Supplemental Movie 4: Animated EVAC streamlines (grey lines), showing the path taken by interstitial fluid, overlaid on vascular perfusion measurements (colour scale, measured in vivo using arterial spin labelling) and the location of blood vessels (yellow structures, measured ex vivo using micro-CT). The data were acquired in an SW1222 colorectal carcinoma tumour xenograft.

Supplemental Movie 3: Animated EVAC streamlines (grey lines), showing the path taken by interstitial fluid, overlaid on vascular perfusion measurements (colour scale, measured in vivo using arterial spin labelling) and the location of blood vessels (yellow structures, measured ex vivo using micro-CT). The data were acquired in an LS174T colorectal carcinoma tumour xenograft.

Supplemental Movie 5: Interstitial convection, measured using EVAC-MRI and animated using a particle simulation (left), compared with MRI signal enhancement with time, following injection with a contrast agent (Gd-DTPA), in an LS174T tumour.

<p>Supplemental Movie 2: Animated EVAC streamlines (grey lines), showing the path taken by interstitial fluid, overlaid on vascular perfusion measurements (colour scale, measured in vivo using arterial spin labelling) and the location of blood vessels (yellow structures, measured ex vivo using micro-CT). The data were acquired in an LS174T colorectal carcinoma tumour xenograft.</p>

Supplemental Movie 1: Schematic movie illustrating the physical principles underpinning the convection-MRI sequence

PDF file - 1763KB, This supplemental material includes S1), a figure with the fit of the VERDICT model to the diffusion data from all the samples, S2), a figure demonstrating the quality of the diffusion images and the signal attenuation from an example sample of each cell line, TS1) a table with the orientation angle estimates of the VERDICT model, S3) a figure to demonstrate that VERDICT performs better than a tensor model, a two-tensor model and an isotropic version of the VERDICT model. We also include details of all the histological procedures as well as extra information about the choice of the vascular compartment in the VERDICT model.

Magnetic hyperthermia (MH) harnesses the heat-releasing properties of superparamagnetic iron oxide nanoparticles (SPIONs) and has potential to stimulate immune activation in the tumor microenvironment whilst sparing surrounding normal tissues. To assess feasibility of localized MH in vivo, SPIONs are injected intratumorally and their fate tracked by Zirconium-89-positron emission tomography, histological analysis, and electron microscopy. Experiments show that an average of 49% (21-87%, n = 9) of SPIONs are retained within the tumor or immediately surrounding tissue. In situ heating is subsequently generated by exposure to an externally applied alternating magnetic field and monitored by thermal imaging. Tissue response to hyperthermia, measured by immunohistochemical image analysis, reveals specific and localized heat-shock protein expression following treatment. Tumor growth inhibition is also observed. To evaluate the potential effects of MH on the immune landscape, flow cytometry is used to characterize immune cells from excised tumors and draining lymph nodes. Results show an influx of activated cytotoxic T cells, alongside an increase in proliferating regulatory T cells, following treatment. Complementary changes are found in draining lymph nodes. In conclusion, results indicate that biologically reactive MH is achievable in vivo and can generate localized changes consistent with an anti-tumor immune response.

There are several limitations with monodrug cancer therapy, including poor bioavailability, rapid clearance and drug resistance. Combination therapy addresses these by exploiting synergism between different drugs against cancer cells. In particular, the combination of epigenetic therapies with conventional chemotherapeutic agents can improve the initial tumour response and overcome acquired drug resistance. Co-encapsulation of multiple therapeutic agents into a single polymeric nanoparticle is one of the many approaches taken to enhance therapeutic effect and improve the pharmacokinetic profile. In this study, different types of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs), matrix and core–shell (CS), were investigated for simultaneous encapsulation of a demethylating drug, decitabine, and a potent anticancer agent, cisplatin. It was shown that by altering the configuration of the CS structure, the release profile could be tuned. In order to investigate whether this could enhance the anticancer effect compared to cisplatin, human ovarian carcinoma cell line (A2780) and its cisplatin resistant variant (A2780cis) were exposed to free cisplatin and the CS–NPs. A better response was obtained in both cell lines (11% and 51% viability of A2780 and A2780cis, respectively) using CS–NPs than cisplatin alone (27%, 82% viability of A2780 and A2780cis, respectively) or in combination with decitabine (22%, 96% viability of A2780 and A2780cis, respectively) at equivalent doses (10 μM).

Induction of apoptosis is often necessary for successful cancer therapy, and the non-invasive monitoring of apoptosis post-therapy could assist in clinical decision making. Isatins are a class of compounds that target activated caspase-3 during apoptosis. Here we report the synthesis of the 5-iodo-1,2,3-triazole (FITI) analog of the PET tracer [ 18 F]ICMT11 as a candidate tracer for imaging of apoptosis with SPECT, as well as PET. Labelling with radioiodine ( 123,125 I) was achieved in 55 ± 12% radiochemical yield through a chelator-accelerated one-pot cycloaddition reaction mediated by copper(I) catalysis. The caspase-3 binding affinity and selectivity of FITI compares favourably to that of [ 18 F]ICMT11 (K i = 6.1 ± 0.9 nM and 12.4 ± 4.7 nM, respectively). In biodistribution studies, etoposide-induced cell death in a SW1222 xenograft model resulted in a 2-fold increase in tumour uptake of the tracer. However, the tumour uptake was too low to allow in vivo imaging of apoptosis with SPECT.

Abstract Purpose: Photoacoustic imaging (PAI) is a novel noninvasive and nonionizing imaging technique that allows longitudinal imaging of tumor vasculature in vivo and monitoring of response to therapy, especially for vascular targeted chemotherapy agents. In this study, we used a novel high-resolution all-optical PAI scanner to observe the pharmacodynamic response to the vascular-disrupting agent OXi4503. Experimental Design: Two models of colorectal carcinoma (SW1222 and LS174T) that possess differing pathophysiologic vascularization were established as subcutaneous tumors in mice. Monitoring of response was performed over a 16-day “regrowth” period following treatment at 40 mg/kg, and at day 2 for a “dose response” study at 40 mg/kg, 10 mg/kg, 1 mg/kg, and sham dose. Results: Qualitative and quantitative changes in PA signal are observed, with an initial decrease followed by a plateau and subsequent return of signal indicating regrowth. Both tumor types exhibited a decrease in signal; however, the more vascularized SW1222 tumors show greater response to treatment. Decreasing the dose of OXi4503 led to a decrease in PA signal intensity of 60%, 52%, and 20% in SW1222 tumors and 30%, 26%, and 4% for LS174T tumors. Conclusions: We have shown for the first time that PAI can observe the pharmacodynamic response of tumor vasculature to drug treatment both longitudinally and at different dose levels. Assessment of differing response to treatment based on vascular pathophysiologic differences among patients has the potential to provide personalized drug therapy; we have demonstrated that PAI, which is clinically translatable, could be a powerful tool for this purpose.

Proteasome inhibitors (PIs) are now standard of care for several cancers, and noninvasive biomarkers of treatment response are critically required for early patient stratification and treatment personalization. The present study evaluated whether chemical exchange (CEST) magnetic resonance imaging (MRI) can provide measurements that can be used as the noninvasive biomarkers of proteasome inhibition, alongside diffusion MRI and relaxometry. The sensitivity of human colorectal carcinoma cells to the PI Ixazomib was assessed via in vitro and in vivo dose-response experiments. Acute in vivo response to Ixazomib was assessed at three dosing concentrations, using CEST MRI (amide, amine, hydroxyl signals), diffusion MRI (ADC) and relaxometry (T1, T2). These responses were further evaluated with the known histological markers for Ixazomib and Bradford assay ex vivo. The CEST signal from amides and amines increased in proportion to Ixazomib dose in colorectal cancer xenografts. The cell lines differed in their sensitivity to Ixazomib, which was reflected in the MRI measurements. A mild stimulation in tumor growth was observed at low Ixazomib doses. Our results identify CEST MRI as a promising method for safely and noninvasively monitoring disrupted tumor protein homeostasis induced by proteasome inhibitor treatment, and for stratifying sensitivity between tumor types.

The magnetic properties and safety of dextran-coated superparamagnetic iron oxide nanoparticles (SPIONs) have facilitated their clinical use as MRI contrast agents and stimulated research on applications for SPIONs in particle imaging and magnetic hyperthermia. The wider clinical potential of SPIONs, however, has been limited by their rapid removal from circulation via the reticuloendothelial system (RES). We explored the possibility of extending SPION circulatory time using fucoidan, a seaweed-derived food supplement, to inhibit RES uptake. The effects of fucoidan on SPION biodistribution were evaluated using ferucarbotran, which in its pharmaceutical formulation (Resovist) targets the RES. Ferucarbotran was radiolabeled at the iron oxide core with technetium-99m (99mTc; t1/2 = 6 h) or zirconium-89 (89Zr; t1/2 = 3.3 days). Results obtained with 99mTc-ferucarbotran demonstrated that administration of fucoidan led to a 4-fold increase in the circulatory half-life (t1/2 slow) from 37.4 to 150 min (n = 4; P < 0.0001). To investigate whether a longer circulatory half-life could lead to concomitant increased tumor uptake, the effects of fucoidan were tested with 89Zr-ferucarbotran in mice bearing syngeneic subcutaneous (GL261) tumors. In this model, the longer circulatory half-life achieved with fucoidan was associated with a doubling in tumor SPION uptake (n = 5; P < 0.001). Fucoidan was also effective in significantly increasing the circulatory half-life of perimag-COOH, a commercially available SPION with a larger hydrodynamic size (130 nm) than ferucarbotran (65 nm). These findings indicate successful diversion of SPIONs away from the hepatic RES and show realistic potential for future clinical applications.

Several distinct fluid flow phenomena occur in solid tumors, including intravascular blood flow and interstitial convection. Interstitial fluid pressure is often raised in solid tumors, which can limit drug delivery. To probe low-velocity flow in tumors resulting from raised interstitial fluid pressure, we developed a novel MRI technique named convection-MRI, which uses a phase-contrast acquisition with a dual-inversion vascular nulling preparation to separate intra- and extravascular flow. Here, we report the results of experiments in flow phantoms, numerical simulations, and tumor xenograft models to investigate the technical feasibility of convection-MRI. We observed a significant correlation between estimates of effective fluid pressure from convection-MRI with gold-standard, invasive measurements of interstitial fluid pressure in mouse models of human colorectal carcinoma. Our results show how convection-MRI can provide insights into the growth and responsiveness to vascular-targeting therapy in colorectal cancers.Significance: A noninvasive method for measuring low-velocity fluid flow caused by raised fluid pressure can be used to assess changes caused by therapy. Cancer Res; 78(7); 1859-72. ©2018 AACR.

Human pancreatic ductal adenocarcinoma (PDAC) involves the dysregulation of multiple signaling pathways. A novel approach to the treatment of PDAC is described, involving the targeting of cancer genes in PDAC pathways having over-representation of G-quadruplexes, using the trisubstituted naphthalene diimide quadruplex-binding compound 2,7-bis(3-morpholinopropyl)-4-((2-(pyrrolidin-1-yl)ethyl)amino)benzo[lmn][3,8]phenanthroline-1,3,6,8(2H,7H)-tetraone (CM03). This compound has been designed by computer modeling, is a potent inhibitor of cell growth in PDAC cell lines, and has anticancer activity in PDAC models, with a superior profile compared to gemcitabine, a commonly used therapy. Whole-transcriptome RNA-seq methodology has been used to analyze the effects of this quadruplex-binding small molecule on global gene expression. This has revealed the down-regulation of a large number of genes, rich in putative quadruplex elements and involved in essential pathways of PDAC survival, metastasis, and drug resistance. The changes produced by CM03 represent a global response to the complexity of human PDAC and may be applicable to other currently hard-to-treat cancers.

The combination of early diagnosis and complete surgical resection offers the greatest prospect of curative cancer treatment. An iodine-124/fluorescein-based dual-modality labeling reagent, 124I-Green, constitutes a generic tool for one-step installation of a positron emission tomography (PET) and a fluorescent reporter to any cancer-specific antibody. The resulting antibody conjugate would allow both cancer PET imaging and intraoperative fluorescence-guided surgery. 124I-Green was synthesized in excellent radiochemical yields of 92 ± 5% (n = 4) determined by HPLC with an improved one-pot three-component radioiodination reaction. The A5B7 carcinoembryonic antigen (CEA)-specific antibody was conjugated to 124I-Green. High tumor uptake of the dual-labeled A5B7 of 20.21 ± 2.70, 13.31 ± 0.73, and 10.64 ± 1.86%ID/g was observed in CEA-expressing SW1222 xenograft mouse model (n = 3) at 24, 48, and 72 h post intravenous injection, respectively. The xenografts were clearly visualized by both PET/CT and ex vivo fluorescence imaging. These encouraging results warrant the further translational development of 124I-Green for cancer PET imaging and fluorescence-guided surgery.

Understanding the uptake of a drug by diseased tissue, and the drug's subsequent spatiotemporal distribution, are central factors in the development of effective targeted therapies. However, the interaction between the pathophysiology of diseased tissue and individual therapeutic agents can be complex, and can vary across tissue types and across subjects. Here, we show that the combination of mathematical modelling, high-resolution optical imaging of intact and optically cleared tumour tissue from animal models, and in vivo imaging of vascular perfusion predicts the heterogeneous uptake, by large tissue samples, of specific therapeutic agents, as well as their spatiotemporal distribution. In particular, by using murine models of colorectal cancer and glioma, we report and validate predictions of steady-state blood flow and intravascular and interstitial fluid pressure in tumours, of the spatially heterogeneous uptake of chelated gadolinium by tumours, and of the effect of a vascular disrupting agent on tumour vasculature.

With clinically-approved proteasome inhibitors now a standard of care for multiple myeloma, and increasing interest in their use in solid tumors, methods for monitoring therapeutic response in vivo are critically required. Here, we show that tumor protein homeostasis can be noninvasively monitored, using chemical exchange (CEST) magnetic resonance imaging (MRI) as a surrogate marker for proteasome inhibition, alongside diffusion MRI and relaxometry. We show that the in vivo CEST signal associated with amides and amines increases in proportion to proteasome inhibitor dose (ixazomib) and the magnitude of therapeutic effect in colorectal cancer xenografts. Moreover, we show that SW1222 and LS174T human colorectal cancer cell lines demonstrate differing sensitivities to ixazomib, which was reflected in our MRI measurements. We also found evidence of a mild stimulation in tumor growth at low ixazomib doses. Our results therefore identify CEST MRI as a promising method for safely and noninvasively monitoring changes in tumor protein homeostasis.

Few chemotherapeutics have had such an impact on cancer management as cis-diamminedichloridoplatinum(II) (CDDP), also known as cisplatin. The first member of the platinum based drug family, CDDP's potent toxicity in disrupting DNA replication has led to its widespread use in multi-drug therapies, with particular benefit in patients with testicular cancers. However, CDDP also produces significant side effects that limit the maximum systemic dose. Various strategies have been developed to address this challenge including encapsulation within micro- or nanocarriers and the use of external stimuli such as ultrasound to promote uptake and release. The aim of this article is to look at these strategies and recent scientific and clinical developments.

Cisplatin forms the basis for many chemotherapy regimens, however the maximum permissible dose is limited by its systemic toxicity. Nanoencapsulation of drugs has been shown to reduce off-target side effects and can potentially improve treatment burden on patients. However, uptake of nanoformulations at tumor sites is minimal without some form of active delivery. We have developed a submicron, polymeric nanoparticle based on biocompatible and degradable poly(lactic-co-glycolic acid) (PLGA) capable of encapsulating cisplatin and which can be bound to the surface of a phospholipid coated microbubble. The acoustic behavior and stability of the resulting nanoparticle loaded microbubbles will be compared with those of unloaded microbubbles. Results will also be presented on the extravasation of particles in a tissue mimicking phantom using a novel long working distance confocal microscope that enables particle distributions to be measured in situ and in real time.

Delivering potent, stable, targeted andin vivoefficacious antibody–drug conjugates (ADCs) using pyridazinedione functional disulfide re-bridging reagents.