Download A proposal for a simple and inexpensive therapeutic cancer

Immunology and Cell Biology (2012) 90, 310–313
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THEORETICAL ARTICLE
A proposal for a simple and inexpensive therapeutic
cancer vaccine
Aude M Fahrer
In this essay, I propose a new method of treating tumours, using an old and inexpensive preparation, that I contend would be
of considerable benefit to patients and their cancer management. My rationale for this treatment initially arose from recent advances
in the understanding of dendritic cell function. (Dendritic cells are key cells of the immune system that are able to either turn
on or turn off T-cell responses.) Evidence to support this approach is found in 100-year-old studies on the immunotherapy of
cancer. Also, I draw on some remarkable, but little-known studies from the 1960s–1990s, demonstrating that the preparation
has already been trialled in humans (although not intratumourally, as I propose), and is considered sufficiently safe to proceed
with clinical trials in cancer volunteers.
Immunology and Cell Biology (2012) 90, 310–313; doi:10.1038/icb.2011.42; published online 24 May 2011
Keywords: cancer immunotherapy; cancer therapeutic; cancer vaccine; Coley’s toxin; Complete Freund’s Adjuvant
The immune system can eliminate cancers. Indeed most people develop
cancerous cells,1 but these are usually held in check without ever
becoming detectable. The immune system is one of the major mechanisms preventing cancer growth. Cancers that do develop have done so
by successfully evading or subverting the immune system.2–4 Nonetheless,
it is possible to stimulate the immune system to eliminate even established cancers. Both adoptive cell therapy (where T cells infiltrating the
tumour are expanded in vitro, and then re-injected into the patient)
and personalised dendritic cell vaccines (where dendritic cells are
removed from a patient, expanded, given cancer antigens and then
re-injected) have provided spectacular remissions in some patients.5–8
A BRIEF SUMMARY OF IMMUNE CELL ACTIVATION
T cells, especially cytotoxic T cells, can directly kill cancer cells. Before
they can do this, however, they must be primed (that is, initially
activated and expanded) by dendritic cells. The function of dendritic
cells is to continuously sample the proteins in a tissue (for example,
those from cancerous cells or from infectious microbes) and then
carry these proteins back to lymph nodes where they can be shown to
T cells.9 Activated dendritic cells can prime T cells specific for the
proteins they display. Once activated and expanded, cytotoxic T cells
then return to the tissues to kill any other cell (in this case, cancer cells
or infected cells), displaying the same protein.
However, it is now clear that the default setting of dendritic cells is
to switch off (tolerise), rather than to prime the immune response.
Dendritic cells are constantly moving to lymph nodes to present
cellular proteins to T cells.9 Most of the time such proteins come
from normal cells, and the dendritic cells have not been activated.
By turning off or deleting any T cells specific for these proteins, nonactivated dendritic cells prevent autoimmune diseases. They can also,
however, prevent anti-cancer responses.
A most effective way of stimulating dendritic cells is to expose them
to bacteria or viruses. Dendritic cells bear pattern recognition receptors, the best studied of these are the Toll-like receptors,10 which
recognise microbial products such as outer membrane constituents,
unmethylated bacterial DNA or double-stranded viral nucleic acids
(sometimes called ‘danger signals’). Exposure to microbes thus activates dendritic cells, allowing them to prime, rather than switch off Tcell responses. This is how successful immune responses are mounted
against infectious diseases.
Co-injection of bacterial products is also the empirical method by
which immunologists, for decades, have elicited immune responses in
experimental animals. The most potent of these immune-stimulating
preparations, routinely used since the 1950s, and readily available
commercially, is Complete Freund’s Adjuvant (CFA).11 It consists of
heat-killed mycobacterium bacteria in oil. Before use, it is vigorously
mixed with an aqueous solution of the protein to be injected, creating
a water-in-oil emulsion. Injection of this emulsion creates a localised
depot of protein and killed bacteria, which are slowly released over
weeks or months. This causes an influx of immune cells to the site. It
can now be understood that activation of dendritic cells is the key to
the remarkable immune responses generated.
THE HYPOTHESIS
My proposal therefore is to inject CFA, emulsified with saline, directly
into the tumour. As well as causing an influx of immune cells into the
Biochemistry and Biomedical Sciences, Research School of Biology, College of Medicine Biology and Environment, The Australian National University, Canberra, Australian Capital
Territory, Australia
Correspondence: Dr AM Fahrer, Biochemistry and Biomedical Sciences, Research School of Biology, College of Medicine Biology and Environment, The Australian National
University, Building #134, Linnaeus Way, Australian Capital Territory 0200, Australia.
E-mail: [email protected]
Received 13 December 2010; revised and accepted 13 April 2011; published online 24 May 2011
Proposal for therapeutic cancer vaccine
AM Fahrer
311
tumour, it will directly activate the resident dendritic cells. As these
dendritic cells are already sampling tumour proteins and presenting
them to T cells, this should achieve priming of a potent anti-cancer
T-cell response.
EVIDENCE SUPPORTING THE HYPOTHESIS
As already stated, successful outcomes from personalised dendritic
cell vaccines, in which the dendritic cells are expanded, loaded with
cancer proteins, and, during this in vitro process, activated, show that
dendritic cells can drive anti-tumour responses. This personalised
approach to cancer therapy, however, is labour intensive (the cells
need to be removed from each patient and cultured in vitro before
being re-injected) and extremely expensive. It will also only work if the
correct subtypes of dendritic cells were initially amplified, and if the
dendritic cells returned to the body successfully home back to lymph
nodes to prime an immune response.12 Activation of the dendritic
cells in situ, as I propose, could overcome all of these hurdles.
Dendritic cell activation can also explain the earlier evidence of
tumour remission following spontaneous or induced infections
(1700s–1900s), and following infections after non-aseptic surgery.13
Thus, bacterial infections fortuitously occurring at or near the site of a
cancer would activate dendritic cells presenting proteins from the
cancer. Indeed, it has been proposed that our much ‘cleaner’ modern
society, with aseptic surgery, and with the widespread use of antibiotics to treat bacterial infections—while evidently leading to a
dramatic decrease in serious disease and death from bacterial diseases—has also led to a dramatic reduction in the rate of spontaneous
cancer remissions.13,14
Dendritic cell activation can also explain the remissions achieved
following repeated injections of bacterial products into, or near the
tumour. This technique was pioneered by William Coley who injected
a mixture of heat-killed Streptococcus pyogenes and Serratia marcescens.15 Even by today’s standards, Coley’s success rate in treating
cancer (in the 1890s to the 1930s) was remarkable. Five-year survival
rates for patients treated with Coley’s toxin were 43% for inoperable
cancers (including carcinomas, melanomas and sarcomas) and 61%
for operable cancers.16 The 5-year survival rates for inoperable
sarcomas was 52%, with 21% of patients remaining disease free for
at least 20 years.17 However, to achieve these results, Coley injected his
patients every day, or every other day for months on end.
Coley used intratumoural injections almost exclusively until 1898,16
but around 1906–1915 began to favour intramuscular, and later
intravenous injections of his toxin.16,18
‘Coley, anxious to prove that his toxins had a systemic rather than a
local reaction such as X-ray, radium and surgery, stopped using
intratumoral injections about 1906, and not until a year or two
before he died did he come to realize the mistake he had made.’18
The bacterial component of CFA is heat-killed mycobacteria. Live
BCG (Bacille Calmette-Guérin, a Mycobacterium bovis strain), as well
as purified subcomponents of BCG, have been used in many human
cancer trials in the 1960s and 1970s (comprehensively reviewed by
Hersh et al.19). Many different routes of BCG administration were
tried. Prolongation of disease-free interval and/or prolonged survival
was achieved in several types of cancer. Hersh et al. concluded that:
‘The requirements for this effect are either intimate contact between
the BCG and tumour cells, or that the tumour and the BCG be in the
same regional lymph node drainage.’
Mycobacterial products are currently used to treat cancers. Live
BCG is a standard treatment for certain forms of superficial bladder
cancer.20 This is an empirically based treatment; the bacteria are delivered
intravesically, in close proximity to the tumour. Mycobacterial extracts
have successfully been used in Japanese cancer patients.21 Mycobacterium
vaccae is being used to treat cancer patients in England.22
Thus, there is compelling evidence that bacterial presence in or near
a tumour can lead to tumour regression. The mechanism of action has
not previously been elucidated, but is discussed in this manuscript.
Furthermore, mycobacteria have been successfully used in cancer
therapy. The advantage of using CFA is that it forms a depot lasting
many weeks. It should therefore achieve the effect of Coley’s toxin, but
only require a single injection (or at most a few, infrequent, injections)
into the tumour. Similarly, CFA would provide a sustained immune
response, when compared with intratumoural live BCG, which is
usually cleared rapidly, and use of CFA eliminates the risk of systemic
infection, a known side effect of the live BCG cancer therapies in a
minority of patients.
Thus, a simple CFA/saline emulsion should be adequate to achieve
the required dendritic cell activation. A somewhat more complex
possibility would evidently be to emulsify Coley’s two bacterial strains
in the aqueous phase of CFA to obtain a preparation containing gram
positive, gram negative and acid-fast bacteria.
UNJUSTIFIED ILL REPUTE OF CFA
CFA, however, currently has a bad reputation. In certain institutions
its use is being discouraged for immunisation of experimental animals.
This is due to local abscess formation at the injection site when
injected intradermally, and the formation of granulomas if injected
more deeply and the potential pain associated with the site of injection.
Most institutions allow animals to be injected only once with CFA,
requiring any booster doses to be given in Incomplete Freund’s Adjuvant
(the same preparation but without the mycobacteria).
In humans, a handful of cases of toxic sequellae after accidental
injection of CFA have been reported,23 contributing to its unfortunate
reputation. The worst of these involved swelling of the limb and pain.
At least some of these reactions can be attributed to the material coinjected with the CFA, rather than to the adjuvant itself.
To be effective, the adjuvant has to be emulsified into a very thick
paste. One of the most readily available methods for doing this
(indeed advocated by Freund himself for creating emulsions24) is to
repeatedly suck the preparation up and down through a syringe and
needle. As the preparation becomes thicker, it becomes harder and
harder to squeeze out of the syringe. It is during this process,
therefore, that countless immunologists (including myself) have
accidentally stuck themselves with this preparation. Thus, the few
published cases of severe reactions are not necessarily representative
of the effect of CFA in the majority of humans.
Incomplete Freund’s Adjuvant has been used in at least 55
human trials, as excellently summarised in the supplemental data
to Miller et al.25
I am aware of three sets of studies in which CFA was used in human
trials, all in cancer patients, although none injected the adjuvant
directly into the cancer.
A 1962 Buffalo, New York, trial involved 232 patients with gynaecological cancers injected with tumour cells or tumour extracts
intradermally in CFA.26 The study’s authors received personal advice
from Freund. Ulcers at the injection site occurred in 79% of patients
healing over the course of several months. Despite the negative
conclusions, 30-month survival rates were 18/232 injected with
vaccine vs 1/139 not injected with vaccine.
A 1970 trial in Queensland, Australia, by Hughes et al.,27 involved
20 patients suffering from malignant melanoma, colon, stomach or
squamous cell vulval carcinoma, or neurofibrosarcoma. Seventeen of
these were injected with tumour extracts in CFA subcutaneously.
Immunology and Cell Biology
Proposal for therapeutic cancer vaccine
AM Fahrer
312
Seven patients developed abscess formation or skin ulceration, but
only after a second or subsequent injection with CFA—no patient
developed these complications after the first injection. Seven of the
17 patients were thought to have benefitted from this treatment.
Finally, and most recently, a series of remarkable trials reported by
Hollinshead et al. used tumour extracts emulsified in CFA. Phase II
trials have been reported for melanoma28 and for colon cancer 29,30
patients. Phase II and III trials have been carried out in lung cancer
patients.29,31–33 In each trial, patients received three intradermal
injections of tumour antigens in CFA, or CFA alone at monthly
intervals. Injections were given into the arms and thighs, not into the
tumour. Side effects reported were ulcers at the site of injection, which
healed over several months and, in some patients, overnight fever
following the injections. The numbers of patients receiving emulsified
CFA totalled at least 245. The results were extraordinarily encouraging:
the lung cancer trials showing differences of around 69 vs 49% 5-year
survival for patients receiving this immunotherapy vs those who did
not. My point in presenting these data, however, is to show that CFA
has already been used in human trials. The side effects—ulcers at the
site of injection and fever—are considerably less oppressive than those
produced by many currently used chemotherapy agents.
SITE OF INJECTION
While the studies mentioned above certainly had beneficial effects in
many patients, none of these studies injected CFA directly into the
tumour. I am arguing that from what we now understand about
dendritic cell priming of the immune system, this should be far more
beneficial, and would also negate the requirement for the preparation
of tumour antigens, from surgically resected tumours, to inject
together with the adjuvant. The notable advantages are that
(1)
The immune response can potentially be directed at all aberrant
cancer proteins within the patient’s tumour rather than to a few
purified proteins, thus maximising the chance of a successful
immune response and minimising the risk of immune escape by
remaining tumour cells.
(2) The treatment is immediately applicable to any solid tumour
and
(3) It is inexpensive.
A successful immune response should be effective both against the
primary tumour, and against any metastases. Injection could therefore
be made into a metastatic tumour, if more accessible than the primary
tumour.
POSSIBLE SIDE EFFECTS
Fever (in the 24 h after injection) is a likely side effect. Pain at the
injection site is also possible, as discussed in a few cases of accidental
injection with CFA. However, it should be noted that Coley reported
that injection of bacteria directly into tumours often rapidly relieved
the pain associated with the tumour, to the point where the patient no
longer required pain killers.16 Furthermore, in the human trials
described above, injections of CFA were well tolerated by patients.
As the CFA remains localised at the site of injection, it can be
excised in the event of any unforeseen or intolerable side effects.
The most serious side effect to consider is the possibility of the
induction of an autoimmune reaction years (or decades) after treatment. This has not been observed in any of the CFA cancer trials,
despite being specifically looked for in Hollinshead’s colon cancer
trials.29 It is also not observed in the millions of experimental animals
injected with CFA, except in specific cases, for example where high
Immunology and Cell Biology
concentrations of myelin proteins emulsified in the adjuvant are used
to induce experimental autoimmune encephalitis.34 Moreover, the
theoretical possibility of autoimmune induction is not likely to be of
major concern to patients in the terminal phase of their disease.
CO-INJECTION WITH OTHER IMMUNOMODULATORY
SUBSTANCES
Evasion of the immune response is a pre-requisite for the establishment of tumours. The treatment I am proposing aims to overcome
this immune evasion by activating dendritic cells. Indeed, sustained
dendritic cell exposure to bacterial lipopolysaccharide (which binds
Toll-like receptor 4) has been shown to overcome T-cell tolerance, and
protect mice from challenge with tumours expressing the tolerising
protein.35
CFA injection is also likely to have a second, independent, antitumour action. The influx of large numbers of immune cells to the site
of injection, and the cytokines produced by these cells will work to
counteract the immunosuppressive milieu established by most
tumours. In the most favourable case, this could lead to rapid reactivation of any quiescent tumour-infiltrating T cells, providing a
second, even more rapid route to tumour destruction. This secondary
aspect of the treatment could be enhanced pharmacologically.
Several potential immunomodulatory antibodies or cytokines have
been proposed for the treatment of cancer patients. Some of these
would be toxic if injected systemically, but may be useful at low doses
injected intratumourally to give very localised immunomodulation.
Examples include anti-interleukin-10, anti-transforming growth factor-b and blocking antibodies against T-cell inhibitory receptors such
as CTLA-4 and PD-1.36 An anti-CTLA-4 monoclonal antibody,
ipilimumab, has recently become available for melanoma treatment.
These could ultimately be injected together with the CFA (although
not as part of the emulsion) to potentiate the treatment.
SELECTION OF PATIENTS, METHODS OF TREATMENT
I envisage two possible scenarios for intratumoural CFA treatment.
(1) Coley had remarkable success treating patients with advanced,
inoperable tumours. This treatment could therefore be tried in
late stage cancer patients. It should be noted, however, that
chemotherapy, and some forms of radiotherapy are highly
immunosuppressive. Therefore, while I suggest that this should
be trialled in any willing patient with solid tumours, who has no
other treatment options remaining, the overall efficacy of the
treatment should be judged especially in the rare patients with
inoperable tumours who have not previously undergone other
treatments.
(2) An equally valid approach would be to trial the treatment in
patients who are facing a wait of at least 3 weeks between
diagnosis and surgery to remove a tumour. This would allow
enough time for induction of an immune response, after which
the tumour (together with the CFA) would be surgically
removed. Pathology examination of the tumour could give
immediate clues as to the success of the immune response
induction.
CONCLUSION
I propose a simple, commercially available, generally applicable and
extremely cheap cancer vaccine that could be trialled immediately:
injection of CFA, emulsified with saline, directly into the tumour. This
could be trialled in the treatment of a wide variety of solid tumours.
Proposal for therapeutic cancer vaccine
AM Fahrer
313
At this stage, the success of such a treatment is hypothetical. I hope
to have convincingly argued that
(1) it has a good chance of being successful in a substantial proportion of patients and
(2) that as CFA has already been used in human cancer patients,
it would be ethical to trial this immediately.
I urge any interested clinicians or medical facilities to investigate this
hypothesis using observational studies in cancer volunteers or through
larger-scale clinical trials.
CONFLICT OF INTEREST
The author declares no conflict of interest.
ACKNOWLEDGEMENTS
I thank Professor Pierre Chapuis, Professor Ian Clark and Dr Iain Wilson
for helpful advice and encouragement. I also thank the Australian Capital
Territory (ACT) Cancer Council for research funding; meeting their volunteers
inspired me to develop and pursue this hypothesis.
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