Milan Makale

Nanoparticle-mediated drug delivery to tumor vasculature suppresses metastasis

Integrin ανβ3 is found on a subset of tumor blood vessels where it is associated with angiogenesis and malignant tumor growth. We designed an ανβ3-targeted nanoparticle (NP) encapsulating the cytotoxic drug doxorubicin (Dox) for targeted drug delivery to the ανβ3-expressing tumor vasculature. We observed real-time targeting of this NP to tumor vessels and noted selective apoptosis in regions of the ανβ3-expressing tumor vasculature. In clinically relevant pancreatic and renal cell orthotopic models of spontaneous metastasis, targeted delivery of Dox produced an antimetastatic effect. In fact, ανβ3-mediated delivery of this drug to the tumor vasculature resulted in a 15-fold increase in antimetastatic activity without producing drug-associated weight loss as observed with systemic administration of the free drug. These findings reveal that NP-based delivery of cytotoxic drugs to the ανβ3-positive tumor vasculature represents an approach for treating metastatic disease.

Quantification of brain lesions using interactive automated software

We developed an interactive program, Analysis of Brain Lesions (ABLe) so that researchers studying the effects of brain lesions on cognition could have a user-friendly tool that could quantitatively characterize such lesions. The program was prepared in Tcl/Tk and will run on any UNIX or PC LINUX platform with the MEDx medical imaging software package. The ABLe is almost completely automated and determines the brain lesion size as well as which cytoarchitectonic brain regions (Brodmann areas) are contained within the boundaries of the lesion. Lesion data from multiple subjects can be grouped together and the degree of lesion overlap displayed. All images are analyzed and displayed within standard Talairach coordinate space, and the precision of the match between the ABLe Brodmann area graphics and the subject/patient brain is easily confirmed. The program is the first easy-to-use software that contains these specific features and is available for interested researchers with a background in lesion analysis.

The Hippocampal System Mediates Logical Reasoning about Familiar Spatial Environments

It has recently been shown that syllogistic reasoning engages two dissociable neural systems. Reasoning about familiar situations engages a frontal-temporal lobe system, whereas formally identical reasoning tasks involving unfamiliar situations recruit a frontal-parietal visuospatial network. These two systems may correspond to the heuristic and formal methods, respectively, postulated by cognitive theory. To determine if this dissociation generalizes to reasoning about transitive spatial relations, we studied 14 volunteers using event-related fMRI, as they reasoned about landmarks in familiar and unfamiliar environments. Our main finding is a task (reasoning and baseline) by spatial content (familiar and unfamiliar) interaction. Modulation of reasoning toward unfamiliar landmarks resulted in bilateral activation of superior and inferior parietal lobules (BA 7, 40), dorsal superior frontal cortex (BA 6), and right superior and middle frontal gyri (BA 8), regions widely implicated in visuospatial processing. By contrast, modulation of the reasoning task toward familiar landmarks, engaged the right inferior/orbital frontal gyrus (BA 11/47), bilateral occipital (BA 18, 19), and temporal lobes. The temporal lobe activation included the right inferior temporal gyrus (BA 37), posterior hippocampus, and parahippocampal gyrus, regions implicated in spatial memory and navigation tasks. These results provide support for the generalization of dual mechanism theory to transitive reasoning and highlight the importance of the hippocampal system in reasoning about familiar spatial environments.

Cellular mechanobiology and cancer metastasis

The primary cause of cancer treatment failure is invasion and metastasis, and invading tumor cells utilize many of the motility patterns that have been documented for normal morphogenesis. Recently, the role of mechanical forces in guiding various tissue and cell movements in embryonic development has been systematically analyzed with new experimental and computational methods. The tissue and cellular mechanobiology approach also holds promise for increasing the understanding of tumor invasion. In fact, the mechanical stiffness of tumors has correlated with invasiveness, and manipulation of extracellular matrix (ECM) stiffness in vitro has suppressed the cancer phenotype. Several important signaling molecules reside on the cytoskeleton, which is affected by external stress imparted by the ECM, and deformation of the nucleus can trigger the activation of certain genes. All these observations suggest that a synthesis of the biology of cancer cell invasion and cellular mechanobiology may offer new targets for the treatment of malignant disease. Accordingly, sensitive and relevant in vivo models and methods to study cancer mechanobiology are needed.

Prostate specific membrane antigen (PSMA) expression in primary gliomas and breast cancer brain metastases

BackgroundPrimary and secondary brain cancers are highly treatment resistant, and their marked angiogenesis attracts interest as a potential therapeutic target. Recent observations reveal that the microvascular endothelium of primary high-grade gliomas expresses prostate specific membrane antigen (PSMA). Breast cancers express PSMA and they frequently form secondary brain tumors. Hence we report here our pilot study addressing the feasibility of PSMA targeting in brain and metastatic breast tumors, by examining PSMA levels in all glioma grades (19 patients) and in breast cancer brain metastases (5 patients).MethodsTumor specimens were acquired from archival material and normal brain tissues from autopsies. Tissue were stained and probed for PSMA, and the expression levels imaged and quantified using automated hardware and software. PSMA staining intensities of glioma subtypes, breast tumors, and breast tumor brain metastases were compared statistically versus normals.ResultsNormal brain microvessels (4 autopsies) did not stain for PSMA, while a small proportion (<5%) of healthy neurons stained, and were surrounded by an intact blood brain barrier. Tumor microvessels of the highly angiogenic grade IV gliomas showed intense PSMA staining which varied between patients and was significantly higher (p < 0.05) than normal brain. Grade I gliomas showed moderate vessel staining, while grade II and III gliomas had no vessel staining, but a few (<2%) of the tumor cells stained. Both primary breast cancer tissues and the associated brain metastases exhibited vascular PSMA staining, although the intensity of staining was generally less for the metastatic lesions.ConclusionsOur results align with and extend previous data showing PSMA expression in blood vessels of gliomas and breast cancer brain metastases. These results provide a rationale for more comprehensive studies to explore PSMA targeted agents for treating secondary brain tumors with PSMA expressing vasculature. Moreover, given that PSMA participates in angiogenesis, cell signaling, tumor survival, and invasion, characterizing its expression may help guide later investigations of the poorly understood process of low grade glioma progression to glioblastoma.

Category-Specific Representations of Social and Nonsocial Knowledge in the Human Prefrontal Cortex

Complex social behavior and the relatively large size of the prefrontal cortex are arguably two of the characteristics that distinguish humans from other animals. Grafman presented a framework concerning how the prefrontal cortex (PFC) controls complex behavior using stored structured event complexes (SECs). We report behavioral and imaging data from a modified go/no-go paradigm in which subjects had to classify words (semantic) and phrases (SEC) according to category. In experimental trials, subjects classified items according to social or nonsocial activity; in control trials, they classified items according to font. Subjects were faster to classify social than nonsocial semantic items, with the reverse pattern evident for the social and nonsocial SEC items. In addition, the conditions were associated with different patterns of PFC activation. These results suggest that there are different psychological and neural substrates for social and nonsocial semantic and SEC representations.

Novel anti-glioblastoma agents and therapeutic combinations identified from a collection of FDA approved drugs

BackgroundGlioblastoma (GBM) is a therapeutic challenge, associated with high mortality. More effective GBM therapeutic options are urgently needed. Hence, we screened a large multi-class drug panel comprising the NIH clinical collection (NCC) that includes 446 FDA-approved drugs, with the goal of identifying new GBM therapeutics for rapid entry into clinical trials for GBM.MethodsScreens using human GBM cell lines revealed 22 drugs with potent anti-GBM activity, including serotonergic blockers, cholesterol-lowering agents (statins), antineoplastics, anti-infective, anti-inflammatories, and hormonal modulators. We tested the 8 most potent drugs using patient-derived GBM cancer stem cell-like lines. Notably, the statins were active in vitro; they inhibited GBM cell proliferation and induced cellular autophagy. Moreover, the statins enhanced, by 40-70 fold, the pro-apoptotic activity of irinotecan, a topoisomerase 1 inhibitor currently used to treat a variety of cancers including GBM. Our data suggest that the mechanism of action of statins was prevention of multi-drug resistance protein MDR-1 glycosylation. This drug combination was synergistic in inhibiting tumor growth in vivo. Compared to animals treated with high dose irinotecan, the drug combination showed significantly less toxicity.ResultsOur data identifies a novel combination from among FDA-approved drugs. In addition, this combination is safer and well tolerated compared to single agent irinotecan.ConclusionsOur study newly identifies several FDA-approved compounds that may potentially be useful in GBM treatment. Our findings provide the basis for the rational combination of statins and topoisomerase inhibitors in GBM.

Intravital imaging and cell invasion.

The main cause of cancer treatment failure is the invasion of normal tissues by cancer cells that have migrated from a primary tumor. An important obstacle to understanding cancer invasion has been the inability to acquire detailed, direct observations of the process over time in a living system. Intravital imaging, and the rodent dorsal skinfold window chamber in particular, were developed several decades ago to address this need. However, it is just recently, with the advent of sophisticated new imaging systems such as confocal and multiphoton microscopy together with the development of a wide range of fluorescent cellular and intracellular markers, that intravital methods and the window chamber have acquired powerful new potential for the study of cancer cell invasion. Moreover, the interaction of various cell signaling pathways with the integrin class of cell surface receptors has increasingly been shown to play a key role in cancer invasion. The window chamber in combination with integrin-knockout rodent models, integrin-deficient tumor cell lines, and integrin antagonists, allows real-time observation of integrin-mediated cancer invasion and angiogenesis. The present review outlines the history, uses, and recent methods of the rodent dorsal skinfold window chamber. The introduction of labeled tumor cells into the chamber is described, and imaging of tumors and angiogenic vessels within chambers using standard brightfield, confocal, and multiphoton microscopy is discussed in detail, along with the presentation of sample images.

Recognition of dextran-superparamagnetic iron oxide nanoparticle conjugates (Feridex) via macrophage scavenger receptor charged domains.

Dextran-coated superparamagnetic iron oxide nanoparticles (dextran-SPIO conjugates) offer the attractive possibility of enhancing MRI imaging sensitivity so that small or diffuse lesions can be detected. However, systemically injected SPIOs are rapidly removed by macrophages. We engineered embryonic cells (HEK293T) to express major macrophage scavenger receptor (SR) subtypes including SR-AI, MARCO, and endothelial receptor collectin-12. These SRs possess a positively charged collagen-like (CL) domain and they promoted SPIO uptake, while the charge neutral lipoprotein receptor SR-BI did not. In silico modeling indicated a positive net charge on the CL domain and a net negative charge on the cysteine-rich (CR) domain of MARCO and SR-AI. In vitro experiments revealed that CR domain deletion in SR-AI boosted uptake of SPIO 3-fold, while deletion of MARCOs CR domain abolished this uptake. These data suggest that future studies might productively focus on the validation and further exploration of SR charge fields in SPIO recognition.

Magnetic resonance and fluorescence imaging of doxorubicin-loaded nanoparticles using a novel in vivo model

We report here the in vivo combined-modality imaging of multifunctional drug delivery nanoparticles. These dextran core-based stealth liposomal nanoparticles (nanosomes) contained doxorubicin, iron oxide for magnetic resonance imaging (MRI) contrast, and BODIPY for fluorescence. The particles were long-lived in vivo because of surface decoration with polyethylene glycol and the incorporation of acetylated lipids that were ultraviolet cross-linked for physical stability. We developed a rodent dorsal skinfold window chamber that facilitated both MRI and non-invasive optical imaging of nanoparticle accumulation in the same tumors. Chamber tumors were genetically labeled with DsRed-2, which enabled co-localization of the MR images, the red fluorescence of the tumor, and the blue fluorescence of the nanoparticles. The nanoparticle design and MR imaging developed with the window chamber were then extended to orthotopic pancreatic tumors expressing DsRed-2. The tumors were MR-imaged using iron oxide-dextran liposomes and by fluorescence to demonstrate the deep imaging capability of these nanoparticles.