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Article title

Biomarkers used to assess radio- and chemotherapy-induced lymphocyte genome instability in a case of cerebral infarction during relapse of a testicular seminoma


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We report a case of a testicular seminoma patient
with relapse who was irradiated after acute cerebral
infarction induced by cisplatin-based chemotherapy.
Lymphocytic genome instability was studied using an
alkaline comet assay, analysis of structural chromosome
aberrations, and cytokinesis-block micronucleus assay
in blood samples collected before and after PET CT
scanning that preceded radiotherapy, as well as before the
administration of the first and after the administration of
the last fraction of 3D conformal radiation. A challengetest
with hydrogen peroxide (H2O2) was performed on
isolated peripheral blood lymphocytes in order to establish
to what extent earlier therapies had modified the response
of the patient’s DNA to external stimuli with a genotoxic
chemical. Levels of primary DNA damage in lymphocytes
increased after diagnostic exposure, lowered prior to
administration of a conformal 3D radiotherapy, and were
the highest at the end of radiotherapy. Ex vivo exposure to
H2O2 caused additional lymphocyte DNA damage, which
gradually increased 15 and 30 minutes after treatment.
Diagnostic and therapeutic exposure to radiation caused
measurable cytogenetic damage that was subjected
to extensive repair. All of the obtained results point to
increased genomic instability in the patient which should
be taken into account in his future medical surveillance.








Physical description


3 - 4 - 2014
9 - 5 - 2014
9 - 6 - 2014


  • Department of
    Oncology, Clinical Hospital Centre Zagreb, Zagreb, Croatia
  • School of Medicine, University of Zagreb, Zagreb,
  • Department of
    Oncology, Clinical Hospital Centre Zagreb, Zagreb, Croatia
  • Department of
    Oncology, Clinical Hospital Centre Zagreb, Zagreb, Croatia
  • Institute for Medical Research
    and Occupational Health, Mutagenesis Unit, Zagreb, Croatia


  • [1] Schmoll H.J., Jordan K., Huddart R., Pes M.P., Horwich A.,Fizazi K., et al., ESMO Guidelines Working Group, Testicularseminoma: ESMO Clinical Practice Guidelines for diagnosis,treatment and follow-up. Ann. Oncol., 2010, 21(Suppl 5):140-146.[Crossref]
  • [2] Cancer Research UK, Testicular cancer incidence statistics,http://www.cancerresearchuk.org/cancer-info/cancerstats/types/testis/incidence/uk-testicular-cancer-incidencestatistics(2013, accessed 30 June 2013)
  • [3] National Comprehensive Cancer Network, Clinical PracticeGuidelines in Oncology, Testicular Cancer, http://www.nccn.org(2013, accessed 30 June 2013)
  • [4] Jones D.A., Ester E.C., Leavitt D., Sweet R., Konety B., JhaG., et al., Adjuvant radiotherapy for synchronous bilateraltesticular seminoma: a case report and a review of thepertinent literature. Case Reports Urol., 2013, http://dx.doi.org/10.1155/2013/241073.[Crossref]
  • [5] Choo R., Quevedo F., Choo C.S., Blute M., Can radiotherapy bea viable salvage treatment option for the relapsed seminomaconfined to the infra-diaphragm region recurring after primarychemotherapy for bulky stage II seminoma? Can. Urol. Assoc.J., 2010, 4, E137-140.
  • [6] Singh N.P., McCoy M.T., Tice R.R., Schneider L.L., A simpletechnique for quantitation of low levels of DNA damage inindividual cells. Exp. Cell. Res., 1988, 175, 184-191.
  • [7] Gamulin M., Kopjar N., Grgić M., Ramić S., Viculin T., PetkovićM., Garaj-Vrhovac V., Cytogenetic follow-up in testicularseminoma patients exposed to adjuvant radiotherapy. Coll.Antropol., 2010, 34, 455-465.
  • [8] Sigma-Aldrich Inc. Histopaque®-1077 (Procedure No. 1077),http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/General_Information/1/1077.Par.0001.File.tmp/1077.pdf (2010,accessed 30 June 2013)
  • [9] Duke R.C., Cohen J.J., Morphological and biochemical assaysof apoptosis. In: Coligan J.E., Kruisbeal A.M. (Eds.), CurrentProtocols in Immunology, John Willey and Sons, New York, 1992
  • [10] Sham A.S.K., Szeto Y.T., Benzie I.F.F., Tan-Un K.C., Preliminarystudy of DNA damage in peripheral lymphocytes from lungcancer patients and healthy subjects, Turk. J. Med. Sci., 2003,33, 149-154.
  • [11] IAEA, PAHO, WHO, Cytogenetic Dosimetry: Applications inPreparedness for and Response to Radiation Emergencies,http://www-pub.iaea.org/MTCD/publications/PDF/EPR-Biodosimetry%202011_web.pdf (2011, accessed 30 June2013)
  • [12] Fenech M., Morley A.A., Measurement of micronuclei inlymphocytes, Mutat. Res., 1985, 147, 29-36.
  • [13] Fenech M., Chang W.P., Kirsch-Volders M., Holland N., BonassiS., Zeiger E., HUMN project: detailed description of the scoringcriteria for the cytokinesis-block micronucleus assay usingisolated human lymphocyte cultures, Mutat. Res., 2003, 534,65-75.
  • [14] Eastmond D.A., Tucker J.D., Identification of aneuploidyinducingagents using cytokinesis-blocked human lymphocytesand antikinetochore antibody, Environ. Mol. Mutagen., 1989,13, 34-43.[Crossref]
  • [15] Bosl G.J., Patil S., Carboplatin in Clinical Stage I Seminoma: Toomuch and too little at the same time, J. Clin. Oncol., 2011, 29,949-956.[Crossref]
  • [16] Gamulin M., Grgić M., Stage I testicular seminoma: Resultsof adjuvant irradiation, detection of patients with relapseddisease and results of relapse therapy, J. Clin. Oncol. (MeetingAbstracts), 2013, 31(No. 15_suppl), e15509.
  • [17] Classen J., Schmidberger H., Meisner C., Winkler C., Dunst J.,Souchon R., et al., Para-aortic irradiation for stage I testicularseminoma: Results of a prospective study in 675 patients. Atrial of the German testicular cancer study group (GTCSG), Br. J.Cancer, 2004, 90, 2305-2311.
  • [18] Martin J.M., Joon D.L., Ng N., Grace M., Gelderen D.V., LawlorM., et al., Towards individualised radiotherapy for stage Iseminoma, Radiother. Oncol., 2005, 76, 251-256.[Crossref]
  • [19] Gerl A., Vascular toxicity associated with chemotherapy fortesticular cancer, Anticancer Drugs, 1994, 5, 607-614.
  • [20] Doehn C., Büttner H., Fornara P., Jocham D, Fatal basilar arterythrombosis after chemotherapy for testicular cancer, Urol. Int.,2000, 65, 43-45.[Crossref]
  • [21] Weijl N.I., Rutten M.F., Zwinderman A.H., Keizer H.J., NooyM.A., Rosendaal F.R., et al., Thromboembolic events duringchemotherapy for germ cell cancer: A cohort study and reviewof the literature, J. Clin. Oncol. 2000, 18, 2169-2178.
  • [22] Russmann S., Winkler A., Lövblad K.O., Stanga Z., Bassetti C.,Lethal ischemic stroke after cisplatin-based chemotherapy fortesticular carcinoma and cannabis inhalation, Eur. Neurol.,2002, 48, 178-180.[Crossref]
  • [23] Nuver J., Smit A.J., van der Meer J., van den Berg M.P., vander Graaf W.T.A., Meinardi M.T., et al., Acute chemotherapyinducedcardiovascular changes in patients with testicularcancer, J. Clin. Oncol., 2005, 23, 9130-9137.[Crossref]
  • [24] Shahab N., Haider S., Doll D.C., Vascular toxicity of antineoplasticagents, Semin. Oncol., 2006, 33, 121-138.[Crossref]
  • [25] Azak A., Oksüzoğlu B., Deren T., Oneç B.M., Zengin N.,Cerebrovascular accident during cisplatin-based combinationchemotherapy of testicular germ cell tumor: an unusual casereport, Anti-Cancer Drugs, 2008, 19, 97-98.
  • [26] Batra R., Davies J.N., Wheatley D., Extensive arterial and venousthrombo-embolism with chemotherapy for testicular cancer:a case report, Cases J., 2009, http://www.casesjournal.com/content/pdf/1757-1626-2-9082.pdf.
  • [27] Meattini I., Scotti V., Pescini F., Livi L., Sulprizio S., Palumbo V.,et al., Ischemic stroke during cisplatin-based chemotherapyfor testicular germ cell tumor: case report and review of theliterature, J. Chemother., 2010, 22, 134-136.
  • [28] Moore R.A., Adel N., Riedel E., Bhutani M., Feldman D.R.,Tabbara N.E., et al, High incidence of thromboembolic eventsin patients treated with cisplatin-based chemotherapy: a largeretrospective analysis, J. Clin. Oncol., 2011,; 29, 3466-3473.[Crossref]
  • [29] Khadjooi K., Adab N., Kenton A., Acute stroke secondary tocarotid artery dissection in a patient with germ cell tumour: didcisplatin play a role?, Onkologie, 2013, 36, 46-48.[Crossref]
  • [30] Dietrich J., Marienhagen J., Schalke B., Bogdahn U.,Schlachetzki F., Vascular neurotoxicity following chemotherapywith cisplatin, ifosfamide, and etoposide, Ann. Pharmacother.,2004, 38, 242-246.
  • [31] Abouassaly R., Fossa S.D., Giwercman A., Kollmannsberger C.,Motzer R.J., Schmoll H.J., Sternberg C.N., Sequelae of treatmentin long-term survivors of testis cancer, Eur. Urol., 2011, 60,516-526.[Crossref]
  • [32] Hall E.J., Wuu C.S., Radiation-induced second cancers: Theimpact of 3D-CRT and IMRT, Int. J. Radiat. Oncol. Biol. Phys.,2003, 56, 83-88.
  • [33] Bokemeyer C., Schmoll H.J., Treatment of testicular cancer andthe development of secondary malignancies, J. Clin. Oncol.,1995, 13, 283-292.
  • [34] Norppa H., Bonassi S., Hansteen I.L., Hagmar L., StrömbergU., Rössner P., et al., Chromosomal aberrations and SCEs asbiomarkers of cancer risk, Mutat. Res., 2006, 600, 37-45.
  • [35] Bonassi S., Znaor A., Ceppi M., Lando C., Chang W.P., HollandN., et al., An increased micronucleus frequency in peripheralblood lymphocytes predicts the risk of cancer in humans,Carcinogenesis, 2007, 28, 625-631.
  • [36] Boffetta P., van der Hel O., Norppa H., Fabianova E., Fucic A.,Gundy S., et al., Chromosomal aberrations and cancer risk:results of a cohort study from Central Europe, Am. J. Epidemiol.,2007, 165, 36-43.
  • [37] Murgia E., Ballardin M., Bonassi S., Rossi A.M., Barale R.,Validation of micronuclei frequency in peripheral bloodlymphocytes as early cancer risk biomarker in a nestedcase-control study, Mutat. Res., 2008, 639, 27-34.
  • [38] Bonassi S., El-Zein R., Bolognesi C., Fenech M., Micronucleifrequency in peripheral blood lymphocytes and cancer risk:evidence from human studies, Mutagenesis, 2011, 26, 93-100.[Crossref]
  • [39] Mohrenweiser H.W., Jones I.M., Variation in DNA repair is afactor in cancer susceptibility: a paradigm for the promises and perils of individual and population risk estimation?, Mutat.Res., 1998, 400, 15-24.
  • [40] Jagetia G.C., Jayakrishnan A., Fernandes D., Vidyasagar M.S.,Evaluation of micronuclei frequency in the cultured peripheralblood lymphocytes of cancer patients before and afterradiation treatment, Mutat. Res., 2001, 491, 9-16.
  • [41] Faust F., Kassie F., Knasmüller S., Boedecker R.H., Mann M.,Mersch-Sundermann V., The use of the alkaline comet assaywith lymphocytes in human biomonitoring studies, Mutat. Res.,2004, 566, 209-229.
  • [42] Moller P, The alkaline Comet assay: towards validation inbiomonitoring of DNA damaging exposures, Basic. Clin.Pharmacol. Toxicol., 2006, 98, 336-345.[Crossref]
  • [43] Wasson G.R., McKelvey-Martin V.J., Downes C.S., The use ofthe comet assay in the study of human nutrition and cancer,Mutagenesis, 2008, 23, 153-162.[Crossref]
  • [44] McKenna D.J., McKeown S.R., McKelvey-Martin V.J., Potentialuse of the comet assay in the clinical management of cancer,Mutagenesis, 2008, 23,183-90.[Crossref]
  • [45] Baciuchka-Palmaro M., Orsière T., Duffaud F., Sari-MinodierI., Pompili J., Bellon L., et al., Acentromeric micronuclei areincreased in peripheral blood lymphocytes of untreated cancerpatients, Mutat. Res., 2002, 520, 189-198.
  • [46] M’Kacher R., Girinsky T., Koscielny S., Dossou J., Violot D.,Béron-Gaillard N., et al., Baseline and treatment-inducedchromosomal abnormalities in peripheral blood lymphocytes ofHodgkin’s lymphoma patients, Int. J. Radiat. Oncol. Biol. Phys.,2003, 57, 321-326.[Crossref]
  • [47] Palyvoda O., Polańska J., Wygoda A., Rzeszowska-Wolny J., DNAdamage and repair in lymphocytes of normal individuals andcancer patients: studies by the comet assay and micronucleustests, Acta Biochim. Pol., 2003, 50, 181-190.
  • [48] Mothersill C., Seymour C.B., Mechanisms and implicationsof genomic instability and other delayed effects of ionizingradiation exposure, Mutagenesis, 1998, 13, 421-426.[Crossref]
  • [49] Hendry J.H., Genomic instability: potential contributions totumour and normal tissue response, and second tumours, afterradiotherapy, Radiother. Oncol., 2001, 59, 117-126.[Crossref]
  • [50] Kopjar N., Želježić D., Garaj-Vrhovac V., Evaluation of DNAdamage in the white blood cells of healthy human volunteersusing the alkaline comet assay and the chromosome aberrationtest, Acta Biochim. Pol., 2006, 53, 321-336.
  • [51] Garaj-Vrhovac V., Đurinec M., Kopjar N., Oreščanin V., A surveyon the cytogenetic status of the Croatian general population byuse of the cytokinesis-block micronucleus assay, Mutat. Res.,2008, 649, 91-100.
  • [52] Kopjar N., Kašuba V., Milić M., Rozgaj R., Želježić D., Gajski G.,et al., Normalne i granične vrijednosti mikronukleus-testa nalimfocitima periferne krvi ispitanika opće populacije RepublikeHrvatske, Arh. Hig. Rada Toksikol., 2010, 61, 219-234.
  • [53] Gamulin M., Katić J., Milić M., Grgić M., Fučić A., Long-termfollow-up study of genome damage elimination in patientswith testicular seminoma exposed to ionising radiation duringradiotherapy. Arh. Hig. Rada Toksikol., 2011, 62, 51-55.
  • [54] Gamulin M., Grgić M., Ramić S., Garaj-Vrhovac V., KopjarN., Rani učinci radioterapije na razini oštećenja genoma ubolesnika liječenih od raka prostate i seminoma testis / Earlyeffects of radiotherapy on genome damage in patients withprostatic cancer and testicular seminoma, In: Knežević Ž.,Majer M., Krajcar Bronić I. (Eds.), Proceedings of the NinthSymposium of the Croatian Radiation Protection Association(10-12 April 2013, Krk, Croatia), Croatian Radiation ProtectionAssociation Zagreb, 2013, 327-333.
  • [55] Wei Z., Lifen J., Jiliang H., Jianlin L., Baohong W., HongpingD., Detecting DNA repair capacity of peripheral lymphocytesfrom cancer patients with UVC challenge test and bleomycinchallenge test, Mutagenesis, 2005, 20, 271-277.[Crossref]
  • [56] Harrison L., Malyarchuk S., Can DNA repair cause enhanced cellkilling following treatment with ionizing radiation?, Pathophysiology,2002, 8, 149-159.[Crossref]
  • [57] Migliore L., Guidotti P., Favre C., Nardi M., Sessa M.R., BrunoriE., Micronuclei in lymphocytes of young patients underantileukemic therapy, Mutat. Res., 1991, 263, 243-248.
  • [58] Tates A.D., van Dam F.J., Natarajan A.T., Zwinderman A.H.,Osanto S., Frequencies of HPRT mutants and micronucleiin lymphocytes of cancer patients under chemotherapy: aprospective study. Mutat. Res., 1994, 307, 293-306.
  • [59] Carbonell E., Demopoulos N.A., Stefanou G., Psaraki K.,Parry K.M., Marcos R., Cytogenetic analysis in peripherallymphocytes of cancer patients treated with cytostatic drugs:results from an EC Collaborative Study, Anticancer Drugs, 1996,7, 514-519.
  • [60] Elsendoorn, T.J., Weijl N.I., Mithoe S., Zwinderman A.H.,Van Dam F., De Zwart F.A., et al., Chemotherapy-inducedchromosomal damage in peripheral blood lymphocytes ofcancer patients supplemented with antioxidants or placebo,Mutat. Res. 2001, 498, 145-158.
  • [61] Kopjar N., Garaj-Vrhovac V., Milas I., Assessment ofchemotherapy-induced DNA damage in peripheral bloodleukocytes of cancer patients using the alkaline comet assay,Teratogen. Carcinogen. Mutagen., 2002, 22, 13-30.[Crossref]
  • [62] Kopjar N., Garaj-Vrhovac V., Milas I., Acute cytogenetic effectsof antineoplastic drugs on peripheral blood lymphocytes incancer patients: chromosome aberrations and micronuclei,Tumori, 2002, 88, 300-312.
  • [63] Kopjar N., Milas I., Garaj-Vrhovac V., Gamulin M., Cytogeneticoutcomes of adjuvant chemotherapy in non-target cells ofbreast cancer patients, Hum. Exp. Toxicol. 2007, 26, 391-399.[Crossref]
  • [64] Liu X., Zhao J., Zheng R., Protection against damaged DNA insingle cell by polyphenols, Die Pharmazie, 2002, 57, 852-854.
  • [65] Liu X., Zhao J., Zheng R., DNA damage of tumor-associatedlymphocytes and total antioxidant capacity in cancerouspatients, Mutat. Res., 2003, 539, 1-8.
  • [66] Slupphaug G., Kavli B., Krokan H.E., The interacting pathwaysfor prevention and repair of oxidative DNA damage, Mutat.Res., 2003, 531, 231-251.
  • [67] Anand S.S., Singh H., Dash A.K., Clinical applications of PETand PET-CT, Med. J. Arm. For. India, 2009, 65, 353-358.
  • [68] Huang B., Law M.W., Khong P.L., Whole-body PET/CT scanning:estimation of radiation dose and cancer risk, Radiology, 2009,251,166-174.
  • [69] Téoule R., Radiation-induced DNA damage and its repair, Int. J.Radiat. Biol. Relat. Stud. Phys. Chem. Med., 1987, 51, 573-89.[Crossref]
  • [70] Li L., Story M., Legerski R.J., Cellular responses to ionizingradiation damage, Int. J. Radiat. Oncol. Biol. Phys., 2001, 49,1157-1162.[Crossref]
  • [71] McMillan T.J., Tobi S., Mateos S., Lemon C., The use of DNAdouble-strand break quantification in radiotherapy, Int. J.Radiat. Oncol. Biol. Phys. 2001, 49, 373-377.[Crossref]
  • [72] Weidner Maluf S., Monitoring DNA damage following radiationexposure using cytokinesis-block micronucleus method andalkaline single-cell gel electrophoresis, Clin. Chim. Acta, 2004,347, 15-24.
  • [73] Jianlin L., Jiliang H., Lifen J., Wei Z., Baohong W., HongpingD., Measuring the genetic damage in cancer patients duringradiotherapy with three genetic end-points, Mutagenesis,2004, 19, 457-464.[Crossref]
  • [74] Cheong N., Zeng Z.C., Wang Y., Iliakis G., Evidence for factrosmodulating radiation-induced G2-delay: potential applicationas radioprotectors, Phys. Med., 2001, Suppl 1, 205-209.
  • [75] Tice R.R., Agurell E., Anderson D., Burlinson B., Hartmann A.,Kobayashi H., et al., Single cell gel/comet assay: guidelinesfor in vitro and in vivo genetic toxicology testing, Environ. Mol.Mutagen., 2000, 35, 206-221.[Crossref]
  • [76] Leprat F., Alapetite C., Rosselli F., Ridet A., Schlumberger M.,Sarasin A., et al., Impaired DNA repair as assessed by the“comet” assay in patients with thyroid tumors after a historyof radiation therapy: a preliminary study, Int. J. Radiat. Oncol.Biol. Phys., 1998, 40, 1019-1026.[Crossref]
  • [77] Müller W.U., Bauch T., Stüben G., Sack H., Streffer C., Radiationsensitivity of lymphocytes from healthy individuals and cancerpatients as measured by the comet assay, Radiat. Environ.Biophys., 2001, 40, 83-89.

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