Reducing Acute and Chronic Hypoxia in Tumours by Combining Nicotinamide with Carbogen Breathing

The ability of nicotinamide and carbogen breathing to improve the radiation response of a C3H mammary carcinoma by reducing both acute and chronic hypoxia was investigated. Using a tumour growth delay assay the response of 200 mm³ foot tumours to local irradiation was found to be increased by either injecting nicotinamide (100-1 000mg/kg) 20 min prior to irradiation, or by allowing mice to breathe carbogen for 10 min before and during the radiation treatment. The greatest radiosensitization occurred when nicotinamide and carbogen were combined. With a histological fluorescent staining technique nicotinamide was shown to prevent transient stoppages in microregional blood flow, and also appeared to improve tumour oxygenation as measured with an Eppendorf oxygen electrode, both effects being consistent with its ability to decrease perfusion limited acute hypoxia. Carbogen had no effect on vessel closure, but it significantly improved tumour oxygenation, which was indicative of it reducing diffusion limited chronic hypoxia.     

Nicotinamide and Other Benzamide Analogs as Agents for Overcoming Hypoxic Cell Radiation Resistance in Tumours

Oxygen deficient hypoxic cells, which are resistant to sparsely ionising radiation, have now been identified in most animal and some human solid tumours and will influence the response of those tumours to radiation treatment. This hypoxia can be either chronic, arising from an oxygen diffusion limitation, or acute, resulting from transient stoppages in microregional blood flow. Although clinical attempts to overcome hypoxia have met with some success, the results have been far from satisfactory, and efforts are still being made to find better methods. Extensive experimental studies, especially in the last decade, have shown that nicotinamide and structurally related analogs can effectively sensitise murine tumours to both single and fractionated radiation treatments and that they do so in preference to the effects seen in mouse normal tissues. The earliest studies suggested that this enhancement of radiation damage was the result of an inhibition of the repair mechanisms, as was well documented in vitro. However, recent studies in mouse tumours have shown that the primary mode of action actually involves a reduction in tumour hypoxia. More specifically, these drugs prevent transient cessations in blood flow, thus inhibiting the development of acute hypoxia. This novel discovery led to the suggestion that the potential role of these agents as radiosensitizers would be when combined with treatments that overcame chronic hypoxia. The first attempt to demonstrate this combined nicotinamide with hyperthermia and found that the enhancement of radiation damage by both agents together was greater than that seen with each agent alone. Similar results were later seen for nicotinamide combined with a perfluorochemical emulsion, carbogen breathing, and pentoxifylline, and in all these studies the effects in tumours were always greater than those seen in appropriate normal tissues. Of all the analogs, it is nicotinamide itself which has been the most extensively studied as a radiosensitizer in vivo and the one that shows the greatest effect in animal tumours. It is also an agent that has been well established clinically for the treatment of a variety of disorders, with daily doses of up to 6 g being considered reasonably safe and associated with a low incidence of side effects. This human dose is equivalent to 100-200 mg/kg in mice and such doses will maximally sensitize murine tumours to radiation. These findings have now resulted in phase I/II clinical trials of nicotinamide, in combination with carbogen breathing, as a potential radiosensitizing treatment.     

Reoxygenation in a C3H Mouse Mammary Carcinoma the importance of chronic rather than acute hypoxia

The role that chronic and acute hypoxia play in tumour reoxygenation after irradiation was investigated in a C3H mouse mammary carcinoma grown in the feet of female CDF1 mice. Tumours at 200 mm³ in size were locally irradiated with a priming dose of 20 Gy and then at various times after given a range of radiation doses under normal or clamped conditions. Local tumour control was determined 90 days later from which the tumour hypoxic fractions were calculated. Untreated tumours contained 23% hypoxic cells. Immediately after 20 Gy this increased to 52% and by 24 h had fallen to 10%. These reoxygenation experiments were repeated, giving either nicotinamide (1000 mg/kg; i.p. injected 30 min before each irradiation) to remove acute hypoxia, or carbogen breathing (for 5 min before and during irradiation) to decrease chronic hypoxia. With nicotinamide the normal hypoxic fraction was reduced to 7%, but after irradiation it had risen to 46% and by 24 h there was full reoxygenation with a value of 5% being observed. Carbogen breathing also decreased the normal hypoxic fraction to 6%, and immediately after irradiation this was increased to 38%. However, by 24 h it was still elevated at around 23%. These results suggest that chronic rather than acute hypoxia is necessary for reoxygenation in this tumour.     

A therapeutic benefit from combining normobaric carbogen or oxygen with nicotinamide in fractionated X-ray treatments.

The ability of normobaric oxygen and carbogen (95% O2 + 5% CO2) combined with nicotinamide to enhance the radiosensitivity of two rodent adenocarcinomas and of mouse skin and kidneys, using a 10 fraction radiation schedule, was compared with the effect of radiation in air with and without the drug. Tumour response was assayed using local control and regrowth delay, and compared with acute skin reactions, decreased renal 51Cr-EDTA clearance and reduction in haematocrit. Nicotinamide increased the radiation sensitivity of CaNT tumours under all three different oxygen concentrations tested (21, 95 and 100% oxygen). The effect was statistically significant for oxygen and carbogen but not for air; the combination of nicotinamide with carbogen gave the greatest increase in tumour radiosensitivity. Relative to treatments in air without the drug, the enhancement ratios (ER) at the TCD50 level were 1.17, 1.65 and 1.83 for CaNT tumours irradiated in air, oxygen or carbogen and injected with nicotinamide 1 h before each fraction. The ER in CaRH tumours irradiated in carbogen plus the drug was 1.83, which was greater, but statistically not significantly different, to that seen with carbogen alone (ER = 1.68). In skin, relative to air without the drug, the increase in radiosensitivity by nicotinamide was greater in oxygen and carbogen than in air (1.29, 1.36 and 1.08, respectively). The ERs for both assays of renal damage were similar and lower than those in skin: less than or equal to 1.07, less than or equal to 1.13 and less than or equal to 1.16 for irradiations done in air, oxygen and carbogen plus nicotinamide, relative to air alone. A comparison of these results in the tumours and normal tissues showed that a significant therapeutic benefit was obtained with normobaric oxygen and carbogen combined with nicotinamide. This benefit is greater than observed with other radiosensitizers tested so far. Toxic side effects of the treatment are unlikely in a clinical situation, since prolonged administration of nicotinamide is well tolerated in man. The combination of normobaric carbogen with nicotinamide could be an effective method of enhancing tumour radiosensitivity in clinical radiotherapy where hypoxia limits the outcome of treatment.     

Alteration of tumour response to radiation by interleukin-2 gene transfer

We have previously shown that BALB/c-derived EMT6 mammary tumours transfected with interleukin (IL)-2 have decreased hypoxia compared to parental tumours, due to increased vascularization. Since hypoxia is a critical factor in the response of tumours to radiation treatment, we compared the radiation response of IL-2-transfected tumours to that of parental EMT6 tumours. Because the IL-2 tumours have an altered host cell composition, which could affect the interpretation of radiation sensitivity as measured by clonogenic cells, we employed flow cytometric analysis to determine the proportion of tumour cells vs host cells in each tumour type. Using this approach, we were able to correct the plating efficiency based on the number of actual tumour cells derived from tumours, making the comparison of the two types of tumours possible. We also excluded the possibility that cytotoxic T-cells present in EMT6/IL-2 tumours could influence the outcome of the clonogenic cell survival assay, by demonstrating that the plating efficiency of cells derived from EMT6/IL-2 tumours remained unchanged after depletion of Thy-1+ cells. The in vivo radiation response results demonstrated that IL-2-transfected tumours were more sensitive to radiation than parental EMT6 tumours. The hypoxic fraction of the EMT6/IL-2 tumours growing in vivo was markedly decreased relative to parental EMT6 tumours thus the increased sensitivity results from the increased vascularity we have previously observed in these tumours. These results indicate the potential therapeutic benefit of combining radiation and immunotherapy in the treatment of tumours.     

Modification of tumour radiation response in vivo by the benzamide analogue pyrazinamide.

Pyrazinamide, the pyrazine analogue of nicotinamide, has been evaluated for its ability to modify the radiation response of hypoxic cells both in vivo and in vitro. Results obtained with three different murine tumour systems EMT6, LLC and SCCVII showed that pyrazinamide at a dose of 0.5 mg g-1 i.p. resulted in enhanced radiation response. Dose modification factors of between 1.3 and 1.6 were observed using in vivo/in vitro clonogenic assays. This enhancement was greater than that obtained in mouse intestine using crypt cell survival as an endpoint (DMF 1.1). In contrast to the tumour data in vivo, the in vitro results indicate that pyrazinamide displays little radiosensitising or toxic properties towards hypoxic CHO cells in culture. These results suggest that pyrazinamide exerts its effects in vivo either by directly perturbing tumour physiology or by being converted to an active metabolite. Blood flow studies performed using laser Doppler flowmetry indicate that pyrazinamide produces a small (32%) increase in overall tumour blood flow in the SCCVII tumour. Based on this finding, additional studies on tumour perfusion at the microregional level were performed in the SCCVII tumour using a histological technique involving injection of fluorescent stains which demarcate functional vasculature. The data show that when compared to saline injected controls, pyrazinamide reduced the number of vessels opening and closing over a 20 min period from 10.2% to 3.8%. This finding suggests that pyrazinamide may exert its effects at least in part by reducing the occurrence of acute hypoxia resulting from dynamic changes in microregional perfusion.     

Further evaluation of nicotinamide and carbogen as a strategy to reoxygenate hypoxic cells in vivo: importance of nicotinamide dose and pre-irradiation breathing time.

The combination of nicotinamide and carbogen breathing is awaiting clinical evaluation as a strategy to overcome tumour hypoxia and thus enhance radiation response. We have continued our evaluation of this approach in the murine SCCVII tumour with the aim of determining the importance of nicotinamide dose and the pre-irradiation breathing time (PIBT) for carbogen. For carbogen breathing alone maximal enhancement of radiation response was observed with PIBT''s of between 5 and 30 min. When nicotinamide (1,000 mg kg-1 IP) was administered 60 min prior to irradiation little or no variation in radiation response was observed for all the PIBT''s examined (5-90 min). Indeed at all PIBT''s the cell survival obtained for the carbogen nicotinamide and radiation combination was indistinguishable from that expected for a fully aerobic response. For PIBT''s of 15 and 60 min we examined the influence of nicotinamide doses between 50 and 1,000 mg kg-1. Significant radiosensitizing effects were observed for all nicotinamide doses tested above 50 mg kg-1. Moreover for doses of 250 mg kg-1 and above the cell survival data was consistent with that expected for a fully aerobic response. No additional benefit accrued from raising the nicotinamide dose above 250 mg kg-1. These results indicate that significant radiosensitization may be expected even with clinically achievable nicotinamide doses when it is combined with carbogen breathing. Furthermore, the use of nicotinamide may reduce the critical importance of PIBT on the radiosensitization observed with carbogen.     

The effects of carbogen and nicotinamide on intravascular oxyhaemoglobin saturations in SCCVII and KHT murine tumours.

Considerable effort has been focused on devising methods for manipulating tumour oxygenation and thereby improving tumour radiosensitivity. The combination of nicotinamide and carbogen has been proposed to oxygenate both chronically and acutely hypoxic cells in tumours. However, results have varied markedly with both tumour model and measurement technique. The current objectives were (1) to determine whether changes in radiosensitivity following oxygen manipulation correlated with changes in tumour oxygenation and (2) to assess whether oxygenation was preferentially improved in specific tumour micro-regions. Using two murine tumour lines, the SCCVII carcinoma and the KHT sarcoma, tumour intravascular HbO2 saturations were measured cryospectrophotometrically following nicotinamide, carbogen or the combination. Generally, nicotinamide had minor effects on oxygenation, arguing against a substantial effect on acute hypoxia, while carbogen and the combination produced marked and equivalent improvements in oxygen availability. These results demonstrate that changes in tumour radiosensitivity may not agree with corresponding changes in oxygenation, even within a given tumour model, and that the efficacy of a given manipulative agent may vary substantially with tumour line. One possible explanation for these findings is that different subpopulations of clonogenic vs non-clonogenic cells may be oxygenated by alternative treatments.     

Energy metabolism in human melanoma cells under hypoxic and acidic conditions in vitro.

The response to treatment and the malignant progression of tumours are influenced by the ability of the tumour cells to withstand severe energy deprivation during prolonged exposure to hypoxia at normal or low extracellular pH (pHe). The objective of the present work was to demonstrate intertumour heterogeneity under conditions of microenvironment-induced energy deprivation and to investigate whether the heterogeneity can be attributed to differences in the capacity of the tumour cells to generate energy in an oxygen-deficient microenvironment. Cultures of four human melanoma cell lines (BEX-c, COX-c, SAX-c, WIX-c) were exposed to hypoxia in vitro at pHe 7.4, 7.0 or 6.6 for times up to 31 h by using the steel-chamber method. High-performance liquid chromatography was used to assess adenylate energy charge as a function of exposure time. Cellular rates of glucose uptake and lactate release were determined by using standard enzymatic test kits. The adenylate energy charge decreased with time under hypoxia in all cell lines. The decrease was most pronounced shortly after the treatment had been initiated and then tapered off. BEX-c and SAX-c showed a significantly higher adenylate energy charge under hypoxic conditions than did COX-c and WIX-c whether the pHe was 7.4, 7.0 or 6.6, showing that tumours can differ in the ability to avoid energy deprivation during microenvironmental stress. There was no correlation between the adenylate energy charge and the rates of glucose uptake and lactate release. Intertumour heterogeneity in the ability to withstand energy deprivation in an oxygen-deficient microenvironment cannot therefore be attributed mainly to differences in the capacity of the tumour cells to generate energy by anaerobic metabolism. The data presented here suggest that the heterogeneity is rather caused by differences in the capacity of the tumour cells to reduce the rate of energy consumption when exposed to hypoxia.     

Hydralazine-induced enhancement of hyperthermic damage in a C3H mammary carcinoma in vivo

Prolonged oxygen deprivation of cells in vitro or in vivo increases the sensitivity of those cells to heat. Hydralazine is a peripheral vasodilator, currently used clinically as an antihypertensive agent, which has been reported to be able to reduce tumour blood flow and increase the degree of tumour hypoxia. We have investigated the potential of hydralazine to enhance the response of a C3H mammary carcinoma to local hyperthermia. The tumour was grown in the foot of mice and its response to treatment assayed by regrowth delay. Our results show that a single intravenous injection of hydralazine (5 mg/kg) significantly enhances the heat damage produced by heating for various times at either 41.5, 42.5, or 43.5 degrees C. This effect was dependent on the time of starting to heat after hydralazine injection, with the greatest enhancement occurring when heat was given within 1 h following drug administration. However, the effect was independent of the hydralazine concentration, at doses above 2.5 mg/kg. Hydralazine also significantly decreased mean arterial blood pressure and the uptake of 86RbCl into tumours. Our results suggest that the observed heat sensitization was primarily a consequence of an increase in tumour hypoxia, probably resulting from a decrease in tumour blood perfusion.     

Clinical outcome and tumour microenvironmental effects of accelerated radiotherapy with carbogen and nicotinamide

Experimental studies have shown an almost 2-fold increase in effectiveness if accelerated radiotherapy combined with carbogen and nicotinamide (ARCON) was compared with standard radiotherapy. This combination was chosen in order to overcome repopulation of clonogens during radiotherapy and to minimize tumour hypoxia. Analysis of microenvironmental parameters is required to identify tumours that can benefit from these new treatment approaches. In this study 124 patients with stage III or IV head and neck squamous cell carcinomas received ARCON treatment. Vascular architecture, perfusion, proliferation and oxygenation were studied in two human laryngeal squamous cell carcinoma xenograft lines and the effects of carbogen and nicotinamide were analysed. Loco-regional control for stage III-IV larynx carcinomas was 85%, for hypopharynx carcinomas 50% and for oral cavity and oropharynx carcinomas 65%. In the experimental studies, carbogen treatment resulted in one tumour line in a decrease of blood perfusion, which was reversed if nicotinamide was added. The other tumour line showed no perfusion changes after carbogen or nicotinamide treatment. Both tumour lines showed a drastic reduction of hypoxia after carbogen breathing only or carbogen breathing plus nicotinamide. The ARCON schedule results in high loco-regional tumour control rates. Analysis of tumour microenvironmental parameters showed differences in response to carbogen and nicotinamide between different tumour lines of similar histology and site of origin. This indicates that it may be advantageous to base the selection of patients for oxygenation modifying treatment on microenvironmental tumour characteristics.     

A pilot study of the effects of mild systemic heating on human head and neck tumour xenografts: Analysis of tumour perfusion,interstitial fluid pressure,hypoxia and efficacy of radiation therapy

Abstract

Purpose: The tumour microenvironment is frequently hypoxic, poorly perfused, and exhibits abnormally high interstitial fluid pressure. These factors can significantly reduce efficacy of chemo and radiation therapies. The present study aims to determine whether mild systemic heating alters these parameters and improves response to radiation in human head and neck tumour xenografts in SCID mice.

Materials and methods: SCID mice were injected with FaDu cells (a human head and neck carcinoma cell line), or implanted with a resected patient head and neck squamous cell carcinoma grown as a xenograft, followed by mild systemic heating. Body temperature during heating was maintained at 39.5?±?0.5?°C for 4?h. Interstitial fluid pressure (IFP), hypoxia and relative tumour perfusion in the tumours were measured at 2 and 24?h post-heating. Tumour vessel perfusion was measured 24?h post-heating, coinciding with the first dose of fractionated radiotherapy.

Results: Heating tumour-bearing mice resulted in significant decrease in intratumoural IFP, increased the number of perfused tumour blood vessels as well as relative tumour perfusion in both tumour models. Intratumoural hypoxia was also reduced in tumours of mice that received heat treatment. Mice bearing FaDu tumours heated 24?h prior to five daily radiation treatments exhibited significantly enhanced tumour response compared to tumours in control mice.

Conclusions: Mild systemic heating can significantly alter the tumour microenvironment of human head and neck tumour xenograft models, decreasing IFP and hypoxia while increasing microvascular perfusion. Collectively, these effects could be responsible for the improved response to radiotherapy.     

Nicotinamide, Fluosol DA and Carbogen: a strategy to reoxygenate acutely and chronically hypoxic cells in vivo.

The effect of Nicotinamide and/or treatment with Fluosol DA and Carbogen breathing on the radiation response of 500-750 mg SCCVII and KHT tumours has been evaluated. Pretreatment with Fluosol DA/Carbogen or Nicotinamide resulted in relatively modest enhancements of radiation damage with enhancement factors of 1.1 and 1.3 being observed using an in vivo/in vitro clonogenic end-point. A combination of Nicotinamide and Fluosol DA/Carbogen resulted in a larger enhancement factor of 1.6 over the radiation dose ranges studied. These modification factors reflect a value close to that expected for a fully aerobic response in this survival range. Growth delay studies in the SCCVII tumour provided similar results. Using a recently developed fluorescence activated cell sorting technique, which utilizes the in vivo pharmacokinetic and DNA binding properties of the bisbenzamide stain Hoechst 33342, the effect of Nicotinamide and/or Fluosol DA/Carbogen schedules on the occurrence of acute hypoxia was assessed. The results clearly show that Nicotinamide significantly reduces the amount of 'acute hypoxia', but has a lesser effect on 'chronic' hypoxic cells. However, combinations of Nicotinamide and Fluosol DA/Carbogen significantly increase the response of both 'acutely' and 'chronically hypoxic' cells. The results provide evidence that a combination of Nicotinamide and Fluosol DA/Carbogen can provide an effective way of reoxygenating both acutely and chronically hypoxic cells.     

The effect of cisplatin on the repair of radiation damage in RIF1 mouse tumours in vivo

The effect of the antitumour agent cisplatin on repair of X-ray-induced damage was studied in RIF1 mouse tumours treated in situ. The response of tumours, assessed by growth delay, to 4 fractions of X-rays given at 5-h intervals was compared with that after single doses. The displacement between the curves was taken as a measure of repair. A single dose of 6 mg.kg-1 cisplatin given 0.5 h before the first fraction resulted in no detectable inhibition of repair despite a significant growth delay caused by drug alone. A dose of 2 mg/kg cisplatin given 0.5 h before each of the X-ray fractions did, however, cause some repair inhibition; a result confirmed by tumour control experiments. The schedule dependence for repair inhibition was the same whether the irradiations were carried out on clamped (fully hypoxic) tumours or under ambient conditions. Significant enhancement of radiation damage was seen after correcting for the effects of drug alone, whether or not repair inhibition occurred. The effects of cisplatin on normal stroma within the tumour (vascular damage) was also investigated by monitoring the regrowth rates of recurrent tumours. In contrast to the effects on tumour cells, no enhancement of damage or inhibition of repair was seen for this assay in the combined treatment schedules.     

Perfusion changes in the RIF-1 tumour and normal tissues after carbogen and nicotinamide, individually and combined.

The strategy of combining carbogen breathing and nicotinamide to overcome chronic and acute hypoxia respectively is being evaluated clinically. The effects of both agents individually and in combination on relative perfusion of 400-700 mm3 RIF-1 tumours and normal tissues were measured by 86Rb extraction. Carbogen breathing alone for 6 min increased relative tumour perfusion by 50-70% compared with control at flow rates of 50 to 200 ml min-1, but the effect was lost at 300 ml min-1. All flow rates also produced similar increases in relative perfusion of lung, of between 36% and 58%, and smaller increases in skin, of between 20% and 34%. The minimum breathing time at 150 ml min-1 to produce a significant increase in relative tumour perfusion was 4.5 min, and the effect was maintained up to 9 min. Nicotinamide alone at 1000 mg kg-1 60 min before assay did not alter relative tumour perfusion. Comparing the combination of nicotinamide with 6 min carbogen breathing at 150 ml min-1 with carbogen breathing alone showed no difference in relative tumour perfusion; increases were of 36% and 42% respectively. Nicotinamide-induced alterations in microcirculation associated with reduction of acute hypoxia have therefore not been detected by 86Rb extraction. The perfusion-enhancing effect of carbogen in this tumour is probably an important component of its radiosensitising ability, in addition to its known ability to increase the oxygen-carrying capacity of the blood, and should be taken into consideration in clinical studies.     

Vascular architecture and microenvironmental parameters in human squamous cell carcinoma xenografts: effects of carbogen and nicotinamide.

BACKGROUND AND PURPOSE: A better understanding of the vascular architecture and the microenvironmental parameters (VAMP) will allow the identification of tumours that can be more effectively treated by intensified fractionated radiotherapy or modifiers of blood flow and oxygenation or combinations of these approaches. MATERIALS AND METHODS: Proliferation (BrdUrd), vascular architecture (endothelial marker), perfusion (Hoechst 33342) and oxygenation (NITP) were studied in two human laryngeal squamous cell carcinoma tumour lines grown as xenografts in nude mice. The effects of carbogen and nicotinamide on these parameters were evaluated. RESULTS: Carbogen treatment resulted in a decrease of the number of perfused blood vessels from 66% to 55% in one of the two tumour lines. In this tumour line nicotinamide prevented this reduction of tumour blood flow by carbogen. In both tumour lines the labelling index (LI) decreased after treatment with carbogen for 1 h, from 11-13% to 5-7%. Both tumour lines showed a drastic reduction of hypoxia by carbogen alone or by carbogen plus nicotinamide. CONCLUSIONS: In both laryngeal squamous cell carcinoma xenograft tumour lines carbogen was very effective in reducing diffusion limited hypoxia. Only in one of the two tested tumour lines carbogen also caused a reduction of tumour blood perfusion, which could be compensated for by nicotinamide. In addition, carbogen reduced tumour cell proliferation. The fact that differences in response to nicotinamide and carbogen were observed and that they can be studied in vivo provides a basis for further development of a 'predictive profile' which will guide the clinician to select the optimal treatment for individual patients or groups of patients.     

The relation of ATP response in CaNT tumours after X-irradiation to varying tumour volumes

The response of adenosine-5'-triphosphate (ATP) levels in implanted murine CaNT tumours, 2.5 h after 10 Gy X-rays (100 kVp) was measured in vivo using various tumour volumes. It is probable that the concentration of ATP, which increased after X-irradiation, plays a major role in energy supply associated with cellular repair processes. Nevertheless, when ATP levels in irradiated tumours were plotted against tumour volume in the range 150-550 mm3, a monotonically decreasing relation was found. This is interpreted as reflecting the degree of hypoxia and changes in the tumour metabolic status. In addition effects of vascular occlusion and artificial hypoxia have been studied after clamping the tumour for 15 min, followed by a radiation dose of 10 Gy and maintaining the clamp for a further 2.5 h. This showed a drastic decrease in the ATP levels 2.5 h after the irradiation, compared with controls in normoxia. It is thought this is due to induction of hypoxia which is produced throughout the tumour volume, resulting in the depletion of cellular ATP. This implies that the clamped tumour cells are less capable of generating a metabolic response following radiation damage.     

ARCON: a novel biology-based approach in radiotherapy

Two mechanisms of radiotherapy resistance which are of major importance in various tumour types are tumour-cell repopulation and hypoxia. ARCON (accelerated radiotherapy with carbogen and nicotinamide) is a new therapeutic strategy that combines radiation treatment modifications, with the aim of counteracting these resistance mechanisms. To limit clonogenic repopulation during therapy, the overall duration of the radiotherapy is reduced, generally by delivering several fractions per day. This accelerated radiotherapy is combined with inhalation of hyperoxic gas to decrease diffusion-limited hypoxia, and nicotinamide, a vasoactive agent, to decrease perfusion-limited hypoxia. Preclinical studies have been done to test the enhancing effects of these three components of ARCON, individually and in combination, in several experimentally induced tumours and normal tissues. In a mouse mammary carcinoma, the tumour-control rate obtained with ARCON was the same as that with conventional treatment, but with a radiation dose almost 50% lower. Phase 1 and 2 clinical trials have shown the feasibility and tolerability of ARCON, and have produced promising results in terms of tumour control. In particular in cancers of the head and neck and bladder, the local tumour-control rates are higher than in other studies, and phase 3 trials for these tumour types are underway. In conjunction with these trials, hypoxia markers detectable by immunohistochemistry are being tested for their potential use in predictive assays to select patients for ARCON and other hypoxia-modifying therapies.     

Hydralazine-induced Changes in Tissue Perfusion and Radiation Response in A C3H Mammary Carcinoma and Mouse Normal Tissues

Hydralazine has been reported to reduce blood perfusion in tumours, thereby increasing hypoxia and subsequently enhancing tumour sensitivity to certain drugs and hyperthermia. We have investigated the ability of hydralazine to induce such changes in a C3H mouse mammary carcinoma and various normal tissues. in tumours, hydralazine (5mg/kg; i.v.) modified the radiation response, measured by a local tumour control assay, producing an effect equivalent to that seen in tumours made fully hypoxic by clamping. This effect was time-dependent and correlated with the decrease in tissue perfusion estimated by the 86-RbCl extraction procedure. Similar effects were seen in normal skin, although the changes were less dramatic and of a shorter duration. Hydralazine also reduced 86-RbCl uptake in liver, kidney, gut and spleen, but not in bladder, muscle and lung, suggesting that it may have the potential to increase the sensitivity of some normal tissues to hypoxic cell cytotoxins.