Associations between radiocurability and interstitial fluid pressure in human tumor xenografts without hypoxic tissue.

PURPOSE: The interstitial fluid pressure (IFP) of the primary tumor is an independent prognostic parameter for cervical cancer patients treated with radiation therapy. The aim of this preclinical study was to investigate whether tumor radiocurability may be associated with IFP through hypoxia-independent mechanisms. EXPERIMENTAL DESIGN: Small A-07 and R-18 melanoma xenografts without hypoxic tissue were used as preclinical tumor models. IFP was measured by using the wick-in-needle method. Radiation dose resulting in 50% local tumor control (TCD(50)), cell density, cell tumorigenicity, plating efficiency in vitro, mitotic index, fraction of Ki67-positive cells, vascular endothelial growth factor-A (VEGF-A) concentration, and radiation-induced endothelial cell apoptosis were assessed in tumors with low and high IFP. RESULTS: TCD(50) was found to be higher for tumors with high IFP than for tumors with low IFP by factors of 1.13 +/- 0.03 (A-07; P < 0.0001) and 1.10 +/- 0.03 (R-18; P < 0.0001). In the A-07 line, tumors with high IFP showed a larger number of clonogenic cells and a higher rate of cell proliferation than tumors with low IFP. In the R-18 line, tumors with high IFP showed a higher concentration of VEGF-A and a lower endothelial cell apoptotic index after irradiation than tumors with low IFP. CONCLUSIONS: The radiation resistance of normoxic tumor tissue with highly elevated IFP may be an indirect consequence of increased tumor cell clonogenicity as well as increased VEGF-A expression, possibly caused by hypertension-induced modifications of signaling pathways regulating cell proliferation, cell survival, and/or angiogenesis.

Assessment of extravascular extracellular space fraction in human melanoma xenografts by DCE-MRI and kinetic modeling.

Tumor aggressiveness and response to therapy are influenced by the extravascular extracellular space fraction (EESF) of the malignant tissue. The EESF may, therefore, be an important prognostic parameter for cancer patients. The aim of this study was to investigate whether gadopentetate dimeglumine (Gd-DTPA)-based dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) can be used to assess the EESF of tumors. Amelanotic human melanoma xenografts (A-07, R-18) were used as preclinical models of human cancer. Images of E.F (E is the initial extraction fraction of Gd-DTPA and F is perfusion) and lambda (the partition coefficient of Gd-DTPA) were obtained by Kety analysis of DCE-MRI data. Our study was based on the hypothesis that lambda is governed by the EESF and is not influenced significantly by microvascular density (MVD) or blood perfusion. To test this hypothesis, we searched for correlations between lambda and E.F, MVD or EESF by comparing lambda images with E.F images, histological preparations from the imaged tissue and the radial heterogeneity in EESF obtained by invasive imaging. Positive correlations were found between lambda and EESF. Thus, median lambda was larger in A-07 tumors than in R-18 tumors by a factor of 4.2 (P<.00001), consistent with the histological observation that EESF is approximately fourfold larger in A-07 tumors than in R-18 tumors. The radial heterogeneity in lambda in A-07 and R-18 tumors was almost identical to the radial heterogeneity in EESF. Moreover, lambda was larger in tissue regions with high EESF than in tissue regions with low EESF in A-07 tumors (P=.048). On the other hand, significant correlations between lambda and MVD or E.F could not be detected. Consequently, Kety analysis of Gd-DTPA-based DCE-MRI series of xenografted tumors provides lambda images that primarily reflect the EESF of the tissue.

Dynamic contrast-enhanced magnetic resonance imaging of human melanoma xenografts with necrotic regions.

PURPOSE: To investigate whether high-resolution images of necrotic regions in tumors can be derived from gadopentetate dimeglumine (Gd-DTPA)-based dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) series. MATERIALS AND METHODS: E-13 human melanoma xenografts were used as preclinical models of human cancer. DCE-MRI was performed at a voxel size of 0.23 x 0.47 x 2.0 mm3 with the use of spoiled gradient recalled sequences. Tumor images of E . F (E is the initial extraction fraction of Gd-DTPA and F is blood perfusion) and lambda (the partition coefficient of Gd-DTPA, which is proportional to extracellular volume fraction) were produced by subjecting DCE-MRI series to Kety analysis, and these images were compared with histological preparations from the imaged slices. RESULTS: Strong correlations were found between fraction of necrotic tissue and fraction of voxels with lambda > lambdaL for lambdaL values of 0.4 to 0.6. Binary lambda images differentiating between lambda values > lambdaL and lambda values < lambdaL were found to mirror necrotic regions well in tumors with large necroses. However, necrotic foci that were small compared with the voxel size were not detectable. CONCLUSION: Clinically relevant images of necrotic tumor regions can be obtained for E-13 melanomas by subjecting Gd-DTPA-based DCE-MRI series to Kety analysis.

Fluctuating and diffusion-limited hypoxia in hypoxia-induced metastasis.

PURPOSE: Most tumors develop regions with hypoxic cells during growth, owing to permanent limitations in oxygen diffusion (chronic or diffusion-limited hypoxia) and/or transient limitations in blood perfusion (acute or fluctuating hypoxia). The aim of this study was to investigate the relative significance of chronic and acute hypoxia in the development of metastatic disease. EXPERIMENTAL DESIGN: D-12 and R-18 human melanoma xenografts were used as models of human cancer. D-12 tumors metastasize to the lungs, whereas R-18 tumors develop lymph node metastases. Fraction of radiobiologically hypoxic cells (HF(Rad)) was measured in individual primary tumors by using a radiobiological assay based on the paired survival curve method. Fraction of immunohistochemically hypoxic cells (HF(Imm)) was assessed in the same tumors by using a pimonidazole-based immunohistochemical assay optimized with respect to achieving selective staining of chronically hypoxic cells. HF(Imm) and the difference between HF(Rad) and HF(Imm), HF(Rad) - HF(Imm), were verified to be adequate variables for fraction of chronically hypoxic cells and fraction of acutely hypoxic cells, respectively. RESULTS: Chronic as well as acute hypoxia were found to promote spontaneous metastasis of D-12 and R-18 tumors. Acute hypoxia influenced metastasis to a greater extent than chronic hypoxia, partly because the fraction of acutely hypoxic cells was larger than the fraction of chronically hypoxic cells in most tumors and partly because acutely hypoxic cells showed a higher metastatic potential than chronically hypoxic cells. CONCLUSIONS: It may be beneficial to focus on fluctuating hypoxia rather than diffusion-limited hypoxia when searching for hypoxia-related prognostic variables and predictive assays.

Fluctuations in pO2 in irradiated human melanoma xenografts.

Several studies have demonstrated that untreated tumors may show significant fluctuations in tissue oxygen tension (pO(2)). Radiation treatment may induce changes in the tumor microenvironment that alter the pO(2) fluctuation pattern. The purpose of the present study was to investigate whether pO(2) fluctuations may also occur in irradiated tumors. A-07 human melanoma xenografts were irradiated with single doses of 0, 5 or 10 Gy. Fluctuations in pO(2) were recorded with OxyLite probes prior to irradiation and 24 and 72 h after the radiation exposure. Radiation-induced changes in the tumor microenvironment (i.e. blood perfusion and extracellular volume fraction) were assessed by dynamic contrast-enhanced magnetic resonance imaging. Seventy-two hours after 10 Gy, tumor blood perfusion had decreased to approximately 40% of that prior to irradiation, whereas the extracellular volume fraction had increased by approximately 25%. Fluctuations in pO(2) were seen in most tumors, irrespective of radiation dose and time after irradiation. The mean pO(2), the number of fluctuations around the mean pO(2), the number of fluctuations around threshold pO(2) values of 1, 2, 3, 5, 7 and 10 mmHg, and the amplitude of the fluctuations were determined for each pO(2) trace. No significant differences were detected between irradiated and unirradiated tumors. The results showed that pO(2) fluctuations may occur in irradiated tumors and that the pO(2) fluctuation pattern in A-07 tumors exposed to 5 or 10 Gy is similar to that in untreated tumors. Consequently, these doses did not induce changes in the tumor microenvironment that were sufficient to cause detectable alterations in the pO(2) fluctuation pattern.

Intratumor heterogeneity in blood perfusion in orthotopic human melanoma xenografts assessed by dynamic contrast-enhanced magnetic resonance imaging.

PURPOSE: To determine the intratumor heterogeneity in blood perfusion of orthotopic human melanoma xenografts by use of gadopentetate dimeglumine (Gd-DTPA)-based dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). MATERIALS AND METHODS: Orthotopic xenografts of an amelanotic human melanoma cell line (A-07) were scanned sagittally, coronally, and axially in three subsequent DCE-MRI sessions, using spoiled gradient recalled sequences, a voxel size of 0.31x0.62x2.0 mm3, and an interleaving acquisition method to avoid slice gaps. Tumor images of E . F (E is initial extraction fraction and F is perfusion) were produced by subjecting the DCE-MRI data to Kety analysis. E . F was used as a parameter for tumor blood perfusion, since E for Gd-DTPA is close to unity in A-07 tumors. RESULTS: All A-07 tumors subjected to investigation showed anisotropic radial heterogeneity in blood perfusion. The blood perfusion was low in the center of the tumors and increased toward the tumor periphery in the cranial, dorsal, caudal, and ventral directions, but not in the lateral and medial directions. In addition, 9 of 10 tumors showed blood perfusion hot spots in central or nonperipheral regions. The hot spots differed significantly between tumors in size, shape, location, and intensity, and appeared to be governed by stochastic processes. This heterogeneity superimposed the radial heterogeneity, but did not overshadow it in any tumor. CONCLUSION: Orthotopic human melanoma xenografts show significant intratumor heterogeneity in blood perfusion. This heterogeneity is made up of two distinctly different components, one stochastic and one nonstochastic radial component. The radial component is anisotropic and dominant and is superimposed by the stochastic component.

Comparison of tumor blood perfusion assessed by dynamic contrast-enhanced MRI with tumor blood supply assessed by invasive imaging.

PURPOSE: To evaluate the potential of Gd-DTPA-based dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) for providing high-resolution tumor blood perfusion images. MATERIALS AND METHODS: Xenografted tumors from two amelanotic human melanoma lines (A-07 and R-18) were used as preclinical models of human cancer. DCE-MRI was performed at a voxel size of 0.5 x 0.2 x 2.0 mm(3) with the use of spoiled gradient recalled sequences. We produced tumor images of E . F (where E is the initial extraction fraction, and F is perfusion) by subjecting the DCE-MRI data to Kety analysis, and then compared those images with images of tumor blood supply. We obtained high-resolution tumor blood supply images using the Bioscope silicon strip detector system to measure the uptake of Na(99m)TcO(4) in histological preparations. We assessed the global blood supply by measuring the tumor uptake of three freely diffusible blood flow tracers: (86)RbCl, [(14)C]IAP, and Na(99m)TcO(4). RESULTS: E . F was found to mirror the blood supply well in A-07 and R-18 tumors. The mean E . F differed between the A-07 and R-18 tumors by a factor of approximately 1.6, and this difference was similar to the difference in the global blood supply. The intratumor heterogeneity in E . F was significant for tumors of both lines, and this heterogeneity was similar to the intratumor heterogeneity in the blood supply. The intratumor heterogeneity in the blood supply differed slightly between the A-07 and R-18 tumors, and even this difference was mirrored by the E . F images. CONCLUSION: E . F images of xenografted tumors reflect blood perfusion. This implies that E . F may be a useful parameter for improving cancer diagnostics and individualizing cancer treatment. This possibility deserves to be investigated thoroughly in clinical studies.