Increased Tumor Perfusion Research Articles

Early vessel normalization improves glioblastoma outcomes

A subset of patients with glioblastoma who respond by showing increased tumour perfusion after early antiangiogenic therapy are more likely to benefit from anti-VEGF treatment combined with radiation and chemotherapy than patients who don’t respond. Furthermore, vascular normalization is associated with longer overall survival in patients with hyperpermeable (but uncompressed) tumours, according to two new studies. Rakesh Jain, senior author on both studies, first hypothesized in 2001 that anti-VEGF therapy works by ‘normalizing’ leaky vessels that surround tumours, improving the delivery of drugs and oxygen, which are also necessary for effective radiotherapy. Drugs that inhibit the VEGF pathway only benefit a subset of patients with solid tumours, and determining biomarkers that can distinguish which patients are more likely to respond to treatment had yet to be identified. In the first study, the researchers used MRI techniques to measure changes in blood tumour vasculature, including vessel architectural imaging, to monitor changes after anti-VEGF therapy. “Early increased tumour perfusion, often seen after one treatment with a pan-VEGFR kinase inhibitor, was associated with a twothirds increase in survival time in patients harbouring glioblastoma, compared with those patients who had no increase in blood perfusion,” explain four authors of the study—Batchelor, Emblem, Duda and Jain—in a joint statement. This effect, seen in half of the patients who underwent anti-VEGF therapy and chemoradiation, was not observed in patients who received chemotherapy alone. Moreover, these changes are associated with improved oxygenation and various plasma biomarkers.

Pharmacokinetic (PK)/pharmacodynamic (PD) results from a phase Ib study of pegylated hyaluronidase PH20 (PEGPH20) in combination with gemcitabine (Gem) in patients with pancreatic cancer.

e15005 Background: Enzymatic degradation of hyaluronan (HA) is a novel strategy to target the desmoplastic stroma of pancreatic cancer. PEGPH20, a pegylated form of recombinant human hyaluronidase PH20, is an investigational drug in clinical trials. Preclinical studies demonstrate that sustained HA removal by PEGPH20 inhibits tumor growth and enhances chemotherapeutic activity in HA-rich xenografts and genetically engineered mouse tumor models. Ph1 PEGPH20 monotherapy studies show increased tumor perfusion by DCE-MRI, metabolic partial responses by FDG-PET, and stromal remodeling in tumor biopsies from selected advanced cancer patients (pts). Methods: This was a dose-escalation study to find the recommended Ph2 dose of PEGPH20 in combination with Gem in pts with Stage IV previously untreated pancreatic cancer. Pts received Gem at 1000 mg/m2 IV qwk for Wks 1-7 plus PEGPH20 at 1, 1.6, or 3 μg/kg IV twice a week for Wks 1-4 and qwk for Wks 5-7. Wk 8 was a rest week. Thereafter, PEGPH20 + Gem were given qwk for 3 wks in 4-wk cycles. Serial plasma samples were collected and hyaluronidase activity measured by an ultrasensitive assay to assess PEGPH20 exposure. Plasma HA catabolites were measured by quantitative HPLC to assess PD. Results: 28 pts were enrolled. Plasma PEGPH20 concentrations were proportional to dose, and kinetics were well-characterized by a 2-compartment PK model. Estimates for clearance (0.5-2 mL/hr/kg) were consistent with long t1/2 (1-2 days) previously seen with single-dose PEGPH20 monotherapy. Wks 1 and 4 PK profiles were similar, suggesting no changes to PEGPH20 clearance mechanisms after multiple doses or effects of Gem on PEGPH20 exposure. Most pretreatment plasma HA levels were <1 μg/mL and increased in a time- and dose-dependent manner after dosing. Circulating HA concentration was >500 µg/mL in several pts given 3 μg/kg PEGPH20. Conclusions: PEGPH20 plasma levels can be predicted using a linear PK model and circulating HA catabolites can be used as a quantitative measure of PEGPH20 PD. Results are consistent with the mechanism of action of hyaluronidase and support further study of PEGPH20 with anticancer agents. Clinical trial information: NCT01453153.

Final analysis of a phase IB/randomized phase II study of gemcitabine (G) plus placebo (P) or vismodegib (V), a hedgehog (Hh) pathway inhibitor, in patients (pts) with metastatic pancreatic cancer (PC): A University of Chicago phase II consortium study.

4012 Background: Sonic Hh (SHh), the ligand for the Hh pathway, is over-expressed in >80% of PC. V had activity in preclinical murine PC models leading to increased tumor perfusion, enhanced tumor delivery of G, and an improvement in survival. Methods: We conducted a placebo-controlled, phase IB/randomized phase II trial of GV or GP. Eligible pts, KPS 80-100, had untreated metastatic PC, or had completed adjuvant therapy > 6 months (mo) prior. Primary endpoint: progression-free survival (PFS). Correlatives: serial SHh serum levels; serial perfusion CT imaging. All pts received G 1000mg/m2over 30 minutes, days (D) 1, 8, 15, Q28D. A lead-in phase IB was performed. Pts, stratified by KPS (80 v 90/100), and disease status (newly diagnosed/recurrent), were randomized to V (150 mg PO daily) or P. For pts on P, cross-over was allowed at progression. Assuming a mPFS of 3.5 months for GP and 5.7 months for GV (HR=0.61), a sample size of 106 subjects (53 per group) provided 85% power to detect this difference, using a one-sided test at the 0.10 significance level. Results: No safety issues were identified in 7 pts enrolled in the phase IB study. The phase II study enrolled 106 evaluable pts (V/P 53/53) at 13 sites 2/10-6/12. Pt characteristics: median age 65/64 (range 52-82/39-83); KPS (% pts) 80: 38/30; 90: 26/38; 100: 36/32; newly diagnosed 91%/91%; recurrent: 9%/9%. Grade 3/4 toxicity (V/P, % pts, >5% in either arm): neutropenia 32/28; lymphopenia 4/15; thrombocytopenia 9/11; anemia 9/23; hyponatremia 4/15; fatigue 13/8; hyperglycemia 23/19; elevated ALT 13/9; hyperbilirubinemia 11/6; nausea 11/11. Response (%): CR 0/2, PR 8/11, SD 51/38. mPFS: 4.0/2.5 mo (95% CI: 2.5-5.3/1.9-3.8; HR 0.81 [0.54-1.21], p=0.30). 22 pts (42%) on GP crossed over to GV at progression. mOS: 6.9/6.1 mo (95% CI:5.8-8.0/5.0-8.0, HR 1.04, [0.69-1.58], p=0.84). Updated laboratory/radiological correlatives will be presented. Conclusions: Toxicity between the groups was similar. The addition of V to G in an unselected cohort does not improve response, PFS, or OS in pts with metastatic PC. Funding NCI N01-CM-62201. Clinical trial information: NCT01064622.

Role of vascular dysfunction in prostate tumor perfusion during exercise

During exercise, blood flow to inactive tissues is reduced through changes in vascular tone elicited largely by the autonomic nervous system. However, in tumors the reduced innervation of feed arteries and possibly blunted adrenergic vasoconstriction may lead to an increased tumor perfusion during exercise. We used control (CON; n=16) and prostate tumor bearing (TB; n=14) Copenhagen rats to test the hypothesis that α‐adrenergic (via norepinephrine; NE) and myogenic vasoconstrictor responses are diminished in the prostatic tumor feed arteries (PTA), resulting in an increased tumor perfusion during exercise. Dunning R‐3327 MatLyLu tumor cells (104) were injected into the ventral prostates of rats. PTA and prostate arteries (CON) were studied in vitro and exposed to cumulative additions of NE (10−9to 10−4M) and increases in intraluminal pressure (myogenic response; 0 to 135 cmH2O). In vivo quantification of perfused tumor vessels during rest and acute exercise (treadmill; 5 min) was determined using Hoechst‐33342 staining. Both α‐adrenergic (7 ± 6% vs. 88 ± 2%, p<0.05) and myogenic vasoconstriction were blunted in PTA versus CON, while the number of PTA perfused increased ~113% with exercise. These data suggest that dysfunction in the PTA results in an inability to augment vascular resistance during exercise, resulting in an enhanced tumor perfusion.Supp.: NIH (AG‐31317) & Florida Biomed. Res. Prog. (1BN‐02).

MR Imaging Assessment of Tumor Perfusion and 3D Segmented Volume at Baseline, during Treatment, and at Tumor Progression in Children with Newly Diagnosed Diffuse Intrinsic Pontine Glioma

DIPG is among the most devastating brain tumors in children, necessitating the development of novel treatment strategies and advanced imaging markers such as perfusion to adequately monitor clinical trials. This study investigated tumor perfusion and 3D segmented tumor volume as predictive markers for outcome in children with newly diagnosed DIPG. Imaging data were assessed at baseline, during, and after RT, and every other month thereafter until tumor progression for 35 patients (ages 2-16 years) with newly diagnosed DIPG enrolled in the phase I clinical study, NCT00472017. Patients were treated with conformal RT and vandetanib, a vascular endothelial growth factor receptor 2 inhibitor. Tumor perfusion increased and tumor volume decreased during combined RT and vandetanib therapy. These changes slowly diminished in follow-up scans until tumor progression. However, increased tumor perfusion and decreased tumor volume during combined therapy were associated with longer PFS. Apart from a longer OS for patients who showed elevated tumor perfusion after RT, there was no association for tumor volume and other perfusion variables with OS. Our results suggest that tumor perfusion may be a useful predictive marker for the assessment of treatment response and tumor progression in children with DIPG treated with both RT and vandetanib. The assessment of tumor perfusion yields valuable information about tumor microvascular status and its response to therapy, which may help better understand the biology of DIPGs and monitor novel treatment strategies in future clinical trials.

Mechanisms of Tumor Vascular Priming by a Nanoparticulate Doxorubicin Formulation

Tumor vascular normalization by antiangiogenic agents may increase tumor perfusion but reestablish vascular barrier properties in CNS tumors. Vascular priming via nanoparticulate carriers represents a mechanistically distinct alternative. This study investigated mechanisms by which sterically-stabilized liposomal doxorubicin (SSL-DXR) modulates tumor vascular properties. Functional vascular responses to SSL-DXR were investigated in orthotopic rat brain tumors using deposition of fluorescent permeability probes and dynamic contrast-enhanced magnetic resonance imaging. Microvessel density and tumor burden were quantified by immunohistochemistry (CD-31) and quantitative RT-PCR (VE-cadherin). Administration of SSL-DXR (5.7 mg/kg iv) initially (3-4 days post-treatment) decreased tumor vascular permeability, k(trans) (vascular exchange constant), vascular endothelial cell content, microvessel density, and deposition of nanoparticulates. Tumor vasculature became less chaotic. Permeability and perfusion returned to control values 6-7 days post-treatment, but intratumor SSL-DXR depot continued to effect tumor vascular endothelial compartment 7-10 days post-treatment, mediating enhanced permeability. SSL-DXR ultimately increased tumor vascular permeability, but initially normalized tumor vasculature and decreased tumor perfusion, permeability, and nanoparticulate deposition. These temporal changes in vascular integrity resulting from a single SSL-DXR dose have important implications for the design of combination therapies incorporating nanoparticle-based agents for tumor vascular priming.

Targeting hyaluronan (HA) in tumor stroma: A phase I study to evaluate the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of pegylated hyaluronidase (PEGPH20) in patients with solid tumors.

2579 Background: HA, a glycosaminoglycan, accumulates in ~25% of solid tumors and is associated with poor prognosis. Tumors that accumulate HA develop high interstitial fluid pressure (IFP) and become resistant to chemotherapy. In animal models, PEGPH20 reduced tumor-associated HA and increased efficacy of chemotherapy. Methods: This ongoing phase 1 study was designed to assess safety, tolerability, PK, PD, and the maximum tolerated dose (MTD) of PEGPH20 as a single agent in patients with treatment-refractory solid tumors. IV PEGPH20 was administered in a twice weekly or a once weekly dosing schedule with oral dexamethasone (dex) pre-and post dose. Biomarkers included tumor histochemistry to detect HA depletion, plasma HA catabolites, and DCE-MRI or FDG-PET. Results: Fifteen patients have been enrolled into the study. Dose administration ranged from 0.5 µg/kg to 5 µg/kg. Musculoskeletal events (MSEs) were the most common and dose limiting toxicities. To date, up to 3 µg/kg twice weekly IV infusions of PEGPH20 were well tolerated with manageable Grade 1 MSEs in most patients. MTD has not been reached. Plasma concentrations of PEGPH20 showed dose proportional exposure with a terminal half life of ~2 days. Enzymatic activity of PEGPH20 was reflected by dose-dependent HA catabolites in plasma. Pre- and post-dose tumor biopsies from 2 colon patients demonstrated 23-60% reduction in tumor HA after 4 weeks of dosing. Serial DCE-MRI and FDG-PET demonstrated increased tumor perfusion (Ktrans) and decreased metabolic activity 8 and 24hrs post-PEGPH20 in one patient each. Doses of ≤3 µg/kg were tolerated with intermittent Grade 1-2 MSE. Enrollment at dose levels > 3 µg/kg is ongoing. Conclusions: Repeated IV PEGPH20 infusion resulted in dose-dependent systemic exposure and is tolerated with concomitant dex treatment. HA catabolites in plasma demonstrated the in vivo enzymatic activity of PEGPH20. Reduced HA in tumor biopsies and serial DCE-MRI and FDG-PET images confirmed PD activity. Increased tumor perfusion by PEGPH20 may enhance drug delivery to tumor. Combination studies of PEGPH20 with cytotoxic chemotherapy are planned.

Effects of cediranib, a VEGF signaling inhibitor, in combination with chemoradiation on tumor blood flow and survival in newly diagnosed glioblastoma.

2009 Background: Anti-angiogenic therapy is hypothesized to synergize with radiation and chemotherapy by improving tumor blood flow. We evaluated the tolerability, efficacy and potential mechanism of action of radiation, temozolomide, and cediranib in newly diagnosed glioblastoma patients. Methods: Newly diagnosed glioblastoma patients were treated with radiation, temozolomide, and cediranib followed by monthly temozolomide for 6 cycles and daily cediranib until tumor progression or toxicity as part of an IRB-approved, Phase Ib/II clinical trial. MRI scans including measurement of cerebral blood flow were performed at baseline, weekly during the 6 weeks of chemoradiation and then monthly. Radiographic response was determined by RANO criteria. Results: Six patients were enrolled in the phase Ib part of the study with cediranib 30 mg daily in combination with temozolomide and radiation. No dose-limiting toxicities were identified. Forty patients were enrolled in the phase II part of the study. Among the entire cohort of 46 patients, median age was 57 (range 35-74), median KPS was 90% (60-100), 36 patients underwent a subtotal resection and 10 underwent biopsy. 26/30 patients taking corticosteroids were able to taper corticosteroids during chemoradiation. Off study reasons included toxicity (14), disease progression (18), and patient preference (2). Five patients remain on study without disease progression and 20 patients have died. Median duration on study was 158 days. Median progression free survival was 288 days (95%CI 240,∞) and median overall survival was 786 days (95%CI 411 ,∞). Best radiographic response in patients who completed chemoradiation was CR in 2 patients, PR in 20 patients, and SD in 15 patients. Patients with increased tumor perfusion during chemoradiation survived nearly 1 year longer (mean OS=611 days) than patients with decreased perfusion (mean OS=269 days). Conclusions: Cediranib was well tolerated and led to improved PFS and OS compared to historical controls, particularly in those with improved perfusion. This combination is being evaluated in an ongoing randomized trial (RTOG 0837).

Targeting hyaluronan (HA) in tumor stroma: Interim safety and translational evaluation of pegylated hyaluronidase (PEGPH20) in patients (pts) with advanced solid tumors—A focus on gastrointestinal malignancies.

249 Background: HA, a glycosaminoglycan in the tumor matrix, accumulates in 20-30% of solid tumors and ≥28%, ≥42% and ≥87% of colon, gastric and pancreatic ductal adenocarcinoma (PDA) tumors, respectively. Tumors that accumulate HA can develop high interstitial fluid pressure (IFP) and are resistant to chemotherapy. HA in human PDA is associated with disease progression and poor prognosis. In animal models, PEGPH20 reduced tumor-associated HA, tumor IFP and tumor growth, and increased efficacy of chemotherapy. Pharmacodynamic (PD) activity of PEGPH20 is being evaluated in Phase 1 (Ph1) trials. Methods: Two Ph1 studies in pts with advanced solid tumors refractory to prior therapy examined the safety, tolerability, PK, PD and MTD of single-agent IV PEGPH20. Dosing in Study 1 was 2x/wk and then reduced to q21d and in Study 2 is 1-2x/wk x 4wks and 1x/wk thereafter with dexamethasone pre-medication regimen. Biomarkers evaluated: Histochemistry to detect depletion of tumor HA, serum HA catabolites, Dynamic Contrast Enhanced (DCE)-MRI and FDG-PET. Results: In Study 1, the first 3 pts demonstrated musculoskeletal events (MSE; 2 pts dosed with 50 µg/kg 2x/wk, severe; 1 pt dosed with 0.5µg/kg 2x/wk, moderate) not predicted from GLP safety studies. The frequency of drug administration was reduced to q21d and the next 10 pts tolerated ≤1.5 µg/kg. In Study 2, dexamethasone pre-medication regimen was evaluated while dosing at 1-2x/wk. Nine pts were dosed ≤5.0 µg/kg. Doses ≤3.0 µg/kg administered 2x/wk were tolerated with intermittent Grade 1-2 MSE. Activity of PEGPH20 was reflected by increased HA serum catabolites. Pre- and post-dose biopsies from 2 colon pts demonstrated a 23-60% reduction in tumor HA after 4 wks of PEGPH20 dosing. DCE-MRI of PDA and FDG-PET of lung metastases increased tumor perfusion (Ktrans) and decreased metabolic activity 8 and 24hrs post-PEGPH20 dose and was still apparent after 4 wks of treatment with PEGPH20. Conclusions: Ongoing Ph1 studies have shown in vivo activity of PEGPH20 through HA plasma catabolites, tumor HA depletion, DCE-MRI and FDG-PET.

Increased Survival of Glioblastoma Patients Who Respond to Antiangiogenic Therapy with Elevated Blood Perfusion

The abnormal vasculature of the tumor microenvironment supports progression and resistance to treatment. Judicious application of antiangiogenic therapy may normalize the structure and function of the tumor vasculature, promoting improved blood perfusion. However, direct clinical evidence is lacking for improvements in blood perfusion after antiangiogenic therapy. In this study, we used MRI to assess tumor blood perfusion in 30 recurrent glioblastoma patients who were undergoing treatment with cediranib, a pan-VEGF receptor tyrosine kinase inhibitor. Tumor blood perfusion increased durably for more than 1 month in 7 of 30 patients, in whom it was associated with longer survival. Together, our findings offer direct clinical evidence in support of the hypothesis that vascular normalization can increase tumor perfusion and help improve patient survival.

Antitumor Actions of Ruthenium(III)-Based Nitric Oxide Scavengers and Nitric Oxide Synthase Inhibitors

The role of endogenous nitric oxide (NO) in the growth and vascularization of a rat carcinosarcoma (P22) has been investigated. Tumor-bearing animals were treated with (i) nitric oxide synthase (NOS) inhibitors, administered via the drinking water, including N(G)-nitro-l-arginine methyl ester (L-NAME), a nonisoform-selective inhibitor, and 2 others that target the inducible (NOS II) enzyme preferentially, namely 1-amino-2-hydroxyguanidine or N-[3-(aminomethyl)benzyl]acetamidine hydrochloride; or (ii) daily injections (intraperitoneally) of 2 Ru(III) polyaminocarboxylates, AMD6221 and AMD6245, both of which are effective NO scavengers. L-NAME, AMD6221, and AMD6245 reduced tumor growth by approximately 60% to 75% of control rates. Tumor sections stained with abs to CD-31/platelet endothelial cell adhesion molecule-1 or NOS III showed that this was associated with a marked reduction (60%-77%) of tumor microvascular densitiy (MVD). Tumors resumed growing promptly when treatment was discontinued, accompanied by partial or complete restoration of MVDs. In contrast, NOS-II selective inhibitors had no effect on tumor growth or vascularization, indicating that both responses require complete blockade of NO production. The results corroborate the view that endogenous NO facilitates tumor development. We suggest that NO deprivation causes tumor feeder vessels to constrict, reducing tumor blood flow. The delivery of oxygen and essential nutrients to the developing tumor is impaired as a consequence, hampering further growth. Normalizing NO levels by withholding treatment causes tumor feeder vessels to dilate, increasing tumor perfusion and reestablishing conditions that allow tumors to begin growing again.

Dynamic Contrast-Enhanced Magnet Resonance Imaging of Sunitinib-Induced Vascular Changes to Schedule Chemotherapy in Renal Cell Carcinoma Xenograft Tumors

In an attempt to develop better therapeutic approaches for metastatic renal cell carcinoma (RCC), the combination of the antiangiogenic drug sunitinib with gemcitabine was studied. Using dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI), we have previously determined that a sunitinib dosage of 20 mg/kg per day increased kidney tumor perfusion and decreased vascular permeability in a preclinical murine RCC model. This sunitinib dosage causing regularization of tumor vessels was selected to improve delivery of gemcitabine to the tumor. DCE-MRI was used to monitor regularization of vasculature with sunitinib in kidney tumors to schedule gemcitabine. We established an effective and nontoxic schedule of sunitinib combined with gemcitabine consisting of pretreatment with sunitinib for 3 days followed by four treatments of gemcitabine at 20 mg/kg given 3 days apart while continuing daily sunitinib treatment. This treatment caused significant tumor growth inhibition resulting in small residual tumor nodules exhibiting giant tumor cells with degenerative changes, which were observed both in kidney tumors and in spontaneous lung metastases, suggesting a systemic antitumor response. The combined therapy caused a significant increase in mouse survival. DCE-MRI monitoring of vascular changes induced by sunitinib, gemcitabine, and both combined showed increased tumor perfusion and decreased vascular permeability in kidney tumors. These findings, confirmed histologically by thinning of tumor blood vessels, suggest that both sunitinib and gemcitabine exert antiangiogenic effects in addition to cytotoxic antitumor activity. These studies show that DCE-MRI can be used to select the dose and schedule of antiangiogenic drugs to schedule chemotherapy and improve its efficacy.

SU‐GG‐I‐05: CT‐Perfusion and FDG‐PET Imaging in Rectal Cancer: New Insights into Tumor Vasculature

Purpose The purpose of this study was to prospectively investigate tumor perfusion in rectal cancer and to determine early changes after hypofractionated radiotherapy. Furthermore, we correlated FDG uptake and tumor perfusion parameters intratumorally. Method and Materials Thirty patients diagnosed with rectal cancer were prospectively included in this study. All patients underwent FDG‐PET and perfusion‐CT imaging prior to the start of radiotherapy treatment. The perfusion‐CT images were analyzed using the extended Kety‐model, quantifying tumor perfusion with three parameters: Ktrans, ve and vp. On the PET‐images, the tumor was delineated using automated SUV‐thresholding with the threshold (percentage of SUVmax within the tumor) depending on the tumor‐to‐background signal ratio. For each patient and for several tumor subrogions, the mean and maximum FDG uptake (SUV) and tumor perfusion (Ktrans) were quantified and correlated using the Spearman's rank correlation coefficient (ρ).Results The median tumors Ktrans values increased significantly after radiotherapy. Also, histogram analysis showed a shift of tumor voxels from lower Ktrans values towards higher Ktrans values. The median Ktrans values were significantly higher for tumor than for muscle tissue on both the pre‐scan and the post‐scan. In contrast, no differences between tumor and muscle tissues before and after radiotherapy were found for ve and vp. When correlating the tumors mean and maximum Ktrans values to the corresponding SUVs, positive correlations were found. Also when correlating the mean and maximum perfusion and FDG uptake within the created tumor subrogions, positive correlations were found. Conclusion Hypofractionated radiotherapy of rectal cancer leads to an increased tumor perfusion as reflected by an elevated Ktrans. FDG uptake was found to correlate with tumor perfusion assessed from dynamic perfusion‐CT images. Highly perfused rectal tumors presented with higher FDG uptake levels when compared to relatively low perfused tumors.

IFN-β Restricts Tumor Growth and Sensitizes Alveolar Rhabdomyosarcoma to Ionizing Radiation

Ionizing radiation is an important component of multimodal therapy for alveolar rhabdomyosarcoma (ARMS). We sought to evaluate the ability of IFN-beta to enhance the activity of ionizing radiation. Rh-30 and Rh-41 ARMS cells were treated with IFN-beta and ionizing radiation to assess synergistic effects in vitro and as orthotopic xenografts in CB17 severe combined immunodeficient mice. In addition to effects on tumor cell proliferation and xenograft growth, changes in the tumor microenvironment including interstitial fluid pressure, perfusion, oxygenation, and cellular histology were assessed. A nonlinear regression model and isobologram analysis indicated that IFN-beta and ionizing radiation affected antitumor synergy in vitro in the Rh-30 cell line; the activity was additive in the Rh-41 cell line. In vivo continuous delivery of IFN-beta affected normalization of the dysfunctional tumor vasculature of both Rh-30 and Rh-41 ARMS xenografts, decreasing tumor interstitial fluid pressure, increasing tumor perfusion (as assessed by contrast-enhanced ultrasonography), and increasing oxygenation. Tumors treated with both IFN-beta and radiation were smaller than control tumors and those treated with radiation or IFN-beta alone. Additionally, treatment with high-dose IFN-beta followed by radiation significantly reduced tumor size compared with radiation treatment followed by IFN-beta. The combination of IFN-beta and ionizing radiation showed synergy against ARMS by sensitizing tumor cells to the cytotoxic effects of ionizing radiation and by altering tumor vasculature, thereby improving oxygenation. Therefore, IFN-beta and ionizing radiation may be an effective combination for treatment of ARMS.

Tumor perfusion increases during hypofractionated short-course radiotherapy in rectal cancer: Sequential perfusion-CT findings

Purpose The purpose of this study was to investigate perfusion of rectal tumors and to determine early responses to short-course hypofractionated radiotherapy (RT). Material and methods Twenty-three rectal cancer patients were included, which underwent perfusion-CT imaging before (pre-scan) and after treatment (post-scan). Contrast-enhancement was measured in tumor and muscle tissues and in the external iliac artery. Perfusion was quantified with three pharmacokinetic parameters: K trans , v e and v p . Perfusion differences between tumor and normal tissue and changes of the pharmacokinetic parameters between both scans were evaluated. Results The median tumors K trans values increased significantly from the pre-scan (0.36 ± 0.11 (min −1)) to the post-scan (0.44 ± 0.13 (min −1)) ( p < 0.001). Also, histogram analysis showed a shift of tumor voxels from lower K trans values towards higher K trans values. Furthermore, the median K trans values were significantly higher for tumor than for muscle tissue on both the pre-scan (0.10 ± 0.05 (min −1), p < 0.001) and the post-scan (0.10 ± 0.04 (min −1), p < 0.001). In contrast, no differences between tumor and muscle tissues were found for v e and v p . Also, no significant differences were observed for v e and v p between the two pCT-imaging time-points. Conclusions Hypofractionated radiotherapy of rectal cancer leads to an increased tumor perfusion as reflected by an elevated K trans , possibly improving the bioavailability of cytotoxic agents in rectal tumors, often administered early after radiotherapy treatment.

Abstract C248: Targeting the tumor microenvironment: Vascular-specific damage by tirapazamine influenced by hypoxia and vascular stability

Abstract Background: Tirapazamine (TPZ; SR 4233; 3-amino-1,2,4-benzotriazine-1,4-di-N-oxide) is a Phase II/III bioreductive anti-cancer agent with greater toxicity to hypoxic vs oxygenated cells in vitro. We have previously reported that TPZ is able to mediate dose-dependent, irreversible central vascular dysfunction in vivo, leaving a hallmark viable rim of surviving peripheral vessels as seen with other vascular targeting agents (VTAs). Purpose: To explore how this agent, which is a bioreductive cytotoxin in vitro, is able to mediate catastrophic damage to tumor endothelium, likely the most oxygenated cell population in solid tumors, we have investigated the role of the tumor microenvironment including tumor hypoxia and vascular function. Methods: Vascular dysfunction may be qualitatively and quantitatively assessed using multiplex immunohistochemistry-based staining of frozen tumor sections for vasculature, perfusion, hypoxia, apoptosis and proliferation. Additional features of the microenvironment such as vascular permeability, architecture and support cells were also assessed using this tumor-mapping approach. Modification of tumor hypoxia was achieved via induction of mild anemia or low oxygen content gas breathing of tumor-bearing mice, and vascular function was increased by over-expression of VEGF in tumor models or systemic administration of nitric oxide (NO), or decreased via inhibition of nitric oxide synthase (NOS). Results: DCE-MRI studies correlated with tumor mapping data have shown that greater pre-treatment perfusion in HCT116 colorectal xenografts predicts for decreased sensitivity to the anti-vascular effects of TPZ. In contrast to observations using DCE-MRI, increasing tumor perfusion and vascular function caused an increase in tumor vessel sensitivity to TPZ, increasing both the magnitude and frequency of response in tumors. Both NOS inhibition and excess NO availability were able to potentiate anti-cancer effects of TPZ through enhancement of its vascular damage. Increasing tumor hypoxia by decreasing blood oxygenation was also found to potentiate the anti-vascular effects of TPZ, with otherwise resistant HT29 colorectal xenograft tumors showing a strong vascular dysfunction effect. Vascular architecture and tumor microenvironmental features were mapped in sensitive HCT116 and resistant HT29 colorectal xenograft models, with vascular maturity, permeability, microregional location of proliferating cells and hypoxia all identified as distinct in the two models. Summary: This work demonstrates that the vascular targeting effects of TPZ are potentiated by both hypoxia and vascular destabilization, and emphasizes the importance of specifically investigating the activity of anti-cancer agents in the context of the tumor microenvironment in vivo. This work was funded by a grant from CIHR &amp; Jennifer Baker is a recipient of a graduate studentship from Michael Smith Foundation for Health Research. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C248.

Dynamic Contrast-Enhanced Magnetic Resonance Imaging of Vascular Changes Induced by Sunitinib in Papillary Renal Cell Carcinoma Xenograft Tumors

To investigate further the antiangiogenic potential of sunitinib for renal cell carcinoma (RCC) treatment, its effects on tumor vasculature were monitored by dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) using an orthotopic KCI-18 model of human RCC xenografts in nude mice. Tumor-bearing mice were treated with various doses of sunitinib, and vascular changes were assessed by DCE-MRI and histologic studies. Sunitinib induced dose-dependent vascular changes, which were observed both in kidney tumors and in normal kidneys by DCE-MRI. A dosage of 10 mg/kg per day caused mild changes in Gd uptake and clearance kinetics in kidney tumors. A dosage of 40 mg/kg per day induced increased vascular tumor permeability with Gd retention, probably resulting from the destruction of tumor vasculature, and also caused vascular alterations of normal vessels. However, sunitinib at 20 mg/kg per day caused increased tumor perfusion and decreased vascular permeability associated with thinning and regularization of tumor vessels while mildly affecting normal vessels as confirmed by histologic diagnosis. Alterations in tumor vasculature resulted in a significant inhibition of KCI-18 RCC tumor growth at sunitinib dosages of 20 and 40 mg/kg per day. Sunitinib also exerted a direct cytotoxic effect in KCI-18 cells in vitro. KCI-18 cells and tumors expressed vascular endothelial growth factor receptor 2 and platelet-derived growth factor receptor β molecular targets of sunitinib that were modulated by the drug treatment. These data suggest that a sunitinib dosage of 20 mg/kg per day, which inhibits RCC tumor growth and regularizes tumor vessels with milder effects on normal vessels, could be used to improve blood flow for combination with chemotherapy. These studies emphasize the clinical potential of DCE-MRI in selecting the dose and schedule of antiangiogenic compounds.

Phase I study of vandetanib (ZD6474) administered during and after irradiation (RT) in children with diffuse intrinsic pontine glioma (DIPG)

10020 Background: Children with DIPG have a dismal prognosis despite use of RT, which is the mainstay of therapy. All chemotherapy regimens used so far demonstrated no benefit. EGFR and VEGFR pathways are considered important in tumorigenesis of DIPG. Methods: We conducted a traditional phase I study combining oral vandetanib (VEGFR-2 and EGFR inhibitor) during and after local RT in children with DIPG. Five dosage levels were tested (50, 65, 85, 110, and 145mg/m2 per day). Vandetanib and RT started on the same day. The first 6 weeks of therapy constituted the dose-limiting toxicity (DLT)-evaluation period. Correlative studies consisted of pharmacokinetic analysis (PK), pharmacodynamic studies in blood, and standard and investigational imaging (before and 1, 3, and 6 weeks after start of therapy). Results: Twenty-one patients were enrolled on study (50 [n = 3], 65 [n = 3], 85 [n = 3], 110 [n = 6], and 145mg/m2 [n = 6]). Two patients experienced DLT consisting of rash/mucositis (level 4) and diarrhea (level 5). The maximum-tolerated dose (MTD) of vandetanib was not reached. Other significant toxicities included lymphopenia grade 3/4 (n = 10), grade 3 neutropenia and hypophosphatemia (one each), grade 2 proteinuria (n = 2), grade 2 hypertension (n = 4), and mild QTc prolongation (n = 7) .Once the phase I component was completed, two extra patients were enrolled at dosage level 5; one of them developed grade 4 seizure secondary to posterior reversible encephalopathy syndrome. PK (n = 21) showed similar drug clearance and volume of distribution compared to adults. However, drug exposure at steady state normalized by dose seemed higher in children. Increased tumor perfusion during the first 6 weeks of therapy was observed in the first 12 patients analyzed. Conclusions: Although MTD was not reached, we recommend administration of vandetanib at a dose of 110mg/m2 per day during and after local RT in children. Further combination studies of vandetanib in children with DIPG are planned. No significant financial relationships to disclose.

Endothelin-1 inhibits prostate cancer growth in vivo through vasoconstriction of tumor-feeding arterioles

The vasoactive peptide endothelin-1 (ET-1) has been implicated in promoting the progression of prostate and other cancers though its precise mechanism(s)-of-action remain unclear. To better define the role of ET-1 in prostate cancer progression, we generated prostate cancer cell lines (PC-3 and 22Rv1) that express elevated levels of ET-1. As anticipated, increased ET-1 lead to modest autocrine growth stimulation of PC-3 cells in monolayer culture and increased colony formation in soft agar by both cell lines. Unexpectedly, however, metastatic colonization of 22Rv1 cells expressing elevated levels of ET-1 was reduced, as was the size of subcutaneous tumors produced by both 22Rv1- and PC-3 cells. Based on these data, we hypothesized that high levels of ET-1 may negatively impact the tumor microenvironment. We found that increased ET-1 expression did not consistently inhibit angiogenesis, indicating that this was not the cause of poor tumor growth. As an alternative explanation, we examined whether elevated ET-1 results in local vasoconstriction and thus reduces the blood supply available to the tumor. Consistent with this hypothesis, treatment of mice bearing PC-3 xenografts with a vasodilator increased tumor perfusion and partially restored tumor growth. Moreover, analysis of tumor vascular casts indicated vasoconstriction of tumor-feeding arterioles. Taken together, our data suggest that the local concentration of the ET-1 peptide is critical for determining a balance between its previously unrecognized tumor growth-suppressing activity (vasoconstriction) and known growth-promoting (mitogenesis, survival, and angiogenesis) activities. These findings may have implications for the modification of current prostate cancer therapies involving ET-1.

Use of dynamic susceptibility-contrast MRI (DSC-MRI) to assess perfusion changes in the ipsilateral brain parenchyma from glioblastoma

We investigated the effect of increased tumor perfusion using dynamic susceptibility-contrast magnetic resonance imaging (DSC-MRI) in glioblastoma (GBM) patients on the surrounding ipsilateral brain tissue with respect to perfusion of the normal, unaffected contralateral brain and of the tumor. DSC-MRI was performed in 11 patients with glioblastoma using a multislice T2*-weighed EPI sequence (TR/TE = 2,000/62 ms; FOV 240 mm; matrix 128 x 128; slice thickness 6 mm) on a standard clinical 1.5 Tesla scanner during intravenous injection of 40 cc Gadolinium-DTPA at a flow rate of 5 cc/s. Maps for relative regional cerebral blood volume (rCBV) and relative regional cerebral blood flow (rCBF) were created and relative values analyzed in relation to the arterial input. Relative CBV and CBF were significantly higher in gray matter than in the respective white matter (paired t-test; P < 0.001) with a high correlation for both perfusion parameters between the gray and the white matter in both ipsilateral and contralateral brain (P < 0.001). The highest values for rCBV and rCBF were found in solid tumor tissue with a significant positive correlation between tumor and the adjacent gray matter (for both rCBV and rCBF; P < 0.001). In GBM patients there is increased metabolism and thus increased rCBV and rCBF within the tumor. This increased perfusion of the tumor is not at the expense of perfusion of the ipsilateral normal brain parenchyma and in fact, the rCBV and rCBF values are linked to tumor-induced changes in rCBV and rCBF.