Dodecafluoropentane Emulsion as a Radiosensitizer in Glioblastoma Multiforme.

Purpose: Glioblastoma multiforme (GBM) is a hypoxic tumor resistant to radiotherapy. The purpose of this study was to assess the safety and efficacy of a novel oxygen therapeutic, dodecafluoropentane emulsion (DDFPe), in chemoradiation treatment of GBM. Experimental Design: In this multicenter phase Ib/II dose-escalation study, patients were administered DDFPe via intravenous infusion (0.05, 0.10, or 0.17 mL/kg) while breathing supplemental oxygen prior to each 2 Gy fraction of radiotherapy (30 fractions over 6 weeks). Patients also received standard-of-care chemotherapy [temozolomide (TMZ)]. Serial MRI scans were taken to monitor disease response. Adverse events were recorded and graded. TOLD (tissue oxygenation level-dependent) contrast MRI was obtained to validate modulation of tumor hypoxia. Results: Eleven patients were enrolled. DDFPe combined with radiotherapy and TMZ was well tolerated in most patients. Two patients developed delayed grade 3 radiation necrosis during dose escalation, one each at 0.1 and 0.17 mL/kg of DDFPe. Subsequent patients were treated at the 0.1 mL/kg dose level. Kaplan-Meier analysis showed a median overall survival of 19.4 months and a median progression-free survival of 9.6 months, which compares favorably to historical controls. Among 6 patients evaluable for TOLD MRI, a statistically significant reduction in tumor T1 was observed after DDFPe treatment. Conclusions: This trial, although small, showed that the use of DDFPe as a radiosensitizer in patients with GBM was generally safe and may provide a survival benefit. This is also the first time than TOLD MRI has shown reversal of tumor hypoxia in a clinical trial in patients. The recommended dose for phase II evaluation is 0.1 mL/kg DDFPe.Trial Registration: NCT02189109. Significance: This study shows that DDFPe can be safely administered to patients, and it is the first-in-human study to show reversal of hypoxia in GBM as measured by TOLD MRI. This strategy is being used in a larger phase II/III trial which will hopefully show a survival benefit by adding DDFPe during the course of fractionated radiation and concurrent chemotherapy.

Construction and validation of a novel aging-related gene signature and prognostic nomogram for predicting the overall survival in ovarian cancer.

BACKGROUND: Ovarian cancer (OC) is the most lethal gynecological malignancy. The objective of this study was to establish and validate an individual aging-related gene signature and a clinical nomogram that can powerfully predict independently the overall survival rate of patients with ovarian cancer. METHODS: Data on transcriptomic profile and relevant clinical information were retrieved from The Cancer Genome Atlas (TCGA) database as a training group, and the same data from three public Gene Expression Omnibus (GEO) databases as validation groups. Univariate Cox regression analysis, lasso regression analysis, and multiple multivariate Cox analysis were analyzed sequentially to select the genes to be included in the aging-associated signature. A risk scoring model was established and verified, the predictive value of the model was evaluated, and a clinical nomogram was established. RESULTS: We found eight genes that were most relevant to prognosis and constructed an eight-mRNA signature. Based on the model, each OC patient's risk score was able to be calculated and patients were split into groups of low and high risks with a distinct outcome. Survival analysis confirmed that the outcome of patients in the high-risk group was dramatically shorter than that of those in the low-risk group, and the eight-mRNA signature can be considered as a powerful and independent predictor that could predict the outcome of OC patient. Additionally, the risk score and age can be used to construct a clinical nomogram as a simpler tool for predicting prognosis. We also explored the association between the risk score and immunity and drug sensitivity. CONCLUSION: This study suggested that the aging-related gene signature could be used as an intervention point and latent prognostic predictor in OC, which may provide new perceptions for postoperative treatment strategies.

CircHIPK3 modulates VEGF through MiR-7 to affect ovarian cancer cell proliferation and apoptosis.

PURPOSE: The purpose of this study was to observe the effects of circHIPK3on the proliferation and apoptosis of ovarian cancer cells, and to further explore the potential mechanism therein. METHODS: CircHIPK3 was determined in the carcinoma tissues, normal adjacent tissues, and also in ovarian cancer cells via RT-PCR. The proliferation and apoptosis of cells were observed via colony-forming assay, 5-ethynyl-2'-deoxyuridine (EdU) staining and Western blotting. Moreover, the effect of the inhibition of circHIPK3 on the in vivo growth of ovarian cancer cells was detected using subcutaneous tumorigenesis assay. Finally, the effect of circHIPK3 on the expression of the micro ribonucleic acid (miR)-7/vascular endothelial growth factor (VEGF) signaling pathway in ovarian cancer cells was examined. RESULTS: CircHIPK3 in the carcinoma tissues was obviously higher than that in normal adjacent tissues. SKOV3 cell lines transfected with circHIPK3 inhibitor exhibited declined number of colonies. The inhibition of circHIPK3 distinctly suppressed the expression of B-cell lymphoma 2 (Bcl-2) and raised that of Bcl-2 associated X protein (Bax). Besides, the inhibition of circHIPK3 obviously weakened the tumorigenicity of ovarian cancer cells subcutaneously transplanted. Finally, it was found that miR-7 declined obviously and VEGF rose distinctly in the carcinoma tissues, and the in vitro assay verified the obvious increase in the expression of miR-7 and the prominently inhibited VEGF protein expression in the ovarian cancer cells with the inhibition of circHIPK3. CONCLUSIONS: CircHIPK3 has an obviously increased expression level in the carcinoma tissues of ovarian cancer patients, and the inhibition of circHIPK3 can activate the miR-7-mediated decline in the expression of VEGF to repress the proliferation and promote the apoptosis of ovarian cancer cells.

Non-Invasive Evaluation of Acute Effects of Tubulin Binding Agents: A Review of Imaging Vascular Disruption in Tumors.

Tumor vasculature proliferates rapidly, generally lacks pericyte coverage, and is uniquely fragile making it an attractive therapeutic target. A subset of small-molecule tubulin binding agents cause disaggregation of the endothelial cytoskeleton leading to enhanced vascular permeability generating increased interstitial pressure. The resulting vascular collapse and ischemia cause downstream hypoxia, ultimately leading to cell death and necrosis. Thus, local damage generates massive amplification and tumor destruction. The tumor vasculature is readily accessed and potentially a common target irrespective of disease site in the body. Development of a therapeutic approach and particularly next generation agents benefits from effective non-invasive assays. Imaging technologies offer varying degrees of sophistication and ease of implementation. This review considers technological strengths and weaknesses with examples from our own laboratory. Methods reveal vascular extent and patency, as well as insights into tissue viability, proliferation and necrosis. Spatiotemporal resolution ranges from cellular microscopy to single slice tomography and full three-dimensional views of whole tumors and measurements can be sufficiently rapid to reveal acute changes or long-term outcomes. Since imaging is non-invasive, each tumor may serve as its own control making investigations particularly efficient and rigorous. The concept of tumor vascular disruption was proposed over 30 years ago and it remains an active area of research.

In vivo hypoxia characterization using blood oxygen level dependent magnetic resonance imaging in a preclinical glioblastoma mouse model.

PURPOSE: Hypoxia measurements can provide crucial information regarding tumor aggressiveness, however current preclinical approaches are limited. Blood oxygen level dependent (BOLD) Magnetic Resonance Imaging (MRI) has the potential to continuously monitor tumor pathophysiology (including hypoxia). The aim of this preliminary work was to develop and evaluate BOLD MRI followed by post-image analysis to identify regions of hypoxia in a murine glioblastoma (GBM) model. METHODS: A murine orthotopic GBM model (GL261-luc2) was used and independent images were generated from multiple slices in four different mice. Image slices were randomized and split into training and validation cohorts. A 7 T MRI was used to acquire anatomical images using a fast-spin-echo (FSE) T2-weighted sequence. BOLD images were taken with a T2*-weighted gradient echo (GRE) and an oxygen challenge. Thirteen images were evaluated in a training cohort to develop the MRI sequence and optimize post-image analysis. An in-house MATLAB code was used to evaluate MR images and generate hypoxia maps for a range of thresholding and ΔT2* values, which were compared against respective pimonidazole sections to optimize image processing parameters. The remaining (n = 6) images were used as a validation group. Following imaging, mice were injected with pimonidazole and collected for immunohistochemistry (IHC). A test of correlation (Pearson's coefficient) and agreement (Bland-Altman plot) were conducted to evaluate the respective MRI slices and pimonidazole IHC sections. RESULTS: For the training cohort, the optimized parameters of "thresholding" (20 ≤ T2* ≤ 35 ms) and ΔT2* (±4 ms) yielded a Pearson's correlation of 0.697. These parameters were applied to the validation cohort confirming a strong Pearson's correlation (0.749) when comparing the respective analyzed MR and pimonidazole images. CONCLUSION: Our preliminary study supports the hypothesis that BOLD MRI is correlated with pimonidazole measurements of hypoxia in an orthotopic GBM mouse model. This technique has further potential to monitor hypoxia during tumor development and therapy.

Developing and validating utility parameters to establish patient-reported outcome-based perioperative symptom management in patients with lung cancer: a multicentre, prospective, observational cohort study protocol.

INTRODUCTION: Patient-reported outcome-based symptom monitoring and alerting have been attractive for patient care after a tumour-removal surgery. However, the implementation parameters of this patient-centred symptom management system in perioperative patients with lung cancer are still lacking. We aim to develop a perioperative symptom scale (PSS) for monitoring, to determine the optimal time points for symptom assessment and to define the alert thresholds for medical intervention. METHODS AND ANALYSIS: This study will prospectively recruit 300 patients undergoing lung cancer surgery in six hospitals. The MD Anderson Symptom Inventory-Lung Cancer Module (MDASI-LC) is used to collect longitudinal symptom data preoperatively, daily postoperatively during in-hospital stay and weekly after discharge until 4 weeks or the start of postoperative oncological therapy. Symptoms that change significantly over time will be generated as the PSS. We will determine the optimal time points for follow-up using the generalised linear mixed-effects models. The MDASI-LC interference-measured functional status will be used as the anchor for the alert thresholds. ETHICS AND DISSEMINATION: Ethics Committee of Sichuan Cancer Hospital approved this study on 16 October 2017 (No. SCCHEC-02-2017-042). The manuscript is based on the latest protocol of Version 3.0, 15 September 2019. The results of this study will be presented at medical conferences and published in peer-reviewed journals. TRIALS REGISTRATION NUMBER: NCT03341377.

Oxygen-sensitive MRI assessment of tumor response to hypoxic gas breathing challenge.

Oxygen-sensitive MRI has been extensively used to investigate tumor oxygenation based on the response (R2 * and/or R1 ) to a gas breathing challenge. Most studies have reported response to hyperoxic gas indicating potential biomarkers of hypoxia. Few studies have examined hypoxic gas breathing and we have now evaluated acute dynamic changes in rat breast tumors. Rats bearing syngeneic subcutaneous (n = 15) or orthotopic (n = 7) 13762NF breast tumors were exposed to a 16% O2 gas breathing challenge and monitored using blood oxygen level dependent (BOLD) R2 * and tissue oxygen level dependent (TOLD) T1 -weighted measurements at 4.7 T. As a control, we used a traditional hyperoxic gas breathing challenge with 100% O2 on a subset of the subcutaneous tumor bearing rats (n = 6). Tumor subregions identified as responsive on the basis of R2 * dynamics coincided with the viable tumor area as judged by subsequent H&E staining. As expected, R2 * decreased and T1 -weighted signal increased in response to 100% O2 breathing challenge. Meanwhile, 16% O2 breathing elicited an increase in R2 *, but divergent response (increase or decrease) in T1 -weighted signal. The T1 -weighted signal increase may signify a dominating BOLD effect triggered by 16% O2 in the relatively more hypoxic tumors, whereby the influence of increased paramagnetic deoxyhemoglobin outweighs decreased pO2 . The results emphasize the importance of combined BOLD and TOLD measurements for the correct interpretation of tumor oxygenation properties.

Examining correlations of oxygen sensitive MRI (BOLD/TOLD) with [18F]FMISO PET in rat prostate tumors.

Hypoxia is regarded as a potential prognostic biomarker for tumor aggressiveness, progression, and response to therapy. The radiotracer 18F-fluoromisonidazole ([18F]FMISO) has been used with positron emission tomography (PET) to reveal tumor hypoxia. Meanwhile, blood oxygen level dependent (BOLD) MRI and tissue oxygen level dependent (TOLD) MRI offer insight into oxygenation based on endogenous signals without the need for radiolabels. Here, we compared BOLD and TOLD MRI with [18F]FMISO uptake using Dunning prostate R3327-AT1 tumor bearing rats. BOLD and TOLD MRI were acquired with respect to an oxygen gas breathing challenge. The following day, dynamic PET was performed up to 90 minutes following IV injection of [18F]FMISO. Tumors showed distinct heterogeneity based on each technique. Correlations were observed between magnitude of mean BOLD or TOLD MRI signal responses to oxygen-breathing challenge and initial distribution of [18F]FMISO. Correlations were observed for whole tumor as well on a regional basis with stronger correlations in the well perfused tumor periphery indicating the strong influence of perfused vasculature. After 90 minutes most correlations with signal intensity became quite weak, but correlations were observed between hypoxic fraction based on FMISO and fractions of tumor showing BOLD or TOLD response in a subset of tumors. This emphasizes the importance of considering regional heterogeneity and responsive fractions, as opposed to simple magnitudes of responses. Although the data represent a small cohort of tumors they present direct correlations between oxygen sensitive MRI and PET hypoxia reporter agents in the same tumors, indicating the potential utility of further investigations.

The effect of flow on blood oxygen level dependent (R*2 ) MRI of orthotopic lung tumors.

PURPOSE: Blood oxygen level dependent (BOLD) MRI based on R2* measurements can provide insights into tumor vascular oxygenation. However, measurements are susceptible to blood flow, which may vary accompanying a hyperoxic gas challenge. We investigated flow sensitivity by comparing R2* measurements with and without flow suppression (fs) in 2 orthotopic lung xenograft tumor models. METHODS: H460 (n = 20) and A549 (n = 20) human lung tumor xenografts were induced by surgical implantation of cancer cells in the right lung of nude rats. MRI was performed at 4.7T after tumors reached 5 to 8 mm in diameter. A multiecho gradient echo MRI sequence was acquired with and without spatial saturation bands on each side of the imaging plane to evaluate the effect of flow on R2* . fs and non-fs R2* MRI measurements were interleaved during an oxygen breathing challenge (from air to 100% O2 ). T2* -weighted signal intensity changes (ΔSI(%)) and R2* measurements were obtained for regions of interest and on a voxel-by-voxel basis and discrepancies quantified with Bland-Altman analysis. RESULTS: Flow suppression affected ΔSI(%) and R2* measurements in each tumor model. Average discrepancy and limits of agreement from Bland-Altman analyses revealed greater flow-related bias in A549 than H460. CONCLUSION: The effect of flow on R2* , and hence BOLD, was tumor model dependent with measurements being more sensitive in well-perfused A549 tumors.

Synthesis of dihydronaphthalene analogues inspired by combretastatin A-4 and their biological evaluation as anticancer agents.

The natural products colchicine and combretastatin A-4 (CA4) have provided inspiration for the discovery and development of a wide array of derivatives and analogues that inhibit tubulin polymerization through a binding interaction at the colchicine site on ß-tubulin. A water-soluble phosphate prodrug salt of CA4 (referred to as CA4P) has demonstrated the ability to selectively damage tumor-associated vasculature and ushered in a new class of developmental anticancer agents known as vascular disrupting agents (VDAs). Through a long-term program of structure activity relationship (SAR) driven inquiry, we discovered that the dihydronaphthalene molecular scaffold provided access to small-molecule inhibitors of tubulin polymerization. In particular, a dihydronaphthalene analogue bearing a pendant trimethoxy aryl ring (referred to as KGP03) and a similar aroyl ring (referred to as KGP413) were potent inhibitors of tubulin polymerization (IC50 = 1.0 and 1.2 µM, respectively) and displayed low nM cytotoxicity against human cancer cell lines. In order to enhance water-solubility for in vivo evaluation, the corresponding phosphate prodrug salts (KGP04 and KGP152, respectively) were synthesized. In a preliminary in vivo study in a SCID-BALB/c mouse model bearing the human breast tumor MDA-MB-231-luc, a 99% reduction in signal was observed with bioluminescence imaging (BLI) 4 h after IP administration of KGP152 (200 mg kg-1) indicating reduced tumor blood flow. In a separate study, disruption of tumor-associated blood flow in a Fischer rat bearing an A549-luc human lung tumor was observed by color Doppler ultrasound following administration of KGP04 (15 mg kg-1).

Tomographic breathing detection: a method to noninvasively assess in situ respiratory dynamics.

Physiological monitoring is a critical aspect of in vivo experimentation, particularly imaging studies. Physiological monitoring facilitates gated acquisition of imaging data and more robust experimental interpretation but has historically required additional instrumentation that may be cumbersome. As frame rates have increased, imaging methods have been able to capture ever more rapid dynamics, passing the Nyquist sampling rate of most physiological processes and allowing the capture of motion, such as breathing. With this transition, image artifacts have also changed their nature; rather than intraframe motion causing blurring and deteriorating resolution, interframe motion does not affect individual frames and may be recovered as useful information from an image time series. We demonstrate a method that takes advantage of interframe movement for detection of gross physiological motion in real-time image sequences. We further demonstrate the ability of the method, dubbed tomographic breathing detection to quantify the dynamics of respiration, allowing the capture of respiratory information pertinent to anesthetic depth monitoring. Our example uses multispectral optoacoustic tomography, but it will be widely relevant to other technologies.

The vascular disrupting agent combretastatin A-4 phosphate causes prolonged elevation of proteins involved in heme flux and function in resistant tumor cells.

Vascular disrupting agents (VDAs) represent a promising class of anti-cancer drugs for solid tumor treatment. Here, we aim to better understand the mechanisms underlying tumor reccurrence and treatment resistance following the administration of a VDA, combretastatin A-4 phosphate (CA4P). Firstly, we used photoacoustic tomography to noninvasively map the effect of CA4P on blood oxygen levels throughout subcutaneous non-small cell lung cancer (NSCLC) tumors in mice. We found that the oxygenation of peripheral tumor vessels was significantly decreased at 1 and 3 hours post-CA4P treatment. The oxygenation of the tumor core reduced significantly at 1 and 3 hours, and reached anoxia after 24 hours. Secondly, we examined the effect of CA4P on the levels of proteins involved in heme flux and function, which are elevated in lung tumors. Using immunohistochemistry, we found that CA4P substantially enhanced the levels of enzymes involved in heme biosynthesis, uptake, and degradation, as well as oxygen-utilizing hemoproteins. Furthermore, measurements of markers of mitochondrial function suggest that CA4P did not diminish mitochondrial function in resistant tumor cells. These results suggest that elevated levels of heme flux and function contribute to tumor regrowth and treatment resistance post-VDA administration.

Oxygenation Imaging by Nuclear Magnetic Resonance Methods.

Oxygen monitoring is a topic of exhaustive research due to its central role in many biological processes, from energy metabolism to gene regulation. The ability to monitor in vivo the physiological distribution and the dynamics of oxygen from subcellular to macroscopic levels is a prerequisite to better understand the mechanisms associated with both normal and disease states (cancer, neurodegeneration, stroke, etc.). This chapter focuses on magnetic resonance imaging (MRI) based techniques to assess oxygenation in vivo. The first methodology uses injected fluorinated agents to provide quantitative pO2 measurements with high precision and suitable spatial and temporal resolution for many applications. The second method exploits changes in endogenous contrasts, i.e., deoxyhemoglobin and oxygen molecules through measurements of T 2* and T 1, in response to an intervention to qualitatively evaluate hypoxia and its potential modulation.

Incorporating Oxygen-Enhanced MRI into Multi-Parametric Assessment of Human Prostate Cancer.

Hypoxia is associated with prostate tumor aggressiveness, local recurrence, and biochemical failure. Magnetic resonance imaging (MRI) offers insight into tumor pathophysiology and recent reports have related transverse relaxation rate (R2*) and longitudinal relaxation rate (R1) measurements to tumor hypoxia. We have investigated the inclusion of oxygen-enhanced MRI for multi-parametric evaluation of tumor malignancy. Multi-parametric MRI sequences at 3 Tesla were evaluated in 10 patients to investigate hypoxia in prostate cancer prior to radical prostatectomy. Blood oxygen level dependent (BOLD), tissue oxygen level dependent (TOLD), dynamic contrast enhanced (DCE), and diffusion weighted imaging MRI were intercorrelated and compared with the Gleason score. The apparent diffusion coefficient (ADC) was significantly lower in tumor than normal prostate. Baseline R2* (BOLD-contrast) was significantly higher in tumor than normal prostate. Upon the oxygen breathing challenge, R2* decreased significantly in the tumor tissue, suggesting improved vascular oxygenation, however changes in R1 were minimal. R2* of contralateral normal prostate decreased in most cases upon oxygen challenge, although the differences were not significant. Moderate correlation was found between ADC and Gleason score. ADC and R2* were correlated and trends were found between Gleason score and R2*, as well as maximum-intensity-projection and area-under-the-curve calculated from DCE. Tumor ADC and R2* have been associated with tumor hypoxia, and thus the correlations are of particular interest. A multi-parametric approach including oxygen-enhanced MRI is feasible and promises further insights into the pathophysiological information of tumor microenvironment.

Tumor physiological changes during hypofractionated stereotactic body radiation therapy assessed using multi-parametric magnetic resonance imaging.

Radiation therapy is a primary treatment for non-resectable lung cancer and hypoxia is thought to influence tumor response. Hypoxia is expected to be particularly relevant to the evolving new radiation treatment scheme of hypofractionated stereotactic body radiation therapy (SBRT). As such, we sought to develop non-invasive tools to assess tumor pathophysiology and response to irradiation. We applied blood oxygen level dependent (BOLD) and tissue oxygen level dependent (TOLD) MRI, together with dynamic contrast enhanced (DCE) MRI to explore the longitudinal effects of SBRT on tumor oxygenation and vascular perfusion using A549 human lung cancer xenografts in a subcutaneous rat model. Intra-tumor heterogeneity was seen on multi-parametric maps, especially in BOLD, T2* and DCE. At baseline, most tumors showed a positive BOLD signal response (%ΔSI) and increased T2* in response to oxygen breathing challenge, indicating increased vascular oxygenation. Control tumors showed similar response 24 hours and 1 week later. Twenty-four hours after a single dose of 12 Gy, the irradiated tumors showed a significantly decreased T2* (-2.9±4.2 ms) and further decrease was observed (-4.0±6.0 ms) after 1 week, suggesting impaired vascular oxygenation. DCE revealed tumor heterogeneity, but showed minimal changes following irradiation. Rats were cured of the primary tumors by 3x12 Gy, providing long term survival, though with ultimate metastatic recurrence.

A role for dynamic contrast-enhanced magnetic resonance imaging in predicting tumour radiation response.

BACKGROUND: Dynamic contrast-enhanced (DCE) MRI may provide prognostic insights into tumour radiation response. This study examined quantitative DCE MRI parameters in rat tumours, as potential biomarkers of tumour growth delay following single high-dose irradiation. METHODS: Dunning R3327-AT1 prostate tumours were evaluated by DCE MRI following intravenous injection of Gd-DTPA. The next day tumours were irradiated (single dose of 30 Gy), while animals breathed air (n=4) or oxygen (n=4); two animals were non-irradiated controls. Growth was followed and tumour volume-quadrupling time (T4) was compared with pre-irradiation DCE assessments. RESULTS: Irradiation caused significant tumour growth delay (T4 ranged from 28 to 48 days for air-breathing rats, and 40 to 75 days for oxygen-breathing rats) compared with the controls (T4=7 to 9 days). A strong correlation was observed between T4 and extravascular-extracellular volume fraction (ve) irrespective of the gas inhaled during irradiation. There was also a correlation between T4 and volume transfer constant (K(trans)) for the air-breathing group alone. CONCLUSIONS: The data provide rationale for expanded studies of other tumour sites, types and progressively patients, and are potentially significant, as many patients undergo contrast-enhanced MRI as part of treatment planning.

Effective Rat Lung Tumor Model for Stereotactic Body Radiation Therapy.

Stereotactic body radiation therapy (SBRT) has found an important role in the treatment of patients with non-small cell lung cancer, demonstrating improvements in dose distribution and even tumor cure rates, particularly for early-stage disease. Despite its emerging clinical efficacy, SBRT has primarily evolved due to advances in medical imaging and more accurate dose delivery, leaving a void in knowledge of the fundamental biological mechanisms underlying its activity. Thus, there is a critical need for the development of orthotropic animal models to further probe the biology associated with high-dose-per-fraction treatment typical of SBRT. We report here on an improved surgically based methodology for generating solitary intrapulmonary nodule tumors, which can be treated with simulated SBRT using the X-RAD 225Cx small animal irradiator and Small Animal RadioTherapy (SmART) Plan treatment system. Over 90% of rats developed solitary tumors in the right lung. Furthermore, the tumor response to radiation was monitored noninvasively via bioluminescence imaging (BLI), and complete ablation of tumor growth was achieved with 36 Gy (3 fractions of 12 Gy each). We report a reproducible, orthotopic, clinically relevant lung tumor model, which better mimics patient treatment regimens. This system can be utilized to further explore the underlying biological mechanisms relevant to SBRT and high-dose-per-fraction radiation exposure and to provide a useful model to explore the efficacy of radiation modifiers in the treatment of non-small cell lung cancer.

Evaluation of tumor ischemia in response to an indole-based vascular disrupting agent using BLI and (19)F MRI.

Vascular disrupting agents (VDAs) have been proposed as an effective broad spectrum approach to cancer therapy, by inducing ischemia leading to hypoxia and cell death. A novel VDA (OXi8007) was recently reported to show rapid acute selective shutdown of tumor vasculature based on color-Doppler ultrasound. We have now expanded investigations to noninvasively assess perfusion and hypoxiation of orthotopic human MDA-MB-231/luc breast tumor xenografts following the administration of OXi8007 based on dynamic bioluminescence imaging (BLI) and magnetic resonance imaging (MRI). BLI showed significantly lower signal four hours after the administration of OXi8007, which was very similar to the response to combretastatin A-4P (CA4P), but the effect lasted considerably longer, with the BLI signal remaining depressed at 72 hrs. Meanwhile, control tumors exhibited minimal change. Oximetry used (19)F MRI of the reporter molecule hexafluorobenzene and FREDOM (Fluorocarbon Relaxometry using Echo Planar Imaging for Dynamic Oxygen Mapping) to assess pO2 distributions during air and oxygen breathing. pO2 decreased significantly upon the administration of OXi8007 during oxygen breathing (from 122 ± 64 to 34 ± 20 Torr), with further decrease upon switching the gas to air (pO2 = 17 ± 9 Torr). pO2 maps indicated intra-tumor heterogeneity in response to OXi8007, though ultimately all tumor regions became hypoxic. Both BLI and FREDOM showed the efficacy of OXi8007. The pO2 changes measured by FREDOM may be crucial for future study of combined therapy.

Assessment of tumor response to oxygen challenge using quantitative diffusion MRI in an animal model.

PURPOSE: To assess tumor response to oxygen challenge using quantitative diffusion magnetic resonance imaging (MRI). MATERIALS AND METHODS: A well-characterized Dunning R3327-AT1 rat prostate cancer line was implanted subcutaneously in the right thigh of male Copenhagen rats (n = 8). Diffusion-weighted images (DWI) with multiple b values (0, 25, 50, 100, 150, 200, 300, 500, 1000, 1500 s/mm(2) ) in three orthogonal directions were obtained using a multishot FSE-based Stejskal-Tanner DWI sequence (FSE-DWI) at 4.7T, while rats breathed medical air (21% oxygen) and with 100% oxygen challenge. Stretched-exponential and intravoxel incoherent motion (IVIM) models were used to calculate and compare quantitative diffusion parameters: diffusion heterogeneity index (α), intravoxel distribution of diffusion coefficients (DDC), tissue diffusivity (Dt), pseudo-diffusivity (Dp), and perfusion fraction (f) on a voxel-by-voxel basis. RESULTS: A significant increase of α (73.9 ± 4.7% in air vs. 78.1 ± 4.5% in oxygen, P = 0.0198) and a significant decrease of f (13.4 ± 3.7% in air vs. 10.4 ± 2.7% in oxygen, P = 0.0201) were observed to accompany oxygen challenge. Correlations between f and α during both air and oxygen breathing were found; the correlation coefficients (r) were -0.90 and -0.96, respectively. Positive correlations between Dt and DDC with oxygen breathing (r = 0.95, P = 0.0003), f and DDC with air breathing were also observed (r = 0.95, P = 0.0004). CONCLUSION: Quantitative diffusion MRI demonstrated changes in tumor perfusion in response to oxygen challenge.

Phosphatidylserine-targeted molecular imaging of tumor vasculature by magnetic resonance imaging.

Phosphatidylserine (PS), normally restricted to the inner leaflet of the plasma membrane, becomes exposed on the outer surface of viable endothelial cells in tumor vasculature, but not in normal blood vessels. In the present study, we report the use of PGN635, a novel human monoclonal antibody that specifically targets PS, for in vivo molecular MRI of tumor vasculature. The F(ab')2 fragments of PGN635 were conjugated to polyethylene glycol (PEG) coated iron oxide nanoparticles (IO). Targeting specificity of the PS-targeted Nanoprobe, IO-PGN635F(ab')2 was first confirmed by in vitro MRI and histological staining. In vivo longitudinal MRI was then performed before and after i.v. injection of IO-PGN635F(ab')2 into mice bearing 4T1 breast tumors. T2-weighted MR images at 9.4 T revealed inhomogeneous signal loss in tumor as early as 2 h post injection. Furthermore, ionizing radiation induced a significant increase in PS exposure on tumor vascular endothelial cells, resulting in significantly enhanced and sustained tumor contrast (p < 0.05). Spatially heterogeneous MRI contrast correlated well with histological staining of tumor vascular endothelium. Our studies suggest that PS exposed within the lumen of tumor vasculature is a highly specific and useful biomarker for targeted MRI contrast agents.