Lise Mari K. Andersen

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

BackgroundAntiangiogenic 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.MethodsA-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.ResultsUntreated A-07 tumors showed higher Ktrans, ve, 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, ve, and ADC were not seen in any of the tumor lines.ConclusionsEarly effects of low dose bevacizumab treatment may be highly heterogeneous within tumors and can be detected with DCE-MRI.

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

PurposeThe 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.MethodsXenografted 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), Ktrans (the volume transfer constant of Gd-DOTA), and ve (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.ResultsDifferentiated tumors showed lower levels of collagen I and collagen IV than non-differentiated tumors. Significant correlations were found between ADC and ve, and both parameters differentiated clearly between collagen-rich non-differentiated tumors and differentiated tumors containing less collagen.ConclusionDifferentiated PDAC xenografts show higher ADC values and higher ve 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 ve most likely have non-differentiated tumors with extensive stroma and, hence, poor prognosis.

Hypoxia biomarkers in squamous cell carcinoma of the uterine cervix

BackgroundThere 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.MethodsTumors 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.ResultsTissue 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.ConclusionBecause 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.

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

BackgroundStudies 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.MethodsR-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.ResultsProperdistatin 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.ConclusionsProperdistatin 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.

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.

Abstract 1801: 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. Citation Format: Jon-Vidar Gaustad, Trude G. Simonsen, Lise Mari K. Andersen, Einar K. Rofstad. Antiangiogenic agents targeting different angiogenic pathways have opposite effects on tumor hypoxia in R-18 human melanoma xenografts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1801. doi:10.1158/1538-7445.AM2017-1801

Abstract A03: Vascular function affected fundamentally different by different antiangiogenic agents

The purpose of this preclinical study was to investigate the effect of two antiangiogenic agents, properdistatin and sunitinib, on the morphology and function of tumor vasculature. Properdistatin is a novel peptide derived from the thrombospondin-1 domain of the plasma protein properdin. Properdistatin treatment has been shown to inhibit angiogenesis and tumor growth in breast cancer xenografts. Sunitinib is a tyrosine kinase inhibitor that interacts with several receptors including vascular endothelial growth factor receptors 1-3, and platelet-derived growth factor receptors α-β. Sunitinib treatment has been shown to inhibit angiogenesis and tumor growth in a number of tumor models, and has been approved clinically for several indications. A-07 human melanoma xenografts grown in dorsal window chambers or as intradermal flank tumors were used as preclinical tumor model. In window chamber tumors, morphologic parameters of tumor vascular networks were assessed from high-resolution transillumination images, and blood supply time and plasma velocities were assessed from first-pass imaging movies recorded after a bolus of 155kDa tetramethylrhodamine isothiocyanate-labeled dextran had been administered intravenously. In flank tumors, parametric images of ADC and Ktrans were produced from diffusion weighted magnetic resonance imaging (DW-MRI) and dynamic contrast-enhanced MRI (DCE-MRI). Tumor hypoxia and necrosis were assessed from immunohistochemical preparations of the imaged tissue, and tumor interstitial fluid pressure (IFP) was assessed with probe measurement. Properdistatin treatment reduced the density of small-diameter vessels, reduced blood supply time, and increased plasma velocities, whereas sunitinib treatment did not alter blood supply time, reduced overall vessel density, induced tumor hypoxia and necrosis, and did not alter tumor IFP. Sunitinib-treated flank tumors showed reduced Ktrans values corresponding to a decrease in vessel density, and increased ADC values corresponding to induction of tumor necrosis. In conclusion, properdistatin treatment improved vascular function, whereas sunitinib treatment did not improve vascular function but induced tumor hypoxia and necrosis in A-07 tumors. This data demonstrates that different antiangiogenic agents may affect vascular function fundamentally different in the same tumor model. DW-MRI and DCE-MRI were sensitive to the sunitinib-induced changes in the tumor microenvironment, demonstrating that these MR-techniques may be used to identify tumors where antiangiogenic treatment does not improve vascular function. Citation Format: Jon-Vidar Gaustad, Trude G. Simonsen, Tord Hompland, Lise Mari K. Andersen, Einar K. Rofstad. Vascular function affected fundamentally different by different antiangiogenic agents. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Angiogenesis and Vascular Normalization: Bench to Bedside to Biomarkers; Mar 5-8, 2015; Orlando, FL. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl):Abstract nr A03.