Blood flow and metabolic microenvironment of brain tumors

Journalof Neuro-Oncology 22: 261-267,1994. 9 1994KluwerAcademic Publ&hers.Printedin the Netherlands. Blood flow and metabolic microenvironment of brain tumors P. Vaupel Department of Physiology and Pathophysiology, Pathophysiology Division, University of Mainz, Duesbergweg 6, D-55099 Mainz, Germany Key words: metabolic microenvironment, blood flow, oxygenation, pH, energy status Introduction Tumor growth and response to nonsurgical treatment modalities, e.g., radio- or chemotherapy, are not only determined by intrinsic or genetic properties of malignant cells, but also by extrinsic epigenetic factors such as the internal milieu ('metabolic microenvironment'). This microenvironment is determined by the distribution of oxygen partial pressures, glucose, lactic acid, and ATP concentrations and by pH. It is critically associated with the efficacy of tumor blood flow including (i) nutritive flow (the flow that is in contact with the cells, 'tissue perfusion') and (ii) shunt flow (blood flowing directly from the arterial to the venous side). Despite the apparent importance of the cellular microenvironment for tumor growth, for early tumor response to treatment, and probably for prediction of long term outcome, reliable data on solid tumors in humans are scarce, although the number of clinical investigations dealing with this subject is rapidly increasing. In the following, it is attempted (i) to summarize the current information concerning blood supply of primary and metastatic brain tumors and flow-related physiological properties, and (ii) to demonstrate that inadequacy and heterogeneous distribution of tumor microcirculation can be a causative factor for the known resistance to radio-/ chemotherapy. Blood flow Blood flow through brain tumors is anything but uniform. Most tumors contain both highly perfused areas which are rapidly growing, as welt as regions with compromised and sluggish perfusion, often associated with the development of necrosis. From experimental brain tumor studies in the rat the following conclusions can be drawn [1-12]: In experimental gliomas there is a tremendous scattering of regional blood flow (tissue perfusion), ranging from 0.1 to 3.8 ml/g/min (Fig. 1). Flow values can be lower than those measured in the normal white matter, or can exceed those of the normal cerebral cortex. When glioma cell lines are implanted subcutaneously, scattering is reduced which emphasizes the great impact of the growth site in experimental systems [6, 7, 9]. Human gliomas [11] and medulloblastomas [12] xenografted into rat exhibited flow values ranging between 0.05 and 1.0 ml/g/min. As a model of metastasis, intracerebral implantation of Walker 256 tumors yielded flow values ranging from 0.15 to 1.75 ml/g/min [1, 4, 8]. These flow data are comparable to those PET-derived values [1319] measured in primary brain tumors or metastatic lesions in patients (Table 1). Also, no differences are observed between brain tumors and other tumor entities outside the brain. In experimental gliomas, blood flow correlated only poorly with morphological and histological features of the tumors (necrosis, cellularity, cytology, location and size). Besides pronounced inter-tumor variability, marked regional heterogeneity was typical for the gliomas investigated [5]. The currently available data can be summarized as follows: 262 Table 1. Blood flow in normal human brain and through primary and metastatic brain lesions (pooled data) Tissue i I ....... i --'~ whitematter [ i :enograffs 0 0.5 1 1.5 2 2.5 regional blood flow (ml.g -1.min -t) Fig. 1. Regional blood flowof intracerebrallyor subcutaneously implanted experimental gliomas, of intracerebrally growing Walker 256 tumors, and of human brain tumor xenograftsin the rat (gray bars). Data are compared with the respectiveflowvalues in the normal rat cortex and white matter (white bars). 1) Blood flow can considerably vary despite similar histological classification. 2) Experimental systems, primary and metastatic brain tumors show flow rates which can be lower than values in the white matter, or can exceed flow rates of the normal cortex. 3) Metastatic lesions show flow rates which are comparable to those of primary brain tumors. 4) Blood flow measured at multiple sites within one tumor shows marked heterogeneity. There is also great inter-tumor variability. 5) Flow in brain tumors does not deviate substantially from that of other tumor entities and may not correlate with tumor grade. Gliomas of different grades show similar mean flow values and great inter-tumor variability within one grade. Oxygenation and oxygen consumption Tissue oxygenation as characterized by the distribution of 0 2 partial pressures (pO2) within a given tissue is the result of oxygen availability and 0 2 consumption of the tumor cells. In normal brain, oxygen supply meets the requirements of the cells, which leads to an adequate tissue oxygenation. In brain tumors, hypoxic (low pO2) tissue areas are found, which can be heterogeneously distributed within the tumor and which are thought to result Blood flow (ml/g/min) Refs. Gliomas Grade I Grade II Grade III Grade IV Brain metastases 0.03-0.43 0.03-1.01 0.26-0.98 0.15-1.02 0.03-0.72 [15, 17, 19] Normal gray matter Normal white matter 0.254).78 0.08-0.33 [141 [15-18] from compromised microcirculation. Mean oxygen tension obtained in experimental brain tumors of the rat (subcutaneously growing gliosarcomas [8]) and in primary human tumors [20] is on average distinctly lower than in normal brain tissue (Figs. 2 and 3). pO 2 values lower than 5 mmHg are considered to be 'hypoxic' in terms of radiation sensitivity. In human glioblastomas, there is no correlation between tumor size and median pO 2 values. Together with our studies on breast and cervix cancers [22, 23], there is a clear indication that median pO 2 values in tumors are distinctly lower than in normal tissue, and that the oxygenation status of individual tumors before therapy cannot be predicted on the basis of tumor size. The lack of predictability is predominantly due to pronounced inter-tumor variability even if tumors of the same pathological stage are compared. Inter-tumor variability is more pronounced than intra-tumor heterogeneity. Oxygen consumption rates of brain tumors exhibit a broad variability, ranging from 2-36 gl/g/min (Fig. 4). There is ample evidence to suggest that the oxygen extraction is not impaired and that the respiratory function of cancer cells is not deficient (Table 2). When considering oxygen extraction data derived from tSO2-PET studies using the 0 2 steady state technique, an underestimation of the mean values has to be considered on the basis of computer simulation studies mimicking tissue heterogeneity [26]. To date published P E T studies may thus not be accurate enough for the assessment of oxygen extraction in tumors. 263 50. 25_ 20 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . normal human brain 40 ............................................ median pO2 = 24 mmHg cerebral cortex 30 o~ v >, 0 . . . . . . . 10 20 30 40 50 60 70 80 90 100 O" 25 20i . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9L gliosarcoma 15e ................................... 5~ 40 1 . . 30 (rat' S'c')". . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 20 30 40 50 60 70 80 90 Fig. 2. Frequency distribution of measured oxygen partial pressures (pO 2 histograms) of the normal rat cerebral cortex (upper panel, ref. [21]) and of an experimental (s.c. growing) rat gliosarcoma (9L tumor, lower panel, ref. [8]). Heart (strenuous exercise) . . . . 300 200 ,oo ~ ~ ---- Skeletal Muscle (strenuous exercise) < 1~ ~ m . . . . . Heart (at rest) - - - . . . . Kidney Liver . . . 10 . 20 30 40 50 60 . . "l . 70 80 90 !00 glioblastomas median pO2 = 7 mmHg . . . . . 30 . . 40 . . . 50 . . . . .. . . 60 . , , , . 70 . . . . 80 . . 90 100 tissue pO2 (mmHg) 100 tissue P02 (mmHg) 40O 20 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 ~o . ......................... . ~ . 10 2012 L 70 . 1r 0 O~ " g -d 0 . 20 ..................................................................... ~176 10s~ ....... _ [ 1.,- . Fig. 3. pO 2 histograms of the normal human brain (upper panel, ref. [14]), and of human glioblastomas (lower panel, ref. [20]). Glucose consumption Over 60 years ago, Warburg [27] studied the glucose turnover in tumor slices incubated aerobically and found high rates of lactic acid production. From these and other experiments, an attitude regarding the high rate of aerobic glycolysis and lactate release became a biochemical 'hallmark' of malignancies. Since that time, more detailed insight into this aspect of tumor energy metabolism has been obtained. Positron emission tomography (PET) and the tracers 18F-2-deoxyglucose (FDG) and 'C-2glucose have been used to measure regional glucose utilization in brain tumors and normal brain in vivo ----Brain 30 ;Bo .2 I Ce[olid Body Spleen lo - - Skelelal Muscle (resting) 2 Fig. 4. Range of oxygen consumption rates (Voz) of human gliomas and brain metastases (black bars), and of normal human gray and white matter (white bars). 02 consumption rates of normal tissues are given for comparison (horizontal lines). Values presented are pooled consumption rates obtained with the 1502 inhalation technique and PET (for a review see ref. [14]). T a b l e 2. Oxygen utilization (02 extraction ratio, 02 extraction fraction) of human brain tumors and normal brain tissue (data derived from PET studies, ~502) Tissue 02 utilization (v/v) Refs. Astrocytomas Brain metastases 0.03-0.52 0.17-0.55 [15-17, 19] Normal brain Whole Gray matter White matter 0.35-0.47 0.30-0.68 0.37-0.67 [24] [16] [15, 17, 19, 25] [15-171 264 Table 3. Glucose uptake rates ('Qg~)of brain tumors and of normal brain tissue (for review see [14]) Tissue 9g, (gmol/g/min) Refs. Gliomas Grades I-II Grades III-IV Whole brain Gray matter White matter Brain stem 0.10-0.93" 0.17-0.88" 0.30-0.40* 0.25-0.51" 0.08-0.28* 0.18-0.28" [28, 29, 36] [19, 30, 36] [31] [32] [33] [34] Exptl. gliomas Exptl. metastases (Walker 256) 0.09-2.11 + 0.34-1.70 + Gray matter White matter 0.30-1.39 + 0.07-0.43 + * H u m a n PET data, § rat data [1, 2, 48, 64]. Exptl. = experimental. (Table 3). Di Chiro etal. [28] found a strong correlation between the glucose consumption rate and the grade of gliomas, with visual 'hot spots' present in all high grade tumors (WHO III and IV) and only in a few cases of low grade gliomas (WHO I and II). In grade I/II gliomas, the glucose consumption rate was on average 0.21 _+0.10 gmol/g/min compared to 0.30 + 0.15 Bmol/g/min in grade II! tumors, and 0.41 + 0.20 gmol/g/min in grade IV tumors. In addition, a depression of glucose consumption in peritumoral regions was found. Meningiomas were also studied by this group [35], where a correlation between metabolic rates of glucose and the tumor growth rate (determined by repeated CT scans) was demonstrated. From these studies, it was concluded that the utilization rate of glucose appears to be at least as reliable as the histological classification of tumors. The differentiation of radiation induced necrosis and glioma recurrence and the detection of meningioma recurrence [30, 35] is also possible using PET and FDG. However, the above mentioned 'strong' correlation between FDG uptake and glioma grade has not been confirmed by other groups [36-38]. Comparing FDG-PET with magnetic resonance spectroscopy (MRS), a correlation between maximum glucose consumption rate and maximum lactate accumulation was demonstrated in human gliomas [36]. pH of brain tumors Besides glucose oxidation, cancer cells intensively metabolize glucose to lactic acid. Since this can lead to acidosis, tumors were thought to have a more acidic intracellular pH (pHi) than most normal tissues. In addition, ATP hydrolysis and glutaminolysis have been discussed as catabolic pathways contributing to tumor acidosis. Using non-invasive techniques like MRS or PET with the tracer 11CDMO [14], studies on human and experimental tumors have shown that pH i is alkaline rather than acidic [14, 39]. Acidic values were only found in bulky, poorly oxygenated, energy- and nutrient-deprived tumors [13]. In experimental brain tumors of the rat and in human brain tumors, pHi values are slightly higher than in normal brain (Fig. 5 [40-51]). Similar data were obtained for pH i of sarcomas when compared with the resting skeletal muscle [14]. In contrast, the extracellular pH (pile) in normal human brain is slightly more alkaline than pHi, whereas in brain tumors pHo is on average more acidic compared to pHi. This difference (0.3-0.5 pH units) has been obtained by electrode measurements in solid tumors [14, 39]. The gradient in proton concentration between 7.5- 7,5- 0 rat brain 9 brain tumors 7.4- 7.4- 7.3- 7.3" 7.2- 7.2" 7.1 7.1- 7.0 7.0- 6.9- 6.9" 6.8" 6.8 6.7. 6.7' I 6.6 pHi pHe 6.6 0 human brain 9 brain tumors \ i pHi pHe Fig. 5. Intracellular (pHi) and extracellular pH (pile) values in normal brain (rat, human), in experimental rat and primary human tumors. Tumor pH~ is higher than that of the tumor extracellular fluid (pHi). This is in contrast to normal tissues (brain, liver, skeletal muscle) in which pH~ is lower than pHe. Values presented are means and the range of pH data measured. 265 intra- and extracellular space in tumors is most probably due to the fact that t u m o r cells, like normal cells, have efficient mechanisms for intracellular p H homeostasis. These mechanisms include a rapid export of anionic lactate- which is accompanied by a m o v e m e n t of protons. The subsequent inadequate removal of protons from the enlarged extracellular space (in tumors, the extracellular space can be enlarged 4-5 fold) due to an inadequate microcirculatory function in m a n y tumors, or at least in some t u m o r areas, may result in an extracellular acidosis. Export of protons into the extracellular space and the resulting proton gradient between the intra- and extracellular space have impact on interrelated ionic gradients. As a result, e.g., the Na + concentration is usually higher in tumor cells than in their normal counterparts, and the K § concentration is distinctly lower [39], which can result in changes of the m e m b r a n e potential. F r o m these data, it can be concluded that on average the bioenergetic stages may be similar in normal brain and in brain tumors. Conclusions Summarizing these in vivo data in the context of brain tumor therapy, the following aspects are of particular importance: Low and heterogeneous tum o r blood flow m a y - in addition to the limiting effects of the blood-brain barrier - result in compromised delivery of drugs from blood to the tissue. Low tumor pO 2 reduces sensitivity to standard radiation and 'OR-dependent' anticancer drugs. Treatm e n t efficacy m a y be further altered by changes of tumor pH. Particularly acidosis can decrease radiation sensitivity and modulate the cytotoxicity of anticancer drugs. In the following presentations, these aspects will be discussed regarding in vivo data obtained with positron emission tomography. Bioenergetic status The m e a s u r e m e n t of regional A T P distributions with quantitative bioluminescence showed that A T P levels in experimental brain tumors are similar to normal brain [52] with slightly lower glucose and substantially higher lactate concentrations in viable tumor tissue. All metabolites showed m a r k e d heterogeneity concerning their regional distribution. In m a n y h u m a n malignancies other than brain tumors high concentrations of p h o s p h o m o n o e s t e r s (precursors of m e m b r a n e lipids), phosphodiesters (metabolites), inorganic phosphate (Pi) and low phospho-creatine (PCr) levels have been reported using 31P-MRS. In contrast, spectra of h u m a n brain tumors often fail to show any significant differences c o m p a r e d to normal brain. The ratios of PCr/P i can be used for characterization of the bioenergetic status of cells. 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