In Situ Vaccination with a TLR9 Agonist and Anti-OX40 Antibody Leads to Tumor Regression and Induces Abscopal Responses in Murine Lymphoma

Background:

Toll-like receptors (TLRs) are components of the innate immune system that recognize pathogen-associated molecular patterns on bacterial, fungal, or viral pathogens. Intratumoral (IT) injection of unmethylated CG-enriched oligodeoxynucleotide (CpG), a TLR9 agonist, results in local tumor eradication but on its own is not able to induce a systemic anti-tumor immune response. OX40 is a potent costimulatory receptor that can potentiate the action of conventional T cells leading to their proliferation, effector function and survival, but can also inhibit or kill T regulatory cells by ADCC. In previous preclinical studies systemic injection of OX40 agonists increased antitumor immunity and improved survival.

Scientific question:

Does local injection of both CpG and low doses of an anti-OX40 agonistic antibody trigger a systemic anti-tumor immune response?

Results:

We implanted the A20 lymphoma tumor bilaterally on opposite sides of the abdomen. After tumors were established we administered microgram quantities of CpG and anti-OX40 antibody into the tumor on one side and monitored both the injected and the uninjected tumor sites. This treatment resulted in both a local and an abscopal effect on the contralateral, untreated tumor. In addition, the animals were protected from a second challenge with A20 cells. This anti-tumor effect was T cell dependent, since depletion of either CD4+ or CD8+ T cells abrogated the therapeutic effect. There was no evidence of toxicity or autoimmunity in the treated animals.

To examine the potential of this maneuver to treat spontaneous, non-transplanted cancers we chose the mouse mammary tumor model- FVB/N-Tg(MMTV-PyVT)634Mul/J. These animals all develop invasive breast cancer tumors in all of their mammary glands by 12 weeks of age. Injections of CpG and anti-OX40 antibody into the first arising tumor not only prevented its growth but significantly reduced the incidence and outgrowth of subsequent tumors at un-injected susceptible mammary glands and reduced the number of lung metastases.

Significance:

TLR9 agonists and anti-OX40 antibodies are currently under clinical development for cancer treatment. We show here that combining anti-OX40 antibody with a TLR9 agonist at a single established tumor is sufficient to trigger a systemic anti-tumor response able to eradicate tumor at distant sites in both transplantable and spontaneously occurring oncogene-driven murine tumors. This anti-tumor effect was long lasting, specific and required T cells.

Impact:

Our current results suggest that CpG and anti-OX40 are sufficient to induce fully protective and curative anti-tumor immune responses, even in spontaneously arising cancer. Anti-OX40 and CpG are both currently in phase-I clinical trials as single agents. Our results provide the rationale for testing these agents clinically in combination as described here, injected locally in low doses in order to induce therapeutic anti-lymphoma immunity.