Debra K. Kelleher

Oxygenation status of malignant tumors: Pathogenesis of hypoxia and significance for tumor therapy

Hypoxic areas are a characteristic property of solid tumors. Hypoxia results from an imbalance between the supply and consumption of oxygen. Major pathogenetic mechanisms for the emergence of hypoxia are (1) structural and functional abnormalities in the tumor microvasculature; (2) an increase in diffusion distances; and (3) tumor- or therapy-associated anemia leading to a reduced O2 transport capacity of the blood. There is pronounced intertumor variability in the extent of hypoxia, which is independent of clinical size, stage, histopathologic type, and grade. Local recurrences have a higher hypoxic fraction than primary tumors. Tumor hypoxia is intensified in anemic patients, especially in tumors with low perfusion rates. Tumor hypoxia is a therapeutic problem, as it makes solid tumors resistant to sparsely ionizing radiation and some forms of chemotherapy. Hypoxia also may modulate the proliferation and cell cycle position of tumor cells and, in turn, the amount of cells destroyed following therapy. Recent clinical studies suggest that hypoxia can enhance malignant progression and increase aggressiveness through clonal selection and genome changes. As a result, loss of differentiation and apoptosis, chaotic angiogenesis, increased locoregional spread, and enhanced metastasis can further increase resistance to therapy and affect long-term prognosis. Hypoxia is a powerful, independent prognostic factor in cervix cancers, carcinomas of the head and neck, and in soft-tissue sarcomas.

Enhanced radiosensitivity in experimental tumours following erythropoietin treatment of chemotherapy-induced anaemia.

The radiosensitivity of solid tumours in anaemic rats treated with recombinant human erythropoietin (rhEPO, epoetin beta) was studied. Anaemia was induced by a single dose of carboplatin (45 mg kg(-1) i.v.), resulting in a reduction in the haemoglobin concentration by 30%. In a second group, the development of anaemia was prevented by rhEPO (1000 IU kg(-1)) administered s.c. three times per week starting 6 days before the carboplatin application. Three days after carboplatin treatment, DS-sarcomas were implanted subcutaneously onto the hind foot dorsum. Neither carboplatin nor rhEPO treatment influenced tumour growth rate. Five days after implantation, tumours were irradiated with a single non-curative dose (10 Gy), resulting in a growth delay with a subsequent regrowth of the tumours. In the rhEPO-treated group, the growth delay lasted significantly longer (9.5 days vs. 4.5 days) and the regrowth was slower (6.0 days vs. 4.1 days) compared with the anaemic group. These data suggest that the correction of chemotherapy-induced anaemia by rhEPO (epoetin beta) treatment increases tumour radiosensitivity, presumably as a result of an improved oxygen supply to tumour tissue.

Modulation of tumor oxygenation

There is a large body of evidence suggesting that deficiencies in the O2 supply of tumors exist due to restrictions (i) in the O2 delivery by perfusion and/or diffusion, and (ii) in the O2 transport capacity. Whereas the former are mostly based on inadequate and heterogeneous microcirculatory functions, the latter are predominantly due to tumor-associated anemia. Possible uses and limitations of measures are discussed which can increase the microvascular O2 content and thus may preferentially serve to enhance diffusion-limited O2 availability. In addition, means are described for improving and increasing the uniformity of microcirculation thus possibly enhancing perfusion-limited O2 delivery. Reducing cellular respiration rate should be of benefit in both pathophysiological conditions. Because both types of O2 limitation coexist in solid tumors, appropriate combinations should be aimed at eradicating tumor hypoxia which is present in at least one third of cancers in the clinical setting.

Pathophysiological and vascular characteristics of tumours and their importance for hyperthermia: Heterogeneity is the key issue

Tumour blood flow before and during clinically relevant mild hyperthermia exhibits pronounced heterogeneity. Flow changes upon heating are not predictable and are both spatially and temporally highly variable. Flow increases may result in improved heat dissipation to the extent that therapeutically relevant tissue temperatures may not be achieved. This holds especially true for tumours or tumour regions in which flow rates are substantially higher than in the surrounding normal tissues. Changes in tumour oxygenation tend to reflect alterations in blood flow upon hyperthermia. An initial improvement in the oxygenation status, followed by a return to baseline levels (or even a drop to below baseline at high thermal doses) has been reported for some tumours, whereas a predictable and universal occurrence of sustained increases in O2 tensions upon mild hyperthermia is questionable and still needs to be verified in the clinical setting. Clarification of the pathogenetic mechanisms behind possible sustained increases is mandatory. High-dose hyperthermia leads to a decrease in the extracellular and intracellular pH and a deterioration of the energy status, both of which are known to be parameters capable of acting as direct sensitisers and thus pivotal factors in hyperthermia treatment. The role of the tumour microcirculatory function, hypoxia, acidosis and energy status is complex and is further complicated by a pronounced heterogeneity. These latter aspects require additional critical evaluation in clinically relevant tumour models in order for their impact on the response to heat to be clarified.

Tumor oxygenation in anemic rats : effects of erythropoietin treatment versus red blood cell transfusion

Anemia was induced in rats by the development of a hemorrhagic ascites. These animals also bore solid tumors (DS-sarcomas) on the hind foot dorsum. The effects of two methods for anemia correction on oxygenation in the solid tumors were compared in this study. Anemia was corrected either chronically by erythropoietin administration (1000 IU/kg) over 14 days (EPO) or acutely by transfusion with red blood cells (TR). Non-anemic and untreated anemic animals served as controls. Tumor oxygenation was determined in anesthetized animals using polarographic needle electrodes and pO2 histography. The reduction in hematocrit and hemoglobin content found in anemic animals could successfully be corrected either by EPO or by TR. Anemia resulted in a worsening of tumor oxygenation which could partially be reversed by EPO or TR in small tumors (< 1.4 ml). In larger tumors (> or = 1.4 ml), neither method of anemia correction resulted in significant changes in tumor oxygenation.

Changes in microregional perfusion, oxygenation, ATP and lactate distribution in subcutaneous rat tumours upon water-filtered IR-A hyperthermia

The effect of hyperthermia on microcirculatory and metabolic parameters in s.c. DS-sarcomas of different sizes on the hind foot dorsum of SD-rats was investigated. Hyperthermia was carried out using a novel water-filtered, infrared-A radiation technique. Heating was performed at a rate of 0.5 degrees C/min until 44 degrees C was achieved in the tumour centre, which was maintained for 60 min. Using a multichannel laser Doppler flowmeter, red blood cell flux could be assessed continuously and at several sites within the tumour tissue simultaneously. Substantial inter-site variations in laser Doppler flux (LDF) were observed during hyperthermia which were independent of tumour size, site of measurement, and temperature at the site of measurement, indicating that single site measurements of tumour LDF are poor predictors of the mean response of a tumour to hyperthermia. When mean LDF was considered, decreases in red blood cell fluxes occurred that were more pronounced the greater the tumour volume. In no case was vascular stasis observed. Hyperthermia did not affect tumour oxygenation substantially. Microregional and global assessment of lactate and ATP concentrations demonstrated increased lactate and decreased ATP levels following hyperthermia. Tumour glucose levels were increased following hyperthermia, possibly due to an enlarged distribution space resulting from development of interstitial oedema. Changes in lactate and ATP levels and the lack of changes in tumour oxygenation suggest a modification of energy metabolism following hyperthermia in the form of increased ATP hydrolysis, intensified glycolysis and impaired oxidative phosphorylation.

Effects of Infrared-A Irradiation on Skin: Discrepancies in Published Data Highlight the Need for an Exact Consideration of Physical and Photobiological Laws and Appropriate Experimental Settings

Skin exposure to infrared (IR) radiation should be limited in terms of irradiance, exposure time and frequency in order to avoid acute or chronic damage. Recommendations aimed at protecting humans from the risks of skin exposure to IR (e.g. ICNIRP, ACGIH) are only defined in terms of acute effects (e.g. heat pain and cardiovascular collapse), whereas the actual exposure conditions (e.g. spectral distribution, exposure geometry, frequency and number of exposures, thermal exchange with the environment, metabolic energy production and regulatory responses) are not taken into consideration. Since the IR component of solar radiation reaching the Earth’s surface is mainly IR‐A, and considering the increased use of devices emitting artificially generated IR‐A radiation, this radiation band is of special interest. A number of in vitro and/or in vivo investigations assessing cellular or tissue damage caused by IR‐A radiation have been undertaken. While such studies are necessary for the development of safety recommendations, the results of measurements undertaken to examine the interaction between skin and IR radiation emitted from different sources presented in this study, together with the detailed examination of the literature reveals a wide spectrum of contradictory findings, which in some instances may be related to methodological shortcomings or fundamental errors in the application of physical and photobiological laws, thus highlighting the need for physically and photobiologically appropriate experiments.

Water-filtered infrared-A radiation: a novel technique for localized hyperthermia in combination with bacteriochlorophyll-based photodynamic therapy.

A novel application of an infrared-A (IR-A) radiation source equipped with a water-filter in the radiation path is described, which allows for tumour treatment with a simultaneous combination of localized hyperthermia (HT) and bacteriochlorophyll-serine (Bchl-ser) based photodynamic therapy (PDT). Using this system, the IR-A radiation was used to heat tumours to 43 degrees C for 60 min, while at the same time activating the Bchl-ser which was injected i.v. at a dose of 20 mg/kg, 10 min following commencement of HT. The growth of tumours undergoing this combined therapy was compared to that of tumours undergoing HT alone or sham-treated controls. Within the 90 day observation period, 100% of tumours in sham-treated animals, 80% in HT-treated animals and only 17% in HT + Bchlser-treated animals reached the end point target volume of 3.5 ml. Thus, the tumour growth inhibition effect of HT can be substantially enhanced by combination with Bchl-ser-PDT. This novel technique has proved to be well-tolerated, easy to apply and should be suitable for treatment of superficial malignancies, especially where hypoxic tumour areas are present.

Nicotinamide exerts different acute effects on microcirculatory function and tissue oxygenation in rat tumors

PURPOSE Nicotinamide has been reported to preferentially radiosensitize tumor tissue, supposedly through a reduction in tumor hypoxia. This may occur as a result of nicotinamide-induced changes in tumor blood flow and therefore the present study was undertaken to evaluate the effect of nicotinamide on circulatory parameters in skeletal muscle and tumor tissue (subcutaneously-implanted DS-sarcomas) of the rat. METHODS AND MATERIALS Mean arterial blood pressure (measured in the common carotid artery using a pressure transducer) and red blood cell flux (as measured by laser Doppler flowmetry) were continuously monitored for 120 min following a single intraperitoneal application of nicotinamide (500 mg/kg). An arterial blood pressure/laser Doppler flux ratio was estimated for tumor and muscle tissue. RESULTS Nicotinamide significantly reduced the mean arterial blood pressure to a minimum value 25% below the pretreatment value 20 min after the commencement of drug administration, with partial recovery thereafter. Red blood cell flux through tumor tissue, following an initial rapid decrease, rose steadily to values 34% above those measured in control animals at t = 60 min, while the arterial blood pressure/laser Doppler flux ratio in tumor tissue fell to values 34% below those of control animals. In skeletal muscle similar trends were seen although the changes were not of the same extent as those seen in tumor tissue. Tumor pO2 was measured 60 min following i.p. application of nicotinamide using polarographic needle electrodes. Despite the significant increase in blood flow following nicotinamide, no significant difference was seen between pO2 histograms obtained in tumors in nicotinamide-treated and control animals. CONCLUSION These findings suggest that nicotinamide preferentially improves tumor microcirculatory function and effectuates a decrease in the arterial blood pressure/laser Doppler flux ratio within tumor tissue, effects which reach their maximum approximately 60 min following nicotinamide administration.

Dynamics of tumor oxygenation and red blood cell flux in response to inspiratory hyperoxia combined with different levels of inspiratory hypercapnia

BACKGROUND AND PURPOSE Increasing arterial oxygen partial pressure (pO2) by breathing hyperoxic gases is an effective means of improving tumor oxygenation, although the efficacy of adding CO2 to the inspiratory gas has been discussed controversially. This study aimed at analyzing the impact of different inspiratory CO2 fractions on the time course of oxygenation and perfusion changes in experimental tumors during and after inspiratory hyperoxia. MATERIAL AND METHODS Perfusion and oxygenation of rat DS-sarcomas were studied during spontaneous breathing of pure oxygen or hyperoxic gas mixtures containing different CO2 fractions (1, 2.5 or 5%). Red blood cell (RBC) flux was assessed as a measure of tumor perfusion using the laser Doppler technique and temporal changes in mean tumor pO2 were measured polarographically. RESULTS Mean tumor pO2 increased 3.6-fold with pure oxygen, approx. 3.3-fold when 1 or 2.5% CO2 was added and 2.7-fold during carbogen breathing. RBC flux also increased by 25-30% with all gases. With pure oxygen and with 1% CO2 (+99% O2), perfusion changes paralleled those of the mean arterial blood pressure whereas with higher CO2 fractions, a decrease in resistance to flow was observed. The differences found with the various gas mixtures were more pronounced after the end of hyperoxia. With pure oxygen, perfusion immediately returned to pretreatment values whereas with higher CO2 fractions perfusion remained elevated for at least 30 min. CONCLUSIONS Higher inspiratory CO2 fractions (2.5 or 5%) lead to a prolonged improvement of tumor perfusion after the end of inspiratory hyperoxia when compared with pure oxygen breathing. Since no principal differences in oxygenation and perfusion were seen between the gases containing 2.5 and 5% CO2, the former may be preferable for inspiratory hyperoxia.