Cellular effects of oncolytic viral therapy on the glioblastoma microenvironment.

The objective of the present study was to evaluate the cellular effects of the oncolytic HSV-1 based vector, G207, on the tumor microenvironment. We established progressively growing intracerebral xenografts in athymic nude rats generated from three different human GBM surgical specimens. The lesions were identified by MRI and subsequently injected with a concentrated vector stock. The animals were killed 10 or 30 days after G207 injection and the tumors were quantitatively evaluated for virus-induced changes in proliferation, apoptosis and vascularity. Moreover, we assessed vector spread as well as the infiltration pattern of CD68-positive inflammatory cells. In all G207-injected lesions, immunostaining identified widespread regions of viral infection and replication (plaques). Proliferation indices were significantly lower, whereas apoptotic counts were significantly elevated in plaques as compared with that in non-infected areas of the same lesions, as well as in corresponding control xenografts. Furthermore, there was a significant decline in the number of blood vessels in the plaques and the vascular area fractions were reduced. CD68-positive inflammatory cells accumulated in the plaques. The present study highlights the favorable cellular responses to G207 treatment seen from a clinical viewpoint, such as reduced tumor cell proliferation, more frequent events of tumor cell death and a strongly attenuated tumor vascular compartment. However, our results suggest that transduction of a significant volume of tumor tissue is essential, as these beneficial changes were only observed in areas of active viral replication, leaving non-transduced tumor tissues unaffected.

Intravital microscopy reveals novel antivascular and antitumor effects of endostatin delivered locally by alginate-encapsulated cells.

The current study describes new, antivascular, and antitumor effects of human endostatin. A novel system for continuous, localized delivery of antiangiogenic compounds to brain tumors was used. The delivery system was composed of endostatin-producing 293 cells encapsulated into immuno-isolating sodium alginate. Intravital multifluorescence microscopy was used to assess vascular and antitumor effects of endostatin in C6 glioma spheroids implanted into an ectopic as well as an orthotopic setting. Analysis of total and functional vascular density, microvascular diameters, vessel perfusion, tumor growth, and tumor cell migration were performed repetitively. Tumor growth was reduced by 35% in treated animals. It was of interest that tumor cell invasion into the surrounding tissue was also inhibited. The total vascular density was reduced by 67.6%, perfusion by 67%, and vessel diameters by 37%. This resulted in a significant reduction in tumor perfusion, although the vessel permeability was not influenced. We have demonstrated that human endostatin not only reduces total vascular density, as shown previously, but also greatly reduces the functionality and the diameters of the vessels. Furthermore, we show that this therapeutic approach also inhibits tumor cell invasion, thus supporting the hypothesis that tumor angiogenesis and invasion represent two interrelated processes. Finally, this work further confirms the new therapeutic concept using alginate cell-encapsulation technology for the localized delivery of therapeutic compounds to central nervous system malignancies.

Circular dichroism of host-guest complexes of achiral pyridino- and phenazino-18-crown-6 ligands with the enantiomers of chiral aralkyl ammonium salts.

Circular dichroism (CD) spectroscopy was used for distinguishing different types of chiral interactions in host-guest complexes of achiral pyridino- and phenazino-18-crown-6 ligands with chiral aralkyl ammonium salts. The general feature of the CD spectra of many homochiral (e.g., (R,R)-host and (R)-guest) and heterochiral (e.g., (R,R)-host and (S)-guest) alpha-(1-naphthyl)ethylamine hydrogenperchlorate salt (NEA) complexes with chiral pyridino- and phenazino-18-crown-6 hosts is exciton interaction. The most interesting example is the coupling of the transitions of the chiral guest NEA with the energetically close transitions of the achiral phenazino-18-crown-6 host 6. The CD spectrum of the complex is predominated by exciton coupling between the (1)B(b) transition of the chiral guest and the (1)B(b) transition of the achiral host. The redshifted intense spectra of the complexes of (R)- or (S)-1-phenylethylamine hydrogenperchlorate salt (PEA) with the achiral diester-pyridino-18-crown-6 host 4 are indicative of merging the pi electron systems into one joint charge transfer chromophore. The appearance of weak bands with alternating sign in the spectrum of PEA complexes of the achiral "parent" pyridino-18-crown-6 host (1) indicates the presence of two or more conformers. The CD spectra of the complexes of achiral phenazino-18-crown-6 host 6 with PEA are also determined by pi-pi interaction. In addition to charge transfer bands, CD bands are also induced in the long-wavelength spectral region of the achiral host. The weak pi-pi interaction between the achiral phenazino-18-crown-6 host and methyl phenylglycinate hydrogenperchlorate (PGMA) or methyl phenylalaninate hydrogenperchlorate (PAMA) does not result in a definite spectral effect in the (1)L(a) region of the spectrum of the chiral guest, but its existence is proven by the weak CD bands induced in the long-wavelength spectral region of the achiral host.

Semipreparative scale enantioseparation of racemic amine and amino ester hydrogen perchlorate salts using a silica gel-bound optically active di-tert-butylpyridino-18-crown-6 ligand.

We report herein results on the enantioseparation of selected racemic amine and amino ester hydrogen perchlorate salts using a silica gel-bound optically pure chiral di-tert-butylpyridino-18-crown-6 ligand (R,R)-1. The effect of solvent composition was studied using appropriate binary and ternary solvent mixtures as eluents. We found that acetonitrile/ methanol/ dichloromethane (MeCN/MeOH/CH2 Cl2 ) ternary solvent mixtures gave better enantioseparations for the racemic salts using chiral stationary phase (R,R)-1 than any of the binary ones. In the present paper we also describe the studies of chromatographic parameters such as loading, flow rate and eluent polarity.

Characterization of chiral host-guest complexation in fast atom bombardment mass spectrometry.

A new method has been developed for the characterization of complexion between host and guest molecules. Adduct formation between chiral crown ethers 1 and 2 and enantiomeric ammonium ions 4 and 5 was examined. The reference compound 3 (achiral host) was chosen to be similar in structure to the chiral crown ethers for quantitative measurements. Our approach is based on a formalism assuming an equilibrium: [chiral host + H](+) + [achiral host + chiral guest](+) ⇌ [chiral host + chiral guest](+) + [achiral host + H](+). The equlibrium constant for this process was calculated using the relative peak intensities of the corresponding species in the FAB mass spectra. It was found that these provide significantly better reproducibility and more reliable results than the relative peak intensity method described before (Sawada, M.; et al. J. Am. Chem. Soc. 1992, 114, 4405; 1993, 115, 7381; Org. Mass Spectrom. 1993, 28, 1525).(1)(-)(3) In the examples studied, the equilibrium constants corresponding to the formation of heterochiral adducts (S,S-R or R,R-S) were higher than those for the formation of homochiral aggregates (S,S-S or R,R-R).