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Clinical evidence for tumor clonogen regeneration: interpretations of the data
Radiotherapy and Oncology, 22 (1991) 161-166 © 1991 Elsevier Science Publishers B.V. All rights reserved. 0167-8140/91/$03.50
161
RADION 00921
Commentary
Clinical evidence for tumor clonogen regeneration" interpretations of the data Soren M. B e n t z e n 1 a n d H o w a r d D. T h a m e s 2 'Department of Experimental Clinical Oncology, Danish Cancer Society, Aarhus, Denmark and 2Department of Biomathematics, U.T. M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A. (Received 17 July 1991, accepted 13 September 1991)
Key words: Radiation Oncology; Tumor control; Overall time; Sqnamous cell carcinoma
Summary A therapeutic gain is expected from accelerated fractionation in radiotherapy because of reduced times for proliferation of tumor clonogens and the likelihood that the late effects of radiation are unaffected by changes in overall time. While there can be no dispute over the existence of the phenomenon, there are questions about the ways clinical data have been interpreted to adduce the influence of proliferation. Moreover, recent clinical and experimental evidence throw some doubt on the assumption that late effects are independent of overall treatment time. It is concluded that some of the issues remain in doubt, and that rather large-sized phase-III trials may be required to show any benefit from reductions in the overall time, especially if this is accompanied by substantial reductions in the total dose.
Introduction Other factors being equal, small tumors are easier to cure by radiotherapy than large ones, on account of there being fewer clonogens to sterilize. If tumor clonogens proliferate appreciably during treatment, it follows that tumors would be easier to cure if the number of clonogens were limited by shortening the overall treatment time; if in addition dose-limiting normal tissues were unaffected by reductions in the treatment time, a therapeutic gain would result. Clearly tumor clonogens have proliferated prior to treatment, and a wealth of retrospective analyses are appearing in the literature suggesting that this also occurs during treatment, even at an accelerated rate. The purpose of this note is not to argue against an influence of time on the results of radiotherapy, but rather to raise a few points about interpretations and especially about the attempts to quantify these effects. These are combined with recent warnings about the influence on late effects of shortening the treatment time
and a consideration of the size of clinical trials that would be required to show a benefit. The arguments are based on a few simple principles concerning the nature of retrospective analyses of radiotherapy data.
Entanglements of dose and time Almost every analysis of the results of radiotherapy is plagued by the very similar information carried by the total dose and the overall treatment time, when treatment delays occur in a relatively small proportion of the patients. This is especially so when no dose-response relationship can be elicited from the data, since then any supposed connection between curative dose and treatment time may be part artifact: when the dose delivered per day is roughly constant, longer treatment times are associated with higher doses. To illustrate these points consider the hypothetical example of a center where patients have been treated to total doses ranging from 45 to 75 Gy, at 2 Gy per
Address for correspondence: Saren M. Bentzen, Department of Experimental Clinical Oncology, Danish Cancer Society, Narrebrogade 44, DK-8000 Aarhus C., Denmark.
162 fraction given 5 times weekly. Delays occur in about 20~o of the treatments, and last an average of one week. A retrospective analysis is carried out on 1000 patients, and patients are stratified by site and T stage. There is no effect of dose on local control, so that within site and T-stage groups the control rate is constant. However, when treatment times are "binned" (i.e. grouped by short intervals) and the corresponding average total doses (which are necessarily isoeffective) are plotted against average times for the bins, a steeply increasing relationship is evident, with "slope" of, say, 0.6 Gy/day. This is somewhat muddled by the patients with treatment delays, but not enough to obscure the "main point". It is happily concluded that tumor clonogen proliferation has been demonstrated through the steep time dependence of the "isoeffect" doses. While the graphs would not be nearly as clean, the same result might occur in an analysis combining the results from multiple institutions, few of which were characterized by a significant dose-response relationship. Some attempt would have to be made to modify the doses to make them "isoeffective" (since control rates vary, more about this below) and other adjustments would be necessary, but the inherent correlation between total dose and overall time would likely strongly influence the appearance of the graph.
Lack of dose-response relationship How can the absence of a statistically significant doseresponse relationship be interpreted? Certainly not by maintaining that dose has no influence on cure (otherwise there would be no need to give a dose of any size). One interpretation of the above hypothetical situation, in line with recent interest in variable radiosensitivity among patients, is that 500 patients were "sensitive" (cured by any dose above 40 Gy) and 500 "resistant" (not cured by any dose below 80 Gy), with random distribution among the doses, sites, T stages, etc., but no clonogen proliferation during treatment. In this situation the average dose would increase steeply with treatment time simply as a result of the practice of dose prescription. Inferences about clonogen proliferation based on this observation would be erroneous. Other types of heterogeneity would also obscure an underlying dose response, e.g. in pretreatment clonogen number [6] (indirectly measured by tumor volume). However, it is likely that patient selection plays the most important role in removing the apparent influence of dose in many clinical series. One example is the practice of "overboosting", where poor responders are given added doses in prolonged times. An illustration is found in the analysis by Bataini et al. [ 1] of primary tumor
control for cancer of the tonsillar region (and the reanalysis of Fowler et al. [ 10]). The data showed a negative dose-response relationship when other factors were uncorrected for, i.e. tumors treated with high doses did worse than tumors treated with lower doses. Another example is when a poor performance status of the patient precludes aggressive short-course therapy; or when patients with large tumors and/or extensive neck disease are treated with larger field sizes, which again may lead to reduced acute tolerance and a consequent treatment break to allow the acute injury to heal. As an example, Pajak et al. [ 13] presented data on the clinical characteristics of 53 patients included in R T O G head and neck radiotherapy trials with major deviations from the planned treatment (of which 44 had a more than 14 days prolongation of treatment time, 74 ~ of these were prolonged because of treatment reactions). These 53 patients tended to have a higher percentage of unfavorable prognostic characteristics like T4, N3, low performance score, high-risk primary sites compared with the other patients in the trials. All of the above-stated effects tend to create a correlation between poor prognosis on one hand and prolonged treatment and high total dose on the other. This of course is not related to repopulation; natural history may thus overshadow the effect of variations in radiotherapy regimen. Still in any of these cases dose seems to have no (or even a negative) influence on control, but prolonged times are associated with treatment failures. Other reasons for lack of dose response include the narrow ranges of total doses and times that are typical of many centers, on account of standardization of treatment, and confounding factors like concurrent disease conditions (e.g. hypertension, anemia) and inadequate staging of disease. Or quite simply that the size of a patient population under study is too small to detect a statistically significant dose-response relationship. Whatever the reason for lack of dose response, a point should be made about its influence on interpretations of the effect of treatment time. Put simply, what can it mean to say that proliferation adversely affects the outcome of treatment if it cannot be shown that dose has any effect? After all, the presumed result of tumor clonogen proliferation is to increase the dose required for tumor sterilization. If the probability of tumor control is independent of dose, how can it depend on overall time? In fact it is likely that local control depends on both dose and time; we merely wish to point out the contradictions inherent in interpretations of analyses where dose is not statistically significant. We now consider some recently published arguments in favor of an important role for tumor clonogen proliferation, in light of the above principles:
163 i
Analyses of the time factor in institutional data The cleanest demonstration of the treatment-time effect stems from the analysis of single-institutional series of patients treated with conventional vs. split-course radiotherapy. Here the effect of repopulation after the onset of treatment (and during the split) is quantified as the dose needed to compensate for each extra day of treatment, Dprolif. In an analysis of data from the University of Florida [ 8 ] this was found to be 1 Gy/day or more depending on the subsite within the head and neck, but with large confidence limits. The results of an analysis of treatments of carcinoma of the oropharynx in Aarhus gave the estimate of Dprolif at 0.68 Gy/day, with 95 ~o confidence interval (0.05,1.3) Gy/day [4]. Withers and colleagues [20] surveyed the literature data on head and neck tumors, and presented the resuits in terms of TCDso (that is, the dose needed to control 50~o of the tumors) as a function of treatment time. In doing so they had to overcome two problems. First, dose fractions of 2 Gy were used in many but not all studies, and so total doses given in the papers had to be converted into biologically equivalent doses given in 2 Gy dose fractions. This was accomplished using the linear-quadratic model with e//~ = 25 Gy. This procedure is not sensitive to the value of ~//~ chosen when it is this high, and the conclusions of the analysis would be similarly unaffected. The second problem was that few of the papers surveyed presented results characterized by a dose-response relationship, and the procedures adopted to circumvent this difficulty likely affected the conclusions, as follows. Generally speaking, the surveyed papers set out only the observed frequency of local control after a given treatment schedule (for example, a certain center may have had 35 ~o local control at 2 years o f T 3 - T 4 oropharynx tumors, and 80~o control of T]-T2 supraglottic tumors). Consequently the biologically equivalent dose in 2-Gy fractions that controlled, say, 35 ~o of T 3 - T 4 oropharynx tumors, had to be converted into an estimate of the dose to control 50~o of these tumors, i.e. the TCDso. This conversion requires an estimate of the steepness of the dose-response curve for the tumors and since for many of the radiotherapy centers and tumors surveyed by Withers et al. [20] there was no statistically significant dose response, an approximate procedure had to be adopted. They assumed that the steepness was characterized by a D O of 5 Gy for all the tumors surveyed, converted all doses to estimates of TCDso, and plotted the results vs. treatment time (reproduced in Fig. 1). There are two ways to view this choice of Do. On the one hand, it is reasonable to assume that the change of
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Fig. l. TCDso as a function of overall treatment time for squamous cell carcinomas of the head and neck. Open symbols are TCDso'S corrected for control rate and dose-fraction size, filled circles are total doses reported in the literature. Stippled line is the rate of increase in TCDso with time assuming a 2 month clonogen doubling rate, solid line is the estimated rate of increase from multivariate analyses of data not included in this figure (from Withers et al. [20]).
control probability with dose for each individual patient is quite steep, and on theoretical grounds a typical mammalian cell-survival curve would suggest that the slope of the survival response to multiple 2 Gy fractions is about 5 Gy. This would describe the expected steepness of the dose response for a patient drawn at random from the population of those treated, and is a consideration that should guide radiotherapists when a change in dose is contemplated for a given patient. Such a steep dose response would not hold, however, for the whole population because of wide distributions in initial tumor clonogen number, radiosensitivity, repopulation rate, etc., which cause the observed dose response to be much shallower than predicted by Do = 5 Gy. Because of this the predicted TCDso would disagree with the most likely institutional experience, based on an average slope for all the patients. Thus a second view is that calculated changes in dose to account for different control probabilities should reflect the actually observed slope for the institutional series under consideration. If there is no dose response for a given series, then no change in dose may be computed, and the best estimate is that the TCDso is the same as the dose that has been determined for a given level of tumor control. How much does the picture set out in Fig. i change, if more realistic estimates of the slopes for institutional series are used? The number of series for which there are significant dose responses is limited (Table I), and for the reasons set out above there is no significant association between tumor cure and dose for a majority of tumors treated at various centers. To avoid confusion
164 TABLE I Inverse slope of clinical dose-response curve (D.) for head and neck tumors• Site
T-category
Various H & N Larynx Larynx Supraglottic Retromolar trig. ant. faucial pillar Oropharynx Base of tongue Tonsillar fossa Base of tongue
Strat. T3 T2 T2 + T3 T 3 + T4 Direct TI + T2 T3 + T4
D a (Gy)
<2 8.2 10 13 14 18 27 28 47
T3 + T4
[11] [17] [17] [18] [18] [4] [18] [18] [18]
Strat., stratified according to subsite and T category; Direct, T category was corrected for in a multivariate ("direct") analysis.
with Do, which has a radiobiological meaning, we have labeled "Da" the apparent (fitted) slope of the clinically observed curves. The range of Da's is wide, and with the exception of one study, in which atypically steep doseresponse relationships were observed, D~'s range from 8 Gy to indefinitely high values: laryngeal carcinomas have D~'s around 10 Gy, whereas other head and neck cancers are characterized by values from 14 Gy up. To get a feeling for the significance of these slopes, the dose to increase the local control probability from 40 to 60 ~o is 10.5 Gy for O a = 18 Gy, as opposed to 2.9 Gy for D~ = 5 Gy. Most clinicians would probably agree that it takes more than 3 Gy to improve the local control rate from 40 to 6 0 ~ for a specific site and T category of otherwise unselected head and neck tumors• In the present context, the point is that when a D~ of, say, 18 Gy is used to adjust doses to estimated TCDso'S the picture is altered significantly (Fig. 2).
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Treatment time (days) Figure 2: Replot of the data in Fig. 1 using an ct/fl = 25 Gy and an apparent slope of the clinical dose-response curve D a = 18 Gy. The area of each data point is proportional to the number of patients in the corresponding series. More details are given in the text.
It is interesting that the approximately 4-week "lag time" until onset of regeneration that was suggested by Withers et al. on basis of Fig. 1 is missing in Fig. 2. An obvious and important consequence of the existence of such a lag would be that shortening treatment below 3-5 weeks, as in the CHART regimen at the Mount Vernon Hospital [ 15 ], would not pay. The presentation in Fig. 2 suggests that the issue is not clear cut. In fact, we checked for a change in the rate of regeneration during treatment by including a time-squared term in a fit of data similar to that shown in Figs. 1 and 2. A number of analyses were done, choosing values of D a ranging from 5 to 20 Gy, but the time-squared term was not significantly different from zero in any of these fits. Withers et al. found the data to be consistent with a slope of 0.6 Gy/day (derived from an analysis of dosetime scatter plots) in fair agreement with the slope of 0.48 Gy/day estimated here. How to interpret these slopes ? As repopulation may be expected to have occurred on purely biological grounds, it is tempting to assume that these slopes reflect this phenomenon, whereby increases in treatment time must more or less uniformly be offset by about 0.5 Gy/day. However, in view of the lack of dose response in many of the series the data cannot unambiguously be interpreted in these terms. A remarkable observation is that if we do the analysis without any correction at all of the total doses, the slope of the regression line remains exactly the same: 0.48 Gy/day. In other words, this slope could simply be interpreted as a consequence of the practice of dose prescription in the various studies: the relationship between the TCDso and overall treatment time would then reflect the fact that higher total doses tend to be delivered in longer overall times ! But why a slope of 0.5 Gy/day? After all, at 10 Gy/week the slope should have been more than 1 Gy/day, if prolonged treatments simply were given as a number of extra 2 Gy dose fractions. Here the patients with delayed treatments will have some influence. It is also important to recall the profound influence of the Ellis NSD formalism from the late 1960s through the early 1980s. This will have shaped the design of many of the clinical regimens whose results are summarized in Figs. 1 and 2 (and in many institutions also the data now available for dose-time-scattergram analyses), such that the increase in total dose with increasing time (and numbers of fractions) would reflect the predictions of the formula. Indeed, when we apply the N S D formula to a schedule with one fraction per day five days per week (remembering that the N SD formula for tumors did not correct explicitly for treatment time), we obtain dD
D . 0.24
dN
N
165 where D = total dose and N = number of fractions (proportional to time). With D/N = 2 Gy per fraction, the right-hand side is equal to 0.48 Gy per extra fraction, i.e. 0.48 Gy per day. These pitfalls can be avoided to some extent by using multivariate analyses and maximum likelihood techniques. In analyses of data from split-course treatments [4,8], a significant dose response was demonstrated, and thus it was possible to interpret the meaning of "dose/day" to offset repopulation. What is to be avoided is having recourse to comparing total doses to treatment times in the manner outlined in Figs. 1 and 2, after the more rigorous likelihood fitting has failed to produce evidence of a significant influence for dose or time.
Prospects for accelerated fractionation The foregoing suggests that the evidence on which rests the prospect for a therapeutic gain from accelerated fractionation is compromised by either wide confidence limits (estimates of Dproliffrom split-course treatments) or multiple interpretations, some of which have nothing to do with repopulation. Worse still, it is to be remembered that the dose-response relationships observed in the clinic are quite shallow so that the "loss" of 1 Gy/day does not necessarily represent a substantial loss of local control probability. Ten days shortening of the treatment was estimated by Bentzen et al. [4] to results in a change from 60 to 70 ~o in local control probability in a typical patient. To detect a change of this magnitude at a significance level of 5~o (one-sided) and with a power of 90 ~o would require a total of 650 patients in a two-arm randomized trial open for intake for 3 years with an additional 3 years of follow-up. This rate of patient accrual will in most situations require a multicenter trial, with the accompanying problems of standardizing treatment. The only way to reduce the number of patients required would be to identify those with a greater potential advantage from treatment acceleration, and for this measurement of the potential doubling time, Tpot, of tumor clonogens has been suggested. Tpo~ was introduced by Steel [16] as the clonogen doubling time that would be measured if cell loss were ignored, i.e. if both daughter cells remained clonogenic after a mitosis. In practice clonogens are lost by any of a number of mechanisms, including differentiation, death, and metastasis, and the net result is that Tpot will be shorter than the actual clonogen doubling time, T~on. It is of course the latter that is of real importance for radiotherapy. Talon may be expressed in terms of Tpo~, the labelling index or S-phase fraction LI, the S-phase transit time Ts, and the cell-loss factor, CLF:
Llon=
~o,
=----.2"~
l
1-CLF
LI
1-CLF
wherein 2; is approximately 0.6. From this relationship it is seen that a shorter Tclon may result from (1) a reduced Ts, (2) an increased LI, or (3) a reduced CLF. Currently the third possibility is favored [20], that the CLF is reduced after the beginning of treatment. If the CLF goes to the limit of zero then the net clonogen doubling time Tolo, would equal the pretreatment Tpot, provided that LI and Ts did not change inappropriately. The attraction of all this is that a pretreatment parameter that can be measured by a single biopsy [3] should be predictive of the extent of accelerated tumor growth after onset of treatment. On the other hand, if LI increased after the cytotoxic insult or the CLF was reduced without being close to zero, and if any of these possibilities happened to a varying degree in various tumors, then the Tpot before treatment might not be predictive for Trion during treatment. On the positive side, preliminary results indicate an association between a short Tpot and possible benefit from accelerated fractionation [2]. More data and longer follow-up are needed to establish the strength of this correlation and to judge whether an individually measured Tpo~ can be used for selective allocation to accelerated treatments. The potential for therapeutic gain from accelerated treatment requires that late normal-tissue injury not be changed for the worse when overall time is shortened. Almost a dogma of normal-tissue radiobiology in the 1980s, the truth of this statement has now been challenged, perhaps because of the necessity of giving more than one treatment per day. The CHART trial [7] has produced four myelopathies, a warning that 6 h is not sufficient for complete repair of sublethal damage in the spinal cord, or a slow-repair process was compromised by shortening the overall time, or other. Ang et al. (pers. comm., 1991) have supported this experimentally by showing that significant recovery took place in the rat spinal cord between 8 and 24 h. Finally, Turesson and Thames [19] showed that late telangiectasia following postmastectomy irradiation was characterized by a repair half-time in excess of 3 h. With regard to acute reactions it has been recognized that acceleration of treatment would lead to their worsening. The attitude has been that as long as the patient could be brought through this phase these reactions did not constitute a therapeutic problem. The experience of Peracchia and Salti [ 14] was one warning that this is not necessarily true. Further, Bentzen and Overgaard [5] showed that after stratification for biologically equivalent dose, a previous history of moist desquamation was a predisposing factor for the
166 development of late telangiectasia. Although this may be a problem unique with the pathogenesis of telangiectasia, it may serve as a warning that more surprises could show up as the treatment schedules approach the limits of acute tolerance. Where does this leave us with respect to clinical trials of accelerated fractionation? First, although there is strong support for the assertion that tumor control will improve with shorter overall times, what remains to be clarified is the magnitude of the anticipated improvement and the fraction of patients who would benefit significantly. As outlined in the preceding, care should be exercised in the interpretation of retrospective analyses. If a sizable proportion of tumors considered candidates for treatment by accelerated treatment was in fact slow-growing, even a minor reduction of total dose would not be compensated by a shorter overall
time. Although the preliminary experience with in vivo measures of cell kinetic parameters like Tpot allows some optimism, it is too early to say if routine use is justified for the allocation of patients for accelerated fractionation. Second, with regard to optimal schedules, there is in our view no clear demonstration of a lag time of 4 weeks before onset of accelerated proliferation of tumor clonogens. Also, if a fixed number of fractions per week is given, then depending on the value of ~/fl for the tumor it could be advantageous to prolong overall treatment time by reducing the fraction size except in tumors that are both radioresistant and fast proliferating [9]. Finally, the interfraction interval required for complete recovery between multiple treatments per day may prove too long to be practical.
References 1 Bataini, J. P., Asselain, B., Jaulerry, Ch., Brunin, F., Bernier, J., Pontvert, D. and Lave, C. A multivariate primary tumour control analysis in 465 patients treated by radical radiotherapy for cancer of the tonsillar region: clinical and treatment parameters as prognostic factors. Radiother. Oncol. 14: 265-277, 1989. 2 Begg, A. C., Holland, I., Moonen, L., Bartelink, H., Schraub, S., Bontemps, P., Le Fur, R., Bogaert, W. van den, Caspers, R., Glabbeke, M. van and Horiot, J.-C. The predictive value of cell kinetics measurements in a European trial of accelerated fractionation in advanced head and neck tumors: an interim report. Int. J. Radiat. Oncol. Biol. Phys. 19: 1449-1453, 1990. 3 Begg, A. C., McNally, N. J., Shrieve, D. C. and Karcher, H. A method to measure the DNA synthesis and the potential doubling time from a single sample. Cytometry 6: 620-626, 1985. 4 Bentzen, S .M., Johansen, L.V., Overgaard, J. and Thames, H. D. Clinical radiobiology of squamous cell carcinoma of the oropharynx. Int. J. Radiat. Oncol. Biol. Phys. 20: 1197-1206, 1991. 5 Bentzen, S. M. and Overgaard, M. Relationship between early and late normal-tissue injury after postmastectomy radiotherapy. Radiother. Oncol. 20: 159-165, 1991. 6 Bentzen, S. M., Overgaard, J., Thames, H. D., Overgaard, M., Hansen, P.V., Maase, H. v o n d e r and Meder, J. Clinical radiobiology of malignant melanoma. Radiother. Oncol. 16: 169-182, 1989. 7 Dische, S. and Saunders, M. I. Continuous, hyperfractionated, accelerated radiotherapy (CHART): an interim report upon late morbidity. Radiother. Oncol. 16: 67-74, 1989. 8 Dubois, J. B., Broquerie, J.L., Delard, R. and Pourquir, H. Analysis of the results of irradiation in the treatment of tonsillar region carcinomas. Int. J. Radiat. Oncol. Biol. Phys. 9: 1195-1203, 1983. 9 Fowler, J. F. How worthwhile are are short schedules in radiotherapy?: a series of exploratory calculations. Radiother. Oncol. 18: 165-181, 1990. 10 Fowler, J. F., Tanner, M. A., Bataini, J. P., Asselain, B., Bernier, J. and Lave, C. Further analysis of the time factor in squamous cell carcinoma of the tonsillar region. Radiother. Oncol. 19: 237-244, 1990.
11 Maciejewski, B., Withers, H. R., Taylor, J. M. G. and Hliniak, A. Dose fractionation and regeneration in radiotherapy for cancer of the oral cavity and oropharynx: tumor dose response and repopulation. Int. J. Radiat. Oncol. Biol. Phys. 16: 831-843, 1989. 12 Overgaard, J., Hjelm-Hansen, M., Johansen, L.V. and Andersen, A. P. Comparison of conventional and split-course radiotherapy as primary treatment in carcinoma of the larynx. Acta Oncol. 27: 147-152, 1988. 13 Pajak, T. F., Laramore, G.E., Marcial, V.A., Fazekas, J.T., Cooper, J., Rubin, P., Curran, W. J. and Davis, L. W. Elapsed treatment days - - a critical item for radiotherapy quality control review in head and neck trials: RTOG report. Int. J. Radiat. Oncol. Biol. Phys. 20: 13-20, 1991. 14 Peracchia, G. and Salti, C. Radiotherapy with thrice-a-day fractionation in a short overall time: clinical experiences. Int. J. Radiat. Oncol. Biol. Phys. 7: 99-104, 1981. 15 Saunders, M. I. and Dische, S. Radiotherapy employing three fractions in each day over a continuous period of 12 days. Br. J. Radiol. 59: 523-525, 1986. 16 Steel, G. G. Growth Kinetics of Tumours, pp. 67-72.Clarendon Press, Oxford, 1977. 17 Stewart, J. G. and Jackson, A. W. The steepness of the dose response curve both for tumor cure and normal tissue injury. Laryngoscope 85:1107-1111, 1975. 18 Thames, H. D., Peters, L.J., Spanos, W. and Fletcher, G.F. Dose response of squamous cell carcinomas of the upper respiratory and digestive tracts. Br. J. Cancer 41, Suppl. IV: 35-38, 1980. 19 Turesson, I. and Thames, H. D. Repair capacity and kinetics of human skin during fractionated radiotherapy: erythema, desquamation, and telangiectasia after 3 and 5 year's follow-up. Radiother. Oncol. 15: 169-188, 1989. 20 Withers, H. R., Taylor, J. M. G. and Maciejewski, B. The hazard of accelerated tumor clonogen repopulation during radiotherapy. Acta Oncol. 27: 131-146, 1988.
161
RADION 00921
Commentary
Clinical evidence for tumor clonogen regeneration" interpretations of the data Soren M. B e n t z e n 1 a n d H o w a r d D. T h a m e s 2 'Department of Experimental Clinical Oncology, Danish Cancer Society, Aarhus, Denmark and 2Department of Biomathematics, U.T. M.D. Anderson Cancer Center, Houston, TX 77030, U.S.A. (Received 17 July 1991, accepted 13 September 1991)
Key words: Radiation Oncology; Tumor control; Overall time; Sqnamous cell carcinoma
Summary A therapeutic gain is expected from accelerated fractionation in radiotherapy because of reduced times for proliferation of tumor clonogens and the likelihood that the late effects of radiation are unaffected by changes in overall time. While there can be no dispute over the existence of the phenomenon, there are questions about the ways clinical data have been interpreted to adduce the influence of proliferation. Moreover, recent clinical and experimental evidence throw some doubt on the assumption that late effects are independent of overall treatment time. It is concluded that some of the issues remain in doubt, and that rather large-sized phase-III trials may be required to show any benefit from reductions in the overall time, especially if this is accompanied by substantial reductions in the total dose.
Introduction Other factors being equal, small tumors are easier to cure by radiotherapy than large ones, on account of there being fewer clonogens to sterilize. If tumor clonogens proliferate appreciably during treatment, it follows that tumors would be easier to cure if the number of clonogens were limited by shortening the overall treatment time; if in addition dose-limiting normal tissues were unaffected by reductions in the treatment time, a therapeutic gain would result. Clearly tumor clonogens have proliferated prior to treatment, and a wealth of retrospective analyses are appearing in the literature suggesting that this also occurs during treatment, even at an accelerated rate. The purpose of this note is not to argue against an influence of time on the results of radiotherapy, but rather to raise a few points about interpretations and especially about the attempts to quantify these effects. These are combined with recent warnings about the influence on late effects of shortening the treatment time
and a consideration of the size of clinical trials that would be required to show a benefit. The arguments are based on a few simple principles concerning the nature of retrospective analyses of radiotherapy data.
Entanglements of dose and time Almost every analysis of the results of radiotherapy is plagued by the very similar information carried by the total dose and the overall treatment time, when treatment delays occur in a relatively small proportion of the patients. This is especially so when no dose-response relationship can be elicited from the data, since then any supposed connection between curative dose and treatment time may be part artifact: when the dose delivered per day is roughly constant, longer treatment times are associated with higher doses. To illustrate these points consider the hypothetical example of a center where patients have been treated to total doses ranging from 45 to 75 Gy, at 2 Gy per
Address for correspondence: Saren M. Bentzen, Department of Experimental Clinical Oncology, Danish Cancer Society, Narrebrogade 44, DK-8000 Aarhus C., Denmark.
162 fraction given 5 times weekly. Delays occur in about 20~o of the treatments, and last an average of one week. A retrospective analysis is carried out on 1000 patients, and patients are stratified by site and T stage. There is no effect of dose on local control, so that within site and T-stage groups the control rate is constant. However, when treatment times are "binned" (i.e. grouped by short intervals) and the corresponding average total doses (which are necessarily isoeffective) are plotted against average times for the bins, a steeply increasing relationship is evident, with "slope" of, say, 0.6 Gy/day. This is somewhat muddled by the patients with treatment delays, but not enough to obscure the "main point". It is happily concluded that tumor clonogen proliferation has been demonstrated through the steep time dependence of the "isoeffect" doses. While the graphs would not be nearly as clean, the same result might occur in an analysis combining the results from multiple institutions, few of which were characterized by a significant dose-response relationship. Some attempt would have to be made to modify the doses to make them "isoeffective" (since control rates vary, more about this below) and other adjustments would be necessary, but the inherent correlation between total dose and overall time would likely strongly influence the appearance of the graph.
Lack of dose-response relationship How can the absence of a statistically significant doseresponse relationship be interpreted? Certainly not by maintaining that dose has no influence on cure (otherwise there would be no need to give a dose of any size). One interpretation of the above hypothetical situation, in line with recent interest in variable radiosensitivity among patients, is that 500 patients were "sensitive" (cured by any dose above 40 Gy) and 500 "resistant" (not cured by any dose below 80 Gy), with random distribution among the doses, sites, T stages, etc., but no clonogen proliferation during treatment. In this situation the average dose would increase steeply with treatment time simply as a result of the practice of dose prescription. Inferences about clonogen proliferation based on this observation would be erroneous. Other types of heterogeneity would also obscure an underlying dose response, e.g. in pretreatment clonogen number [6] (indirectly measured by tumor volume). However, it is likely that patient selection plays the most important role in removing the apparent influence of dose in many clinical series. One example is the practice of "overboosting", where poor responders are given added doses in prolonged times. An illustration is found in the analysis by Bataini et al. [ 1] of primary tumor
control for cancer of the tonsillar region (and the reanalysis of Fowler et al. [ 10]). The data showed a negative dose-response relationship when other factors were uncorrected for, i.e. tumors treated with high doses did worse than tumors treated with lower doses. Another example is when a poor performance status of the patient precludes aggressive short-course therapy; or when patients with large tumors and/or extensive neck disease are treated with larger field sizes, which again may lead to reduced acute tolerance and a consequent treatment break to allow the acute injury to heal. As an example, Pajak et al. [ 13] presented data on the clinical characteristics of 53 patients included in R T O G head and neck radiotherapy trials with major deviations from the planned treatment (of which 44 had a more than 14 days prolongation of treatment time, 74 ~ of these were prolonged because of treatment reactions). These 53 patients tended to have a higher percentage of unfavorable prognostic characteristics like T4, N3, low performance score, high-risk primary sites compared with the other patients in the trials. All of the above-stated effects tend to create a correlation between poor prognosis on one hand and prolonged treatment and high total dose on the other. This of course is not related to repopulation; natural history may thus overshadow the effect of variations in radiotherapy regimen. Still in any of these cases dose seems to have no (or even a negative) influence on control, but prolonged times are associated with treatment failures. Other reasons for lack of dose response include the narrow ranges of total doses and times that are typical of many centers, on account of standardization of treatment, and confounding factors like concurrent disease conditions (e.g. hypertension, anemia) and inadequate staging of disease. Or quite simply that the size of a patient population under study is too small to detect a statistically significant dose-response relationship. Whatever the reason for lack of dose response, a point should be made about its influence on interpretations of the effect of treatment time. Put simply, what can it mean to say that proliferation adversely affects the outcome of treatment if it cannot be shown that dose has any effect? After all, the presumed result of tumor clonogen proliferation is to increase the dose required for tumor sterilization. If the probability of tumor control is independent of dose, how can it depend on overall time? In fact it is likely that local control depends on both dose and time; we merely wish to point out the contradictions inherent in interpretations of analyses where dose is not statistically significant. We now consider some recently published arguments in favor of an important role for tumor clonogen proliferation, in light of the above principles:
163 i
Analyses of the time factor in institutional data The cleanest demonstration of the treatment-time effect stems from the analysis of single-institutional series of patients treated with conventional vs. split-course radiotherapy. Here the effect of repopulation after the onset of treatment (and during the split) is quantified as the dose needed to compensate for each extra day of treatment, Dprolif. In an analysis of data from the University of Florida [ 8 ] this was found to be 1 Gy/day or more depending on the subsite within the head and neck, but with large confidence limits. The results of an analysis of treatments of carcinoma of the oropharynx in Aarhus gave the estimate of Dprolif at 0.68 Gy/day, with 95 ~o confidence interval (0.05,1.3) Gy/day [4]. Withers and colleagues [20] surveyed the literature data on head and neck tumors, and presented the resuits in terms of TCDso (that is, the dose needed to control 50~o of the tumors) as a function of treatment time. In doing so they had to overcome two problems. First, dose fractions of 2 Gy were used in many but not all studies, and so total doses given in the papers had to be converted into biologically equivalent doses given in 2 Gy dose fractions. This was accomplished using the linear-quadratic model with e//~ = 25 Gy. This procedure is not sensitive to the value of ~//~ chosen when it is this high, and the conclusions of the analysis would be similarly unaffected. The second problem was that few of the papers surveyed presented results characterized by a dose-response relationship, and the procedures adopted to circumvent this difficulty likely affected the conclusions, as follows. Generally speaking, the surveyed papers set out only the observed frequency of local control after a given treatment schedule (for example, a certain center may have had 35 ~o local control at 2 years o f T 3 - T 4 oropharynx tumors, and 80~o control of T]-T2 supraglottic tumors). Consequently the biologically equivalent dose in 2-Gy fractions that controlled, say, 35 ~o of T 3 - T 4 oropharynx tumors, had to be converted into an estimate of the dose to control 50~o of these tumors, i.e. the TCDso. This conversion requires an estimate of the steepness of the dose-response curve for the tumors and since for many of the radiotherapy centers and tumors surveyed by Withers et al. [20] there was no statistically significant dose response, an approximate procedure had to be adopted. They assumed that the steepness was characterized by a D O of 5 Gy for all the tumors surveyed, converted all doses to estimates of TCDso, and plotted the results vs. treatment time (reproduced in Fig. 1). There are two ways to view this choice of Do. On the one hand, it is reasonable to assume that the change of
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Fig. l. TCDso as a function of overall treatment time for squamous cell carcinomas of the head and neck. Open symbols are TCDso'S corrected for control rate and dose-fraction size, filled circles are total doses reported in the literature. Stippled line is the rate of increase in TCDso with time assuming a 2 month clonogen doubling rate, solid line is the estimated rate of increase from multivariate analyses of data not included in this figure (from Withers et al. [20]).
control probability with dose for each individual patient is quite steep, and on theoretical grounds a typical mammalian cell-survival curve would suggest that the slope of the survival response to multiple 2 Gy fractions is about 5 Gy. This would describe the expected steepness of the dose response for a patient drawn at random from the population of those treated, and is a consideration that should guide radiotherapists when a change in dose is contemplated for a given patient. Such a steep dose response would not hold, however, for the whole population because of wide distributions in initial tumor clonogen number, radiosensitivity, repopulation rate, etc., which cause the observed dose response to be much shallower than predicted by Do = 5 Gy. Because of this the predicted TCDso would disagree with the most likely institutional experience, based on an average slope for all the patients. Thus a second view is that calculated changes in dose to account for different control probabilities should reflect the actually observed slope for the institutional series under consideration. If there is no dose response for a given series, then no change in dose may be computed, and the best estimate is that the TCDso is the same as the dose that has been determined for a given level of tumor control. How much does the picture set out in Fig. i change, if more realistic estimates of the slopes for institutional series are used? The number of series for which there are significant dose responses is limited (Table I), and for the reasons set out above there is no significant association between tumor cure and dose for a majority of tumors treated at various centers. To avoid confusion
164 TABLE I Inverse slope of clinical dose-response curve (D.) for head and neck tumors• Site
T-category
Various H & N Larynx Larynx Supraglottic Retromolar trig. ant. faucial pillar Oropharynx Base of tongue Tonsillar fossa Base of tongue
Strat. T3 T2 T2 + T3 T 3 + T4 Direct TI + T2 T3 + T4
D a (Gy)
<2 8.2 10 13 14 18 27 28 47
T3 + T4
[11] [17] [17] [18] [18] [4] [18] [18] [18]
Strat., stratified according to subsite and T category; Direct, T category was corrected for in a multivariate ("direct") analysis.
with Do, which has a radiobiological meaning, we have labeled "Da" the apparent (fitted) slope of the clinically observed curves. The range of Da's is wide, and with the exception of one study, in which atypically steep doseresponse relationships were observed, D~'s range from 8 Gy to indefinitely high values: laryngeal carcinomas have D~'s around 10 Gy, whereas other head and neck cancers are characterized by values from 14 Gy up. To get a feeling for the significance of these slopes, the dose to increase the local control probability from 40 to 60 ~o is 10.5 Gy for O a = 18 Gy, as opposed to 2.9 Gy for D~ = 5 Gy. Most clinicians would probably agree that it takes more than 3 Gy to improve the local control rate from 40 to 6 0 ~ for a specific site and T category of otherwise unselected head and neck tumors• In the present context, the point is that when a D~ of, say, 18 Gy is used to adjust doses to estimated TCDso'S the picture is altered significantly (Fig. 2).
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Treatment time (days) Figure 2: Replot of the data in Fig. 1 using an ct/fl = 25 Gy and an apparent slope of the clinical dose-response curve D a = 18 Gy. The area of each data point is proportional to the number of patients in the corresponding series. More details are given in the text.
It is interesting that the approximately 4-week "lag time" until onset of regeneration that was suggested by Withers et al. on basis of Fig. 1 is missing in Fig. 2. An obvious and important consequence of the existence of such a lag would be that shortening treatment below 3-5 weeks, as in the CHART regimen at the Mount Vernon Hospital [ 15 ], would not pay. The presentation in Fig. 2 suggests that the issue is not clear cut. In fact, we checked for a change in the rate of regeneration during treatment by including a time-squared term in a fit of data similar to that shown in Figs. 1 and 2. A number of analyses were done, choosing values of D a ranging from 5 to 20 Gy, but the time-squared term was not significantly different from zero in any of these fits. Withers et al. found the data to be consistent with a slope of 0.6 Gy/day (derived from an analysis of dosetime scatter plots) in fair agreement with the slope of 0.48 Gy/day estimated here. How to interpret these slopes ? As repopulation may be expected to have occurred on purely biological grounds, it is tempting to assume that these slopes reflect this phenomenon, whereby increases in treatment time must more or less uniformly be offset by about 0.5 Gy/day. However, in view of the lack of dose response in many of the series the data cannot unambiguously be interpreted in these terms. A remarkable observation is that if we do the analysis without any correction at all of the total doses, the slope of the regression line remains exactly the same: 0.48 Gy/day. In other words, this slope could simply be interpreted as a consequence of the practice of dose prescription in the various studies: the relationship between the TCDso and overall treatment time would then reflect the fact that higher total doses tend to be delivered in longer overall times ! But why a slope of 0.5 Gy/day? After all, at 10 Gy/week the slope should have been more than 1 Gy/day, if prolonged treatments simply were given as a number of extra 2 Gy dose fractions. Here the patients with delayed treatments will have some influence. It is also important to recall the profound influence of the Ellis NSD formalism from the late 1960s through the early 1980s. This will have shaped the design of many of the clinical regimens whose results are summarized in Figs. 1 and 2 (and in many institutions also the data now available for dose-time-scattergram analyses), such that the increase in total dose with increasing time (and numbers of fractions) would reflect the predictions of the formula. Indeed, when we apply the N S D formula to a schedule with one fraction per day five days per week (remembering that the N SD formula for tumors did not correct explicitly for treatment time), we obtain dD
D . 0.24
dN
N
165 where D = total dose and N = number of fractions (proportional to time). With D/N = 2 Gy per fraction, the right-hand side is equal to 0.48 Gy per extra fraction, i.e. 0.48 Gy per day. These pitfalls can be avoided to some extent by using multivariate analyses and maximum likelihood techniques. In analyses of data from split-course treatments [4,8], a significant dose response was demonstrated, and thus it was possible to interpret the meaning of "dose/day" to offset repopulation. What is to be avoided is having recourse to comparing total doses to treatment times in the manner outlined in Figs. 1 and 2, after the more rigorous likelihood fitting has failed to produce evidence of a significant influence for dose or time.
Prospects for accelerated fractionation The foregoing suggests that the evidence on which rests the prospect for a therapeutic gain from accelerated fractionation is compromised by either wide confidence limits (estimates of Dproliffrom split-course treatments) or multiple interpretations, some of which have nothing to do with repopulation. Worse still, it is to be remembered that the dose-response relationships observed in the clinic are quite shallow so that the "loss" of 1 Gy/day does not necessarily represent a substantial loss of local control probability. Ten days shortening of the treatment was estimated by Bentzen et al. [4] to results in a change from 60 to 70 ~o in local control probability in a typical patient. To detect a change of this magnitude at a significance level of 5~o (one-sided) and with a power of 90 ~o would require a total of 650 patients in a two-arm randomized trial open for intake for 3 years with an additional 3 years of follow-up. This rate of patient accrual will in most situations require a multicenter trial, with the accompanying problems of standardizing treatment. The only way to reduce the number of patients required would be to identify those with a greater potential advantage from treatment acceleration, and for this measurement of the potential doubling time, Tpot, of tumor clonogens has been suggested. Tpo~ was introduced by Steel [16] as the clonogen doubling time that would be measured if cell loss were ignored, i.e. if both daughter cells remained clonogenic after a mitosis. In practice clonogens are lost by any of a number of mechanisms, including differentiation, death, and metastasis, and the net result is that Tpot will be shorter than the actual clonogen doubling time, T~on. It is of course the latter that is of real importance for radiotherapy. Talon may be expressed in terms of Tpo~, the labelling index or S-phase fraction LI, the S-phase transit time Ts, and the cell-loss factor, CLF:
Llon=
~o,
=----.2"~
l
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LI
1-CLF
wherein 2; is approximately 0.6. From this relationship it is seen that a shorter Tclon may result from (1) a reduced Ts, (2) an increased LI, or (3) a reduced CLF. Currently the third possibility is favored [20], that the CLF is reduced after the beginning of treatment. If the CLF goes to the limit of zero then the net clonogen doubling time Tolo, would equal the pretreatment Tpot, provided that LI and Ts did not change inappropriately. The attraction of all this is that a pretreatment parameter that can be measured by a single biopsy [3] should be predictive of the extent of accelerated tumor growth after onset of treatment. On the other hand, if LI increased after the cytotoxic insult or the CLF was reduced without being close to zero, and if any of these possibilities happened to a varying degree in various tumors, then the Tpot before treatment might not be predictive for Trion during treatment. On the positive side, preliminary results indicate an association between a short Tpot and possible benefit from accelerated fractionation [2]. More data and longer follow-up are needed to establish the strength of this correlation and to judge whether an individually measured Tpo~ can be used for selective allocation to accelerated treatments. The potential for therapeutic gain from accelerated treatment requires that late normal-tissue injury not be changed for the worse when overall time is shortened. Almost a dogma of normal-tissue radiobiology in the 1980s, the truth of this statement has now been challenged, perhaps because of the necessity of giving more than one treatment per day. The CHART trial [7] has produced four myelopathies, a warning that 6 h is not sufficient for complete repair of sublethal damage in the spinal cord, or a slow-repair process was compromised by shortening the overall time, or other. Ang et al. (pers. comm., 1991) have supported this experimentally by showing that significant recovery took place in the rat spinal cord between 8 and 24 h. Finally, Turesson and Thames [19] showed that late telangiectasia following postmastectomy irradiation was characterized by a repair half-time in excess of 3 h. With regard to acute reactions it has been recognized that acceleration of treatment would lead to their worsening. The attitude has been that as long as the patient could be brought through this phase these reactions did not constitute a therapeutic problem. The experience of Peracchia and Salti [ 14] was one warning that this is not necessarily true. Further, Bentzen and Overgaard [5] showed that after stratification for biologically equivalent dose, a previous history of moist desquamation was a predisposing factor for the
166 development of late telangiectasia. Although this may be a problem unique with the pathogenesis of telangiectasia, it may serve as a warning that more surprises could show up as the treatment schedules approach the limits of acute tolerance. Where does this leave us with respect to clinical trials of accelerated fractionation? First, although there is strong support for the assertion that tumor control will improve with shorter overall times, what remains to be clarified is the magnitude of the anticipated improvement and the fraction of patients who would benefit significantly. As outlined in the preceding, care should be exercised in the interpretation of retrospective analyses. If a sizable proportion of tumors considered candidates for treatment by accelerated treatment was in fact slow-growing, even a minor reduction of total dose would not be compensated by a shorter overall
time. Although the preliminary experience with in vivo measures of cell kinetic parameters like Tpot allows some optimism, it is too early to say if routine use is justified for the allocation of patients for accelerated fractionation. Second, with regard to optimal schedules, there is in our view no clear demonstration of a lag time of 4 weeks before onset of accelerated proliferation of tumor clonogens. Also, if a fixed number of fractions per week is given, then depending on the value of ~/fl for the tumor it could be advantageous to prolong overall treatment time by reducing the fraction size except in tumors that are both radioresistant and fast proliferating [9]. Finally, the interfraction interval required for complete recovery between multiple treatments per day may prove too long to be practical.
References 1 Bataini, J. P., Asselain, B., Jaulerry, Ch., Brunin, F., Bernier, J., Pontvert, D. and Lave, C. A multivariate primary tumour control analysis in 465 patients treated by radical radiotherapy for cancer of the tonsillar region: clinical and treatment parameters as prognostic factors. Radiother. Oncol. 14: 265-277, 1989. 2 Begg, A. C., Holland, I., Moonen, L., Bartelink, H., Schraub, S., Bontemps, P., Le Fur, R., Bogaert, W. van den, Caspers, R., Glabbeke, M. van and Horiot, J.-C. The predictive value of cell kinetics measurements in a European trial of accelerated fractionation in advanced head and neck tumors: an interim report. Int. J. Radiat. Oncol. Biol. Phys. 19: 1449-1453, 1990. 3 Begg, A. C., McNally, N. J., Shrieve, D. C. and Karcher, H. A method to measure the DNA synthesis and the potential doubling time from a single sample. Cytometry 6: 620-626, 1985. 4 Bentzen, S .M., Johansen, L.V., Overgaard, J. and Thames, H. D. Clinical radiobiology of squamous cell carcinoma of the oropharynx. Int. J. Radiat. Oncol. Biol. Phys. 20: 1197-1206, 1991. 5 Bentzen, S. M. and Overgaard, M. Relationship between early and late normal-tissue injury after postmastectomy radiotherapy. Radiother. Oncol. 20: 159-165, 1991. 6 Bentzen, S. M., Overgaard, J., Thames, H. D., Overgaard, M., Hansen, P.V., Maase, H. v o n d e r and Meder, J. Clinical radiobiology of malignant melanoma. Radiother. Oncol. 16: 169-182, 1989. 7 Dische, S. and Saunders, M. I. Continuous, hyperfractionated, accelerated radiotherapy (CHART): an interim report upon late morbidity. Radiother. Oncol. 16: 67-74, 1989. 8 Dubois, J. B., Broquerie, J.L., Delard, R. and Pourquir, H. Analysis of the results of irradiation in the treatment of tonsillar region carcinomas. Int. J. Radiat. Oncol. Biol. Phys. 9: 1195-1203, 1983. 9 Fowler, J. F. How worthwhile are are short schedules in radiotherapy?: a series of exploratory calculations. Radiother. Oncol. 18: 165-181, 1990. 10 Fowler, J. F., Tanner, M. A., Bataini, J. P., Asselain, B., Bernier, J. and Lave, C. Further analysis of the time factor in squamous cell carcinoma of the tonsillar region. Radiother. Oncol. 19: 237-244, 1990.
11 Maciejewski, B., Withers, H. R., Taylor, J. M. G. and Hliniak, A. Dose fractionation and regeneration in radiotherapy for cancer of the oral cavity and oropharynx: tumor dose response and repopulation. Int. J. Radiat. Oncol. Biol. Phys. 16: 831-843, 1989. 12 Overgaard, J., Hjelm-Hansen, M., Johansen, L.V. and Andersen, A. P. Comparison of conventional and split-course radiotherapy as primary treatment in carcinoma of the larynx. Acta Oncol. 27: 147-152, 1988. 13 Pajak, T. F., Laramore, G.E., Marcial, V.A., Fazekas, J.T., Cooper, J., Rubin, P., Curran, W. J. and Davis, L. W. Elapsed treatment days - - a critical item for radiotherapy quality control review in head and neck trials: RTOG report. Int. J. Radiat. Oncol. Biol. Phys. 20: 13-20, 1991. 14 Peracchia, G. and Salti, C. Radiotherapy with thrice-a-day fractionation in a short overall time: clinical experiences. Int. J. Radiat. Oncol. Biol. Phys. 7: 99-104, 1981. 15 Saunders, M. I. and Dische, S. Radiotherapy employing three fractions in each day over a continuous period of 12 days. Br. J. Radiol. 59: 523-525, 1986. 16 Steel, G. G. Growth Kinetics of Tumours, pp. 67-72.Clarendon Press, Oxford, 1977. 17 Stewart, J. G. and Jackson, A. W. The steepness of the dose response curve both for tumor cure and normal tissue injury. Laryngoscope 85:1107-1111, 1975. 18 Thames, H. D., Peters, L.J., Spanos, W. and Fletcher, G.F. Dose response of squamous cell carcinomas of the upper respiratory and digestive tracts. Br. J. Cancer 41, Suppl. IV: 35-38, 1980. 19 Turesson, I. and Thames, H. D. Repair capacity and kinetics of human skin during fractionated radiotherapy: erythema, desquamation, and telangiectasia after 3 and 5 year's follow-up. Radiother. Oncol. 15: 169-188, 1989. 20 Withers, H. R., Taylor, J. M. G. and Maciejewski, B. The hazard of accelerated tumor clonogen repopulation during radiotherapy. Acta Oncol. 27: 131-146, 1988.