Cancer gene therapy of adenovirus-mediated anti-4-1BB scFv in immunocompetent mice

Abstract

The use of immunostimulatory molecule genes aiming at enhancing anti-tumor immunity has emerged as a new approach to treat cancers. 4-1BB signaling, an important costimulatory pathway delivering a signal for T cell activation, survival and growth, has become one of the most promising targets for cancer immunotherapy. In this work, a recombinant nonreplicative adenovirus (Ad.4-1BB scFv) carrying a single-chain Fv fragments (scFv) specific for 4-1BB gene (anti-4-1BB scFv) was generated, haracterized and explored for its stimulation of anti-tumor immunity in immunocompetent C57BL/6 mice. Ad.4-1BB scFv could efficiently infect murine hepatoma Hepa 1-6 cells and induce anti-4-1BB scFv expression on the cell surface. Moreover, Ad.4-1BB scFv did not cause obvious cytotoxicity effect on human and murine tumor cell lines (A549, PLC/PRF/5, Hepa 1-6 and TC-1) even at a high MOI, which suggested Ad.4-1BB scFv had no direct effect on tumor cells. Intratumoral injection of Ad.4-1BB scFv to established Hepa 1-6 tumors significantly suppressed the tumor growth in C57BL/6 mice. The anti-tumor effect might be mainly attributed to the anti-4-1BB scFv-mediated immune activity, as evidenced by enhanced interferon-gamma-producing splenic cells and increased lymphocytes infiltration in the tumor microenvironment. These results indicated that nonreplicative adenovirus carrying the anti-4-1BB scFv gene possessed powerful in vivo anti-tumor efficacy and might be a valuable tool for cancer immunotherapy.

Therapeutic potential of anti-CD137 (4-1BB) monoclonal antibodies

“…These anti-tumor effects of anti-4-1BB ScFv were believed to result from low-dose cyclophosphamide-mediated depletion of Tregs [101]. Similar results were obtained in studies carried out by Liu et al [102]. These authors observed that intratumoral injection of anti-4-1BB ScFV inhibited the growth of Hepa 1 --6 tumors, and this protective effect was shown to depend on increased IFN-g and increased tumor infiltration by T cells [102].…”

Potent inhibition of OKT3-induced T cell proliferation and suppression of CD147 cell surface expression in HeLa cells by scFv-M6-1B9

“…The reasoning behind our scFv format was to preserve the genetic source of the antibody fragment by cloning it into a phagemid vector and utilizing relatively simple production in bacteria. Furthermore, the small molecular size of scFv should provide high tissue penetration efficiency (Kioi et al, 2008;Liu et al, 2008;Stirnemann et al, 2008), and a lack of significant toxicity (Korn et al, 2004;Yang et al, 2007a;Nam et al, 2008) while giving rapid plasma clearance (Mao et al, 1999;Hamilton et al, 2002;Olafsen et al, 2004;Pavoni et al, 2006), which makes such an antibody a potentially suitable candidate for clinical application.…”

Anti-4-1BB scFv immunogene therapy and low dose cyclophosphamide exhibit a synergistic antitumor effect in established murine lung tumors

“…37 In our previous work, adenoviruses carrying the anti-4-1BB scFv gene (Ad.4-1BB scFv) were used as an immunotherapy method to treat Hepa 1-6 tumors in C57BL/6 mice. 17 Although delay of tumor growth was observed, the antitumor efficacy was still not satisfactory. tumors after Ad.4-1BB scFv or Ad.4-1BB scFv plus CTX treatment (Fig.…”

“…20 reported previously. 17 The anti-4-1BB scFv was the variable region from the anti-4-1BB hybridoma 1D8. For cell surface expression of scFv and anti-4-1BB scFv detection, the transmembrane domain and cytoplasmic tail from CD80 (hCD80 Tm and cytoplasmic domain) were used and a human immunoglobulin Fc fragment (hIgG Fc) following the scFv was added.…”

“…15,16 Our previous study also showed that intratumoral injection of Ad.4-1BB scFv, an adenovirus carrying a single-chain Fv fragments (scFv) specific for 4-1BB gene (anti-4-1BB scFv), delayed the established Hepa 1-6 tumor growth in C57BL/6 mice. 17 However, administration of agonistic 4-1BB scFv does not always induce meaningful tumor regression in some tumor models. 18 This may be because immune response to malignant tumors is often weak and ineffective, and tumors evolve multiple mechanisms to escape immune surveillance including induction of T-cell tolerance.…”