TLR Agonists for Cancer Immunotherapy: Tipping the Balance between the Immune Stimulatory and Inhibitory Effects

The agonists of toll-like receptors (TLRs) have been actively pursued for their antitumor potentials, either as monotherapy or as adjuvants to vaccination or other therapeutic modalities (1). A search on ClinicalTrials.gov using the key words “TLR” and “cancer” returned 34 listings. The idea of using TLR agonist to provide a “danger signal” and break tolerance to tumor antigens has been well embraced by tumor immunologists. However, the promise of TLR agonists-based immunotherapy remains to be realized in the clinic, and only very few TLR agonists have been approved by the FDA. For example, bacillus Calmette–Guerin (BCG) and imiquimod have been approved as standalone therapies, whereas monophosphoryl lipid A (MPL) was approved as a vaccine component. A review of recently published literature on the use of TLR agonists in cancer setting revealed a common mechanism that might have explained the underperformance of TLR agonists as cancer therapeutics: induction of immune suppressive factors that put a break on the TLR agonists-induced inflammation. As shown in Figure 1, TLR agonists have immune stimulatory effects through the induction of costimulatory molecules (CD80, CD86, and CD40) on dendritic cells (DCs) and inflammatory cytokines (TNF-α and IL12) that polarize Th1 immune response. On the other hand, TLR agonists have immune inhibitory effects as evidenced by the induction of several immune suppressive factors, including IL-10, T regulatory cells (Treg), and PD-L1, all of which could dampen anti-tumor immunity. The following is a brief summary on TLR agonistsinduced self-regulatory feedback and the indication for cancer immunotherapy. INDUCTION OF IL-10 IL-10 is an immune suppressive cytokine that inhibits the activity of Th1 cells, thus impeding viral clearance and anti-tumor Th1 immunity. IL-10 could be secreted by different immune cells, including Treg (2), CD4 T cells (3, 4), monocytes, and macrophages (5). The induction of IL-10 by TLR agonists has been demonstrated in infectious disease setting as well as tumor setting. For example, TLR4 signaling with LPS was shown to activate innate IL-10 production in response to Bordetella pertussis, which both directly, and by promoting the induction of IL-10-secreting type 1 regulatory T cells (Tr1), inhibit Th1 responses and limit inflammatory pathology in the lungs during infection with B. pertussis (6). TLR2 ligation and induction of IL-10 were also shown to suppress immunity against Candida albicans (7). Induction of IL-10 also caused the persistence of lymphocytic choriomeningitis virus (LCMV), and IL-10 blockade using a neutralizing antibody can restore T cell immunity and lead to viral clearance (3). IL-10 induction by TLR agonists has been observed in mouse models of breast cancer and melanoma. In MMTV/neu-transgenic mice, a model of human HER2 breast cancer, topical treatment with TLR7 agonist imiquimod induced IL-10, and the major source of IL-10 was Tr1 cells (4). IL-10 induction was also observed in a mouse model of implanted TSA breast cancer, where topical imiquimod was shown to synergize with radiation and low-dose cyclophosphamide in inhibiting tumor growth (8). In a mouse model of B16 melanoma, the induction of IL-10 has also been shown to limit the anti-tumor effects of TLR2 agonist Pam2 lipopeptide (9). Altogether these publications suggest IL-10 induction is probably a common regulatory mechanism that dampens TLR agonists-induced antitumor immunity. IL-10 blockade using anti-IL-10 neutralizing mAb significantly enhanced the anti-tumor effects of topical imiquimod (4). The addition of antiIL-10R to TLR9 agonist CpG also exhibited robust anti-tumor activity exceeding by far that of CpG alone, and elicited antitumor immune memory (10). Thus IL-10 blockade holds promise to augment the anti-tumor effects of TLR agonists.