Photoactivation of lysosomally sequestered sunitinib after angiostatic treatment causes vascular occlusion and enhances tumor growth inhibition

The angiogenesis inhibitor sunitinib is a tyrosine kinase inhibitor that acts mainly on the VEGF and PDGF pathways. We have previously shown that sunitinib is sequestered in the lysosomes of exposed tumor and endothelial cells. This phenomenon is part of the drug-induced resistance observed in the clinic. Here, we demonstrate that when exposed to light, sequestered sunitinib causes immediate destruction of the lysosomes, resulting in the release of sunitinib and cell death. We hypothesized that this photoactivation of sunitinib could be used as a vaso-occlusive vascular-targeting approach to treating cancer. Spectral properties of sunitinib and its lysosomal accumulation were measured in vitro. The human A2780 ovarian carcinoma transplanted onto the chicken chorioallantoic membrane (CAM) and the Colo-26 colorectal carcinoma model in Balb/c mice were used to test the effects of administrating sunitinib and subsequently exposing tumor tissue to light. Tumors were subsequently resected and subject to immunohistochemical analysis. In A2780 ovarian carcinoma tumors, treatment with sunitinib+light resulted in immediate specific angio-occlusion, leading to a necrotic tumor mass 24 h after treatment. Tumor growth was inhibited by 70% as compared with the control group (**P<0.0001). Similar observations were made in the Colo-26 colorectal carcinoma, where light exposure of the sunitinib-treated mice inhibited tumor growth by 50% as compared with the control and by 25% as compared with sunitinib-only-treated tumors (N≥4; P=0.0002). Histology revealed that photoactivation of sunitinib resulted in a change in tumor vessel architecture. The current results suggest that the spectral properties of sunitinib can be exploited for application against certain cancer indications.Angiogenesis inhibitors are currently firmly implemented in the clinical management of cancer. For example, sunitinib has been approved for the treatment of advanced renal cell carcinoma (RCC),¹ gastrointestinal stromal tumors (GIST)² and pancreatic neuroendocrine tumors.³ Studies assessing its activity against other tumor types are currently underway.⁴ Sunitinib was developed as an angiogenesis inhibitor⁵ and revolutionized the management of advanced RCC and GIST. Its mode of action is based on the suppression of the tyrosine kinase activity of several growth factor receptors, mainly VEGFR2 and PDGFR (alpha and beta).⁶ It has been previously shown that sunitinib is sequestered by tumor cells in their lysosomal compartments.⁷ This sequestration, which limits the exposure of other parts of the cell to sunitinib, is part of a drug-induced resistance mechanism that has also been clinically observed.⁷ The sequestration and accumulation of sunitinib in the lysosomes is similar to a phenomenon that has been described for certain other chemotherapeutics^{8, 9} or photosensitizing compounds,¹⁰ and depends, at least to some extent, on the hydrophobic and weak basic features of the molecule. We discovered a particular characteristic of sunitinib based on its optical properties, which may be helpful in the treatment of certain cancers. On the basis of the spectral features of sunitinib, we hypothesized that the drug may have photosensitizer-like properties.¹¹ If so, exposure to light of an appropriate wavelength could lead to the disruption of the lysosomal membrane, the release of sunitinib and re-exposure of the cell to this molecule. This could lead to further cell damage and eventually cell death. In this study, we show that the exposure of sunitinib-treated cells to light of an appropriate wavelength excites the molecule, which leads to the generation of reactive oxygen species (ROS). We show in two tumor models, that is, human ovarian carcinoma (A2780) xenografted on the chorioallantoic membrane of the chicken embryo (chicken chorioallantoic membrane (CAM)) and in colorectal carcinoma in Balb/c mice, that treatment with sunitinib and its subsequent photoexcitation leads to significant occlusion of the vasculature and inhibition of tumor growth. Our results indicate that additional antitumor activity can be obtained after the normal use of sunitinib through its use as a photosensitizer and the application of light. The combination of classical sunitinib-induced angiostasis with the re-exposure of tumor cells to sunitinib after the destruction of lysosomes and photodynamic vessel obstruction can lead to a strategy that may be applicable in patients.     

Synthesis and Characterization of Novel Quinazoline Type Inhibitors for Mutant and Wild-Type EGFR and RICK Kinases

The development of selective protein kinase inhibitors has become an important area of drug discovery for the treatment of different diseases. We report the synthesis and characterization of a series of novel quinazoline derivatives against three therapeutically important and pharmacologically related kinases: 1) epidermal growth factor receptor (EGFR; wild type and mutant) in the field of cancer, 2) receptor-interacting caspase-like apoptosis-regulatory kinase (RICK) in the field of inflammation, and 3) pUL97 of human cytomegalovirus (HCMV). For reference purpose we have synthesized the four clinically relevant quinazolines, including the lead compounds, which we previously identified for RICK and pUL97. A total of 52 quinazoline derivatives were synthesized and tested on the basis of these leads to specifically target the hydrophobic pocket of the ATP-binding site. Selected compounds were tested on wild-type and mutant forms of EGFR, RICK, and pUL97 kinases; their logP and logS values for assessing suitability as drugs were calculated and hit or lead compounds identified.     

Dynamic localisation of Ran GTPase during the cell cycle

Background  

Ran GTPase has multiple functions during the cell division cycle, including nucleocytoplasmic transport, mitotic spindle assembly and nuclear envelope formation. The activity of Ran is determined by both its guanine nucleotide-bound state and its subcellular localization.     

Optimization of physical factors affecting the production of thermo-stable organic solvent-tolerant protease from a newly isolated halo tolerant Bacillus subtilis strain Rand

Background  

Many researchers have reported on the optimization of protease production; nevertheless, only a few have reported on the optimization of the production of organic solvent-tolerant proteases. Ironically, none has reported on thermostable organic solvent-tolerant protease to date. The aim of this study was to isolate the thermostable organic solvent-tolerant protease and identify the culture conditions which support its production. The bacteria of genus Bacillus are active producers of extra-cellular proteases, and the thermostability of enzyme production by Bacillus species has been well-studied by a number of researchers. In the present study, the Bacillus subtilis strain Rand was isolated from the contaminated soil found in Port Dickson, Malaysia.     

Transcriptomic response to differentiation induction

Background  

Microarrays used for gene expression studies yield large amounts of data. The processing of such data typically leads to lists of differentially-regulated genes. A common terminal data analysis step is to map pathways of potentially interrelated genes.     

Regulation of peptidoglycan synthesis by outer-membrane proteins

Growth of the mesh-like peptidoglycan (PG) sacculus located between the bacterial inner and outer membranes (OM) is tightly regulated to ensure cellular integrity, maintain cell shape, and orchestrate division. Cytoskeletal elements direct placement and activity of PG synthases from inside the cell, but precise spatiotemporal control over this process is poorly understood. We demonstrate that PG synthases are also controlled from outside of the sacculus. Two OM lipoproteins, LpoA and LpoB, are essential for the function, respectively, of PBP1A and PBP1B, the major E.?coli bifunctional PG synthases. Each Lpo protein binds specifically to its cognate PBP and stimulates its transpeptidase activity, thereby facilitating attachment of new PG to?the sacculus. LpoB shows partial septal localization, and our data suggest that the LpoB-PBP1B complex contributes to OM constriction during cell?division. LpoA/LpoB and their PBP-docking regions are restricted to γ-proteobacteria, providing models for niche-specific regulation of sacculus growth.