Estrogens and qrowth of malignant melanoma
Myra K. Jacobsohn and Gert M. Jacobsohn
Department of Biological Chemistry, Hahnemann University,
Philadelphia, PA 19102, USA
Melanin, the brown-black polymer in pigment cells, is being recognized increasingly as
a non-specific binding agent for various organic substances. This binding is in lieu of,
or in addition to, receptor protein binding which has a relatively higher specificity than
the melanin. We have found that melanin is capable of binding estrogens;
2-hydroxyestradiol, a metabolite of estradiol produced in the liver, brain and uterus,
binds to a somewhat higher extent than estradiol (Jacobsohn et al. 1988). Melanin which is
being actively synthesized from dopa binds more estradiol than preformed melanin
(Jacobsohn et al. 1988). The binding is not a unique phenomenon. A number of aromatic
substances have the ability to bind to melanin and this phenomenon has been used to
explain the localization of drugs at their site of action or the concentration of toxins
in specific locales where they may exercise a damaging effect. The binding may also remove
harmful substance from causing damage elsewhere. The successful treatment of tinnitus by
lidocaine injection was thought by Lyttkens (1986) to be due to the drug accumulating on
melanin of the inner ear. He found lidocaine, bupivacaine and chlorpromazine, a
phenothiazine derivarive, to associate with melanin-containing structures of the eye and
of the inner ear (Lyttkens et al, 1979). Melanin of the eye, hair and skin of pigmented
mice can take up serotoni, dopamine, epinephrin, and norepinephrin (Lindquist, 1973).
Selegiline and other amphetamines, chloroquine, dopamine, norepinephrin, and the morphine
derivative MPTP, can bind to melanin granules (Bathory et al,1987; Stepien et al, 1987;
Lindquist 1973; Lindquist et al, 1987). Debing et al (1988) reported the binding of 15
aromatic drugs to calf eye melanosomes and to synthetic dopa-melanin.
The binding of estrogens to melanin of melanoma could be responsible for the favorable
effect associated with estrogens during the later course of development of the malignancy
in women.
Women with melanoma, especially during the child-bearing age, have a better survival rate
than men. The effect may be due to oxidative reactions of estradiol leading to formation
of ortho-quinones which, by acting as melanocytotoxic agents, could provide the host with
a mechanism for defense. Precedence exists for the action of quinones as cytotoxic
intermediaries. A specific strategy for treatment of melanoma is based upon tyrosinase
activation of pro-drugs with selectivity conferred by the enzyme (Wick 1983; Pezzuto et
al, 1988). Quinones and semiquinones are generated and it is assumed that semiquinones act
biologically via generation of superoxide (Pezzuto et al, 1988). The cytotoxicity of
4-hydroxyanisole to malignant melanoma was proposed to be due to its oxidation to the
ortho-quinone and subsequent reverse dismutation to the semiquinone, but it is uncertain
whether the quinone or the semiquinone is the actual cytotoxic product (Riley 1985; Nilges
et al, 1984). In studies on malignant cells in culture, estradiol, 2-hydroxyestradiol, and
2-methoxyestradiol were found to be cytotoxic to dividing MCF-7 and HeLa cells (Seegers et
al, 1989). On exposure af these cells to high concentrations (³
1 x 10-6 M) of the steroids, they showed increased mitosis with formation of
abnormal and fragmented polar bodies and disoriented microtubule arrangements (Seegers et
al. 1989).
Tyrosinase, the enzyme responsible for melanin formation by way of successive oxidations
of tyrosine and its products, has the ability to oxidize estrogens (Jellinck et al, 1971;
Jellinck et al, 1963; Jacobsohn et al, 1988; Jacobsohn et al, 1984). Mushroom tyrosinase
has been used for some time in the laboratory preparation of 2-hydroxyestradiol from
estradiol (Jellinck et al, 1971; Hersey et al, 1981), and we found that the enzyme
can oxidize the hydroxylated product in stoichiometric relationship to molecular oxygen in
presence of catechol (Jacobsohn et al, 1984). The enzyme used in this work originated from
mushrooms but it is likely that the mammalian enzyme can oxidize estrogens as well.
Incubations of 2,4,6,7-3H-estradiol with tvrosinase isolated from B-16 melanoma
cells yielded 3H-water (Ewaskiewicz and Jacobsohn, in prepn.) by methodology
analogous to the Pomerantz assay for tyrosinase activity (Pomerantz, 1969). Incubations of
[14C ] - estradiol or [14C]-2-hydroxyestradiol, together with DOPA
and tyrosinase from the B16 mouse melanoma cell line, produced melanin with the estrogen
label firmly attached to the pigment (Jacobsohn et al, 1988). The steroids were attached
to melanin in such a way that they were resistant to extraction with organic solvents such
as ethyl ether or hot methanol. This was taken as evidence that the unextractable portion
of steroids had been oxidized and bound to melanin in covalent linkage; similar
experiments with the mushroom enzyme have already shown that the 2-hydroxy estrogen can be
oxidized and incorporated into melanin (Jacobsohn et al, 1988; Jacobsohn et al, 1984). An
alternate but remote possibility is that the estrogens are bound to inner spaces within
the pigmented polymer where the solvent cannot reach them. In more recent experiments, it
was possible to isolate and identify the 2,3- orthoquinone of estradiol from incubation
mixtures of the fungal enzyme and 2-hydroxyestradiol (Jacobsohn et al. 1989; Jacobsohn et
al, 1990). Incubations of the quinone with murine tyrosinase and dopa have shown that the
quinone can be incorporated into melanin, perhaps by additional enzyme-catalyzed steps or
through reverse dismutation with dopa-melanin intermediates (Jacobsohn et al, 1990). It
should be noted that melanin itself, in the absence of tyrosinase, may function as an
oxidant for estrogens, because exposure of melanin to estradiol or 2-hydroxyestradiol can
cause physiologically significant amounts of steroid to be retained by the pigment
(Jacobsohn et al, 1988; Jacobsohn et al,1990).
Estrogens are thought to be co-carcinogens because they are able to exacerbate the
activity of true carcinogens but do not appear to initiate growth of a malignancy by
themselves. After the growth of melanoma cells has been triggered by external events,
estrogens may intensify the malignancy by binding to melanin of affected cells. This may
explain the often-noted higher incidence of melanoma in women compared to men. As the
disease progresses and increased amounts of estrogens are concentrated in melanoma cells,
their conversion to cytotoxic oxidation products may present a more favourable outcome. We
hope that this hypothesis will stimulate additional research on the relationship of
estrogens to growth and development of pigment celIs.
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