Abstract
Tumors develop when infiltrating immune cells contribute growth stimuli, and cancer cells are selected to survive within such a cytotoxic microenvironment. One possible immune-escape mechanism is the upregulation of PI-9 (Serpin B9) within cancer cells. This serine proteinase inhibitor selectively inactivates apoptosis-inducing granzyme B (GrB) from cytotoxic granules of innate immune cells. We demonstrate that most classical Hodgkin lymphoma (cHL)-derived cell lines express PI-9, which protects them against the GrB attack and thereby renders them resistant against GrB-based immunotherapeutics. To circumvent this disadvantage, we developed PI-9-insensitive human GrB mutants as fusion proteins to target the Hodgkin-selective receptor CD30. In contrast to the wild-type GrB, a R201K point-mutated GrB construct most efficiently killed PI-9-positive and -negative cHL cells. This was tested in vitro and also in vivo whereby a novel optical imaging-based tumor model with HL cell line L428 was applied. Therefore, this variant, as part of the next generation immunotherapeutics, also named cytolytic fusion proteins showing reduced immunogenicity, is a promising molecule for (targeted) therapy of patients with relapsing malignancies, such as cHL, and possibly other PI-9-positive malignancies, such as breast or lung carcinoma.
Highlights
A major problem of conventional cancer therapy is its lack of specificity
The best studied and the most promising one is granzyme B (GrB),2–8 an effector molecule stored in the granules of cytotoxic immune cells to kill transformed tumor or virusinfected cells
We recently generated a variety of GrB mutants, which showed a reduced proteinase inhibitor-9 (PI-9)-inhibitor binding in the first experimental studies
Summary
A major problem of conventional cancer therapy is its lack of specificity. more specific approaches are under investigation to exclusively kill abnormal cells. As plant- or bacteria-derived toxins can cause severe side effects, the generation of less immunogenic compounds is essential This led to the fourth generation of immunotoxins, first so coined by Mathew and Verma, which include humanization of both the cellbinding ligand as well as the pro-apoptotic enzyme. The best studied and the most promising one is granzyme B (GrB), an effector molecule stored in the granules of cytotoxic immune cells to kill transformed tumor or virusinfected cells.. As a very high number of positive tumor cells predict unfavorable clinical outcome, these might be positively selected upon immune surveillance.. As a very high number of positive tumor cells predict unfavorable clinical outcome, these might be positively selected upon immune surveillance.12,16 Killing these treatment-resistant cells may strongly increase the therapeutic success and possibly lifespan. The R201K GrB mutant demonstrated the best antitumor efficacy against both PI-9-positive and -negative tumor cells
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