Creatine and Creatinine Metabolism - Physiological Reviews

1158 MARKUS WYSS AND RIMA KADDURAH-DAOUK Volume 80
cCr on the phosphorylation patterns of MAP, on the function
of the katanin proteins, as well as on the ATP levels
in responsive and nonresponsive tumors.
In another set of experiments, the potential relationships
between cellular swelling and antitumor activity
were evaluated (857, 858). C6 rat glioma multicellular
spheroids were mapped by magnetic resonance spectroscopy,
and sets of images demonstrated increased diffusion
of water into the viable rim upon treatment with cCr.
It was suggested that uptake of cCr is accompanied by
cotransport of sodium, which leads to water accumulation
(944). No cellular swelling was observed in multicellular
spheroids of OC 238 human ovarian carcinoma cells
exposed to cCr. Whereas the C6 line expressed CK activity
and effectively built up PcCr, very little CK activity and
no PcCr were found in the OC 238 cell line. Unexpectedly,
both cell lines were growth inhibited by cCr, suggesting
that induction of cellular swelling does not account for
the antitumor activity of cCr. Also, there was no significant
change in the nucleoside triphosphate pool in both
cell lines, suggesting that growth inhibition may not be
due to disturbances of general cellular energetics. The
authors proposed that there might be more than one
mechanism operative for cCr; it could potentially act on
the cell surface or in a mitochondrial environment that is
not visible by 31 P-NMR.
These conclusions were corroborated in part by a
study in nude mice carrying a human colon adenocarcinoma
(LS174T; CK activity 2.12 U/mg protein), where
feeding for 2 wk with 2.5–5% Cr or 0.1–0.5% cCr significantly
inhibited tumor growth (512). Both substances
were equally potent, and the best correlation was observed
between tumor growth inhibition and the total Cr
or (Cr � cCr) concentration in the tumor tissue. The
antiproliferative effect of Cr and cCr was not related to
energy deficiency or to the proportion of PCr and PcCr
but was associated with acidosis. Cr and cCr did not
induce excessive water accumulation and had no systemic
effects like induction of weight loss or hypoglycemia
that might have caused tumor growth inhibition.
Ara et al. (28) evaluated the effects of Cr analogs on
pancreatic hormones and glucose metabolism. Rats bearing
the 13762 mammary carcinoma were treated with cCr
(GPA; PCr) on days 4–8 and 14–18 after tumor implantation
with or without the addition of sugars. Tumor
growth delays increased from 9.3 (1.6; 7.6) days in animals
not receiving sugars to 15.0 (6.3; 12.6) days in animals
drinking sugar water. This could be a result of (direct or
indirect) stimulation of tissue uptake of creatine/cCr by
sugars, an effect that was noted previously (see sect. XI).
Interestingly, blood glucose levels decreased over the
course of the treatment, whereas the skeletal muscle
glucose transporter protein GLUT-4 increased 1.5- to
2-fold. Plasma insulin concentration was decreased by
75–80%, plasma glucagon slightly elevated, and plasma
somatostatin increased three- to fourfold. These hormones
are known to modulate tumor growth (for references,
see Ref. 28), with insulin being an established
growth factor for several tumors and somatostatin being
a potent antiproliferative agent. Changes in hormonal profiles
induced by Cr analogs may therefore be part of the
mechanism of their antitumor action.
cCr has recently been evaluated in a phase I/II clinical
study in terminal cancer patients. Safety and pharmacokinetic
profiles were established (O’Keefe et al. and
Schimmel et al., unpublished data). In a dose escalation
study over a period of 10 wk, cCr was administered
continuously by intravenous infusion at dose levels ranging
from 10 to 150 mg/kg. Hypoglycemia and fluid retention
were noted as dose-dependent and reversible side
effects. The maximum tolerated dose was determined to
be 80 mg/kg. No significant hematological, liver, or renal
changes were observed. The good tolerance of high levels
of cCr also in animal models and its unique mechanism of
action make it a potentially attractive addition to cancer
chemotherapy.
In addition to cCr, a series of other Cr and PCr
analogs were evaluated for antitumor activity against the
ME-180 cervical carcinoma, the MCF-7 breast adenocarcinoma,
and the HT-29 colon adenocarcinoma cell lines
(60). Several analogs, exhibiting thermodynamic and kinetic
properties distinct from those of Cr and PCr, and
thus impacting the rate of ATP production through CK
(see sect. VIIIB), inhibited the growth of the established
tumor cell lines in culture with IC 50 values in the low
millimolar range. The compounds that were active in vitro
were also shown to be active in vivo and substantially
delayed the growth of subcutaneously implanted rat
mammary adenocarcinomas. From the Cr analogs tested,
cCr and phosphinic cyclocreatine were most active. The
antitumor activity appeared to require rapid phosphorylation
and buildup of new stable phosphagens that are
less efficient than PCr in regenerating ATP. Compounds
like GPA and hcCr became active after longer exposure
times. This suggests that poor substrates will become
active as antitumor agents if allowed sufficient time to
build up their phosphorylated counterparts. Cr was inactive
in these assays both in vitro and in vivo.
From the phosphorylated series of compounds, PCr
and PcCr were most active, and several others were modestly
active (60). Unlike the nonphosphorylated series,
there was no obvious relationship between the antitumor
activity of these compounds and their ability to generate
ATP through CK. Furthermore, these phosphorylated
molecules are poorly taken up by cells and may therefore
modify a target distinct from CK, possibly at the membrane
level. However, PcCr was shown to exhibit similar
specificity and potency as cCr against high CK expressing
tumor cells (859). Interestingly, CK activity seemed to be
required at least in part for mediating the compound’s