The Search for the Human Breast Cancer Virus
Elisabeth
Rieping, Gemuender Str.15, D-50937 Köln
Until today the search for a human breast cancer virus did not result in
a reproducible method of isolation. Analysis of successful transmissions show
that there might be an agent that can become transmitted cell-bound causing
lymphoma in mice. It is not unlikely that the agent is a provirus harbored by
mononuclear blood cells quite similar to HTLV-1 and BLV.
Ever since the discovery that breast neoplasms of the mouse are
transmitted by a virus, scientists have tried to discover a similar agent
responsible for the disease in women.
The
standard procedure to find a transmissible agent, a transfection experiment to
another person, was not used, of course, for ethical reasons.
Instead researchers tried to
transmit human breast cancer to mice. But in the animals infected with human
breast cancer material, the incidence of mammary tumors increased only slightly
from less then 1% to 3%. However, a significantly increased incidence of
lymphoma in the infected mice was observed [i].
Aside from the
abovementioned research, no other work concerning the transfection of human
breast cancer to animals could be found. Presumably, this is not because nobody
tried it, but because nobody tried it successfully. So why was this group at
least partially successful?
The usual procedure
microbiologists use to distinguish between bacterial and viral agents is to
filtrate the material. Of cause,
this filtration also blocks the transmission of cells.
But these successful researchers prepared their human breast cancer material differently. They homogenized it, added antibiotics to kill contaminating bacteria, centrifuged it and extracted the interphase between cells and fat. Using this procedure, their material will have contained enough cells to transmit some latent proviruses integrated in the DNA of these cells. If they had used the usual procedure of filtering, cell-bound viruses would not have been transmitted.
It is likely that other
researchers who were not successful in transmitting the breast cancer virus used
filters as this was then standard procedure. At that time it was not known that
some retroviruses, for example the bovine leukemia virus BLV, seem to try to
escape eradication by producing little virus in vivo [ii].
Viruses like BLV or the human immune deficiency virus, HIV, which are
transmitted by live cells, were not yet well known in 1977.
More recently, in 1995, the
frequent development of murine T-cell lymphomas after implantation of
human inflammatory breast cancer cells in nude mice was demonstrated [iii].
These two experiments hint
at the existence of a transmissible agent in breast cancer material. Probably
both experiments worked because actual cells were used for the transfection, although in the first
experiment cells were probably used unintentionally.
Already in 1986, monocytes from breast cancer patients had been
discovered to form giant cells [iv].
As giant cell formation may be a sign of viral infection, attempts were made to
find retroviral particles and to test them for reverse transcriptase activity.
After
cultivating mononuclear cells for some days, particles could be harvested from
97% of all breast cancer patients and from about 11% of healthy control group
participants. It is possible that this 11% represents women who were carriers, but who had not yet developed
cancer.
The reverse transcriptases
of retroviruses work best with
magnesium ions, while cellular enzymes prefer manganese ions. The enzyme of the
particles from cultivated monocytes of breast cancer cells preferred magnesium
ions. The particles were described to be slightly smaller than the murine
mammary tumor virus MMTV. Its size was described to be more similar to that of
HIV [v].
The work of this group from
Liverpool was not continued because two other groups had unsuccessfully tried to
confirm the work. They both failed in different ways.
One group could neither
cultivate the monocytes in a way to produce multinuclear giant cells, nor could
they find reverse transcriptase activity[vi].
These test, done with the same primer that had been used in the original work,
showed neither reverse transcrpitase activity with the material from cultivated
monocytes of human breast cancer cells, nor with HIV-1 or with HTLV-1 which were
used as positive controls. No information was given by the researchers to
elucidated why this was so.
Contrary to these results, the other group found significant multinuclear giant cell formation. But they found them not only in cultured monocytes from breast cancer patients, but also in those of control subjects with no or with benign breast disease [vii].
Fortunately this publication
was very detailed. One change in the design of their study in comparison to the
original work, they wanted to reproduce, was the use of a different tissue
culture medium : pooled female AB serum. In the successful experiments published in
1988 calves serum had been used. Unpooled serum derived from one animal raised
in a controled environment is much less likely to be contaminated.
As the original Liverpool
study had shown that the retrovirus like particles could be cultivated from the
blood of about 11% of healthy
controls, possibly women who were
already infected, but who did not yet have a tumor big enough to be clinically
detectable, pooled human serum might contain material from theses virus carriers.
An additional result from
this study is very interesting. In spite of giant cell formation, a reverse
transcriptase activity could not be demonstrated. The crude supernatant from the
cell cultures even blocked the activity of MMTV reverse transcriptase, which had
been used as a positive control.
As the originally described
particles could be observed only after several days of cultivation in non-human,
serum conditioned tissue culture medium, it is possible that by growing the
cells with the help of pooled human serum, the viruses produced by or released
to the medium were inhibited or that their reverse transcriptase activity - like
that of the MMTV particles - was blocked.
In a recent review on a possible retroviral etiology of human breast
cancer [viii]
the author, probably stimulated by her experience with bone disease [ix],
[x]
but also by examples of other animal [xi],[xii]
and human retroviruses[xiii]
pointed out that blood mononuclear cells should be examined first when searching
for a human breast cancer virus and that a possibly causative retrovirus must
not necessarily be MMTV or a very close relative.
Unlike MMTV which is
difficult to perceive as an agent accessing the human food chain (or another way
of transmittance) bovine leukemia virus BLV is a syncytia or giant cell forming virus
contained in milk used for human nutrition, especially for nutrition of human
and pet animal infants in Western, but today also in non -Western societies. It
has been suspected since its discovery, but with old methods, antibodies against
it could not be found in human sera.
This has changed however
[xiv].
Using immunoblotting to test the sera of 257 humans for antibodies of four
isotypes (IgG1, IgM, IgA, and IgG4) to the BLV capsid antigen p24, at least one
antibody isotype reactive with BLV was detected in 74% of the human sera tested.
The specificity of the reactivity was strongly suggested by competition studies
and by ruling out cross-reacting antibodies to other chronic human viruses.
Independently and earlier
than the above mentioned work, antibodies against BLV in humans had been
demonstrated by an other group[xv]
They had looked for antibodies against BLV in patients with multiple sclerosis
and found them both in patients and in controls. Therefore they discontinued
this work.
Changes which BLV might
induce in bovine mammary tissue were also sought. Altered growth properties of
bovine mammary epithelial cell lines containing BLV DNA were found. The cell
lines showed reduced population-doubling time, higher saturation density, and
increased longevity, features that are typical for tumor cells [xvi].
The evidence that BLV could produce such changes in bovine mammary cells
inspired a search for BLV DNA in human breast cancer.
DNA of BLV was found
in tissue of breast cancer patients. The study is going on [xvii].
If BLV is the cause of human
breast cancer and if the infection is transmitted by infant food contaminated
with a bovine born virus, [xviii],
it should be traceable in about every tenth person in Western society since
about one in ten women gets breast cancer.
In NewYork healthy blood
donors are checked for human T-cell lymphothropic viruses HTLV-1 and HTLV-II,
although the city is not an area in which this viruses are endemic. About 8,6%
of the blood donors diagnosed to be free of HTLV-I and II, as they lack
antibodies against viral structural proteins, were shown to have antibodies
against a Tax protein homologous to the HTLV Tax Protein. DNA homologous to TAX
DNA was found by two independent groups as well [xix],[xx].
As New York is not an area in which HTLV is endemic, this was a surprising
result.
It would not be surprising
if this protein, homologous to HTLV-1 Tax, is the BLV Tax protein. BLV is a
virus closely related to the HTLV-I and is contained in the bovine milk used as
food by humans.
BLV DNA is integrated in the
DNA of bovine milk lymphocytes. It is likely that it is also in the pooled
bovine milk used for the production of baby food. As DNA is not destroyed by
cooking, the integrated BLV DNA will not be destroyed by the heat based
procedures used to make infant food more safe. DNA is sufficient for
transfecting the virus.
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