Biomedical Engineering Reference
In-Depth Information
19.6 FUTURE DIRECTIONS
TABLE 19.5 Summary of T
reg
and Fusion Protein
Combination Therapy in COLON 26-Bearing BALB/C
Mice at 19 Days Post-tumor Implantation
The overall objective of this chapter was to evaluate the
efficacy of genetically engineered fusion proteins developed
in our laboratory for the immunotherapy of solid tumors.
The use of chTNT-3, which targets necrotic areas of solid
tumors regardless of the species or origin of the tumors,
enabled us to compare by the same targeting antibody, the
relative therapeutic potential of selected cytokines, chemo-
kines, and co-stimulatory molecules in tumor-bearing
immunocompetent mice. This point cannot be overempha-
sized since there are few antibodies available that target
murine tumor models. By our studies, it has become clear
that the targeting of specific but not all immune modulators
to the tumor microenvironment can cause effective destruc-
tion of established tumors. As controls, soluble nontargeting
Fc fusion proteins were also prepared for most of these
immunoregulatory molecules, and to our surprise, many of
these appear to be as effective as the corresponding antibody
fusion proteins that target tumor. It appears that when
administered in high enough concentrations, soluble Fc
fusion proteins can also reverse immune tolerance to tumor.
Various cytokines, chemokines, and co-stimulatory mole-
cules are highly potent proteins, requiring a much lower dose
than antibodies [31]. This is seen clearly in dosing studies in
which a threshold amount of soluble Fc is required for the
induction of tumor suppression. Threshold amounts were
less often seen in those mice receiving antibody fusion
proteins. Although very potent in their antitumor action, it
remains to be seen if these soluble Fc fusion proteins are more
toxic in humans than corresponding antibody fusion proteins
or if they are effective against larger, well-established tumors.
The other major finding of these studies, namely the role of
T
reg
cells in tumor growth, illustrates the importance of
understanding existing mechanisms of immune tolerance in
cancer patients. For instance, do all cases of a given tumor
type evoke a similar mechanism of tumor escape, and do
metastatic lesions display the same mechanism as primary
lesions in the same patient? These types of questions should
be answered prior to proceeding with the testing of specific
immunotherapeutic reagents to maximize the therapeutic
potential of a given reagent. We should also be aware that
histopathologic diagnosis may not predict which immuno-
suppressive mechanisms are at play in individual patients.
Instead, a comprehensive analysis of the immune phenotype
in patients may be necessary to identify correctly which
immunotherapeutic approach will be effective [109].
Tumor Reduction (%)
Antibody
Fusion
Protein
Dose
(ug)
CD25
Depleted
CD8
Depleted
CD4
Depleted Nondepleted
B7.1/NHS76
30
100
<
5
100
40
B7.1-Fc
10
70
NA
NA
45
0
a
chTNT-
3/muCD137L
30
NA
80
80
0
a
Fc-muCD137L
30
NA
85
70
Fc-muGITRL
90
NA
45
100
90
LEC/chTNT-3
30
100
<
5
100
55
a
Tumors grew faster than nontreated controls.
B7.1-Fc immunotherapy is dependent on the induction of
IFN-
g
, activated CD8
T cells, and partially by the use of
perforin for tumor cell killing. When used in combination
with T
reg
cell depletion, memory cells are generated to
assure the complete destruction of tumor in the host.
In order to assess the role of CD4
þ
T cells in
CD137L mediated immunotherapy, T-subset depletion stud-
ies were performed with cytotoxic antibodies against CD4
þ
and CD8
þ
þ
and CD8
cells. Anti-CD4 antibody (Clone GK1.5) and
anti-CD8 antibody (Clone H35) were injected i.p. on the 5th
day after tumor implantation. Importantly, CD8
þ
T cell
depletion completely abrogated the antitumor effects of
TNT-3/CD137L (Figure 19.1) and Fc-CD137L (Table 19.5),
indicating that tumor suppression was dependent on this
subpopulation of T cells [68]. Depletion studies were also
performed in mice treated with Fc-GITRL. Groups that
received soluble Fc-GITRL or DTA-1 (agonist rat anti-
murine GITR) treatment alone resulted in a 90% reduction
in tumor volume at day 19 posttumor implantation compared
to the control LFA-Fc group, respectively (Table 19.5).
The anti-CD4
þ
alone treatment group resulted in a 96%
decrease in tumor volume compared with LFA-Fc, while the
anti-CD8
þ
alone treatment group resulted in no significant
difference in tumor volume compared to control LFA-Fc
treated mice. The group treated with anti-CD4
þ
in combi-
nation with Fc-GITRL resulted in the same percent tumor
reduction as the group treated with anti-CD4
þ
alone. In
contrast, the group that received Fc-GITRL in combination
with anti-CD8
þ
resulted in an 89% increase in tumor
volume compared to Fc-GITRL treatment alone. These
results demonstrate that CD8
þ
T cells are critical for tumor
destruction by Fc-GITRL as with the other fusion proteins
described above. Since the Fc-GITRL was by itself so
potent, it is difficult to show the contribution of anti-T
reg
deletion in these mice. Additionally, Fc-GITRL may by
itself suppress T
reg
function thereby contributing to its out-
standing therapeutic effects in vivo.
þ
19.7 CONCLUSION
In conclusion, in the last 10 years, we have generated a panel
of cytokine, chemokine, and co-stimulatory fusion proteins
[110] and tested them similarly in multiple solid tumor
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