Vascular toxicity in dogs associated with overdoses of a novel benzodiazepine receptor partial agonist.

Necrotizing arteritis and periarteritis were found in Beagle and German Shepherd dogs treated for 13 or 52 weeks with the novel benzodiazepine receptor (BZR) partial agonist Ro 16-6028 (generic name bretazenil). Eight male and one female out of a total of 20 dogs treated with 40-60 mg/kg/day Ro 16-6028 developed the arteritis, predominantly in the heart or the epididymis. Two of these animals died prematurely following treatment at the initial dosing levels of 80 and 55 mg/kg/day; one of these two dogs was asymptomatic and in good general condition until death. Clinically, all but one of the dogs showed sedation, ataxia, stiff gait, body weight-loss and a deterioration of the general condition as well as changes of some laboratory parameters. No signs of arteritis and untoward clinical or laboratory findings were seen at lower doses. Possible aetiologies, as well as the mechanisms involved in arteritis in general and the genetic disposition of beagles in particular for this type of effect, are discussed. Reflections on the potential risk to man of this so far unknown finding after oral treatment with 1,4-benzodiazepines (BZs) are presented.

Promoter activity and expression of sequences from Ti-plasmid stably maintained in mammalian cells.

Sequences of the plant-pathogenic Ti-plasmid were found to be constitutively expressed in LTK- and in HeLa-cells. Activity of the nopaline-synthase (nos) promoter in these cells was demonstrated by directing expression of G418 resistance from a connected neomycin-phosphotransferase II (NPT II) gene. Control transfections with the widely used thymidine-kinase (TK) promoter gave comparable transfection rates as found for the nos-promoter with NPT II. The function of the nos-promoter was also confirmed by assaying neomycin-phosphotransferase synthesized in cells containing a plasmid with the NPT II-gene under control of this promoter. Several LTK+ clones stably transfected with Ti-plasmid propagated the total Ti-plasmid DNA in a colinear state presumably as an episomal unit. Dot blot analysis and polymerase chain reaction showed predominant transcription of Ti-sequences from the T-DNA area reflecting transcriptional activity of this region not only in plant cells but also in animal cells. These results provide new information about promoter functions in systems unrelated to their natural environment.

The HBV X-ORF encodes a transactivator: a potential factor in viral hepatocarcinogenesis.

We report here on a transactivating function of HBV DNA. The effect is shown by stimulation of transient expression of pSV2cat DNA in cotransfected human liver CCL13 cells. Transfection experiments with plasmid constructs containing different HBV DNA fragments and Northern analyses of RNA from cells transfected with these recombinant plasmids indicate that a transactivating function is encoded within the X-ORF. A frameshift mutation within the X gene causes loss of activity thus demonstrating requirement of a protein. The increase in the level of CAT-specific RNA suggests that the transactivation is by transcriptional enhancement. Constitutive expression of the transactivator function was also observed in cells stably transfected with HBV DNA. A number of eukaryotic promoters, SV40-early, HSV-TK, HTLV-I and RSV LTRs were responsive to transactivation by HBV DNA. However, the MMTV LTR and the human metallothionein promoter (MTIIA) were considerably less responsive than the others. The transactivational potential of HBV DNA was much higher in human cells and cells of higher primates than in rodent cells, thereby indicating interacting cellular factors. These results introduce additional considerations for the role of HBV in the development of hepatocellular tumors.