Search

International audience; This roadmap outlines the potential roles of metallic nanoparticles (MNPs) in the field of radiation therapy. MNPs made up of a wide range of materials (from Titanium, Z = 22, to Bismuth, Z = 83) and a similarly wide spectrum of potential clinical applications, including diagnostic, therapeutic (radiation dose enhancers, hyperthermia inducers, drug delivery vehicles, vaccine adjuvants, photosensitizers, enhancers of immunotherapy) and theranostic (combining both diagnostic and therapeutic), are being fabricated and evaluated. This roadmap covers contributions from experts in these topics summarizing their view of the current status and challenges, as well as expected advancements in technology to address these challenges.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

We have previously demonstrated that endothelial targeting of gold nanoparticles followed by external beam irradiation can cause specific tumor vascular disruption in mouse models of cancer. The induced vascular damage may lead to changes in tumor physiology, including tumor hypoxia, thereby compromising future therapeutic interventions. In this study, we investigate the dynamic changes in tumor hypoxia mediated by targeted gold nanoparticles and clinical radiation therapy (RT). By using noninvasive whole-body fluorescence imaging, tumor hypoxia was measured at baseline, on day 2 and day 13, post-tumor vascular disruption. A 2.5-fold increase (P

Effective drug delivery is restricted by pathophysiological barriers in solid tumors. In human pancreatic adenocarcinoma, poorly-permeable blood vessels limit the intratumoral permeation and penetration of chemo or nanotherapeutic drugs. New and clinically viable strategies are urgently sought to breach the neoplastic barriers that prevent effective drug delivery. Here, we present an original idea to boost drug delivery by selectively knocking down the tumor vascular barrier in a human pancreatic cancer model. Clinical radiation activates the tumor endothelial-targeted gold nanoparticles to induce a physical vascular damage due to the high photoelectric interactions. Active modulation of these tumor neovessels lead to distinct changes in tumor vascular permeability. Noninvasive MRI and fluorescence studies, using a short-circulating nanocarrier with MR-sensitive gadolinium and a long-circulating nanocarrier with fluorescence-sensitive nearinfrared dye, demonstrate more than two-fold increase in nanodrug delivery, post tumor vascular modulation. Functional changes in altered tumor blood vessels and its downstream parameters, particularly, changes in Ktrans (permeability), Kep (flux rate), and Ve (extracellular interstitial volume), reflect changes that relate to augmented drug delivery. The proposed dual-targeted therapy effectively invades the tumor vascular barrier and improve nanodrug delivery in a human pancreatic tumor model and it may also be applied to other nonresectable, intransigent tumors that barely respond to standard drug therapies.

A cost-effective method for serial in vivo imaging of tumor microvasculature has been developed. We evaluated acoustic angiography (AA) for visualizing and assessing non-small cell lung tumor (A549) microvasculature in mice prior to and following tumor vascular disruption by vascular-targeted gold nanoparticles (GNPs) and radiotherapy. Standard B-mode and microbubble-enhanced AA images were acquired at pre- and post-treatment time points. Using these modes, a new metric, 50% Vessel Penetration Depth (VPD(50)) was developed to characterize the 3D spatial heterogeneity of microvascular networks. We observed an increase in tumor perfusion after radiation-induced vascular disruption, relative to control animals. This was also visualized in vessel morphology mode, which showed a loss in vessel integrity. We found that tumors with poorly perfused vasculature at day 0 exhibited a reduced growth rate over time. This suggested a new method for reducing in-group treatment response variability by using pre-treatment microvessel maps to objectively identify animals for study removal.

Effective drug delivery is severely restricted by the presence of complex pathophysiological barriers in solid tumors. In human pancreatic adenocarcinoma, mature and hypopermeable tumor blood vessels limit the permeation and penetration of chemo or nanotherapeutics to cancer cells and substantially reduce the treatment efficacy. New, clinically-viable strategies are therefore sought to breach the neoplastic barriers that prevent optimal tumor-specific drug delivery. Here, we present an original idea to boost targeted drug delivery by selectively knocking down the tumor vascular barrier in a poorly permeable human pancreatic cancer model. For the first time, we demonstrate that clinical irradiation (10 Gy, 6 MV) can induce tumor vascular modulation when combined with tumor endothelial-targeting gold nanoparticles. Active disruption of tumor blood vessels by nanoparticle-combined radiotherapy led to increased vessel permeability and improved tumor uptake of two prototypical model nanodrugs: i) a short-circulating nanocarrier with MR-sensitive gadolinium (Gad-NC; 8 kDa; t1/2=1.5 h) and ii) a long-circulating nanocarrier with fluorescence-sensitive NIR dye (FL-NC; 30 kDa; t1/2=25 h). Functional changes in the altered tumor vessel dynamics, measured by relative changes in permeability (Ktrans), flux rate (Kep) and extracellular interstitial volume (Ve) were consistent with the concomitant increase in nanodrug delivery. This combination of radiation-induced antivascular and nanodrug-mediated anti-tumor treatment offers high therapeutic benefit for tumors with pathophysiology that restricts efficient drug delivery.

Selective killing of cancer cells while minimizing damage to healthy tissues is the goal of clinical radiation therapy. This therapeutic ratio can be improved by image-guided radiation delivery and selective radiosensitization of cancer cells. Here, we have designed and tested a novel trimodal theranostic nanoparticle made of bismuth and gadolinium for on-site radiosensitization and image contrast enhancement to improve the efficacy and accuracy of radiation therapy. We demonstrate in vivo magnetic resonance (MR), computed tomography (CT) contrast enhancement, and tumor suppression with prolonged survival in a non-small cell lung carcinoma model during clinical radiation therapy. Histological studies show minimal off-target toxicities due to the nanoparticles or radiation. By mimicking existing clinical workflows, we show that the bismuth-gadolinium nanoparticles are highly compatible with current CT-guided radiation therapy and emerging MR-guided approaches. This study reports the first in vivo proof-of-principle for image-guided radiation therapy with a new class of theranostic nanoparticles.

Purpose Tumor vascular targeted gold nanoparticles induce tumor vascular disruption when combined with external beam radiation therapy. Although effective in suppressing tumor growth and improving progression-free survival, tumor hypoxia may be a potential challenge in this anti-vascular therapy. Here, we investigate, the dynamic changes in tumor hypoxia pre- and post-radiation therapy using a gold nanoparticle-based tumor vascular disrupting agent. Materials and methods 4-6 weeks old female nude-FOXn1 mice were subcutaneously inoculated with human A549 cells (~3×106) into the left flank. Functionalized gold nanoparticles (AuNP) were used to target the tumor blood vessels and tumor vascular disruption was induced via radiation. 10 Gy radiation treatment was delivered using a clinical radiation beam (6 MV) and HypoxiSense680 based in vivo fluorescence imaging was performed to visualize changes in tumor hypoxia at 24 h, 48 h and an extended period of 10 days. Tumor hypoxia was confirmed via immunohistochemistry. Mice were divided into four treatment groups: control, AuNP only, IR only, and AuNP+IR. Following treatment, tumor progression and overall survival was measured. Further analysis of nanoparticle biodistribution and toxicity were accessed using TEM Imaging, IC-PMS, and immunohistochemistry. Results Longitudinal changes in tumor hypoxia were observed in all radiation-based treatment conditions. Combining gold nanoparticle and radiation resulted in an increase in tumor hypoxia at 48 h (p < 0.05) and a return to baseline in 10 days. In contrast, the ‘radiation only’ group showed an increase in tumor hypoxia by a factor of 0.5 at 48 h post-IR compared to baseline while 10 days later the tumor hypoxia remained stable. Quantitative variation in the hypoxia blood factor, CA9 increased 24 h post IR in the AuNP+IR, followed by a decrease in hypoxic by day 10 in accordance with in vivo imaging data. No change was observed with the IR only group. These findings were confirmed with representative pimonidazole staining that showed an increase in tumor hypoxia a few hours after AuNP+IR treatment. The mean relative reduction in tumor size post-treatment was a factor of 5.2 (p < 0.05) in the AuNP+IR group compared to the control and 3.5 compared to the IR only group, nearly 80 days post-treatment. Overall survival showed an average gain of up to 24 days in the AuNP+IR group compared to all other treatments. Almost 100 days post-treatment, 50% survival was observed in the AuNP+IR group compared to 20% in the IR-only groups (p < 0.05). Conclusions Noninvasive imaging showed that AuNP+IR results in a transient increase and subsequent decline in mean tumor hypoxia, leading to substantial tumor regression and an overall increase in tumor survival. High radiation-induced vascular damage may lead to better tumor reduction and prolonged survival in the human non-small cell lung cancer model. Citation Format: Olivia J. Kelada, Sijumon Kunjachan, Needa A. Virani, Alexandre Detappe, Jennifer Hayashi, Thomas Ireland, Douglas E. Biancur, Rajiv Kumar, Srinivas Sridhar, Mike Makrigiorgos, Ross I. Berbeco. Noninvasive imaging of tumor hypoxia during radiation-induced tumor vascular disruption [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 85.

Drug delivery in the CNS is limited by endothelial tight junctions forming the impermeable blood-brain barrier. The development of new treatment paradigms has previously been hampered by the restrictiveness of the blood-brain barrier to systemically administered therapeutics. With recent advances in stereotactic localization and noninvasive imaging, we have honed the ability to modulate, ablate, and rewire millimetric brain structures to precisely permeate the impregnable barrier. The wide range of focused radiations offers endless possibilities to disrupt endothelial permeability with different patterns and intensity following 3-dimensional coordinates offering a new world of possibilities to access the CNS, as well as to target therapies. We propose a review of the current state of knowledge in targeted drug delivery using noninvasive image-guided approaches. To this end, we focus on strategies currently used in clinics or in clinical trials such as targeted radiotherapy and magnetic resonance guided focused ultrasound, but also on more experimental approaches such as magnetically heated nanoparticles, electric fields, and lasers, techniques which demonstrated remarkable results both in vitro and in vivo. We envision that biodistribution and efficacy of systemically administered drugs will be enhanced with further developments of these promising strategies. Besides therapeutic applications, stereotactic platforms can be highly valuable in clinical applications for interventional strategies that can improve the targetability and efficacy of drugs and macromolecules. It is our hope that by showcasing and reviewing the current state of this field, we can lay the groundwork to guide future research in this realm., SCOPUS: re.j, info:eu-repo/semantics/published

International audience; Radiation therapy is a major treatment regimen for more than 50% of cancer patients. The collateral damage induced on healthy tissues during radiation and the minimal therapeutic effect on the organ-of-interest (target) is a major clinical concern. Ultra-small, renal clearable, silica based gadolinium chelated nanoparticles (SiGdNP) provide simultaneous MR contrast and radiation dose enhancement. The high atomic number of gadolinium provides a large photoelectric cross-section for increased photon interaction, even for high-energy clinical radiation beams. Imaging and therapy functionality of SiGdNP were tested in cynomolgus monkeys and pancreatic tumor-bearing mice models, respectively. A significant improvement in tumor cell damage (double strand DNA breaks), growth suppression, and overall survival under clinical radiation therapy conditions were observed in a human pancreatic xenograft model. For the first time, safe systemic administration and systematic renal clearance was demonstrated in both tested species. These findings strongly support the translational potential of SiGdNP for MR-guided radiation therapy in cancer treatment.

Nanomedicines are sub-micrometer-sized carrier materials designed to improve the biodistribution of i.v. administered (chemo-) therapeutic agents. In recent years, ever more efforts in the nanomedicine field have employed optical imaging (OI) techniques to monitor biodistribution and target site accumulation. Thus far, however, the longitudinal assessment of nanomedicine biodistribution using OI has been impossible, due to limited light penetration (in the case of 2D fluorescence reflectance imaging; FRI) and to the inability to accurately allocate fluorescent signals to nonsuperficial organs (in the case of 3D fluorescence molecular tomography; FMT). Using a combination of high-resolution microcomputed tomography (μCT) and FMT, we have here set out to establish a hybrid imaging protocol for noninvasively visualizing and quantifying the accumulation of near-infrared fluorophore-labeled nanomedicines in tissues other than superficial tumors. To this end, HPMA-based polymeric drug carriers were labeled with Dy750, their biodistribution and tumor accumulation were analyzed using FMT, and the resulting data sets were fused with anatomical μCT data sets in which several different physiologically relevant organs were presegmented. The robustness of 3D organ segmentation was validated, and the results obtained using 3D CT-FMT were compared to those obtained upon standard 3D FMT and 2D FRI. Our findings convincingly demonstrate that combining anatomical μCT with molecular FMT facilitates the noninvasive assessment of nanomedicine biodistribution.

Over the past few decades, many different types of nanomedicines have been evaluated, both in vitro and in vivo. In general, nanomedicines are designed to improve the in vivo properties of low-molecular-weight (chemo-) therapeutic drugs, i.e. their biodistribution and the target site accumulation, and to thereby improve the balance between their efficacy and toxicity. A significant number of studies have also addressed the in vitro properties of nanomedicines, showing e.g. their ability to overcome cellular multidrug resistance (MDR). Particularly promising results in this regard have been reported for ‘pharmacologically active’ carrier materials, such as Pluronics, which are able to directly inhibit drug efflux pumps and other cellular detoxification mechanisms. In the present report, we have set out to evaluate the ability of classical (and pharmacologically inactive) carrier materials to overcome MDR. To this end, four different drug-sensitive and drug-resistant cancer cell lines were treated with increasing concentrations of free doxorubicin, of polymer-bound doxorubicin, of micellar doxorubicin and of liposomal doxorubicin, and resistance indices (IC50 in resistant cells/IC50 in sensitive cells) were determined. In addition, the cellular uptake of the four formulations was evaluated using fluorescence microscopy. It was found that the carrier materials did manage to overcome MDR to some extent, but that the overall benefit was quite small; only for polymer-bound doxorubicin in A431 cells, a significant (4-fold) reduction in the resistance index was observed. These findings indicate that the ability of classical nanomedicines to overcome cellular MDR should not be overestimated.

Macrophages are important drug targets as they mediate a wide variety of infectious diseases. Visceral leishmaniasis (VL), schistosomiasis, brucellosis, and salmonellosis are some of the well-known infectious diseases in which macrophages play a prominent pathophysiological role. For instance, VL parasites exclusively house in the macrophages of liver and spleen. They are resistant to lysosomal degradation by unknown mechanisms, they survive and thrive safely within macrophages, they multiply, and they ultimately affect visceral organs, leading to severe pathological and sometimes even fatal conditions. The majority of routinely used drugs administered in free form distribute all over the body via systemic circulation, leading to relatively low therapeutic activity and a certain degree of toxicity. Unlike for nonmicrobial diseases, targeting parasites procuring resistance and ineffective therapeutic outcome can be obviously speculated in case of infectious disease. The preferential uptake by macrophages, intended to improve the balance between efficacy and toxicity, can be achieved by the use of nanomedicines, i.e. submicron-sized macromolecular or particulate drug delivery systems. This insight has stimulated researchers to use nanomedicines--which tend to be recognized by macrophages as 'foreign' and consequently are taken up by the intended target cells much more effectively than their free drug counterparts--to improve the treatment of infectious diseases. The literature reports extensively on such approaches; however, there are several constraints that limit the application of nanomedicine in macrophage-mediated drug targeting. Here, we briefly describe the strategies that are used to achieve effective drug targeting to macrophages, using VL as a model disease, and we also put forth an understanding of the most important limiting factors. Various physicochemical and biological factors used by researchers as reported in the literature are addressed, and the most important mechanisms and modes by which macrophage-specific drug targeting can be achieved are summarized. Based on the evidence obtained to date, it can be concluded that targeting macrophages is a valuable and validated strategy for improving the treatment of infectious diseases.

The potential of chitosan microparticles as a carrier of doxorubicin for the treatment of visceral leishmaniasis was evaluated by macrophage-mediated drug targeting approach. Cationic charge of doxorubicin was masked by complexing it with dextran sulphate (a poly anion) in order to facilitate its incorporation into cationic chitosan microparticles. Prior to in vitro and in vivo studies, characterization studies were carried out systematically: particle size (∼1.049 µm), surface morphology (fluorescence microscopy - spherical structured microparticles), Fourier transform infrared spectroscopy (to characterize effective cross-linking) and differential scanning calorimetry. In vitro studies were carried out in J774.1 in order to check the effective endocytotic uptake of microparticles by macrophages. In vivo studies were conducted in Syrian golden hamsters as per well-established protocols and the results drawn from in vivo studies displayed substantial reduction in leishmanial parasite load for doxorubicin-encapsulated chitosan microparticles: ∼78.2 ± 10.4%, when compared to the control (free doxorubicin): 33.3 ± 2.4%.

Introduction: More than 50% of cancer patients receive radiation therapy at some point during their care. Gold nanoparticles (GNPs) can amplify the radiation dose by facilitating the ejection of low-energy photoelectrons, resulting in increased DNA damage. One of the main challenges of radiation therapy in cancer is to sustain this damage for longer durations. DNA single-strand breaks (SSBs) are repaired by base excision repair, which utilizes Poly(ADP-ribose) polymerase (PARP). PARP inhibition during radiotherapy provides an attractive alternative in maximizing treatment outcomes. Here, we explore a strategy to combine the radiosensitizing effect of GNPs with the DNA-repair inhibiting ability of NanoTalazoparib (nTLZ), a liposomal formulation of the PARP inhibitor, talazoparib (TLZ). Methods: GNPs were synthesized by reducing AuCl4 using Tetrakis(hydroxymethyl)phosphonium chloride and further PEGylating using heterobifunctional PEGs. A liposomal formulation of TLZ was synthesized using the Nanoassemblr, a microfluidics-based device. Physicochemical characterization of GNPs and nTLZ was carried out using TEM, DLS and release kinetics studies. In vitro studies were carried out to assess the toxicity of the GNPs using MTT assay in non-small cell lung cancer (NSCLC) cell line Calu-6. The therapeutic efficacy of combination treatment using GNPs, nTLZ and radiation was done using clonogenic survival assays. Clonogenic assay was carried out using Calu 6 cells, which were sequentially treated with GNPs (1mg/mL), TLZ (0.5 uM) and nTLZ (0.5 uM) with and without radiation. The radiation doses varied from 0-10Gy for each set of treatments. Results: PEGylated GNP's showed a hydrodynamic diameter of ~10-12 nm with a spherical morphology whereas nTLZ size was 70 nm encapsulating TLZ at a concentration of ~200 ug/ml. MTT assay showed no toxicity for PEGylated GNPs treated upto a concentration of 3.0 mg/mL. Cells treated with either GNPs, TLZ or nTLZ did not show significant reduction in colony formation, but were reduced with increasing doses of radiation. The survival plots showed a highly additive antiproliferative effect for the GNP + nTLZ combination at all radiation doses, while the free TLZ + GNPs combination was not as effective at inhibiting colony formation. In vivo experiments assessing the combination therapy in a subcutaneous xenograft mice model using Calu 6 are currently under way. Conclusions: The preliminary in vitro results indicate that the combination of radiosensitizing GNPs with a potent DNA repair enzyme inhibitor, TLZ, has an immense potential as a complimentary combination therapy in conjunction with radiation therapy in treatment of lung cancer. This work is supported by the CaNCURE program (grant #1CA174650-02), American Lung Association, Dana-Farber Cancer Institute and Brigham and Women's Hospital. Citation Format: Ana G. Vazquez-Pagan, Paige Baldwin, Ravina M. Ashtaputre, Sijumon Kunjachan, Srinivas Sridhar, Rajiv Kumar, Ross Berbeco. Nanoparticle-mediated concomitant radiation dose amplification and PARP inhibition in lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3710.

International audience

As nanoparticle solutions move towards human clinical trials in radiation therapy, the influence of key clinical beam parameters on therapeutic efficacy must be considered. In this study, we have investigated the clinical radiation therapy delivery variables that may significantly affect nanoparticle-mediated radiation dose amplification. We found a benefit for situations which increased the proportion of low energy photons in the incident beam. Most notably, “unflattened” photon beams from a clinical linear accelerator results in improved outcomes relative to conventional “flat” beams. This is measured by significant DNA damage, tumor growth suppression, and overall improvement in survival in a pancreatic tumor model. These results, obtained in a clinical setting, clearly demonstrate the influence and importance of radiation therapy parameters that will impact clinical radiation dose amplification with nanoparticles.

International audience; AGuIX are gadolinium-based nanoparticles developed mainly for imaging due to their MR contrast properties. They also have a potential role in radiation therapy as a radiosensitizer. We used MRI to quantify the uptake of AGuIX in pancreatic cancer cells, and TEM for intracellular localization. We measured the radiosensitization of a pancreatic cancer cell line in a low-energy (220 kVp) beam, a standard 6 MV beam (STD) and a flattening filter free 6 MV beam (FFF). We demonstrated that the presence of nanoparticles significantly decreases cell survival when combined with an X-ray beam with a large proportion of low-energy photons (close to the k-edge of the nanoparticles). The concentration of nanoparticles in the cell achieves its highest level after 15 min and then reaches a plateau. The accumulated nanoparticles are mainly localized in the cytoplasm, inside vesicles. We found that the 6 MV FFF beams offer the best trade-off between penetration depth and proportion of low-energy photons. At 10 cm depth, we measured a DEF20 % of 1.30 ± 0.47 for the 6 MV FFF beam, compared to 1.23 ± 0.26 for the 6 MV STD beam. Additional measurements with un-incubated nanoparticles provide evidence that chemical processes might also be contributing to the dose enhancement effect.

Aim: To enable multimodal in vivo and ex vivo optical imaging of the biodistribution and tumor accumulation of core-crosslinked polymeric micelles (CCPMs). Materials & methods: mPEG-b-p(HPMAm-Lac)-based polymeric micelles, core-crosslinked via cystamine and covalently labeled with two different fluorophores (Dy-676/488), were synthesized. The CCPMs were intravenously injected into CT26 tumor-bearing mice. Results: Upon intravenous injection, the CCPMs accumulated in CT26 tumors reasonably efficiently, with values reaching approximately 4%ID at 24 h. Ex vivo two-photon laser scanning microscopy confirmed efficient extravasation of the image-guided CCPMs out of tumor blood vessels and relatively deep penetration into the tumor interstitium. Conclusion: CCPMs were labeled with multiple fluorophores, and the results obtained exemplify that combining several different in vivo and ex vivo optical imaging techniques is highly useful for analyzing the biodistribution and tumor accumulation of nanomedicines.

Purpose: MR-guided radiation therapy is a current and emerging clinical reality. We have designed and tested a silica-based gadolinium chelates nanoparticle (AGuIX) for integration with MR-guided radiation therapy. The AGuIX nanoparticles used in this study are a dual-modality probe with radiosensitization properties and better MRI contrast than current FDA-approved gadolinium chelates. In advance of an approved Phase I clinical trial, we report on the efficacy and safety in multiple animal models and clinically relevant radiation conditions. By modeling our study on current clinic workflows, we show compatibility with modern patient care, thus heightening the translational significance of this research. Methods: The dual imaging and therapy functionality of AGuIX was investigated in mice with clinical radiation beams while safety was evaluated in mice, and nonhuman primates after systemic injection of 0.25 mg/g of nanoparticles. MRI/ICP-MS were used to measure tumor uptake and biodistribution. Due to their small size (2–3 nm), AGuIX have good renal clearance (t1/2=19min). We performed in vitro cell uptake quantification and radiosensitization studies (clonogenic assays and DNA damage quantification). In vivo radiation therapy studies were performed with both 6MV and 6MV-FFF clinical radiation beams. Histology was performed to measure the increase in DNA damage in the tumor and to evaluate the toxicity in healthy tissues. Results: In vitro and in vivo results demonstrate statistically significant increase (P < 0.01) in DNA damage, tumor growth supression and survival (+100 days) compared to radiation alone. Negligible toxicity was observed in all of the animal models. The combination of 6MV-FFF/AGuIX demonstrated a substantial dose enhancement compared to 6MV/AGuIX (DEF = 1.36 vs. 1.22) due to the higher proportion of low energy photons. Conclusion: With demonstrated efficacy and negligible toxicity in mice and non-human primates, AGuIX is a biocompatible nanoplatform with strong translational potential for MR-guided radiation therapy.

Theranostics 4(10), 960-971 (2014). doi:10.7150/thno.9293, Published by Ivyspring, Wyoming, NSW

The Enhanced Permeability and Retention (EPR) effect is extensively used in drug delivery research. Taking into account that EPR is a highly variable phenomenon, we have here set out to evaluate if contrast-enhanced functional ultrasound (ceUS) imaging can be employed to characterize EPR-mediated passive drug targeting to tumors. Using standard fluorescence molecular tomography (FMT) and two different protocols for hybrid computed tomography-fluorescence molecular tomography (CT-FMT), the tumor accumulation of a ~ 10 nm-sized near-infrared-fluorophore-labeled polymeric drug carrier (pHPMA-Dy750) was evaluated in CT26 tumor-bearing mice. In the same set of animals, two different ceUS techniques (2D MIOT and 3D B-mode imaging) were employed to assess tumor vascularization. Subsequently, the degree of tumor vascularization was correlated with the degree of EPR-mediated drug targeting. Depending on the optical imaging protocol used, the tumor accumulation of the polymeric drug carrier ranged from 5 to 12% of the injected dose. The degree of tumor vascularization, determined using ceUS, varied from 4 to 11%. For both hybrid CT-FMT protocols, a good correlation between the degree of tumor vascularization and the degree of tumor accumulation was observed, within the case of reconstructed CT-FMT, correlation coefficients of ~ 0.8 and p-values of < 0.02. These findings indicate that ceUS can be used to characterize and predict EPR, and potentially also to pre-select patients likely to respond to passively tumor-targeted nanomedicine treatments.

The use of nanoparticles with high atomic (Z) number have been known to attenuate X-rays and the unique properties associated with gold nanoparticles makes them as potent radiosensitizers for enhancing Radiotherapy (RT) treatments. The interaction of high Z materials with the X-rays results in photoelectric absorption which leads to generation of photoelectrons. These low energy photoelectrons can deliver lethal energy in the close proximity. The success of cancer radiation therapy relies heavily on the effective delivery of radiation dose to the tumor site sparing the surrounding normal tissues. To overcome the limitations associated with increasing the radiation dose, due to normal tissue toxicities, the feasibility of radiosensitizing the tumor using gold nanoparticles provide a promising alternative. Targeting gold nanoparticles based formulations to the tumor prior to radiation therapy will result in radiation dose enhancement, by generating secondary photoelectrons, locally inside the tumor and thereby minimizing the dose dependent toxicity to non-specific neighboring tissues. Here, we have developed different Gold nanoparticles based formulations to locally radiosensitize the tumor cells in three different cancer models. For targeting pancreatic cancers, we have fabricated a new generation of GNPs formulation which are 3-4 nm in diameter and surface passivated with hetero-bifunctional PEG, fluorophore AlexaFlour 647 and peptide RGD. This formulation showed a 2.8-fold in vitro cell kill enhancement with X-rays, as demonstrated by clonogenic survival assays. In vivo studies confirmed the highly specific tumor uptake in tumor endothelial cells in orthotopic pancreatic tumor mice model. The combined treatment in animals treated with GNPs and radiation (10Gy) showed the maximum endothelial cells damage, as confirmed with confocal imaging of the tumor sections and CD31 immunostaining. For targeting the lung cancer, we have used the same GNPs formulation but adopted a inhalation/instillation (INH) route for administering the nanoparticles in vivo in transgenic lung cancer mice model as opposed to customary intravenous (i.v) route. Fluorescence imaging and ex-vivo electron microcopy results showed a 4.7 times higher concentration of GNPs in the lung tumors of mice when using INH delivery compared to i.v. approach. The survival studies with these animals are currently underway. For using these radiosensitizing nanoparticles in brachytherapy applications, we have incorporated GNPs in modified brachytherapy spacers to enhance radiation dose locally in tumor without any additional surgical intervention. These spacers are normally placed inside the tumor to spatially distribute the radioactive seeds. The incorporation of GNPs in these spacers allowed for enhancing the radiation dose by the GNPs released from the spacers. These spacers have same morphology (5 mm in length and 0.8 mm in diameter) as commercial spacers with additional radiosensitizing properties because of doped GNPs. We have shown the time dependent release of the GNPs as a function of size of GNPs from the spacers into the tumor by optical imaging. We have further incorporated the GNPs based brachytherapy spacer with the immunoadjuvants, anti-CD40 to combine the radiation and immunotherapy in a single platform. The preliminary results indicate that gold nanoparticles based nanoplatform shows promise as a potential theranostic radiosensitizer which allows for combining multiple therapies in a single platform. This work is supported by ARMY/ W81XWH-12-1-0154, NSF DGE 0965843, HHS/5U54CA151881-02, NCI R03CA164645, NCI1 K01CA17247801, the Electronics Materials Research Institute at Northeastern University, and Brigham and Women's Hospital. Citation Format: Rajiv Kumar, Wilfred Ngwa, Vinit Joshi, Sijumon Kunjachan, Ross Berbeco, Mike Makrigiorgos, Srinivas Sridhar. Gold nanoparticles based platforms for localized radiosensitization in cancer radiation therapy. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr B41.

Multidrug (MDR) resistance is a pathophysiological phenomenon employed by cancer cells which limits the prolonged and effective use of chemotherapeutic agents. MDR is primarily based on the over-expression of drug efflux pumps in the cellular membrane. Prominent examples of such efflux pumps, which belong to the ATP-binding cassette (ABC) superfamily of proteins, are Pgp (P-glycoprotein) and MRP (multidrug resistance-associated protein), nowadays officially known as ABCB1 and ABCC1. Over the years, several strategies have been evaluated to overcome MDR, based not only on the use of low-molecular-weight MDR modulators, but also on the implementation of 1-100(0) nm-sized drug delivery systems. In the present manuscript, after introducing the most important physiological principles of MDR, we summarize prototypic nanomedical strategies to overcome multidrug resistance, including the use of carrier materials with intrinsic anti-MDR properties, the use of nanomedicines to modify the mode of cellular uptake, and the co-formulation of chemotherapeutic drugs together with low- and high-molecular-weight MDR inhibitors within a single drug delivery system. While certain challenges still need to be overcome before such constructs and concepts can be widely applied in the clinic, the insights obtained and the progress made strongly suggest that nanomedicine formulations hold significant potential for improving the treatment of multidrug-resistant malignancies.

Macrophages are important drug targets as they mediate a wide variety of infectious diseases. Visceral leishmaniasis (VL), schistosomiasis, brucellosis, and salmonellosis are some of the well-known infectious diseases in which macrophages play a prominent pathophysiological role. For instance, VL parasites exclusively house in the macrophages of liver and spleen. They are resistant to lysosomal degradation by unknown mechanisms, they survive and thrive safely within macrophages, they multiply, and they ultimately affect visceral organs, leading to severe pathological and sometimes even fatal conditions. The majority of routinely used drugs administered in free form distribute all over the body via systemic circulation, leading to relatively low therapeutic activity and a certain degree of toxicity. Unlike for nonmicrobial diseases, targeting parasites procuring resistance and ineffective therapeutic outcome can be obviously speculated in case of infectious disease. The preferential uptake by macrophages, intended to improve the balance between efficacy and toxicity, can be achieved by the use of nanomedicines, i.e. submicron-sized macromolecular or particulate drug delivery systems. This insight has stimulated researchers to use nanomedicines--which tend to be recognized by macrophages as 'foreign' and consequently are taken up by the intended target cells much more effectively than their free drug counterparts--to improve the treatment of infectious diseases. The literature reports extensively on such approaches; however, there are several constraints that limit the application of nanomedicine in macrophage-mediated drug targeting. Here, we briefly describe the strategies that are used to achieve effective drug targeting to macrophages, using VL as a model disease, and we also put forth an understanding of the most important limiting factors. Various physicochemical and biological factors used by researchers as reported in the literature are addressed, and the most important mechanisms and modes by which macrophage-specific drug targeting can be achieved are summarized. Based on the evidence obtained to date, it can be concluded that targeting macrophages is a valuable and validated strategy for improving the treatment of infectious diseases.

Nanomedicine formulations are considered to be superior to standard low-molecular-weight drugs because of an increased drug accumulation at the pathological site and a decreased localization to healthy non-target tissues, together leading to an improved balance between the efficacy and the toxicity of (chemo-) therapeutic interventions. To better understand and further improve nanomedicine-mediated drug targeting, it is important to design systems and strategies which are able to provide real-time feedback on the localization, the release and the therapeutic efficacy of these formulations. The advances made over the past few years with regard to the development of novel imaging agents and techniques have provided a broad basis for the design of theranostic nanomedicine materials, i.e. multicomponent carrier constructs in which drugs and imaging agents are combined, and which can be used to address issues related to drug localization, drug release and drug efficacy. Here, we summarize several recent efforts in this regard, and we show that theranostic systems and strategies hold significant potential for monitoring and improving nanomedicine-mediated drug targeting.

Purpose: AGuIX are gadolinium-based nanoparticles, initially developed for MRI, that have a potential role in radiation therapy as a radiosensitizer. Our goal is to demonstrate that these nanoparticles can both be used as an MRI contrast agent, as well as to obtain local dose enhancement in a pancreatic tumor when delivered in combination with an external beam irradiation. Methods: We performed in vitro cell uptake and radiosensitization studies of a pancreatic cancer cell line in a low energy (220kVp) beam, a standard clinical 6MV beam (STD) and a flattening filter free clinical 6MV beam (FFF). After injection of 40mM of nanoparticles, a biodistribution study was performed in vivo on mice with subcutaneous xenograft pancreatic tumors. In vivo radiation therapy studies were performed at the time point of maximum tumor uptake. Results: The concentration of AGuIX nanoparticles in Panc-1 pancreatic cancer cells, determined in vitro by MRI and ICPMS, peaks after 30 minutes with 0.3% of the initial concentration (5mg/g). Clonogenic assays show a significant effect (p