Biomedical Engineering Reference
In-Depth Information
The optimal doxorubicin-loaded egg (E)-derived phospholipids
and cholesterol (EPC/EPG/Cholesterol/TCS 7:3:4:0.2) OLV-DOX
formulation showed reasonable shelf life and good physical stability
in plasma (<5% release in 1 h at 37
°
C). However, today, we know
that this way of testing was misleading, as the dilution of the OLV-
DOX used was small.
12.2.3
Cytotoxicity to Cells in Culture (
in vitro
)
In vitro
testing of this formulation (OLV-DOX) indicated that the drug
encapsulation in the OLV has almost no eff ect on the IC
50
(compare
1.1
×
10
-8M
for the liposomal drug with 1.8
×
10
-8M
for the free drug)
in the two cell lines studied [J-6456 lymphoma and P388 leukemia,
(Gabizon and Barenholz, 1988)]. At the time the experiment was
performed, we did not understand that this should be a warning
signal of the “viability” of this formulation as it suggested unwanted
relatively fast dilution-dependent major drug leakage from the
liposomes to the cell growth medium. At a later stage, when we
evaluated why this formulation failed in humans (see below),
we found that no growth inhibition of the cells (no cytotoxicity)
was observed in the presence of the cation exchanger Dowex-50
[which selectively and efficiently binds the positively charged free
doxorubicin, but not liposome-associated drug (Druckmann et al.,
1989; Amselem et al., 1990)]. The same removal of drug cytotoxicity
by the cation exchanger was obtained for free doxorubicin (F-DOX),
indicating that in both cases, drug enters the cells in its free
(nonliposomal) form.
12.2.4
Pharmacokinetics and Biodistribution Studies in
Mice
Preclinical PK and BD studies of the above optimal OLV-DOX in
normal and tumor-bearing mice met all of our objectives and
expectations. Biodistribution studies showed that the encapsulation
of DOX into these OLV increased liver (the target organ) and spleen
DOX uptake concomitantly with reducing DOX levels in the heart,
kidneys, and small intestine, as well as reducing organ and plasma
clearance of the drug (Gabizon et al., 1982, 1983). In mice bearing
liver metastasis, the DOX level after i.v. injection of OLV-DOX reached
a level severalfold higher than after injection of an equal amount of
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