{"id":4722,"date":"2023-02-16T10:31:49","date_gmt":"2023-02-16T16:31:49","guid":{"rendered":"https:\/\/www.crpa-acrp-bulletin.ca\/?p=4722"},"modified":"2023-04-17T19:33:25","modified_gmt":"2023-04-18T01:33:25","slug":"single-cell-dna-sequencing-a-potential-dosimetric-tool","status":"publish","type":"post","link":"https:\/\/www.crpa-acrp-bulletin.ca\/fr\/2023\/02\/16\/single-cell-dna-sequencing-a-potential-dosimetric-tool\/","title":{"rendered":"Single-Cell DNA Sequencing \u2014 A Potential Dosimetric Tool"},"content":{"rendered":"

\"\"<\/h4>\n

F. Mathew, J. Manalad, L. Galarneau, N. Ybarra, and J. Kildea, and P. N. Tonin, McGill University, Montreal, QC
\nJ. Yeo, National University of Singapore
\nY.C. Wang, and I. Ragoussis, McGill Genome Centre, Montreal, QC<\/h4>\n
\n

R\u00e9sum\u00e9 :<\/h4>\n
\n

Chaque ann\u00e9e, le comit\u00e9 des \u00e9tudiants et des jeunes professionnels de l\u2019ACRP organise le concours de communications \u00e9tudiantes Anthony J. MacKay. La ou le laur\u00e9at re\u00e7oit le troph\u00e9e Anthony J. MacKay, un prix de 250 $ en esp\u00e8ces et son article est publi\u00e9 dans le Bulletin<\/em> de l\u2019ACRP. L\u2019ann\u00e9e derni\u00e8re, c\u2019est Felix Mathew qui a remport\u00e9 le concours. Felix a pr\u00e9sent\u00e9 son article intitul\u00e9 \u00ab\u2009Single-Cell DNA Sequencing \u2014 A Potential Dosimetric Tool\u2009\u00bb \u00e0 l\u2019ICRP 2021+1.<\/p>\n

Les rayonnements ionisants peuvent introduire dans les cellules humaines normales des mutations susceptibles de les transformer en cellules canc\u00e9reuses au fil du temps. Les diff\u00e9rents types de rayonnements ont un potentiel canc\u00e9rig\u00e8ne diff\u00e9rent, mais la mani\u00e8re dont ils affectent l\u2019int\u00e9grit\u00e9 du g\u00e9nome humain normal n\u2019a pas encore \u00e9t\u00e9 caract\u00e9ris\u00e9e. L\u2019\u00e9quipe de recherche de Felix tente d\u2019\u00e9tudier et de caract\u00e9riser le type et la dose de rayonnement et leur impact sur le g\u00e9nome humain.<\/p>\n<\/div>\n<\/div>\n

 <\/p>\n

Introduction<\/h2>\n

It is well known that ionizing radiation (IR) \u2014 radiation with enough energy to ionize an atom \u2014 can introduce mutations in normal human cells that may transform them into cancer cells over time. It has also been established that different types of radiation have different carcinogenic potentials, though how this affects the integrity of the normal human genome has not yet been characterized. In our research program, we are trying to find a way to investigate and characterize radiation type and dose and their impact on the genomes of radiation-exposed human cells.<\/p>\n

In 2016, Behjati et al. reported differences in the mutation patterns of tumours that could differentiate radiation-associated tumours from radiation-naive tumours [1]<\/a>. Motivated by the Behjati et al. findings, we set out to investigate whether similar mutational information could be used to learn about radiation action in normal human cells. Specifically, we wanted to find out if, by analyzing the mutations induced in vitro<\/em> in a radiation-exposed group of human cells, we could discern the initiating radiation type and dose. We focused our analyses on two types of radiation: neutrons and photons.<\/p>\n

Given that neutrons ionize matter more densely than photons due to their high linear energy transfer (LET), we hypothesized that the mutational patterns of neutrons in exposed cells would be different from those of photons. To test our hypothesis, we investigated mutational effects by single-cell whole genome sequencing (ScWGS) a reference normal human cell line that was exposed to radiation [2]<\/a>.<\/p>\n

 <\/p>\n

Methods<\/h2>\n

This research was carried out in two parts: (1) experimental measurements using human cells, and (2) Monte Carlo simulations of single-cell irradiations to supplement the experimental work.<\/p>\n

Experimental measurements<\/h3>\n

We designed a simple experiment to perform our study as shown in Figure 1. Our plan was to expose human B-lymphoblastoid cells to equal doses of photons and neutrons separately and then compare the induced mutations to evaluate the differences.<\/p>\n

\"\"<\/p>\n

Figure 1:<\/strong> Pictorial representation of our experiment design.<\/p>\n

As a first step, we irradiated our cell samples with various doses of 6 MV photons from a medical linear accelerator. Irradiated cells were incubated for 24 hours to allow one DNA damage repair cycle, and subsequently subjected to ScWGS. Large genomic deletion alterations induced by radiation in the genomes of individual cells were identified and quantified.<\/p>\n

Monte Carlo simulations<\/h3>\n

Our group has developed a single-cell nuclear DNA model (shown in Figure 2) [3]<\/a> using TOPAS-nBio, a Monte Carlo simulation toolkit [4]<\/a>. To complement our experimental work in part 1, we used this model to computationally examine radiation-induced DNA damage that could potentially lead to the types of large genomic deletion mutations that we can identify in our experimental work.<\/p>\n

This DNA model was irradiated with the same doses of radiation (photon or neutron) as in our experiment in part 1. The number of double-strand break (DSB) cluster damages introduced by radiation was measured from the simulations. A DSB cluster refers to a group of DNA damages, within 40 base pairs distance of each other, with a double-strand break also being a part of the group.<\/p>\n

\"\"<\/p>\n

Figure 2:<\/strong> Single-cell nuclear DNA model that we use for Monte Carlo simulations in the TOPAS-nBio simulation toolkit.<\/p>\n

 <\/p>\n

Results<\/h2>\n

In Figure 3, the mean number of large deletion mutations observed in individual cells exposed to 6 MV photons in our experiment is qualitatively compared to the number of DSB clusters measured in cells, as obtained from photon and neutron irradiation simulations. These preliminary experimental results show good agreement with the simulations. Although experimental exposure to neutrons has not yet been undertaken, simulation results confirm our hypothesis that we should expect to see distinct differences between photon- and neutron-induced mutations, at least in terms of dose dependency.<\/p>\n

\"\"<\/p>\n

Figure 3:<\/strong> Qualitative comparison of our results from both experiment and simulation of single-cell irradiations with photons and neutrons. To date, the experiment has only been performed with 6 MV photons.<\/p>\n

 <\/p>\n

Conclusion<\/h2>\n

Our preliminary results suggest that ScWGS can be used to quantify the dose dependence of radiation-induced mutations for ionizing photons and neutrons. Our experimental data showed good agreement with our Monte Carlo results, which is promising. However, we have yet to validate our initial findings with repeated experiments. If confirmed, we believe our strategy of examining radiation-induced mutations with single-cell whole-genome sequencing will open up new avenues for radiation biodosimetry.<\/p>\n

 <\/p>\n

References<\/h2>\n

[1] <\/a>Behjati, S., Gundem, G., Wedge, D. C., Roberts, N. D., Tarpey, P. S., Cooke, S. L., <\/span>et al.\u00a0(2016). Mutational signatures of ionizing radiation in second malignancies.\u00a0<\/span>Nature communications<\/span><\/i>,\u00a07<\/i>, 12605. https:\/\/doi.org\/10.1038\/ncomms12605<\/a><\/span><\/p>\n

[2]<\/a> Li, R., Couturier, C., Savage, P., Monlong, J., Bourque, G., Petrecca, K., Park, M., & Ragoussis, J. (2018). Abstract 2177: Sensitive single cell copy number profiling using a novel microfluidic droplet based platform. Cancer Research, 78<\/em>(13_Supplement), 2177\u20132177. https:\/\/doi.org\/10.1158\/1538-7445.am2018-2177<\/a><\/p>\n

[3]<\/a> Montgomery, L., Lund, C. M., Landry, A., & Kildea, J. (2021). Towards the characterization of neutron carcinogenesis through direct action simulations of clustered DNA damage. Physics in Medicine & Biology, 66<\/em>(20), 205011. https:\/\/doi.org\/10.1088\/1361-6560\/ac2998<\/a><\/p>\n

[4]<\/a> Schuemann, J., McNamara, A. L., Ramos-M\u00e9ndez, J., Perl, J., Held, K. D., Paganetti, H., et al. (2018). Topas-nBio: An extension to the TOPAS Simulation Toolkit for cellular and sub-cellular radiobiology. Radiation Research, 191<\/em>(2), 125. https:\/\/doi.org\/10.1667\/rr15226.1<\/a><\/p>\n

 <\/p>\n

 <\/p>\n

\n

\"\"Felix Mathew<\/h2>\n

Felix est \u00e9tudiant au doctorat au d\u00e9partement de g\u00e9nie biologique et biom\u00e9dical de l\u2019Universit\u00e9 McGill. Dans le cadre de ses recherches, il tente de d\u00e9terminer les signatures mutationnelles des rayonnements \u00e0 haute intensit\u00e9, en particulier les neutrons, par le biais du s\u00e9quen\u00e7age du g\u00e9nome entier d\u2019une seule cellule. Il fait partie de l\u2019\u00e9quipe de recherche sur les effets canc\u00e9rig\u00e8nes induits par les neutrons du Dr John Kildea en tant qu\u2019\u00e9tudiant dipl\u00f4m\u00e9 du programme de physique des rayonnements m\u00e9dicaux.<\/p>\n<\/div>\n

 <\/p>\n

\n

Vous voulez lire d\u2019autres articles comme celui-ci ?<\/h2>\n

Le\u00a0Bulletin<\/em> de l\u2019Association canadienne de la radioprotection (ACRP) est une publication essentielle \u00e0 tout professionnel de la radioprotection du Canada. Son contenu \u00e9ditorial procure aux professionnels de la radioprotection les enseignements, l\u2019information, les conseils et les solutions utiles, tous n\u00e9cessaires pour demeurer \u00e0 l\u2019avant-garde de la profession.<\/p>\n

Abonnez-vous aujourd\u2019hui pour que nous vous envoyions un courriel chaque fois qu\u2019un nouveau num\u00e9ro est mis en ligne. Revisitez souvent le site entre chaque num\u00e9ro pour obtenir les mises \u00e0 jour et consulter de nouveaux articles.<\/p>\n

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Chaque ann\u00e9e, le comit\u00e9 des \u00e9tudiants et des jeunes professionnels de l\u2019ACRP organise le concours de communications \u00e9tudiantes Anthony J. MacKay. La ou le laur\u00e9at re\u00e7oit le troph\u00e9e Anthony J. MacKay, un prix de 250 $ en esp\u00e8ces et son article est publi\u00e9 dans le Bulletin de l\u2019ACRP. L\u2019ann\u00e9e derni\u00e8re, c\u2019est Felix Mathew qui a remport\u00e9 le concours. Felix a pr\u00e9sent\u00e9 son article intitul\u00e9 \u00ab\u2009Single-Cell DNA Sequencing \u2013 A Potential Dosimetric Tool\u2009\u00bb \u00e0 l\u2019ICRP 2021+1.<\/p>\n","protected":false},"author":121,"featured_media":4707,"comment_status":"open","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"_monsterinsights_skip_tracking":false,"footnotes":""},"categories":[29],"tags":[],"class_list":["post-4722","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-articles-fr"],"yoast_head":"\nSingle-Cell DNA Sequencing \u2014 A Potential Dosimetric Tool - CRPA-ACRP Bulletin<\/title>\n<meta name=\"description\" content=\"Le gagnant du concours 2022 de communications \u00e9tudiantes Anthony J. 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