02 BOOK OF ABSTRACTS .indd

The effect of Doxorubicin and Vinblastine in EAT tumours
in CBA mice
Maja ^ema`ar, Ana Poga~nik, Veronika Kloboves-Prevodnik, Gregor Ser{a,
Marija Auersperg
Institute of Oncology, Zalo{ka 2, SI-1000 Ljubljana, Slovenia
The design of most combined chemotherapeutic schedules used in patients is based
on the data from preclinical studies, and phase I and II clinical studies. Little attention
is paid to the timing of drugs or possible interaction of drugs in a particular combined
schedule. Both these factors could be crucial for the clinical effect of chemotherapy.
The primary objective of our study was to explore whether antitumour scheduledependency
exists for the combination of doxorubicin (Doxo) and vinblastine (VLB).
The second objective was to find out whether after pretreatment with VLB the
accumulation of Doxo within the tumor cells could be increased.
Intraperitoneal (i.p.) Ehrlich ascites tumours (EAT) syngeneic to CBA mice were used
in experiments. Three days after tumour transplantation, animals were treated with
VLB (0.006 mg/kg) or Doxo (0.9 mg/kg) alone, VLB followed by Doxo, Doxo followed
by VLB and both drugs given simultaneously. The time interval between i.p. injections
of the drugs was 24 h or 48h. The cell number was obtained by counting viable cells
using Trypan Blue exclusion assay. The autofluorescence of Doxo was observed
under fluorescence microscopy and measured by flow cytometer. After all various
treatment schedules, the DNA distribution pattern was also determined using flow
cytometry. In addition, morphology of the tumour samples from the animals treated
with various treatment combinations was evaluated. Furthermore, animal survival was
determined.
Combinations of Doxo and VLB administered at 48 h, but not at 24 h interval,
regardless
78p7
of the sequence of drugs resulted in significantly reduced cell number
in ascites in comparison with all other treatments. After treatment with VLB alone,
morphologically cells were moderately enlarged and multinucleated. After treatment
with Doxo alone, cells, as well as nuclei and nucleoli were enlarged. In the combined
treatment schedules, the dominant morphological changes observed in the samples
were dependent on the first drug applied. DNA histograms showed increased G 2
M
compartment of samples from animals treated with Doxo alone or when Doxo
preceded VLB or when the drugs were given simultaneously. In contrast, after VLB
or when VLB preceded Doxo, DNA histograms showed less distinctive peaks, high
DNA values depassing G 2
M peak were also observed. Regardless of the sequence and
timing of the treatment, median survival times of animals did not significantly differ
between the treatment groups neither at 24 h nor at 48 h interval
The combination of VLB and Doxo is schedule dependent. For the effect was crucial
time interval, but not the sequence between the drug administrations. When translating
the results of preclinical studies to clinical setting, we need to be cautious, since
different tumor models used can yield controversial results. Nevertheless, the data
from preclinical studies should be taken into consideration when planning combined
treatment schedules in clinical situation.