Role of noninvasive molecular imaging in determining response

The intersection of immunotherapy and radiation oncology is a rapidly evolving area of preclinical and clinical investigation. The strategy of combining radiation and immunotherapy to enhance local and systemic antitumor immune responses is intriguing yet largely unproven in the clinical setting because the mechanisms of synergy and the determinants of therapeutic response remain undefined. In recent years, several noninvasive molecular imaging approaches have emerged as a platform to interrogate the tumor immune microenvironment. These tools have the potential to serve as robust biomarkers for cancer immunotherapy and may hold several advantages over conventional anatomic imaging modalities and contemporary invasive tissue acquisition techniques. Given the key and expanding role of precision imaging in radiation oncology for patient selection, target delineation, image guided treatment delivery, and response assessment, noninvasive molecular-specific imaging may be uniquely suited to evaluate radiation/immunotherapy combinations. Herein, we describe several experimental imaging-based strategies that are currently being explored to characterize in vivo immune responses, and we review a growing body of preclinical data and nascent clinical experience with immuno-positron emission tomography molecular imaging as a putative biomarker for cancer immunotherapy. Finally, we discuss practical considerations for clinical translation to implement noninvasive molecular imaging of immune checkpoint molecules, immune cells, or associated elements of the antitumor immune response with a specific emphasis on its potential application at the interface of radiation oncology and immuno-oncology

Monitoring of Immune Cell Response to B Cell Depletion Therapy and Nerve Root Injury Using Spio Enhanced MRI

Magnetic resonance (MR) is a robust platform for non-invasive, high-resolution anatomical imaging. However, MR imaging lacks the requisite sensitivity and contrast for imaging at the cellular level. This represents a clinical impediment to greater diagnostic accuracy. Recent advances have allowed for the in vivo visualization of populations and even of individual cells using superparamagnetic iron oxide (SPIO) MR contrast agents. These nanoparticles, commonly manifested as a core of a single iron oxide crystal or cluster of crystals coated in a biocompatible shell, function to shorten proton relaxation times. In MR imaging these constructs locally dephase protons, resulting in a decrease in signal (hypointensity) localized to the region of accumulation of SPIO. In the context of immune cell imaging, SPIO can provide insight into the cellular migration patterns, trafficking, temporal dynamics and progression of diseases and their related pathological states. Furthermore, by visualizing the presence and activity of immune cells, SPIO-enabled cellular imaging can help evaluate the efficacy of therapy in immune disorders. This thesis examines the production, modification and application of SPIO in a range of in vitro and in vivo immune-response-relevant cellular systems. The role of different nanoparticle characteristics including diameter, surface charge and concentration are investigated in the labeling of T cells in culture. Following optimization of SPIO loading conditions for lymphocytes, the effect these particles have on the activation of primary B cells are elucidated. B cells are tracked using a variety of modalities, with and without the application of B cell depleting therapy. This is to evaluate the efficacy of SPIO as in vivo marker for B cell distribution. Unmodified SPIO were applied to monitor macrophage infiltration in a transient nerve root compression model, with implications for neck pain diagnosis and treatment. Nanoparticle accumulation and MR hypointensity was correlated to the presence of activated macrophage at the site of injury. Taken together, the application of SPIO to study nanoparticle uptake in vitro and visualization of immune cells in vivo provide a basis for advanced study and diagnosis of diverse pathologies

Size, charge and concentration dependent uptake of iron oxide particles by non-phagocytic cells

A promising new direction for contrast-enhanced magnetic resonance (MR) imaging involves tracking the migration and biodistribution of superparamagnetic iron oxide (SPIO)-labeled cells in vivo. Despite the large number of cell labeling studies that have been performed with SPIO particles of differing size and surface charge, it remains unclear which SPIO configuration provides optimal contrast in non-phagocytic cells. This is largely because contradictory findings have stemmed from the variability and imprecise control over surface charge, the general need and complexity of transfection and/or targeting agents, and the limited number of particle configurations examined in any given study. In the present study, we systematically evaluated the cellular uptake of SPIO in non-phagocytic T cells over a continuum of particle sizes ranging from 33 nm to nearly 1.5 μm, with precisely controlled surface properties, and without the need for transfection agents. SPIO labeling of T cells was analyzed by flow cytometry and contrast enhancement was determined by relaxometry. SPIO uptake was dose-dependent and exhibited sigmoidal charge dependence, which was shown to saturate at different levels of functionalization. Efficient labeling of cells was observed for particles up to 300 nm, however, micron-sized particle uptake was limited. Our results show that an unconventional highly cationic particle configuration at 107 nm maximized MR contrast of T cells, outperforming the widely utilized USPIO (\u3c50 nm)

Superparamagnetic Iron Oxide Nanoparticle Probes for Molecular Imaging

The field of molecular imaging has recently seen rapid advances in the development of novel contrast agents and the implementation of insightful approaches to monitor biological processes non-invasively. In particular, superparamagnetic iron oxide nanoparticles (SPIO) have demonstrated their utility as an important tool for enhancing magnetic resonance contrast, allowing researchers to monitor not only anatomical changes, but physiological and molecular changes as well. Applications have ranged from detecting inflammatory diseases via the accumulation of non-targeted SPIO in infiltrating macrophages to the specific identification of cell surface markers expressed on tumors. In this article, we attempt to illustrate the broad utility of SPIO in molecular imaging, including some of the recent developments, such as the transformation of SPIO into an activatable probe termed the magnetic relaxation switch

Preclinical efficacy of hK2 targeted [177Lu]hu11B6 for prostate cancer theranostics

Androgen ablating drugs increase life expectancy in men with metastatic prostate cancer, but resistance inevitably develops. In a majority of these recurrent tumors, the androgen axis is reactivated in the form of increased androgen receptor (AR) expression. Targeting proteins that are expressed as a down-stream effect of AR activity is a promising rationale for management of this disease. The humanized IgG1 antibody hu11B6 internalizes into prostate and prostate cancer (PCa) cells by binding to the catalytic cleft of human kallikrein 2 (hK2), a prostate specific enzyme governed by the AR-pathway. In a previous study, hu11B6 conjugated with Actinium-225 (225Ac), a high linear energy transfer (LET) radionuclide, was shown to generate an AR-upregulation driven feed-forward mechanism that is believed to enhance therapeutic efficacy. We assessed the efficacy of hu11B6 labeled with a low LET beta-emitter, Lutetium-177 (177Lu) and investigated whether similar tumor killing and AR-enhancement is produced. Moreover, single-photon emission computed tomography (SPECT) imaging of 177Lu is quantitatively accurate and can be used to perform treatment planning. [177Lu]hu11B6 therefore has significant potential as a theranostic agent. Materials and Methods: Subcutaneous PCa xenografts (LNCaP s.c.) were grown in male mice. Biokinetics at 4-336 h post injection and uptake as a function of the amount of hu11B6 injected at 72 h were studied. Over a 30 to 120-day treatment period the therapeutic efficacy of different activities of [177Lu]hu11B6 were assessed by volumetric tumor measurements, blood cell counts, molecular analysis of the tumor as well as SPECT/CT imaging. Organ specific mean absorbed doses were calculated, using a MIRD-scheme, based on biokinetic data and rodent specific S-factors from a modified MOBY phantom. Tumor tissues of treated xenografts were immunohistochemically (IHC) stained for Ki-67 (proliferation) and AR, SA-β-gal activity (senescence) and analyzed by digital autoradiography (DAR). Results: Organ-to-blood and tumor-to-blood ratios were independent of hu11B6 specific activity except for the highest amount of antibody (150 µg). Tumor accumulation of [177Lu]hu11B6 peaked at 168 h with a specific uptake of 29 ± 9.1 percent injected activity per gram (%IA/g) and low accumulation in normal organs except in the submandibular gland (15 ± 4.5 %IA/g), attributed to a cross-reaction with mice kallikreins in this organ, was seen. However, SPECT imaging with therapeutic amounts of [177Lu]hu11B6 revealed no peak in tumor accumulation at 7 d, probably due to cellular retention of 177Lu and decreasing tumor volumes. For [177Lu]hu11B6 treated mice, tumor decrements of up to 4/5 of the initial tumor volume and reversible myelotoxicity with a nadir at 12 d were observed after a single injection. Tumor volume reduction correlated with injected activity and the absorbed dose. IHC revealed retained expression of AR throughout treatment and that Ki-67 staining reached a nadir at 9-14 d which coincided with high SA- β-gal activity (14 d). Quantification of nuclei staining showed that Ki-67 expression correlated negatively with activity uptake. AR expression levels in cells surviving therapy compared to previous timepoints and to controls at 30 d were significantly increased (p = 0.017). Conclusions: This study shows that hu11B6 labeled with the low LET beta-emitting radionuclide 177Lu can deliver therapeutic absorbed doses to prostate cancer xenografts with transient hematological side-effects. The tumor response correlated with the absorbed dose both on a macro and a small scale dosimetric level. Analysis of AR staining showed that AR protein levels increased late in the study suggesting a therapeutic mechanism, a feed forward mechanism coupled to AR driven response to DNA damage or clonal lineage selection, similar to that reported in high LET alpha-particle therapy using 225Ac labeled hu11B6, however emerging at a later timepoint

Tumour cell labelling by magnetic nanoparticles with determination of intracellular iron content and spatial distribution of the intracellular iron

Abstract: Magnetically labelled cells are used for in vivo cell tracking by MRI, used for the clinical translation of cell-base therapies. Studies involving magnetic labelled cells may include separation of labelled cells, targeted delivery and controlled release of drugs, contrast enhanced MRI and magnetic hyperthermia for the in situ ablation of tumours. Dextran-coated super-paramagnetic iron oxide (SPIO) ferumoxides are used clinically as an MR contrast agents primarily for hepatic imaging. The material is also widely used for in vitro cell labelling, as are other SPIO-based particles. Our results on the uptake by human cancer cell lines of ferumoxides indicate that electroporation in the presence of protamine sulphate (PS) results in rapid high uptake of SPIO nanoparticles (SPIONs) by parenchymal tumour cells without significant impairment of cell viability. Quantitative determination of cellular iron uptake performed by colorimetric assay is in agreement with data from the literature. These results on intracellular iron content together with the intracellular distribution of SPIONs by magnetic force microscopy (MFM) following in vitro uptake by parenchymal tumour cells confirm the potential of this technique for clinical tumour cell detection and destruction

Prostate cancer theranostics - An overview

Metastatic prostate cancer is incurable, and novel methods to detect the disease earlier and to direct definitive treatment are needed. Molecularly specific tools to localize diagnostic and cytotoxic radionuclide payloads to cancer cells and the surrounding microenvironment are recognized as a critical component of new approaches to combat this disease. The implementation of theranostic approaches to characterize and personalize patient management is beginning to be realized for prostate cancer patients. This review article summarized clinically translated approaches to detect, characterize, and treat disease in this rapidly expanding field

Platelet aggregation induced by polystyrene and platinum nanoparticles is dependent on surface area

Nanoparticles are key components underlying recent technological advances in various industrial and medical fields, and thus understanding their mode of interaction with biological systems is essential. However, while several nanoparticle systems have been shown to interact with blood platelets, many questions remain concerning the mechanisms of platelet activation and the role that the physicochemical properties of nanoparticles play in inducing platelet aggregation. Here, using negatively charged polystyrene nanoparticles with sizes of 25, 50, 119, 151, 201 nm and negatively charged platinum nanoparticles with sizes of 7 and 73 nm, we show that it is not the size of the nanoparticles but rather the nanoparticle surface area that is critical in mediating the effects on platelet activation. The nanoparticles stimulate platelet aggregation through passive (agglutination) and activation of integrin αIIbβ3 through a pathway regulated by Src and Syk tyrosine kinase