Bone marrow somatic mutation after genotoxic cancer therapy

We report here some results from studies carried out on Thai temporary labourers in Israel. All had come to Israel within the past year and they were young (20-35), healthy, and free of STD. All were HIV seronegative. They had very high levels of serum IgE and IgG and their TNF systems were clearly activated, as seen by the high levels of serum soluble TNF receptors type II (p75) and by the high level of surface membrane expression of these receptors on lymphocytes (table). Serum levels of interferon-’Y were low and not different from those observed in controls (not

shown). immune activation in this group of The pattern has some of the characteristics of activation of type 2 T-helper cells (Th2)-namely, high IgE and IgG and normal interferon-y-plus an activated TNF system which does not clearly belong to either Thl-type or Th2-type activation. We suggest that this pattern makes these individuals more susceptible to HIV infection once exposed. Observations by Maggi and colleagues3 support this nor n, which also fits in with the ideas of Clerici and Shearer4on the role ofThl/Th2 activation in HIV infection.

These

findings indicate

healthy, HIV-seronegative Thais.

Alexander Kalinkovich, Shlomo Maayan, Ziva Weisman, Nurit Harpaz, Zvi Bentwich R Ben-Ari Institute of Clinical Immunology, Kaplan Hospital, 76100 Rehovot, Israel; and Department of Clinical Microbiology and Infectious Diseases, Hebrew University Medical School, Hadassah Hospital, Jerusalem

1 Mastro TD, Satten GA, Nopkesorn T, Sangkharomya S, Longini IM Jr. Probability of female-to-male transmission of HIV-1 in Thailand. Lancet 1994; 343: 204-07. 2 Bentwich Z, Wainberg A. Host immunity and the course of HIV infection. Isr J Med Sci 1993; 29 (suppl): 34-36. 3 Maggi E, Mazzetti M, Ravina A, et al. HIV does not induce a TH1/TH2 switch but favors the development of, and preferentially replicates in, CD4+ T cell clones producing TH2-type cytokines. 3rd international conference on cytokines: basic principles and practical applications (Florence, March, 1994): 82. 4 Clenci M, Shearer GM. A TH1→TH2 switch is a critical step in the etiology of HIV infection. Immunol Today 1993; 14: 107-11.

Bone

marrow

genotoxic

somatic mutation after

cancer

after localised bone marrow exposure contrast with our observations following more moderate and homogeneous whole-body radiation exposures-such as those received by atomic bomb survivors and victims of the caesium-137 accident in Brazil and the nuclear reactor incident at Chernobyl-where the GPA assay yielded significant dosedependent increases in variant cell frequencies (Vf) in samples obtained years to decades after the exposures. 1,2 However, study of a small number of patients with moderate partit body exposure from pelvic implants for benign gynaec.. 19ical disease revealed a non-significant, but persisten. increase in GPA Vf.’ By conl st with their results with radiotherapy patients, Mott et al ’emonstrated a significant induction of GPA Vf by mutagenic chemotherapy. These data are also consistent with our studies in adults-specifically, breast cancer patients treated with adjuvant chemotherapy that included cyclophosphamide and doxorubicin3 and germ cell tumour patients who received cisplatin-based chemotherapy.4 We

evidence of post-therapy genetic damage beyond one red-blood-cell lifetime in the cyclophosphamide/doxorubicin patients; however, as with the patients reported by Mott et al, our data for the cisplatin-treated patients provide evidence for significant persistence in samples obtained up to seven years post-therapy. These three studies demonstrate that long-term persistence of chemical mutagen exposure, detectable by the GPA assay, will depend critically on the mode of action of particular agents and their differential mutagenic effects on committed erythroid progenitors versus long-lived pluripotent stem cells. We have previously discussed the potential prognostic value of the GPA assay, and of other established biomarkers, for survival in childhood cancer.’ We believe that the risk of secondary cancers associated with genotoxic therapy for a primary malignancy will depend on the magnitude of the initial mutagenic response and the level of persistent effects. Thus both will need to be evaluated for a given exposure before either the risk in groups of patients treated with a defined regimen or the prospective significance of assay results in individual patients can be estimated. The GPA assay directly measures in-vivo somatic mutation and segregation occurring by any of several saw no

therapy

SIR-The report by Mott and

colleagues (April 2, p 828) of A somatic mutation in paediatric cancer glycophorin (GPA) patients treated with chemotherapy and/or radiotherapy is an important contribution to our understanding of the induction and persistence of chemotherapy-induced in-vivo somatic mutation and adds additional data on the lack of response of the assay to localised high-dose radiotherapy. These observations confirm the capability of the GPA assay as a cumulative biodosimeter of genotoxic exposure and point to the utility and the limitations of this technique as a clinical biomonitor of genotoxic therapy. Our experience with the GPA assay in adult patients after genotoxic medical exposures is detailed in the table (some of these data have previously appeared in summary form only). The lack of response of the GPA assay in the paediatric radiotherapy patients reported on by Mott et al confirms our observations in adults treated with conventional radiotherapy.’ These observations are best explained by an essentially total ablation of bone marrow stem cells (the ultimate progenitors of the GPA variant erythrocytes detected in the peripheral circulation) by the localised highdose radiotherapy with negligible exposure to surrounding marrow. This conclusion is also consistent with the observation of low relative risk of malignancies, primary and secondary, associated with such treatments. Mott’s results

cancers of following types: adenocarcinoma, bone metastases, brain, breast, colon, germ cell, head and neck, Hodgkin’s and non-Hodgkin’s lymphoma, malignant melanoma, metastatic renal carcinoma, neuroblastoma, ovarian carcinoma,

*Patients had

sarcoma, small cell cancer of liver or lung, squamous cell carcinoma of cervix, thoracic. tpelvic radium for benign gynaecological disorders (treated 1937-49).

include: doxorubicin, aminoglutethimide, bleomycin, cyclophosphamide, dacarbazme, dactinomycin, 4’-deoxydoxorub!C!n, oestradiol, etoposide, fluorouracil, leucovorin, lomustine, mechlorethamine, melphalan, methotrexate, mitomycin C, prednisone, procarbazme, tamoxifen, vinblastine, vincristine.

tAgents

§Not significantly different from control population (p>0’05 by t and U tests). Slgnlflcantly different from both control and pre-therapy populations (p<0’05 by

t and

U tests).

Table:

Frequencies

of GPA variants

following genotoxic medical

exposure

1507

molecular mechanisms implicated in oncogenesis. It has the advantage of integrating environmental and genetic factors when evaluating the potential long-term health effects of medical exposures. Ultimately, application of this and other intermediate biomarkers of carcinogenesis will permit individualised design and modification of the therapeutic regimen and better prognostic assessment in all types of cancer.

Stephen

G

Grant, William L Bigbee

Department of Environmental and Occupational Health, Graduate School of Public Health, University of Pittsburgh, Pittsburgh, PA 152 38, USA

1

2

3

4

Grant SG, Bigbee WL, Langlois RG, Jensen RH. Methe for the detection of mutational and segregational events: relevance to the monitoring of survivors of childhood cancer. In: Green DM, D’Angio GJ, eds. Late effects of treatment for childhood cancer. New York: Wiley-Liss, 1992: 133-50. Jensen RH, Langlois RG, Grant SG, Bigbee WL. Biodosimetry of ionizing radiation in humans using the glycophorin A genotoxicity assay. In: Dewey WC, Edington M, Fry RJM, et al, eds. Radiation research: a twentieth-century perspective, vol II (congress proceedings). New York: Academic Press, 1993: 172-76. Bigbee WL, Wyrobek AJ, Langlois RG, Jensen RH, Everson RB. The effect of chemotherapy on the in vivo frequency of glycophorin A "null" variant erythrocytes. Mutat Res 1990: 240: 165-75. Perera FP, Motzer RJ, Tang D, et al. Multiple biological markers in germ cell tumor patients treated with platinum-based chemotherapy. Cancer Res 1992; 52: 3558-65.

patients who also need anticoagulant therapy. In his accompanying commentary, White makes the point that these patients received a higher level of anticoagulation than in the other trials and in SPAF I. An important omission from the report was the individual prothrombin times and international normalised ratios at the actual time of the haemorrhage. Furthermore, how many patients were actually taking warfarin at the time of the intracerebral bleeds? It would also be of interest to establish the circumstances of the haemorrhages. For instance, were these related to trauma? In SPAF I there were 4 subdural haemorrhages, which we presume were due to trauma. Michael D

1

2

3

4

5

Stroke Prevention in Atrial Fibrillation II Study SiR-The SPAF II investigators (March 19, p 687) present their results in the setting of five placebo-controlled trials including their own, and one secondary prevention trial, convincingly demonstrating that warfarin is better than placebo and better than aspirin in reducing stroke in patients with atrial fibrillation (refs 1-5, and the EAFT trial, Nov 20, p 1255). Why then is there a discrepancy between their report and previous publications? The conclusion from SPAF II is that in patients under age 75, the event rate on aspirin is low enough to warrant its use rather than warfarin; in patients over 75, the intracerebral bleed rate cancels out any beneficial effect. Both these conclusions need clarification. The under 75 group seems to consist of patients who were at low risk. Patients were re-randomized from SPAF I-ie, survivors of the placebo group and responders in the treatment group would be assigned, at least in the shortterm, to a low-risk category. This assumption is supported by the low event rate in both the warfarin and aspirin groups, even in comparison with SPAF I in which the event rates (per year) were 3-6% and 2-3%, compared with 1-9% and 1-3% for warfarin and aspirin, respectively. In addition, the placebo-controlled trials showed early benefit for warfarin, and in SPAF I for patients with atrial fibrillation, longer follow-up is needed to demonstrate significant differences between warfarin and aspirin. Furthermore, many patients were not on warfarin at the time of their events, and the non-treatment differences more closely resemble those of the other trials. For warfarin-treated patients over 75, an intracerebral bleed rate of 1-8% per year was found in SPAF II as opposed to an estimated mean rate of 0-3% for all ages in the other trials . Patients with blood pressures of 180/100 mm Hg were allowed into the study, and although the mean blood pressures were well within the normal range, the study does not exclude the possibility that poor control of blood pressure, especially in the elderly, may have been a contributing factor. An important finding of this study may be that even tighter blood control is necessary in elderly 1508

Ezekowitz, Kenneth E James

Yale University, Yale University School of Medicine, Section of Cardiovascular Medicine, PO Box 20817, New

Haven, CT 06520-8017, USA

Petersen P, Boysen G, Godtfredsen J, Andersen ED, Andersen B. Placebo-controlled, randomised trial of warfarin and aspirin for prevention of thromboembolic complications in chronic atrial fibrillation: the Copenhagen AFASAK Study. Lancet 1989; i: 175-79. Stroke Prevention in Atrial Fibrillation Investigators. Stroke Prevention in Atrial Fibrillation Study: final results. Circulation 1991; 84: 527. The Boston Area Anticoagulation Trial for Atrial Fibrillation Investigators. The effect of low-dose warfarin on the risk of stroke in patients with nonrheumatic atrial fibrillation. N Engl J Med 1990; 323: 1505. Connolly SJ, Laupacis A, Gent M, Roberts RS, Cairns JA, Joyner C, for the CAFA Study Coinvestigators. Canadian Atrial Fibrillation Anticoagulation (CAFA) Study. J Am Coil Cardiol 1991; 18: 349. Ezekowitz MD, Bridgers SL, James KE, et al. Warfarin in the prevention of stroke associated with nonrheumatic atrial fibrillation. N Engl J Med 1992; 327: 1406.

Authors’

reply

SiR-Ezekowitz and James ask for clarifications of the results of the SPAF II trial. Although space constraints preclude specific responses to all their concerns, we shall address the relevant points. The low rate of primary events in patients on aspirin under age 75 years was not due to inclusion of rerandomised survivors. Indeed, as stated repeatedly in our report, this subset had a slightly higher rate of stroke than the others and, without these cases, the event rate would be even lower. Patients free of stroke after little more than a year without active therapy are not an atypical low-risk subgroup, as Ezekowitz and James suggest, but in fact make up the majority of participants in recent clinical trials of antithrombotic medication for atrial fibrillation. The SPAF II cohort under 75 years of age consisted of 715 patients entered in fourteen different cities, and are representative of atrial fibrillation patients in these trials. A detailed analysis of bleeding complications and particularly their relation to intensity of anticoagulation and blood pressure will be reported separately. Briefly, we found that advanced age and hypertension at entry were predictive of intracerebral haemorrhage, and anticoagulation intensity was related to major bleeding in elderly patients. The 1-8% per year rate of intracranial bleeding is high in comparison with other trials; this could be attributable either to chance or to inclusion of elderly patients with hypertension or other co-morbidity. Most participants over 75 years entered SPAF II after the results of placebo-controlled trials made warfarin the recommended therapy for patients of all ages with atrial fibrillation. In this respect, our cohort of older patients may better reflect current practice than those who entered clinical trials before the value of warfarin was established. It is noteworthy that the rate of intracranial haemorrhage in aspirin-treated elderly patients in the SPAF II cohort (0-8% per year) was also higher than that for anticoagulated patients in other recent atrial fibrillation trials, suggesting