Emerging hyperthermia applications for pediatric oncology.

Local application of hyperthermia has a myriad of effects on the tumor microenvironment as well as the host's immune system. Ablative hyperthermia (typically > 55 °C) has been used both as monotherapy and adjuvant therapy, while mild hyperthermia treatment (39-45 °C) demonstrated efficacy as an adjuvant therapy through enhancement of both chemotherapy and radiation therapy. Clinical integration of hyperthermia has especially great potential in pediatric oncology, where current chemotherapy regimens have reached maximum tolerability and the young age of patients implies significant risks of late effects related to therapy. Furthermore, activation of both local and systemic immune response by hyperthermia suggests that hyperthermia treatments could be used to enhance the anticancer effects of immunotherapy. This review summarizes the state of current applications of hyperthermia in pediatric oncology and discusses the use of hyperthermia in the context of other available treatments and promising pre-clinical research.

Are Current Technical Exclusion Criteria for Clinical Trials of Magnetic Resonance-Guided High-Intensity Focused Ultrasound Too Restrictive?: Early Experiences at a Pediatric Hospital.

Certain technical criteria must be met to ensure the treatment safety of magnetic resonance-guided high-intensity focused ultrasound. We retrospectively reviewed how our enrollment criteria were applied from 2014 to 2017 in a clinical trial of magnetic resonance-guided high-intensity focused ultrasound ablation of recurrent malignant and locally aggressive benign solid tumors. Among the 36 screened patients between 2014 and 2017, more than one-third were excluded for technical exclusion criteria such as the anatomic location and proximity to prosthetics. Overall, patients were difficult to accrue for this trial, given the incidence of these tumors. To increase potential accrual, screening exclusion criteria could be more generalized and centered on the ability to achieve an acceptable treatment safety margin, rather than specifically excluding on the basis of general anatomic areas.

Feasibility of targeting canine soft tissue sarcoma with MR-guided high-intensity focused ultrasound.

PURPOSE: Magnetic resonance imaging-guided high-intensity-focused ultrasound (MR-HIFU) is a non-invasive treatment modality that precisely focuses ultrasound energy within a tumour and can be customised to result in a wide range of local bioeffects. The purpose of this study was to determine the feasibility of using MR-HIFU to treat soft tissue sarcoma (STS) in dogs. MATERIALS AND METHODS: Medical records of dogs admitted to the Virginia-Maryland College of Veterinary Medicine from 1 January 2012 to 31 December 2016 were searched for a diagnosis of sarcoma with available cross-sectional imaging of the tumour (MRI or CT). Fifty-three (53) dogs were eligible for inclusion. Tumor tissue (in bone as well as in soft tissue) was considered targetable unless: (1) the ultrasound path was completely obstructed by bone or gas and (2) the MR-HIFU target was within the spinal cord or less than 1 cm from the margin of the spinal cord. Tumors were categorised as <50% targetable, ≥50% targetable or non-targetable. RESULTS: Eighty-one percent of STS (81.1%, 43/53) were targetable. The head/spine tumour sites had the highest proportion of non-targetable tumours (36%, 9/25). The majority of truncal and axillary tumours were ≥50% targetable (88.9%, 16/18) ,and all extremity tumours were considered ≥50% targetable (100%, 5/5). CONCLUSIONS: The majority of STS were targetable. This is the first study to evaluate MR-HIFU targetability of canine STS. HIFU has potential as a therapeutic modality for treating STS in dogs, and this veterinary application is a possible model for treatment of naturally occurring STS in humans.

Mechanical fractionation of tissues using microsecond-long HIFU pulses on a clinical MR-HIFU system.

PURPOSE: High intensity focussed ultrasound (HIFU) can non-invasively treat tumours with minimal or no damage to intervening tissues. While continuous-wave HIFU thermally ablates target tissue, the effect of hundreds of microsecond-long pulsed sonications is examined in this work. The objective of this study was to characterise sonication parameter-dependent thermomechanical bioeffects to provide the foundation for future preclinical studies and facilitate clinical translation. METHODS AND MATERIALS: Acoustic power, number of cycles/pulse, sonication time and pulse repetition frequency (PRF) were varied on a clinical magnetic resonance imaging (MRI)-guided HIFU (MR-HIFU) system. Ex vivo porcine liver, kidney and cardiac muscle tissue samples were sonicated (3 × 3 grid pattern, 1 mm spacing). Temperature, thermal dose and T2 relaxation times were quantified using MRI. Lesions were histologically analysed using H&E and vimentin stains for lesion structure and viability. RESULTS: Thermomechanical HIFU bioeffects produced distinct types of fractionated tissue lesions: solid/thermal, paste-like and vacuolated. Sonications at 20 or 60 Hz PRF generated substantial tissue damage beyond the focal region, with reduced viability on vimentin staining, whereas H&E staining indicated intact tissue. Same sonication parameters produced dissimilar lesions in different tissue types, while significant differences in temperature, thermal dose and T2 were observed between the parameter sets. CONCLUSION: Clinical MR-HIFU system was utilised to generate distinct types of lesions and to produce targeted thermomechanical bioeffects in ex vivo tissues. The results guide HIFU research on thermomechanical tissue bioeffects, inform future studies and advice sonication parameter selection for direct tumour ablation or immunomodulation using a clinical MR-HIFU system.

Magnetic Resonance Imaging-guided High-intensity Focused Ultrasound Applications in Pediatrics: Early Experience at Children's National Medical Center.

Magnetic resonance imaging-guided high-intensity focused ultrasound (MR-HIFU) is a novel technology that integrates magnetic resonance imaging with therapeutic ultrasound. This unique approach provides a completely noninvasive method for precise thermal ablation of targeted tissues with real-time imaging feedback. Over the past 2 decades, MR-HIFU has shown clinical success in several adult applications ranging from treatment of painful bone metastases to uterine fibroids to prostate cancer and essential tremor. Although clinical experience in pediatrics is relatively small, the advantages of a completely noninvasive and radiation-free therapy are especially attractive to growing children. Unlike elderly patients, young children must deal with an entire lifetime of negative effects related to collateral tissue damage associated with invasive surgery, side effects of chemotherapy, and risk of secondary malignancy due to radiation exposure. These reasons provide a clear rationale and strong motivation to further advance clinical utility of MR-HIFU in pediatrics. We begin with an introduction to MR-HIFU technology and the clinical experience in adults. We then describe our early institutional experience in using MR-HIFU ablation to treat symptomatic benign, locally aggressive, and metastatic tumors in children and young adults. We also review some limitations and challenges encountered in treating pediatric patients and highlight additional pediatric applications which may be feasible in the near future.

Technical aspects of osteoid osteoma ablation in children using MR-guided high intensity focussed ultrasound.

BACKGROUND: Osteoid osteoma (OO) is a painful bone tumour occurring in children and young adults. Magnetic resonance imaging-guided high intensity focussed ultrasound (MR-HIFU) allows non-invasive treatment without ionising radiation exposure, in contrast to the current standard of care treatment with radiofrequency ablation (RFA). This report describes technical aspects of MR-HIFU ablation in the first 8 paediatric OO patients treated in a safety and feasibility clinical trial (total enrolment of up to 12 patients). MATERIALS AND METHODS: OO lesions and adjacent periosteum were treated with MR-HIFU ablation in 5-20 sonications (sonication duration = 16-48 s, frequency = 1.2 MHz, acoustic power = 20-160 W). Detailed treatment workflow, patient positioning and coupling strategies, as well as temperature and tissue perfusion changes were summarised and correlated. RESULTS: MR-HIFU ablation was feasible in all eight cases. Ultrasound standoff pads were shaped to conform to extremity contours providing acoustic coupling and aided patient positioning. The energy delivered was 10 ± 7 kJ per treatment, raising maximum temperature to 83 ± 3 °C. Post ablation contrast-enhanced MRI showed ablated volumes ranging 0.46-19.4 cm3 extending further into bone (7 ± 4 mm) than into soft tissue (4 ± 6 mm, p = 0.01, Mann-Whitney). Treatment time ranged 30-86 min for sonication and 160 ± 40 min for anaesthesia. No serious treatment-related adverse events were observed. Complete pain relief with no medication occurred in 7/8 patients within 28 days following treatment. CONCLUSIONS: MR-HIFU ablation of painful OO appears technically feasible in children and it may become a non-invasive and radiation-free alternative for painful OO. Therapy success, efficiency, and applicability may be improved through specialised equipment designed more specifically for extremity bone ablation.

Comparison of Noninvasive High-Intensity Focused Ultrasound with Radiofrequency Ablation of Osteoid Osteoma.

OBJECTIVE: To evaluate clinical feasibility and safety of magnetic resonance imaging-guided high-intensity focused ultrasound (MR-HIFU) treatment of symptomatic osteoid osteoma and to compare clinical response with standard of care treatment. STUDY DESIGN: Nine subjects with radiologically confirmed, symptomatic osteoid osteoma were treated with MR-HIFU in an institutional review board-approved clinical trial. Treatment feasibility and safety were assessed. Clinical response was evaluated in terms of analgesic requirement, visual analog scale pain score, and sleep quality. Anesthesia, procedure, and recovery times were recorded. This MR-HIFU group was compared with a historical control group of 9 consecutive patients treated with radiofrequency ablation. RESULTS: Nine subjects (7 male, 2 female; 16 ± 6 years) were treated with MR-HIFU without technical difficulties or any serious adverse events. There was significant decrease in their median pain scores 4 weeks within treatment (6 vs 0, P < .01). Total pain resolution and cessation of analgesics were achieved in 8 of 9 patients after 4 weeks. In the radiofrequency ablation group, 9 patients (8 male, 1 female; 10 ± 6 years) were treated in routine clinical practice. All 9 demonstrated complete pain resolution and cessation of medications by 4 weeks with a significant decrease in median pain scores (9 vs 0, P < .001). One developed a second-degree skin burn, but there were no other adverse events. Procedure times and treatment charges were comparable between the 2 groups. CONCLUSION: This pilot study shows that MR-HIFU treatment of osteoid osteoma refractory to medical therapy is feasible and can be performed safely in pediatric patients. Clinical response is comparable with standard of care treatment but without any incisions or exposure to ionizing radiation. TRIAL REGISTRATION: ClinicalTrials.govNCT02349971.

CEM43°C thermal dose thresholds: a potential guide for magnetic resonance radiofrequency exposure levels?

OBJECTIVE: To define thresholds of safe local temperature increases for MR equipment that exposes patients to radiofrequency fields of high intensities for long duration. These MR systems induce heterogeneous energy absorption patterns inside the body and can create localised hotspots with a risk of overheating. METHODS: The MRI + EUREKA research consortium organised a "Thermal Workshop on RF Hotspots". The available literature on thresholds for thermal damage and the validity of the thermal dose (TD) model were discussed. RESULTS/CONCLUSIONS: The following global TD threshold guidelines for safe use of MR are proposed: 1. All persons: maximum local temperature of any tissue limited to 39 °C 2. Persons with compromised thermoregulation AND (a) Uncontrolled conditions: maximum local temperature limited to 39 °C (b) Controlled conditions: TD < 2 CEM43°C 3. Persons with uncompromised thermoregulation AND (a) Uncontrolled conditions: TD < 2 CEM43°C (b) Controlled conditions: TD < 9 CEM43°C The following definitions are applied: Controlled conditions A medical doctor or a dedicated trained person can respond instantly to heat-induced physiological stress Compromised thermoregulation All persons with impaired systemic or reduced local thermoregulation KEY POINTS: • Standard MRI can cause local heating by radiofrequency absorption. • Monitoring thermal dose (in units of CEM43°C) can control risk during MRI. • 9 CEM43°C seems an acceptable thermal dose threshold for most patients. • For skin, muscle, fat and bone,16 CEM43°C is likely acceptable.

Reduction of peak acoustic pressure and shaping of heated region by use of multifoci sonications in MR-guided high-intensity focused ultrasound mediated mild hyperthermia.

PURPOSE: Ablative hyperthermia (>55 °C) has been used as a definitive treatment for accessible solid tumors not amenable to surgery, whereas mild hyperthermia (40-45 °C) has been shown effective as an adjuvant for both radiotherapy and chemotherapy. An optimal mild hyperthermia treatment is spatially accurate, with precise and homogeneous heating limited to the target region while also limiting the likelihood of unwanted thermal or mechanical bioeffects (tissue damage, vascular shutoff). Magnetic resonance imaging-guided high-intensity focused ultrasound (MR-HIFU) can noninvasively heat solid tumors under image-guidance. In a mild hyperthermia setting, a sonication approach utilizing multiple concurrent foci may provide the benefit of reducing acoustic pressure in the focal region (leading to reduced or no mechanical effects), while providing better control over the heating. The objective of this study was to design, implement, and characterize a multifoci sonication approach in combination with a mild hyperthermia heating algorithm, and compare it to the more conventional method of electronically sweeping a single focus. METHODS: Simulations (acoustic and thermal) and measurements (acoustic, with needle hydrophone) were performed. In addition, heating performance of multifoci and single focus sonications was compared using a clinical MR-HIFU platform in a phantom (target = 4-16 mm), in normal rabbit thigh muscle (target = 8 mm), and in a Vx2 tumor (target = 8 mm). A binary control algorithm was used for real-time mild hyperthermia feedback control (target range = 40.5-41 °C). Data were analyzed for peak acoustic pressure and intensity, heating energy efficiency, temperature accuracy (mean), homogeneity of heating (standard deviation [SD], T10 and T90), diameter and length of the heated region, and thermal dose (CEM(43)). RESULTS: Compared to the single focus approach, multifoci sonications showed significantly lower (67% reduction) peak acoustic pressures in simulations and hydrophone measurements. In a rabbit Vx2 tumor, both single focus and multifoci heating approaches were accurate (mean = 40.82±0.12 °C [single] and 40.70±0.09 °C [multi]) and precise (standard deviation = 0.65±0.05 °C [single] and 0.64±0.04 °C [multi]), producing homogeneous heating (T(10-90) = 1.62 °C [single] and 1.41 °C [multi]). Heated regions were significantly shorter in the beam path direction (35% reduction, p < 0.05, Tukey) for multifoci sonications, i.e., resulting in an aspect ratio closer to one. Energy efficiency was lower for the multifoci approach. Similar results were achieved in phantom and rabbit muscle heating experiments. CONCLUSIONS: A multifoci sonication approach was combined with a mild hyperthermia heating algorithm, and implemented on a clinical MR-HIFU platform. This approach resulted in accurate and precise heating within the targeted region with significantly lower acoustic pressures and spatially more confined heating in the beam path direction compared to the single focus sonication method.The reduction in acoustic pressure and improvement in spatial control suggest that multifoci heating is a useful tool in mild hyperthermia applications for clinical oncology.

Mild hyperthermia with magnetic resonance-guided high-intensity focused ultrasound for applications in drug delivery.

PURPOSE: Mild hyperthermia (40-45 °C) is a proven adjuvant for radiotherapy and chemotherapy. Magnetic resonance guided high intensity focused ultrasound (MR-HIFU) can non-invasively heat solid tumours under image guidance. Low temperature-sensitive liposomes (LTSLs) release their drug cargo in response to heat (>40 °C) and may improve drug delivery to solid tumours when combined with mild hyperthermia. The objective of this study was to develop and implement a clinically relevant MR-HIFU mild hyperthermia heating algorithm for combination with LTSLs. MATERIALS AND METHODS: Sonications were performed with a clinical MR-HIFU platform in a phantom and rabbits bearing VX2 tumours (target = 4-16 mm). A binary control algorithm was used for real-time mild hyperthermia feedback control (target = 40-41 °C). Drug delivery with LTSLs was measured with HPLC. Data were compared to simulation results and analysed for spatial targeting accuracy (offset), temperature accuracy (mean), homogeneity of heating (standard deviation (SD), T10 and T90), and thermal dose (CEM43). RESULTS: Sonications in a phantom resulted in better temperature control than in vivo. Sonications in VX2 tumours resulted in mean temperatures between 40.4 °C and 41.3 °C with a SD of 1.0-1.5 °C (T10 = 41.7-43.7 °C, T90 = 39.0-39.6 °C), in agreement with simulations. 3D spatial offset was 0.1-3.2 mm in vitro and 0.6-4.8 mm in vivo. Combination of MR-HIFU hyperthermia and LTSLs demonstrated heterogeneous delivery to a partially heated VX2 tumour, as expected. CONCLUSIONS: An MR-HIFU mild hyperthermia heating algorithm was developed, resulting in accurate and homogeneous heating within the targeted region in vitro and in vivo, which is suitable for applications in drug delivery.

Targeted drug delivery by high intensity focused ultrasound mediated hyperthermia combined with temperature-sensitive liposomes: computational modelling and preliminary in vivovalidation.

PURPOSE: To develop and validate a computational model that simulates (1) tissue heating with high intensity focused ultrasound (HIFU), and (2) resulting hyperthermia-mediated drug delivery from temperature-sensitive liposomes (TSL). MATERIALS AND METHODS: HIFU heating in tissue was simulated using a heat transfer model based on the bioheat equation, including heat-induced cessation of perfusion. A spatio-temporal multi-compartment pharmacokinetic model simulated intravascular release of doxorubicin from TSL, its transport into interstitium, and cell uptake. Two heating schedules were simulated, each lasting 30 min: (1) hyperthermia at 43 °C (HT) and (2) hyperthermia followed by a high temperature (50 °C for 20 s) pulse (HT+). As preliminary model validation, in vivo studies were performed in thigh muscle of a New Zealand White rabbit, where local hyperthermia with a clinical magnetic resonance-guided HIFU system was applied following TSL administration. RESULTS: HT produced a defined region of high doxorubicin concentration (cellular concentration ∼15-23 µg/g) in the target region. Cellular drug uptake was directly related to HT duration, with increasing doxorubicin uptake up to ∼2 h. HT+ enhanced drug delivery by ∼40% compared to HT alone. Temperature difference between model and experiment within the hyperthermia zone was on average 0.54 °C. Doxorubicin concentration profile agreed qualitatively with in vivo fluorescence profile. CONCLUSIONS: Computational models can predict temperature and delivered drug from combination of HIFU with TSL. Drug delivery using TSL may be enhanced by prolonged hyperthermia up to 2 h or by local cessation of vascular perfusion with a high temperature pulse following hyperthermia.

Image-guided drug delivery with magnetic resonance guided high intensity focused ultrasound and temperature sensitive liposomes in a rabbit Vx2 tumor model.

Clinical-grade doxorubicin encapsulated low temperature sensitive liposomes (LTSLs) were combined with a clinical magnetic resonance-guided high intensity focused ultrasound (MR-HIFU) platform to investigate in vivo image-guided drug delivery. Plasma pharmacokinetics were determined in 3 rabbits. Fifteen rabbits with Vx2 tumors within superficial thigh muscle were randomly assigned into three treatment groups: 1) free doxorubicin, 2) LTSL and 3) LTSL + MR-HIFU. For the LTSL + MR-HIFU group, mild hyperthermia (40-41 °C) was applied to the tumors using an MR-HIFU system. Image-guided non-invasive hyperthermia was applied for a total of 30 min, completed within 1h after LTSL infusion. High-pressure liquid chromatography (HPLC) analysis of the harvested tumor and organ/tissue homogenates was performed to determine doxorubicin concentration. Fluorescence microscopy was performed to determine doxorubicin spatial distribution in the tumors. Sonication of Vx2 tumors resulted in accurate (mean = 40.5 ± 0.1 °C) and spatially homogenous (SD = 1.0 °C) temperature control in the target region. LTSL + MR-HIFU resulted in significantly higher tumor doxorubicin concentrations (7.6- and 3.4-fold greater compared to free doxorubicin and LTSL respectively, p<0.05, Newman-Keuls). This improved tumor concentration was achieved despite heating <25% of the tumor volume. Free doxorubicin and LTSL treatments appeared to deliver more drug in the tumor periphery as compared to the tumor core. In contrast, LTSL + MR-HIFU treatment suggested an improved distribution with doxorubicin found in both the tumor periphery and core. Doxorubicin bio-distribution in non-tumor organs/tissues was fairly similar between treatment groups. This technique has potential for clinical translation as an image-guided method to deliver drug to a solid tumor.

Thresholds for thermal damage to normal tissues: an update.

The purpose of this review is to summarise a literature survey on thermal thresholds for tissue damage. This review covers published literature for the consecutive years from 2002-2009. The first review on this subject was published in 2003. It included an extensive discussion of how to use thermal dosimetric principles to normalise all time-temperature data histories to a common format. This review utilises those same principles to address sensitivity of a variety of tissues, but with particular emphasis on brain and testis. The review includes new data on tissues that were not included in the original review. Several important observations have come from this review. First, a large proportion of the papers examined for this review were discarded because time-temperature history at the site of thermal damage assessment was not recorded. It is strongly recommended that future research on this subject include such data. Second, very little data is available examining chronic consequences of thermal exposure. On a related point, the time of assessment of damage after exposure is critically important for assessing whether damage is transient or permanent. Additionally, virtually no data are available for repeated thermal exposures which may occur in certain recreational or occupational activities. For purposes of regulatory guidelines, both acute and lasting effects of thermal damage should be considered.

Formulation and characterisation of magnetic resonance imageable thermally sensitive liposomes for use with magnetic resonance-guided high intensity focused ultrasound.

PURPOSE: Objectives of this study were to: 1) develop iLTSL, a low temperature sensitive liposome co-loaded with an MRI contrast agent (ProHance® Gd-HP-DO3A) and doxorubicin, 2) characterise doxorubicin and Gd-HP-DO3A release from iLTSL and 3) investigate the ability of magnetic resonance-guided high intensity focused ultrasound (MR-HIFU) to induce and monitor iLTSL content release in phantoms and in vivo. METHODS: iLTSL was passively loaded with Gd-HP-DO3A and actively loaded with doxorubicin. Doxorubicin and Gd-HP-DO3A release was quantified by fluorescence and spectroscopic techniques, respectively. Release with MR-HIFU was examined in tissue-mimicking phantoms containing iLTSL and in a VX2 rabbit tumour model. RESULTS: iLTSL demonstrated consistent size and doxorubicin release kinetics after storage at 4°C for 7 days. Release of doxorubicin and Gd-HP-DO3A from iLTSL was minimal at 37°C but fast when heated to 41.3°C. The magnitude of release was not significantly different between doxorubicin and Gd-HP-DO3A over 10 min in HEPES buffer and plasma at 37°, 40° and 41.3°C (p > 0.05). Relaxivity of iLTSL increased significantly (p < 0.0001) from 1.95 ± 0.05 to 4.01 ± 0.1 mMs⁻¹ when heated above the transition temperature. Signal increase corresponded spatially and temporally to MR-HIFU-heated locations in phantoms. Signal increase was also observed in vivo after iLTSL injection and after each 10-min heating (41°C), with greatest increase in the heated tumour region. CONCLUSION: An MR imageable liposome formulation co-loaded with doxorubicin and an MR contrast agent was developed. Stability, imageability, and MR-HIFU monitoring and control of content release suggest that MR-HIFU combined with iLTSL may enable real-time monitoring and spatial control of content release.

Comparative effects of thermosensitive doxorubicin-containing liposomes and hyperthermia in human and murine tumours.

PURPOSE: In previous reports, laboratory-made lysolecithin-containing thermosensitive liposome encapsulating doxorubicin (LTSL-DOX) showed potent anticancer effects in FaDu human squamous cell carcinoma. To further study the spectrum of LTSL-DOX activity, the efficacy of its commercial formulation was re-examined in FaDu and compared in HCT116, PC3, SKOV-3 and 4T07 cancer cell lines. Factors that may influence differences in HT-LTSL-DOX efficacy were also examined. METHODS: Anticancer effect was measured using standard growth delay methods. We measured doubling time and clonogenic survival after doxorubicin exposure in vitro, and interstitial pH and drug concentrations in vivo. RESULTS: In all five tumour types, HT-LTSL-DOX increased median tumour growth time compared with untreated controls (p < 0.0006) and HT alone (p < 0.01), and compared with LTSL-DOX alone in FaDu, PC-3 and HCT-116 (p < 0.0006). HT-LTSL-DOX yielded significantly higher drug concentrations than LTSL-DOX (p < 0.0001). FaDu was most sensitive (p < 0.0014) to doxorubicin (IC(50) = 90 nM) in vitro, compared to the other cell lines (IC(50) = 129-168 nM). Of the parameters tested for correlation with efficacy, only the correlation of in vitro doubling time and in vivo median growth time was significant (Pearson r = 0.98, p = 0.0035). Slower-growing SKOV-3 and PC-3 had the greatest numbers of complete regressions and longest tumour growth delays, which are clinically important parameters. CONCLUSIONS: These results strongly suggest that variations in anti-tumour effect of HT-LTSL-DOX are primarily related to in vitro doubling time. In the clinic, the rate of tumour progression must be considered in design of treatment regimens involving HT-LTSL-DOX.

Effect of pazopanib on tumor microenvironment and liposome delivery.

Pathologic angiogenesis creates an abnormal microenvironment in solid tumors, characterized by elevated interstitial fluid pressure (IFP) and hypoxia. Emerging theories suggest that judicious downregulation of proangiogenic signaling pathways may transiently "normalize" the vascular bed, making it more suitable for drug delivery and radiotherapy. In this work, we investigate the role of pazopanib, a small-molecule inhibitor of vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) receptors, on tumor IFP, angiogenesis, hypoxia, and liposomal drug delivery. Nude mice bearing A549 human non-small cell lung cancer xenografts were treated with 100 mg/kg pazopanib (n = 20) or vehicle (n = 20) through oral gavage for 8 days, followed by a one-time intravenous dose of 10 mg/kg Doxil (liposomal doxorubicin). Pazopanib treatment resulted in significant reduction of tumor IFP and decreased vessel density, assessed by CD31 staining. Despite these trends toward normalization, high-performance liquid chromatography revealed no differences in doxorubicin concentration between pazopanib-treated and control tumors, with Doxil penetration from microvessels being significantly reduced in the pazopanib group. Additionally, tumor hypoxia, evaluated by CA-IX immunostaining and confirmed in a second study by EF5 expression (n = 4, 100 mg/kg pazopanib; n = 4, vehicle), was increased in pazopanib-treated tumors. Our results suggest that the classic definition of tumor "normalization" may undermine the crucial role of vessel permeability and oncotic pressure gradients in liposomal drug delivery, and that functional measures of normalization, such as reduced IFP and hypoxia, may not occur in parallel temporal windows.

Application of mixed spin iMQCs for temperature and chemical-selective imaging.

The development of accurate and non-invasive temperature imaging techniques has a wide variety of applications in fields such as medicine, chemistry and materials science. Accurate detection of temperature both in phantoms and in vivo can be obtained using iMQCs (intermolecular multiple quantum coherences), as demonstrated in a recent paper. This paper describes the underlying theory of iMQC temperature detection, as well as extensions of that work allowing not only for imaging of absolute temperature but also for imaging of analyte concentrations through chemically-selective spin density imaging.

Quantitative diffuse reflectance and fluorescence spectroscopy: tool to monitor tumor physiology in vivo.

This study demonstrates the use of optical spectroscopy for monitoring tumor oxygenation and metabolism in response to hyperoxic gas breathing. Hemoglobin saturation and redox ratio were quantified for a set of 14 and 9 mice, respectively, measured at baseline and during carbogen breathing (95% O(2), 5% CO(2)). In particular, significant increases in hemoglobin saturation and fluorescence redox ratio were observed upon carbogen breathing. These data were compared with data obtained concurrently using an established invasive technique, the OxyLite partial oxygen pressure (pO(2)) system, which also showed a significant increase in pO(2). It was found that the direction of changes were generally the same between all of the methods, but that the OxyLite system was much more variable in general, suggesting that optical techniques may provide a better assessment of global tumor physiology. Optical spectroscopy measurements are demonstrated to provide a reliable, reproducible indication of changes in tumor physiology in response to physiologic manipulation.

An in vitro system to evaluate the effects of ischemia on survival of cells used for cell therapy.

Maintaining cell viability is a major challenge associated with transplanting cells into ischemic myocardium to restore function. A likely contributor to significant cell death during cardiac cell therapy is hypoxia/anoxia. We developed a system that enabled quantification and association of cell survival with oxygen and nutrient values within in vitro constructs. Myoblasts were suspended in 2% collagen gels in 1 cm diameter x 1 cm deep constructs. At 48 +/- 3 h post-seeding, oxygen levels were measured using microelectrodes and gels were snap-frozen. Bioluminescence metabolite imaging and TUNEL staining were performed on cryosections. Oxygen and glucose consumption and lactate production rates were calculated by fitting data to Fick's second law of diffusion with Michaelis-Menten kinetics. Oxygen levels dropped to 0 mmHg and glucose levels dropped from 4.28 to 3.18 mM within the first 2000 mum of construct depth. Cell viability dropped to approximately 40% over that same distance and continued to drop further into the construct. We believe this system provides a reproducible and controllable test bed to compare survival, proliferation, and phenotype of various cell inputs (e.g., myoblasts, mesenchymal stem cells, and cardiac stem cells) and the impact of different treatment regimens on the likelihood of survival of transplanted cells.

Magnetic resonance imaging of temperature-sensitive liposome release: drug dose painting and antitumor effects.

BACKGROUND: In preclinical studies, lysolipid-based temperature-sensitive liposomes (LTSLs) containing chemotherapy drugs administered in combination with local hyperthermia have been found to increase tumor drug concentrations and improve antitumor efficacy of the drugs. We used a novel magnetic resonance imaging (MRI) method to measure the temporal and spatial patterns of drug delivery in a rat fibrosarcoma model during treatment with LTSLs containing doxorubicin and an MRI contrast agent (manganese) (Dox/Mn-LTSLs) administered at different times with respect to hyperthermia. METHODS: Rats bearing 10- to 12-mm fibrosarcomas (n = 6-7 per group) were treated with Dox/Mn-LTSLs (at a dose of 5 mg doxorubicin/kg body weight) before and/or during 60 minutes of local tumor hyperthermia administered via a catheter inserted at the center of the tumor. Drug distribution was monitored continuously via MRI. Magnetic resonance changes were used to calculate intratumoral doxorubicin concentrations throughout treatment. Tumors were monitored until they reached five times their volume on the day of treatment or 60 days. Doxorubicin concentrations and times for tumors to reach five times their volume on the day of treatment were analyzed using the Kruskal-Wallis test and the Kaplan-Meier product-limit method, respectively. All statistical tests were two-sided. RESULTS: Administration of Dox/Mn-LTSLs before, during, and both before and during hyperthermia yielded central, peripheral, and uniform drug distributions, respectively. Doxorubicin accumulated more quickly and reached higher concentrations in the tumor when Dox/Mn-LTSLs were administered during hyperthermia than when administered before hyperthermia (rate: 9.8 versus 1.8 microg/min, difference = 8.0 microg/min, 95% confidence interval [CI] = 6.8 to 12.8 microg/min, P = .003; concentration: 15.1 versus 8.0 ng/mg, difference = 7.1 ng/mg, 95% CI = 3.6 to 10.6 ng/mg, P = .028). LTSL administered during hyperthermia also yielded the greatest antitumor effect, with a median time for tumors to reach five times their volume on the day of treatment of 34 days (95% CI = 30 days to infinity) compared with 18.5 days (95% CI = 16 to 23 days) for LTSL before hyperthermia and 22.5 days (95% CI = 15 to 25 days) for LTSL before and during hyperthermia. CONCLUSIONS: In this rat fibrosarcoma model, LTSLs were most effective when delivered during hyperthermia, which resulted in a peripheral drug distribution.