Abstract:
Thalidomide, renowned for causing birth defects in the late 1950s when used for the relief
of morning sickness, has attracted new interest for the treatment of inflammatory
conditions such as erythema nodosum leprosum and human malignancies such as multiple
myeloma. Different species have different sensitivities to thalidomide that could be related
to differences in its metabolism. In this study, methodologies using liquid chromatographymass
spectrometry were developed to identify thalidomide metabolites formed in vivo and
in vitro in liver microsomes from mice, rabbits and humans, firstly to seek explanations for
inter-species differences in sensitivity, and secondly to determine whether thalidomide or
its metabolite(s) is the active agent.
Four hydrolysis products were detected in plasma and urine samples from multiple
myeloma patients (MMPs) on thalidomide therapy, and mice and rabbits after oral
administration of thalidomide. Six hydroxylated metabolites were detected in mice and
rabbits, but not in plasma and urine from MMPs. In vitro studies confirmed that murine
and rabbit liver microsomes catalysed the hydroxylation of thalidomide efficiently, but
significant production of hydroxylation of thalidomide was not observed using human liver
microsomes. The degree of hydroxylation both in vivo and in vitro was highest in mice and
lowest in humans with rabbits in between. It is unlikely that hydroxylated metabolites are
responsible for the effects of thalidomide in the treatment of multiple myeloma, since they
were not present in quantifiable amounts in patients who were responding to the treatment.
The three major hydrolysis products that were detected in patients were compared with
thalidomide for their ability to inhibit tube formation in an in vitro angiogenesis assay, to
inhibit TNF production induced with LPS in human peripheral blood leucocytes, and to
modulate DMXAA-induced TNF production and antitumour activity in mice. One of the
three, N-(o-carboxybenzoyl)glutamic acid imide (CG) was found to be as active as
thalidomide in all the assays at concentrations (1-2 μg/ml) that are achievable in MMPs.
Since CG has been shown by other laboratories to be non-teratogenic, the studies in this thesis indicate that CG would be a more favourable, non-teratogenic approach to cancer
therapy compared with thalidomide.