[The development of pharmacophore models for thymidine kinase-dependent virostatic nucleoside analogs]. / Zur Entwicklung eines Pharmakophormodells für thymidinkinaseabhängige, nukleosidanaloge Virostatika.

The most potent drugs in the therapy of Herpes Simplex virus infections belong to the class of guanosine derivatives which bear an acyclic ether substitutent at N-9 (I,II). The initial step in the mode of action was shown to be a monophosphorylation at the primary hydroxyl group of the side chain. The selectivity of the antiviral activity results from the fact that only the virus encoded thymidine kinase is able to bind the acyclic guanosine derivatives. In order to understand this mechanism we try to describe the common pharmacophoric pattern of pyrimidine and purine substrates by molecular modeling methods. It can be shown that a specific binding of guanine could be provided by the enzyme and that flexible side chains are needed instead of deoxyribose to fulfill the requirements of the pharmacophore.

Attempts to standardize the screening for antiviral drugs by in vitro tests.

Plaque reduction (PR) and persistent infection cell culture (PICC) tests were used as a standard procedure for the selection of effective and well-tolerated antiviral compounds. Antiviral activity and cytotoxicity as well as other characteristics of the test compound, i.e., the development of drug resistance or the potential for activating the multiplication of other viruses can be examined by this method. In the described system analogues of 2'-deoxyuridine were tested against herpes simplex virus.

Karyotype alterations of BHK-21 cell lines by persistent virus infections I. Development of the cell lines and conventional chromosome analysis.

From BHK-21 cell cultures inoculated with different viruses cell lines with persistent infection were developed and examined for alterations in their karyotypes. In all cell lines studied, even in those no longer producing infective virus chromosomal breaks and interchanges were observed. Besides these effects probably induced directly by the virus infection, a shift in the number of chromosomes was found. Based on certain properties of the BHK-21 cell lines carrying a virus infection a hypothesis is discussed concerning the cooperation of various cellular and viral factors which enable a virus to persist.

Karotype alterations of BHK-21 cell lines by persistent virus infections. II. Analysis of the banding patterns.

Results of banding pattern analysis of the chromosome set of cell lines derived from BHK-21 by persistent infection with different viruses (Schwöbel et al., 1977) are reported. Almost all metaphases contained several structurally altered chromosomes which could not be identified. On the other hand, for each cell line analysed a characteristic karyotype could be established. The possible relevance of the results with respect to persistent virus infections in vivo is discussed.

Use of cell cultures with persistent virus infections to test the efficacy of antiviral compounds.

BHK-21 cells persistently infected with either vaccinia or foot and mouth disease virus were used to study the efficacy of antiviral compounds. The results of the persistent infection cell culture (PICC) test were compared with those obtained by the plaque reduction (PR) test. The comparison showed that: (1) the PICC test is more informative than the PR test; (2) stimulative as well as inhibitory activities of compounds are detectable, and (3) since the PICC test can be carried on for several weeks or even months this test is especially well suited to study the problem of drug resistance in cell cultures.

Changes of the cell type of BHK-21 lines by persistent Sindbis virus.

Persistent infection by Sindbis virus (SV) can be induced in cultures of BHK-21 cells. In a persistently infected cell line only 5% of the cells produced virus. Virus titers in the medium reached 10(5) PFU per ml. A persistent infection could be eliminated from cultures by seeding at most 100 cells per bottle. Compared with the original line decontaminated lines differed in several properties. After infection with SV ten times less virus was released into the medium, and an infection persisted without producing periodic cell destructions as observed in establishing a persistent infection in the original line. In decontaminated lines plaques formed by different viruses were either not visible or smaller than those in normal cells. Persistently infected as well as decontaminated lines had lost three chromosomes. The altered cell type of persistently infected lines apparently originated by selection of a cell mutant which was not destroyed by SV. Results suggested that a small number of cells susceptible to the virus continuously arises from the population of altered cells, and that virus infection is transmitted and maintained in the cultures by these sensitive cells.