Cancer Immune Therapy Edited by G. Stuhler and P. Walden ...

13.1 History of Cancer Immunotherapy with Bacterial Extracts and Nucleic Acids
mor responses. Endotoxin became an obvious candidate for mediating this toxicity,
especially as it was recognized to be a trigger for the production of the hopefully
named tumor necrosis factor (TNF). Although lipopolysaccharides (LPSs) have
many remarkable immune effects, they clearly cannot be the sole explanation for the
antitumor activities of ªColey's toxinsº. One reason is simply that the first patient
treated by Coley was treated only with streptococcus, which of course has no endotoxin.
Second, endotoxins are very toxic, which appears to preclude their human clinical
development, with the exception of detoxified forms such as monophosphorylipid,
which has shown activity as an adjuvant.
In order to determine the active component of BCG, Tokunaga et al. fractionated it
into the various components and assayed each of these for their antitumor activity
[6]. Surprisingly, these studies demonstrated that only the DNA fraction of BCG contained
antitumor activity. BCG DNA was further shown to induce natural killer (NK)
cell activity, and the production of type 1 and type 2 interferons (IFNs) [7]. Tokunaga's
discovery led to human clinical trials of a BCG DNA preparation in Japan, with some
encouraging responses, but no further clinical development [8]. By cloning mycobacterial
genes and synthesizing constituent oligodeoxynucleotides (ODN), these investigators
concluded that the immune stimulatory effects of BCG DNA could be attributed
to the presence of certain self-complementary palindromes in these ODN [9].
All of the active palindromes were noted to contain at least one CpG dinucleotide
and to be more common in the genomes of bacteria compared to humans [10]. The
mechanism of immune recognition of the palindromes was thought to depend on
their secondary or tertiary structure and methylation of the CpGs was reported to
have no influence on the immune stimulatory activities of the DNA [10].
In independent studies, Pisetsky et al. reported that bacterial DNA (bDNA) induced
murine B cell proliferation and immunoglobulin secretion, but vertebrate DNA did
not [11]. This effect was shown to be independent of any LPS contamination of the
bDNA because it was abolished by nuclease digestion of the DNAs and was not affected
by polymixin. bDNA was not unique in being immune stimulatory ± Pisetsky
et al. also demonstrated that synthetic poly(dC 7 dG) was a B cell mitogen and that
this mitogenicity was abolished by methylation of the cytosines [12]. However, these
investigators also did not associate CpG methylation with different immune activities
of bacterial and vertebrate DNAs. Instead, they suggested that the immune stimulatory
effects resulted from unique higher-ordered structures of the DNA molecules
that were disrupted by methylation.
Whiletheimmune stimulatoryeffectsofthese DNAmoleculeswereunexpected, they
should not be totally surprising. After all, nucleic acids are polyanions, and can form
hydrogen bonds with other biological molecules, which in principle could give them
drug-like effects. In fact, certain double-stranded RNA structures have been known
for many years to have immune stimulatory effects, which are thought to result from
an immune defense mechanism triggered by viral double-stranded RNAs [13]. This
immune activation can be mimicked by poly(rI 7 rC), which induces NK cell activity
and Th1-like immune activation with interleukin (IL)-12, IFN-a and IFN-g production
in both murine and human cells [14, 15]. In mice and primates, poly(rI 7rC) has prophylactic
and therapeutic activity against otherwise lethal viral infections [16, 17], and
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