DCE-MRI and Quantitative Histology Reveal Enhanced Vessel Maturation but Impaired Perfusion and Increased Hypoxia in Bevacizumab-Treated Cervical Carcinoma.

PURPOSE: This study had a dual purpose: to investigate (1) whether bevacizumab can change the microvasculature and oxygenation of cervical carcinomas and (2) whether any changes can be detected with dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). METHODS AND MATERIALS: Two patient-derived xenograft models of cervical cancer (BK-12 and HL-16) were included in the study. Immunostained histologic preparations from untreated and bevacizumab-treated tumors were analyzed with respect to microvascular density, vessel pericyte coverage, and tumor hypoxia using CD31, α-SMA, and pimonidazole as markers, respectively. DCE-MRI was performed at 7.05 T, and parametric images of Ktrans and ve were derived from the data using the Tofts pharmacokinetic model. RESULTS: The tumors of both models showed decreased microvascular density, increased vessel pericyte coverage, and increased vessel maturation after bevacizumab treatment. Bevacizumab-treated tumors were more hypoxic and had lower Ktrans values than untreated tumors in the BK-12 model, whereas bevacizumab-treated and untreated HL-16 tumors had similar hypoxic fractions and similar Ktrans values. Significant correlations were found between median Ktrans and hypoxic fraction, and the data for untreated and bevacizumab-treated tumors were well fitted by the same curve in both tumor models. CONCLUSIONS: Bevacizumab-treated tumors show less abnormal microvessels than untreated tumors do, but because of treatment-induced vessel pruning, the overall function of the microvasculature might be impaired after bevacizumab treatment, resulting in increased tumor hypoxia. DCE-MRI has great potential for monitoring bevacizumab-induced changes in tumor hypoxia in cervical carcinoma.

Diffusion-Weighted MRI Is Insensitive to Changes in the Tumor Microenvironment Induced by Antiangiogenic Therapy.

Antiangiogenic treatment (AAT) used in combination with radiation therapy or chemotherapy is a promising strategy for the treatment of several cancer diseases. The vascularity and oxygenation of tumors may be changed significantly by AAT, and consequently, a noninvasive method for monitoring AAT-induced changes in these microenvironmental parameters is needed. The purpose of this study was to evaluate the potential usefulness of diffusion-weighted magnetic resonance imaging (DW-MRI). DW-MRI was conducted with a Bruker Biospec 7.05-T scanner using four diffusion weightings and diffusion sensitization gradients in three orthogonal directions. Maps of the apparent diffusion coefficient (ADC) were calculated by using a monoexponential diffusion model. Two cervical carcinoma xenograft models (BK-12, HL-16) were treated with bevacizumab, and two pancreatic carcinoma xenograft models (BxPC-3, Panc-1) were treated with sunitinib. Pimonidazole and CD31 were used as markers of hypoxia and blood vessels, respectively, and fraction of hypoxic tissue (HFPim) and microvascular density (MVD) were quantified by analyzing immunohistochemical preparations. MVD decreased significantly after AAT in BK-12, HL-16, and BxPC-3 tumors, and this decrease was sufficiently large to cause a significant increase in HFPim in BK-12 and BxPC-3 tumors. The ADC maps of treated tumors and untreated control tumors were not significantly different in any of these three tumor models, suggesting that the AAT-induced microenvironmental changes were not detectable by DW-MRI. DW-MRI is insensitive to changes in tumor vascularity and oxygenation induced by bevacizumab or sunitinib treatment.

DCE-MRI of Sunitinib-Induced Changes in Tumor Microvasculature and Hypoxia: A Study of Pancreatic Ductal Adenocarcinoma Xenografts.

The purpose of this study was dual: to investigate (a) whether sunitinib may induce changes in tumor microvasculature and hypoxia in pancreatic ductal adenocarcinoma (PDAC) and (b) whether any changes can be detected by DCE-MRI. Sunitinib-treated and untreated control tumors of two PDAC xenograft models (BxPC-3 and Panc-1) were subjected to DCE-MRI before the imaged tumors were prepared for quantitative analysis of immunohistochemical preparations. Pimonidazole was used as a hypoxia marker, and fraction of hypoxic tissue (HFPim), density of CD31-positive microvessels (MVDCD31), and density of αSMA-positive microvessels (MVDαSMA) were measured. Parametric images of Ktrans and ve were derived from the DCE-MRI data by using the Tofts pharmacokinetic model. BxPC-3 tumors showed increased HFPim, decreased MVDCD31, unchanged MVDαSMA, and increased vessel maturation index (VMI = MVDαSMA/MVDCD31) after sunitinib treatment. The increase in VMI was seen because sunitinib induced selective pruning rather than maturation of αSMA-negative microvessels. Even though the microvessels in sunitinib-treated tumors were less abnormal than those in untreated tumors, this microvessel normalization did not improve the function of the microvascular network or normalize the tumor microenvironment. In Panc-1 tumors, HFPim, MVDCD31, MVDαSMA, and VMI were unchanged after sunitinib treatment. Median Ktrans increased with increasing MVDCD31 and decreased with increasing HFPim, and the correlations were similar for treated and untreated BXPC-3 and Panc-1 tumors. These observations suggest that sunitinib may induce significant changes in the microenvironment of PDACs, and furthermore, that Ktrans may be an adequate measure of tumor vascular density and hypoxia in untreated as well as sunitinib-treated PDACs.

Increasing aggressiveness of patient-derived xenograft models of cervix carcinoma during serial transplantation.

Four patient-derived xenograft (PDX) models (BK-12, ED-15, HL-16, LA-19) of carcinoma of the uterine cervix have been developed in our laboratory, and their stability during serial transplantation in vivo was investigated in this study. Two frozen cell stocks were established, one from xenografted tumors in passage 2 (early generation) and the other from xenografted tumors transplanted serially in mice for approximately two years (late generation), and the biology of late generation tumors was compared with that of early generation tumors. Late generation tumors showed higher incidence of lymph node metastases than early generation tumors in three models (ED-15, HL-16, LA-19), and the increased metastatic propensity was associated with increased tumor growth rate, increased microvascular density, and increased expression of angiogenesis-related and cancer stem cell-related genes. Furthermore, late generation tumors showed decreased fraction of pimonidazole-positive tissue (i.e., decreased fraction of hypoxic tissue) in two models (HL-16, LA-19) and decreased fraction of collagen-I-positive tissue (i.e., less extensive extracellular matrix) in two models (ED-15, HL-16). This study showed that serially transplanted PDXs may not necessarily mirror the donor patients' diseases, and consequently, proper use of serially transplanted PDX models in translational cancer research requires careful molecular monitoring of the models.

Diffusion-weighted MRI-derived ADC values reflect collagen I content in PDX models of uterine cervical cancer.

Apparent diffusion coefficient (ADC) values derived from diffusion-weighted magnetic resonance imaging (DW-MRI) are known to reflect the cellular environment of biological tissues. However, emerging evidence accentuates the influence of stromal elements on ADC values. The current study sought to elucidate whether a correlation exists between ADC and the fraction of collagen I-positive tissue across different tumor models of uterine cervical cancer. Early and late generation tumors of four patient-derived xenograft (PDX) models of squamous cell carcinoma (BK-12, ED-15, HL-16, and LA-19) were included. DW-MRI was performed with diffusion encoding constants (b) of 200, 400, 700, and 1000 s/mm2 and diffusion gradient sensitization in three orthogonal directions. The fraction of collagen I-positive connective tissue was determined by immunohistochemistry. Mono-exponential decay curves, from which the ADC value of tumor voxels was calculated, yielded good fits to the diffusion data. A significant inverse correlation was detected between median tumor ADC and collagen I fraction across the four PDX models, indicating that collagen fibers in the extracellular space have the ability to inhibit the movement of water molecules in these xenografts. The results encourage further exploration of DW-MRI as a non-invasive imaging method for characterizing the stromal microenvironment of tumors.

Vascular abnormalities and development of hypoxia in microscopic melanoma xenografts.

BACKGROUND: Studies investigating the oxygenation status and the development of hypoxia in microscopic tumors are sparse. The purpose of this study was to measure the extent of hypoxia in microscopic melanoma xenografts and to search for possible mechanisms leading to the development of hypoxia in these tumors. METHODS: A-07, D-12, R-18, and U-25 human melanoma xenografts grown in dorsal window chambers or as flank tumors were used as preclinical tumor models. Morphologic and functional parameters of vascular networks were assessed with intravital microscopy, and the expression of angiogenesis-related genes was assessed with quantitative PCR. Microvessels, pericytes, and the extent of hypoxia were assessed by immunohistochemistry in microscopic tumors by using CD31, αSMA, and pimonidazole as markers, and the extent of radiobiological hypoxia was assessed in macroscopic flank tumors. RESULTS: Macroscopic R-18 and U-25 tumors showed extensive hypoxia, whereas macroscopic A-07 and D-12 tumors were less hypoxic. R-18 and U-25 tumors developed hypoxic regions before they reached a size of 2-3 mm in diameter, whereas A-07 and D-12 tumors of similar size did not show hypoxic regions. The development of hypoxic regions was not caused by low vessel density, but was rather a result of inadequate vascular function. Inadequate vascular function was not caused by low vessel diameters or long vessel segments, but was associated with poor vascular pericyte coverage. Poor pericyte coverage was associated with the expression of eight angiogenesis-related genes. CONCLUSIONS: Two of the four investigated melanoma models developed hypoxic regions in microscopic tumors, and the development of hypoxia was associated with poor vascular pericyte coverage and inadequate vascular function.

DCE-MRI of patient-derived xenograft models of uterine cervix carcinoma: associations with parameters of the tumor microenvironment.

BACKGROUND: Abnormalities in the tumor microenvironment are associated with resistance to treatment, aggressive growth, and poor clinical outcome in patients with advanced cervical cancer. The potential of dynamic contrast-enhanced (DCE) MRI to assess the microvascular density (MVD), interstitial fluid pressure (IFP), and hypoxic fraction of patient-derived cervical cancer xenografts was investigated in the present study. METHODS: Four patient-derived xenograft (PDX) models of squamous cell carcinoma of the uterine cervix (BK-12, ED-15, HL-16, and LA-19) were subjected to Gd-DOTA-based DCE-MRI using a 7.05 T preclinical scanner. Parametric images of the volume transfer constant (K trans) and the fractional distribution volume (v e) of the contrast agent were produced by pharmacokinetic analyses utilizing the standard Tofts model. Whole tumor median values of the DCE-MRI parameters were compared with MVD and the fraction of hypoxic tumor tissue, as determined histologically, and IFP, as measured with a Millar catheter. RESULTS: Both on the PDX model level and the single tumor level, a significant inverse correlation was found between K trans and hypoxic fraction. The extent of hypoxia was also associated with the fraction of voxels with unphysiological v e values (v e > 1.0). None of the DCE-MRI parameters were related to MVD or IFP. CONCLUSIONS: DCE-MRI may provide valuable information on the hypoxic fraction of squamous cell carcinoma of the uterine cervix, and thereby facilitate individualized patient management.

Metastatic pathway and the microvascular and physicochemical microenvironments of human melanoma xenografts.

BACKGROUND: Malignant melanoma of the skin can metastasize through blood vessels and lymphatics. The primary tumor develops a vascular microenvironment characterized by abnormal blood vessels and lymphatics and a physicochemical microenvironment characterized by low oxygen tension, regions with hypoxic tissue, and high interstitial fluid pressure (IFP). This study aimed at identifying relationships between the metastatic route of melanomas and characteristic features of the microvascular and physicochemical microenvironments of the primary tumor. METHODS: Two patient-derived xenograft (PDX) models (E-13, N-15) and four cell line-derived xenografts (CDX) models (C-10, D-12, R-18, T-22) of human melanoma were included in the study. Tumors were transplanted to an orthotopic site in BALB/c-nu/nu mice, and when the tumors had grown to a volume of 500-600 mm3, the IFP of the primary tumor was measured and the hypoxia marker pimonidazole was administered before the host mouse was euthanized. The primary tumor, lungs, and six pairs of lymph nodes were evaluated by examining hematoxylin/eosin-stained and immunostained histological preparations. The expression of angiogenesis-related genes was assessed by quantitative PCR. RESULTS: C-10, D-12, and E-13 tumors disseminated primarily by the hematogenous route and developed pulmonary metastases. These tumors showed high angiogenic activity and high expression of the F3 gene as well as ANGPT2 and TIE1, genes encoding proteins of the angiopoietin-tie system. N-15, R-18, and T-22 tumors disseminated mainly by the lymphogenous route and developed metastases in draining lymph nodes. These tumors had highly elevated IFP and showed high expression of NRP2, a gene encoding neuropilin-2. CONCLUSION: The primary metastatic route of orthotopic human melanoma xenografts and the development of lung and lymph node metastases are influenced significantly by the microvascular and physicochemical microenvironments of the primary tumor.

Antiangiogenic agents targeting different angiogenic pathways have opposite effects on tumor hypoxia in R-18 human melanoma xenografts.

BACKGROUND: Studies comparing the effect of antiangiogenic agents targeting different angiogenic pathways are sparse. The purpose of this study was to compare the effect of properdistatin and sunitinib treatment in a preclinical model of malignant melanoma. Properdistatin is a small peptide derived from the thrombospondin-1 domain of the plasma protein properdin, and sunitinib is a tyrosine kinase inhibitor targeting several receptors including the vascular endothelial growth factor receptors. METHODS: R-18 human melanoma xenografts growing in dorsal window chambers were treated with properdistatin, sunitinib, or vehicle. Parameters describing the morphology of tumor vasculature were assessed from high-resolution transillumination images, and BST (blood supply time; the time needed for arterial blood to flow from the main supplying artery to downstream microvessels) was assessed from first-pass imaging movies recorded after a bolus of fluorescence-labeled dextran had been administered intravenously. Tumor hypoxia was assessed from immunohistochemical preparations of the imaged tissue by using pimonidazole as a hypoxia marker. RESULTS: Properdistatin treatment selectively removed small-diameter vessels and reduced BST, whereas sunitinib treatment reduced the density of small- and large-diameter vessel similarly and did not change BST. These observations imply that properdistatin treatment reduced geometric resistance to blood flow and improved vascular function, whereas sunitinib treatment did not affect vascular function. Accordingly, sunitinib-treated tumors showed higher hypoxic fractions than properdistatin-treated tumors. CONCLUSIONS: Properdistatin and sunitinib both inhibited angiogenesis, but had distinctly different effects on vascular morphology, vascular function, and extent of hypoxia in R-18 human melanoma xenografts.

Lymph node metastasis and the physicochemical micro-environment of pancreatic ductal adenocarcinoma xenografts.

Pancreatic ductal adenocarcinoma (PDAC) patients develop lymph node metastases early and have a particularly poor prognosis. The poor prognosis has been shown to be associated with the physicochemical microenvironment of the tumor tissue, which is characterized by desmoplasia, abnormal microvasculature, extensive hypoxia, and highly elevated interstitial fluid pressure (IFP). In this study, we searched for associations between lymph node metastasis and features of the physicochemical microenvironment in an attempt to identify mechanisms leading to metastatic dissemination and growth. BxPC-3 and Capan-2 PDAC xenografts were used as preclinical models of human PDAC. In both models, lymph node metastasis was associated with high IFP rather than high fraction of hypoxic tissue or high microvascular density. Seven angiogenesis-related genes associated with high IFP-associated lymph node metastasis were detected by quantitative PCR in each of the models, and these genes were all up-regulated in high IFP/highly metastatic tumors. Three genes were mutual for the BxPC-3 and Capan-2 models: transforming growth factor beta, angiogenin, and insulin-like growth factor 1. Further comprehensive studies are needed to determine whether there is a causal relationship between the up-regulation of these genes and high IFP and/or high propensity for lymph node metastasis in PDAC.

Dynamic contrast-enhanced MRI of the microenvironment of pancreatic adenocarcinoma xenografts.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease with poor outcome. Resistance to treatment is associated with impaired vascularity, extensive hypoxia, and interstitial hypertension. In this study, the potential of dynamic contrast-enhanced (DCE)-MRI as a method for assessing the microvascular density (MVD), the fraction of hypoxic tissue, and the interstitial fluid pressure (IFP) of PDACs was investigated. MATERIAL AND METHODS: Intramuscular BxPC-3, Capan-2, MIAPaCa-2, and Panc-1 PDAC xenografts were used as preclinical models of human PDACs. DCE-MRI with Gd-DOTA as contrast agent was conducted with a 7.05-T scanner, and the DCE-MRI series were analyzed voxelwise by using the Tofts pharmacokinetic model. Tumor MVD and hypoxia were measured in histological preparations by using pimonidazole as a hypoxia marker and CD31 as a marker of endothelial cells. IFP was measured with a Millar catheter. RESULTS: Ktrans (the volume transfer constant of Gd-DOTA) increased with increasing MVD and decreased with increasing hypoxic fraction, but was not associated with IFP. Any association between ve (the fractional distribution volume of Gd-DOTA) and MVD, hypoxic fraction, or IFP could not be detected. CONCLUSIONS: This study shows that DCE-MRI is a useful modality for assessing important features of the microenvironment of PDAC xenografts and thus provides the basis for future preclinical and clinical DCE-MRI investigations of PDAC.

The Effect of Sunitinib Treatment in Human Melanoma Xenografts: Associations with Angiogenic Profiles.

The effect of antiangiogenic agents targeting the vascular endothelial growth factor A (VEGF-A) pathway has been reported to vary substantially in preclinical studies. The purpose of this study was to investigate the effect of sunitinib treatment on tumor vasculature and oxygenation in melanoma xenografts with different angiogenic profiles. A-07, U-25, D-12, or R-18 melanoma xenografts were grown in dorsal window chambers and given daily treatments of sunitinib (40 mg/kg) or vehicle. Morphologic parameters of tumor vascular networks were assessed from high-resolution transillumination images, and tumor blood supply times (BSTs) were assessed from first-pass imaging movies. Tumor hypoxia was assessed with immunohistochemistry by using pimonidazole as hypoxia marker, and the gene expression and the protein secretion rate of angiogenic factors were assessed by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. The melanoma lines differed substantially in the expression of VEGF-A, VEGF-C, and platelet-derived growth factor A. Sunitinib treatment reduced vessel densities and induced hypoxia in all melanoma lines, and the magnitude of the effect was associated with the gene expression and protein secretion rate of VEGF-A. Sunitinib treatment also increased vessel segment lengths, reduced the number of small-diameter vessels, and inhibited growth-induced increases in the diameter of surviving vessels but did not change BST. In conclusion, sunitinib treatment did not improve vascular function but reduced vessel density and induced hypoxia in human melanoma xenografts. The magnitude of the treatment-induced effect was associated with the VEGF-A expression of the melanoma lines.

Properdistatin inhibits angiogenesis and improves vascular function in human melanoma xenografts with low thrombospondin-1 expression.

In this study, the effect of properdistatin, a novel peptide derived from the thrombospondin 1 (TSP-1) domain of properdin, was investigated in three melanoma xenograft models with different TSP-1 expression. The tumors were grown in dorsal window chambers and were treated with 80 mg/kg/day properdistatin or vehicle. Morphological parameters of the tumor vasculature were assessed from high resolution transillumination images. Blood supply time (i.e., the time required for arterial blood to flow from a supplying artery to downstream microvessels) and plasma velocities were assessed from first-pass imaging movies recorded after a bolus of fluorescence-labeled dextran had been administered intravenously. Gene and protein expression of TSP-1 were assessed with quantitative PCR and immunohistochemistry, respectively. Properdistatin treatment inhibited angiogenesis in low TSP-1 expressing tumors but did not alter the vasculature in high TSP-1 expressing tumors. In low TSP-1 expressing tumors, properdistatin selectively removed small-diameter capillaries, but did not change the morphology of tumor arterioles or tumor venules. Properdistatin also reduced blood supply times and increased plasma velocities, implying that the treatment reduced the geometric resistance to blood flow and improved vascular function.

Functional intratumoral lymphatics in patient-derived xenograft models of squamous cell carcinoma of the uterine cervix: implications for lymph node metastasis.

Studies of cell line-derived human tumor xenografts have suggested that the lymphatics seen in immunohistochemical preparations from non-peripheral regions of tumors are nonfunctional. In this investigation, lymphangiogenesis, hemangiogenesis, and lymph node metastasis were studied in patient-derived xenograft (PDX) models of carcinoma of the uterine cervix. Lymph vessel density (LVD) and blood vessel density (BVD) were measured in immunohistochemical preparations. The expression of angiogenesis-related genes was investigated by quantitative PCR. Lymphatic functionality was assessed with the ferritin assay, and tumor interstitial fluid pressure (IFP) was measured with a Millar catheter. The PDX models mirrored the angiogenesis and aggressiveness of the donor patients' tumors, and two highly aggressive models developed functional lymphatics within the tumor mass. Tumors with functional intratumoral lymphatics showed low IFP, high LVD, high BVD, high expression of a large number of angiogenesis-related genes, and high incidence of lymph node metastases. LVD correlated with BVD, and lymph node metastasis was associated with high LVD and high BVD. Nine angiogenesis-related genes associated with the development of functional intratumoral lymhatics were identified. High expression of these genes, high LVD, and high BVD may be important biomarkers for poor outcome in cervix carcinoma.

Diffusion-weighted and dynamic contrast-enhanced MRI of pancreatic adenocarcinoma xenografts: associations with tumor differentiation and collagen content.

PURPOSE: The aggressiveness of pancreatic ductal adenocarcinoma (PDAC) is highly dependent on the level of differentiation and the composition of the stroma. In this preclinical study, we investigated the potential of diffusion-weighted magnetic resonance imaging (DW-MRI) and dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) as noninvasive methods for providing information on the differentiation and the stroma of PDACs. METHODS: Xenografted tumors initiated from four PDAC cell lines (BxPC-3, Capan-2, MIAPaCa-2, and Panc-1) were included in the study. DW-MRI and DCE-MRI were carried out on a 7.05-T MR scanner, and tumor images of ADC (the apparent diffusion coefficient), K (trans) (the volume transfer constant of Gd-DOTA), and v e (the fractional distribution volume of Gd-DOTA) were produced. The level of differentiation and the amount and structure of collagen I and collagen IV were determined by examining histological preparations. RESULTS: Differentiated tumors showed lower levels of collagen I and collagen IV than non-differentiated tumors. Significant correlations were found between ADC and v e, and both parameters differentiated clearly between collagen-rich non-differentiated tumors and differentiated tumors containing less collagen. CONCLUSION: Differentiated PDAC xenografts show higher ADC values and higher v e values than their non-differentiated counterparts. This observation supports the application of parametric MR images as tumor biomarkers in PDAC. Patients showing low values of ADC and v e most likely have non-differentiated tumors with extensive stroma and, hence, poor prognosis.

Patient-derived xenograft models of squamous cell carcinoma of the uterine cervix.

Patient-derived xenograft (PDX) models of cancer are considered to reflect the biology and treatment response of human tumors to a larger extent than xenograft models initiated from established cell lines. The characterization of a panel of four novel PDX models of cervical carcinoma of the uterine cervix is described in this communication. The outcome of treatment differed substantially among the donor patients, and the PDX models were found to mirror the histology, aggressiveness, and metastatic propensity of the donor patients' tumors. Two of the models (BK-12 and LA-19) were highly metastatic, one model (ED-15) was poorly metastatic, and one model (HL-16) was non-metastatic. The primary tumors of the two highly metastatic models showed high density of intratumoral lymphatics, whereas the other two models did not develop intratumoral lymphatics. The potential of the models to metastasize to lymph nodes was associated with high expression of both angiogenesis-related genes and cancer stem cell-related genes. The models may be highly valuable for studying mechanisms linking lymph node metastasis to lymphangiogenesis, hemangiogenesis, and the presence of cancer stem cells.

Early effects of low dose bevacizumab treatment assessed by magnetic resonance imaging.

BACKGROUND: Antiangiogenic treatments have been shown to increase blood perfusion and oxygenation in some experimental tumors, and to reduce blood perfusion and induce hypoxia in others. The purpose of this preclinical study was to investigate the potential of dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) and diffusion weighted MRI (DW-MRI) in assessing early effects of low dose bevacizumab treatment, and to investigate intratumor heterogeneity in this effect. METHODS: A-07 and R-18 human melanoma xenografts, showing high and low expression of VEGF-A, respectively, were used as tumor models. Untreated and bevacizumab-treated tumors were subjected to DCE-MRI and DW-MRI before treatment, and twice during a 7-days treatment period. Tumor images of Ktrans (the volume transfer constant of Gd-DOTA) and ve (the fractional distribution volume of Gd-DOTA) were produced by pharmacokinetic analysis of the DCE-MRI data, and tumor images of ADC (the apparent diffusion coefficient) were produced from DW-MRI data. RESULTS: Untreated A-07 tumors showed higher Ktrans, v e, and ADC values than untreated R-18 tumors. Untreated tumors showed radial heterogeneity in Ktrans, i.e., Ktrans was low in central tumor regions and increased gradually towards the tumor periphery. After the treatment, bevacizumab-treated A-07 tumors showed lower Ktrans values than untreated A-07 tumors. Peripherial tumor regions showed substantial reductions in Ktrans, whereas little or no effect was seen in central regions. Consequently, the treatment altered the radial heterogeneity in Ktrans. In R-18 tumors, significant changes in Ktrans were not observed. Treatment induced changes in tumor size, v e, and ADC were not seen in any of the tumor lines. CONCLUSIONS: Early effects of low dose bevacizumab treatment may be highly heterogeneous within tumors and can be detected with DCE-MRI.

Hypoxia biomarkers in squamous cell carcinoma of the uterine cervix.

BACKGROUND: There is significant evidence that severe tumor hypoxia may cause resistance to chemoradiotherapy and promote metastatic spread in locally advanced carcinoma of the uterine cervix. Some clinical investigations have suggested that high expression of hypoxia-inducible factor-1α (HIF-1α) and/or its target gene carbonic anhydrase IX (CAIX) may be useful biomarkers of tumor hypoxia and poor outcome in cervical cancer. Here, we challenged this view by investigating possible associations between HIF-1α expression, CAIX expression, fraction of hypoxic tissue, and lymph node metastasis in experimental human tumors. METHODS: Tumors of two cervical carcinoma xenograft lines (CK-160 and TS-415) were included in the study. Pimonidazole was used as a hypoxia marker, and tumor hypoxia, HIF-1α expression, and CAIX expression were detected by immunohistochemistry. Metastatic status was assessed by examining external lymph nodes in the inguinal, axillary, interscapular, and submandibular regions and internal lymph nodes in the abdomen and mediastinum. RESULTS: Tissue regions staining positive for pimonidazole, HIF-1α, or CAIX were poorly colocalized, both in CK-160 and TS-415 tumors. The expression of HIF-1α or CAIX did not correlate with the fraction of hypoxic tissue in any of the two tumor lines. Furthermore, clinically relevant associations between HIF-1α or CAIX expression and lymph node metastasis were not found. CONCLUSION: Because significant associations between HIF-1α expression, CAIX expression, fraction of hypoxic tissue, and incidence of lymph node metastases could not be detected in any of two preclinical models of human cervical cancer, it is not realistic to believe that high expression of HIF-1α or CAIX can be useful biomarkers of tumor hypoxia and poor outcome in a highly heterogeneous disease like cervical carcinoma.

Thrombospondin-1 domain-containing peptide properdistatin improves vascular function in human melanoma xenografts.

Properdistatin is a novel peptide derived from the thrombospondin-1 domain of the plasma protein properdin. The purpose of this study was to investigate the effect of properdistatin treatment on the morphology and function of tumor vasculature. A-07 human melanoma xenografts grown in dorsal window chambers were used as preclinical model. Tumors were treated with 80 mg/kg/day properdistatin or vehicle for 4 days. Morphologic parameters of tumor vascular networks were assessed from high-resolution transillumination images, and tumor blood supply time and plasma velocities were assessed from first-pass imaging movies recorded after a bolus of 155 kDa tetramethylrhodamine isothiocyanate-labeled dextran had been administered intravenously. Properdistatin-treated tumors showed reduced density of small-diameter vessels, reduced blood supply time, and increased plasma velocities. In conclusion, properdistatin treatment inhibited angiogenesis and improved vascular function in A-07 tumors.