the Limited Test improved treatment methods. Ban Treaty from 1963 was implemented, the atmos- The bomb-pulse technique has to the present pheric 14C concentration commenced decreasing authors’ knowledge not yet been used to study mainly due to uptake of 14CO into the oceans and 2 cancer. Understanding of the time cause for the also into the biosphere (see Figure 1). development of human cancer tissue is of utmost Already in the early 1970s, Harkness and importance for improved prevention and treatment Walton(8) realised the potential of using bomb-14C strategies. So far, certain assumptions have had to as a kinetic tracer in humans. Since then, the well- be made in order to estimate how long a cancer has known decreasing atmospheric 14C concentration grown. There are theoretical models to describe has provided useful information in several fields in tumour growth rates, such as an exponential growth the medical sciences, see e.g. Falso and Buchholz(9) model or the Gompertz Model, where the latter and Spalding et al.(10). For instance, this so called takes into consideration a slower growth rate as the 14C bomb pulse has served in studies of turnover (11) tumour size increases. These models have been rates in human cells and tissues (e.g. in eye lenses , (17) (12) (13) applied to cancer in general and to breast cancer fat cells and Achilles tendon ). The develop- – in particular(18 20). These models have been shown ment of various diseases has also been investigated to fit experimental and clinical data, such as obser- vations of the doubling of tumour volume on serial †These authors contributed equally to this work. mammograms. From these studies, the average © The Author 2016. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. 14C AND STABLE ISOTOPES IN BREAST CANCER tumour volume doubling time for human breast can- cer has been estimated to be between 105 and 327 days(21–29). Theoretically, this means that a breast can- cer that is 10-mm large at detection (30 volume dou- blings) has been growing for 9–27 years. However, these studies are limited due to the lack of direct observations of tumour age. The main purpose of this study was to perform an initial investigation of the possibility to determine the growth rate of breast cancer with 14C bomb-pulse dating. An additional aim was to highlight the limita- tions of the 14C bomb-pulse technique of human tis- sues due to dietary variations. Furthermore, the paper pays attention to how dietary information can be obtained from stable isotope ratio analysis of carbon and nitrogen, also providing means to increase the accuracy of bomb-pulse dating. Stable isotope ratio 14 Figure 1. C specific activity in atmospheric CO2 and analysis may also give important information relating oceans representative for the northern hemisphere cancer development to diet. (expressed in units of Fraction Modern, F14C(4);F14Cis ~1.0 before the nuclear weapons tests, corresponding to the natural 14C/12C ratio of about 10−10%(5–7)). Bomb-pulse dating and stable isotope analysis The 14C bomb-pulse technique uses 14Cdatafrom 14 14 fi atmospheric clean-air CO2 to translate the C spe- having lower Cspeci c activity than clean air, thus cific activity of the sample into a calendar date (so contributing to producing too young CaliBomb dates (33) called CaliBomb dates, see ‘Material and Methods’ (at the declining bomb-pulse curve) . 14 section). For humans, carbon enters the body mainly C bomb-pulse dates reported in the literature 14 through the diet. Thus, C data from clean-air CO2 usually do not take into account that there is a delay may not be fully representative for humans. time between production and consumption of the (33, 34) A previous study by Georgiadou et al.(30) used food . Neither is it commonly considered that human blood serum samples from a biobank to esti- marine foodstuffs can influence the accuracy of the mate the accuracy of bomb-pulse dating on human calibration (see the difference in marine and atmos- material. Blood serum samples collected from residents pheric calibration curve in Figure 1). However, since of Malmö (Sweden) in 1978 exhibited CaliBomb dates a large consumption of marine foodstuffs has the between 3.0 ± 0.4 years before the collection date and potential to affect the calibration curve (see Figure 1) 0.2 ± 0.5 years after the collection date (the average on an individual basis, information about the diet deviation from collection date was −1.5 ± 0.7 years). may be important in bomb-pulse dating of human tis- Two major effects associated with the age deviation, sue samples. Additionally, the individual diet is of competing in opposite directions, were identified: (1) interest for studies of the development of various dis- (35) delay time between production and consumption eases, including breast cancer . of foodstuffs, which can explain CaliBomb dates One technique to assess the diet of an organism obtained before the collection date and (2) excessive is analysis of the ratios of stable isotopes of carbon 13 15 consumption of marine food products, which has the and nitrogen (expressed as δ Candδ Nin‰, δ = − × potential of producing CaliBomb dates after the col- where X [(Rsample/Rstandard) 1] 1000, and R 13 12 lection date. is the ratio of heavy to light isotope, i.e. C/ Cor 15 14 (36) 13 15 As discussed in the study by Georgiadou et al.(30), N/ N) (see e.g. Schoeller ). δ Candδ Npro- there are also other factors that have the possibility to vide dietary information since the stable isotope influence the obtained CaliBomb date, however, for ratios in tissues and organs of the consumer reflect the majority of the population probably to a more those of the diet with a small shift. This discrimin- limited extent than delay time and marine food con- ation of one of the stable isotopes occurs in each sumption. Anthropogenic 14C released from nuclear step of the food chain. The difference in δ-value power plants or from research laboratories, industry between an organism and its diet, referred to as the 13 or hospitals using 14C as a tracer has the potential of discrimination factor, is generally about 1‰ for δ C 15 (37) producing too old CaliBomb dates (at the declining and 3‰ for δ N . The discrimination factor, bomb-pulse curve)(31, 32). On the other hand, fossil however, also depends on various factors such as fuel-based products in food industry (e.g. in CO used species, age, metabolic processes and environmental 2 (37) in cultivation in greenhouses) as well as food grown conditions (see e.g. Caut et al. and O’Connell (38) in heavily industrialised areas may lead to foodstuffs et al. ). Thus, different trophic levels (positions in 159 K. LÅNG ET AL.