Dose-Response Modifiers in Radiation Therapy

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Chapter 3 Dose-Response Modifiers in Radiation Therapy

Michael R. Horsman, Jacob C. Lindegaard, Cai Grau, Marianne Nordsmark, Jens Overgaard

When cancer patients undergo radiation therapy, there is a clear dose-response relationship between the dose delivered and the response of the tumor to the radiation (Fig. 3-1). Unfortunately, damage to normal tissue also increases as the radiation dose increases, and this complication limits the total radiation dose that can be given. Substantial effort has been made to modify these dose-response relationships and increase the separation between tumor tissue and normal tissue dose-response curves. The approach has been to either selectively increase the radiation damage to tumor tissue without affecting normal tissues, or protect normal tissues without protecting tumor tissue to the same extent.

Figure 3-1 Schematic illustration of the proportion of patients cured and patients with normal tissue complications as a function of the total radiation dose received.

Agents capable of enhancing radiation response include certain conventional chemotherapeutic agents, the halogenated pyrimidines, and agents that specifically overcome radioresistance resulting from hypoxic cells that occur because of environmental conditions within most solid tumors. The most widely investigated method applied to the hypoxia problem is radiosensitization of the hypoxic cells by using either electron-affinic sensitizing drugs or hyperthermia. Another approach often used to reduce hypoxia—especially in experimental systems—involves increasing oxygen availability by (1) having the patient breathe high-oxygen-content gas, (2) introducing perfluorochemical emulsions into the vascular system to increase the oxygen-carrying capacity of the blood, (3) using agents that affect hemoglobin, or (4) using drugs that increase tumor blood perfusion. Many experimental studies have also demonstrated that hypoxic cells can be preferentially destroyed by bioreductive drugs that are active under reduced oxygen conditions or by again using hyperthermia; each of these hypoxic-cell cytotoxins improve the radiation response of tumors. Preliminary data suggest that a new group of agents, the so-called vascular targeting agents, may enhance radiation damage. This group includes drugs that inhibit angiogenesis, the process by which tumors develop their own vascular supply, and drugs that preferentially damage the already established tumor vessels.

Radiation protectors fall into several categories. There are the true radiation protectors, in particular sulfhydryl compounds, which primarily appear to interact with radicals that form after radiation exposure. Another group of agents decrease oxygen delivery to tissues, and as such would be expected to reduce the amount of radiation damage induced. Finally, there are various cytokines, growth factors, prostaglandins, and essential fatty acids that are capable of radioprotection, although their mechanisms of action are not entirely clear.

Radiosensitization by conventional chemotherapeutic agents (e.g., cisplatin, 5-fluorouracil, mitomycin C), halogenated pyrimidines (e.g., 5-bromodeoxyuridine, 5-iododeoxyuridine), or hyperthermia is discussed in detail elsewhere in this book. In this chapter, the focus will be on hypoxic cell modifiers, vascular targeting drugs, and radioprotectors.

The Hypoxia Problem

The Importance of Oxygen

In 1909, Gottwald Schwarz,¹ in a simple but elegant experiment, demonstrated that the radiation response of skin was markedly decreased if the blood flow in the irradiated area was reduced by compression. Although he did not acknowledge that the phenomenon was the result of a lack of oxygen, his study was probably the first radiobiologically oriented clinical study implicating the importance of environmental parameters in the outcome of radiotherapy. This finding was used to introduce the concept of kompressionsanämie, by which the skin was made anemic, thereby allowing a higher dose to be given to deeply situated tumors. Following the work of Schwarz, Müller,² in 1910, reported that tissues in which the blood flow was stimulated by diathermia showed a more prominent response to radiation. This early study not only demonstrated the importance of the oxygen supply in radiotherapy but was also the first clinical approach showing how resistance could be overcome by using hyperthermia. Subsequently, sporadic clinical and experimental observations indicated the importance of a sufficient blood supply in securing an adequate radiation response. These observations led Gray and colleagues³ in the early 1950s to postulate that oxygen deficiency or hypoxia was a major source of radiation resistance.

The first clinical indication that hypoxia existed in tumors was made around the same time by Thomlinson and Gray ⁴ when, from histologic observations in carcinoma of the bronchus, they reported seeing viable tumor regions surrounded by a vascular stroma from which the tumor cells obtained their nutrients and oxygen. As the tumors grew, the viable regions expanded and areas of necrosis appeared at the center. The thickness of the resulting shell of viable tissue was 100 to 180 µm, which was within the same range as the calculated diffusion distance for oxygen in respiring tissues. It was thereby suggestive that as oxygen diffused from the stroma, it was consumed by the cells and, although cells beyond the diffusion distance were unable to survive, the cells immediately bordering the necrotic area might be viable but hypoxic. In 1968, Tannock⁵ described an inverted version of the Thomlinson and Gray picture, with functional blood vessels surrounded by cords of viable tumor cells outside of which areas of necrosis were seen. This “corded” structure, illustrated in Figure 3-2, is the more typical picture found in most solid tumors. It arises because the tumor blood vessels, which are derived from the normal tissue vessels by angiogenesis, are inadequate to meet the needs of the rapidly growing tumor cells. This hypoxia is commonly called chronic hypoxia. It was also suggested that hypoxia in tumors could be acute in nature. Chaplin and colleagues⁶ confirmed the existence of acutely hypoxic cells in tumors and demonstrated that these cells resulted from transient stoppages in tumor blood flow (see Fig. 3-2). To date, these temporary cessations in blood flow have been observed in mouse and rat tumors as well as in human tumor xenografts, with 4% to 8% of the total functional vessels involved,⁷ although the exact causes of these stoppages are not known. The current use of “chronic” and “acute” to explain hypoxia in tumors is probably an oversimplification of the real situation. Chronic hypoxia generally refers to prolonged and reduced oxygen concentrations that influence radiation response, but there is evidence that oxygen concentrations that are higher but still below normal physiologic levels are often found. Furthermore, reduced perfusion can be either partial or total, and although cells in the former condition would be oxygen deprived, cells in the latter condition would be starved of oxygen and nutrients and, therefore, their survival and response to therapy would be expected to be different.

Figure 3-2 Schematic representation of the interrelationship between tumor cells and the vascular supply. On the left, cells are seen growing as a “corded” structure around a functional vessel from which the cells receive their oxygen supply. As oxygen diffuses out from the vessel it is used up; therefore, the outermost viable cells (shown by the shading) are oxygen deprived or chronically hypoxic. A similar arrangement is seen on the right, but here flow through the vessel is transiently stopped, making all the cells oxygen deprived.

Reprinted from Horsman MR: Measurement of tumor oxygenation, Int J Radiat Oncol Biol Phys 42:701-714, 1998.

Evidence for Hypoxia in Tumors

In experimental tumors it is not only relatively easy to identify hypoxia but one can also quantitatively estimate the percentage of cells that are hypoxic. Three major techniques are routinely used,⁸ including the paired survival curve, the clamped tumor growth delay, and the clamped tumor control assays. Each involves comparing the response of tumors irradiated under normal air breathing conditions with that of tumors artificially made hypoxic by clamping. Using these procedures, hypoxia has been directly identified in most animal solid tumors, with the values ranging from less than 1% to well over 50% of the total viable cell population.⁸ Unfortunately, none of these procedures can be applied in clinical situations. One must, therefore, rely on indirect techniques.

Tumor hypoxia has previously been estimated clinically by various methods,^9,¹⁰ including measurement of tumor vascularization by using such endpoints as intercapillary distance, vascular density, and the distance from tumor cells to the nearest blood vessel; measurement of tumor metabolic activity by using biochemical, high-performance liquid chromatography, bioluminescent, and magnetic resonance techniques; and estimation of the degree of DNA damage by using the Comet assay. The more popular techniques currently under investigation include measurement of the binding of exogenous markers identified by immunohistochemical techniques from histologic sections (e.g., pimonidazole, EF5) or by noninvasive positron emission tomography (PET) (e.g., fluorine-18 [¹⁸F]-labeled misonidazole, FAZA), single-photon emission computed tomography (SPECT) (e.g., iodine-123 [¹²³I]-labeled azomycin arabinoside), or magnetic resonance spectroscopy (e.g., SR4554). Also, the up-regulation of endogenous proteins either immunohistochemically (e.g., carbonic anhydrase IX, GLUT-1, HIF-1) or from blood samples (e.g., osteopontin) and the measurement of partial pressure of oxygen (PO₂) distributions with polarographic electrodes can be used. The latter method has become by far the most popular. The results of an international multicenter study in head and neck cancer patients are illustrated in Figure 3-3. In this study, the tumor PO₂ was measured before radiation therapy and was found to correlate with the overall survival rate in that patients with a lower tumor oxygenation status did significantly worse.¹¹

Figure 3-3 Oxygen levels were measured with Eppendorf electrodes before radiation therapy in 397 patients with squamous cell carcinomas of the head and neck. Tumors were stratified by whether the fraction of PO₂ values ≤2.5 mm Hg (HP 2.5) were above or below the median value for the whole group (i.e., 19%). The lines show Kaplan-Meier estimates of actuarial overall survival probability for patients with less hypoxic tumors (HP 2.5 ≤19%; red line) compared with more hypoxic tumors (HP 2.5 >19%; blue line), p = 0.006.

From Nordsmark M, Bentzen SM, Rudat V, et al: Prognostic value of tumor oxygenation in 397 head and neck tumors after primary radiation therapy. An international multi-center study, Radiother Oncol 77:18-24, 2005, with permission.

Probably the best evidence for the existence of hypoxia in human tumors comes from the large number of clinical trials in which hypoxic modification has shown some benefit.¹² The latter situation constitutes a circular argument: if hypoxic modification shows an improvement, then hypoxic clonogenic cells must have been present in tumors. It is, however, likely that even tumors with the same histologic makeup and of the same type have substantial heterogeneity with respect to the extent of hypoxia. It must be admitted that today, almost a century after the first clinical description, the importance of hypoxia and its influence on the outcome of radiotherapy is still the subject of substantial debate. However, we will now discuss in detail how the different hypoxic modifiers have been used to modify the radiation dose response of tumors.

Overcoming Tumor Hypoxia

Breathing High-Oxygen-Content Gas

Because the oxygen supply to tumors is insufficient to meet the needs of all of the tumor cells, radiation-resistant hypoxia develops; therefore, an obvious solution for improving the tumor’s radiation response would be to increase the oxygen supply. This has been tried, both experimentally and clinically, by simply allowing the tumor-bearing host to breathe high-oxygen-content gas mixtures before and during irradiation.

Early experimental studies reported that breathing either oxygen or carbogen (95% O₂ + 5% CO₂) could substantially enhance the response of murine tumors to radiation and that the best effect was generally seen when the gases were inhaled under hyperbaric (typically 3 atmospheres [atm]) rather than normobaric conditions.¹³ This is not surprising because hyperbaric conditions would be expected to saturate the blood with more oxygen. However, more recent studies have indicated that the radiosensitization produced by normobaric oxygen or carbogen are quite substantial¹⁴; because it is quicker and easier to breathe gas under normobaric conditions, the use of cumbersome, expensive, complex hyperbaric chambers is probably not necessary.

Clinically, the use of high-oxygen-content gas breathing, specifically under hyperbaric conditions, was introduced relatively early.¹⁵ Most trials were fairly small and suffered from the applications of unconventional fractionation schemes, but it appeared that the effect of hyperbaric oxygen was superior to that of radiotherapy given in air, especially when few and large fractions were applied. In the large, multicenter clinical trials conducted by the Medical Research Council (Table 3-1), the results from both uterine cervix and advanced head and neck tumors showed a significant benefit in local tumor control and subsequent survival rates. The same findings were not observed in bladder cancer nor were they seen in a number of smaller studies. In retrospect, the use of hyperbaric oxygen was stopped somewhat prematurely. This was partly the result of the introduction of hypoxic radiosensitizers and partly because of problems with patient compliance; it has been claimed that hyperbaric treatment caused significant suffering, but the discomfort associated with such a treatment must be considered minor compared with the often life-threatening complications associated with chemotherapy, which is used with less restrictive indications.

TABLE 3-1 Multicenter Randomized Trials with Hyperbaric Oxygen (HBO)

The use of high-oxygen-content gas breathing under normobaric conditions to radiosensitize human tumors has also been tried clinically, but it failed to show any dramatic improvement.¹⁶ In the most recent study this may have been the result of size limitation, but in earlier studies it may have been caused by the failure to achieve the optimal preirradiation gas breathing time. Experimental studies have shown that the amount of time is critical for the enhancement of radiation damage and that it can vary from tumor to tumor.¹⁷

Hypoxic Cell Radiosensitizers

An alternative approach to the hypoxia problem is the use of chemical agents that mimic oxygen and preferentially sensitize the resistant population to radiation. The advantage of these drugs over oxygen is that they are not rapidly metabolized by the tumor cells through which they diffuse and thus the drugs can penetrate further than oxygen and so reach all the tumor cells. In the early 1960s, researchers found that the efficiency of radiosensitization was directly related to electron affinity,¹⁸ and that finding ultimately led to in vitro studies demonstrating preferential radiosensitization of hypoxic cells by highly electron-affinic nitroaromatic compounds.¹⁹ Several of these compounds were later shown to be effective at enhancing radiation damage in tumors in vivo,²⁰ and as a result they underwent clinical testing.

The drugs reaching clinical evaluation include metronidazole, misonidazole, benznidazole, desmethylmisonidazole, etanidazole, pimonidazole, nimorazole, ornidazole, sanazole, and doranidazole. Initial clinical studies were with metronidazole in brain tumors and were followed, in the latter part of the 1970s, by a boom in clinical trials exploring the potential of misonidazole as a radiosensitizer.²⁰ The results from the multicenter randomized trials are summarizsed in Table 3-2. Most of the trials with misonidazole were unable to generate any significant improvement in radiation response, although a benefit was seen in some trials, especially the second Danish Head and Neck Cancer study (DAHANCA 2), which found a highly significant improvement in the stratification subgroup of pharynx tumors but not in the prognostically better glottic carcinomas. The overall impression of the “misonidazole era” was a prolongation of the inconclusive experience from the hyperbaric oxygen trials, namely that the problems related to hypoxia had not been ruled out definitively. Therefore, the search for more efficient or less toxic hypoxic sensitizers continues. Furthermore, the experience from the misonidazole trials has been taken into account to select a more homogeneous tumor population in which hypoxia is more likely to be present.

TABLE 3-2 Multicenter Randomized Trials with Nitroimidazoles

Results from subsequent randomized trials with other nitroaromatic compounds have been conflicting. The European pimonidazole trial in uterine cervical cancer was very disappointing, whereas the two other multicenter trials in head and neck cancer, using etanidazole, showed no benefit. On the other hand, studies with the low-toxicity drug nimorazole given to patients with supraglottic and pharyngeal carcinomas (DAHANCA 5) showed a highly significant benefit in terms of improved locoregional tumor control and disease-free survival rates ²¹ (Fig. 3-4), thereby confirming the result of the DAHANCA 2 study. A more recent International Atomic Energy Agency (IAEA) trial with the 3-nitrotriazole compound sanazole (AK 2123), in uterine cervical cancer, also demonstrated a significant improvement in both local tumor control and overall survival rates.²²

Figure 3-4 Actuarial estimated locoregional tumor control and disease-specific survival rate in patients randomized to receive nimorazole or placebo in conjunction with conventional radiotherapy for carcinoma of the pharynx and supraglottic larynx.

Reprinted from Overgaard J, Sand Hansen H, Overgaard M, et al: A randomised double-blind phase III study of nimorazole as a hypoxic radiosensitizer of primary radiotherapy in supraglottic larynx and pharynx carcinoma. Results of the Danish head and neck cancer study [DAHANCA] protocol 5-85, Radiother Oncol 46:135-146, 1998, with permission from Elsevier Science.

A meta-analysis of all randomized clinical studies in which hypoxic radiosensitizers were given with primary radiotherapy, involving more than 11,000 patients in 91 randomized trials, showed that radiosensitizer modification of tumor hypoxia significantly improved the locoregional tumor control after radiotherapy with an odds ratio of 0.78 (0.72 to 0.85; 95% confidence intervals). This treatment benefit was mostly the result of an improved response in head and neck cancer with an odds ratio of 0.74 (0.65 to 0.83) and to a lesser extent in bladder tumors. No significant effect was observed in other tumor sites (e.g., cervix, lung, central nervous system, esophagus). The overall survival rate was also improved with an overall odds ratio of 0.83 (0.77 to 0.90). In conclusion, the hypoxic radiosensitizer era has clearly demonstrated an overall significant benefit for both local control and survival rates in certain tumor sites. The overall observed gain was an improvement of 5% to 10 % in local control. Although small, such gains are certainly relevant to pursue, especially because they are apparently associated with a similar improvement in survival rates. On this basis there is no need for a feeling of disappointment at the apparent lack of a major clinical benefit. The nonsignificant outcome of most clinical trials is not due to a biologic lack of importance of hypoxia but, in most cases, is considered a consequence of poor clinical trial methodology with an overly optimistic study design and an expected treatment gain that goes far beyond what is reasonable. Overall, the results with nitroimidazoles add to the general consensus that if a nontoxic hypoxic modification can be applied then such treatments may certainly be relevant as a baseline therapy together with radiotherapy for cancers such as advanced head and neck cancer. Such a strategy has been adopted in Denmark, where nimorazole has become part of the standard radiotherapy treatment in head and neck cancers.