Resolvin E 1 br It may also be possible to repolarize TAMs u
It may also be possible to repolarize TAMs using drug-free inorganic NPs. High levels of intracellular iron can modulate cytokine signaling and cell proliferation by increasing oxidative stress, particularly in the oxidative environment of the TME. Ferumoxytol is a superparamagnetic iron-oxide NP (SPION) approved for the treatment of chronic iron deficiency anemia. These particles are readily cleared by TAMs in the TME without specific targeting moieties. In vitro, they have no effect on macrophage proliferation or survival108. However, they demonstrate clear polarization towards an M1 phenotype (as evidence by increased expression of TNF-α and CD86) and away from the M2 phenotype (decreased expression of CD206 and IL10). In vivo, ferumoxytol delayed local progression and significantly decreased metastatic spread in MMTV-PYMT mouse models of breast cancer.
In addition to altering the polarization of TAMs, targeted NPs have also been used to deplete M2 macrophages in the TME (Figure 4). In one study, mannose-conjugated lipid-coated NPs encapsulating calcium zolendronate (a bisphosphonate that demonstrates selective cytotoxicity to TAMs109) effectively depleted established S180 tumor xenografts of inhibitory TAMs and markedly enhanced tumor growth delay. TAMs also express high levels of sialic Resolvin E 1 receptors110. Zhou et al. have shown that sialic acid-coated liposomes loaded with epirubicin can enhance selective clearance of M2 TAMs and improve rejection of murine xenograft sarcoma models without significant systemic toxicity111. Collectively, these studies demonstrate that functionalized NPs can be engineered to efficiently repolarize or deplete inhibitory TAMs in the TME to improve responsiveness to cancer immunotherapies including T-cell checkpoint inhibitors.
Th2 CD4+ regulatory T-cells are another potent immunosuppressive cell population commonly found in the TME. These cells can suppress local immune responses through inhibitory cytokine secretion (including TGF-β and IL-4,6,10) or by releasing perforin or granzymes to eliminate activated CD8+ T-cells and APCs within the TME. Several surface receptors are overexpressed on tumoral CD4+ T-regs including glucocorticoid-induced TNF receptor (GITR) and the Nrp1 receptor. Sacchetti et al. generated GITR-conjugated PEG-modified single-walled carbon nanotubes and demonstrated selective and efficient intracellular transport of targeted NPs in tumoral Th2 cells112. Another group more recently generated multi-layer hybrid NPs which included GITR-labeled polymeric cores to selectively deliver imatinib to Th2 T-regs113. The functionalized particles inhibited suppressive cytokine signaling from Th2 cells and improved activation of infiltrating CD8+ T-cells.
Preclinical work from non-oncologic studies has demonstrated that the physical characteristics of NPs can also affect the relative Th1 to Th2 response to immune adjuvants. One study using diphtheria toxins showed that whereas micro and nanosize formulations of aluminum adjuvants stimulate both Th1 and Th2 responses, calcium-phosphate nanoparticles preferentially stimulated Th1 but not Th2 responses. Antigen and adjuvant containing NPs have been used to selectively modulate the relative activity of Th1 and Th2 lymphocytes in numerous non-oncologic disease models including hepatitis B, asthma, and environmental hyperimmunities. Lessons learned from these systems can be used to improve modulation of the TME to a more immunostimulatory phenotype.
TAFs are another potently immunosuppressive population of cells in the TME. TAFs are believed to result from the transformation of mesenchymal fibroblasts by cytokine signaling in tumors. TAFs can suppress tumor-specific immunity in several ways114-116. In some tumor types, including pancreatic cancer, TAFs are also responsible for the formation of dense, largely acellular desmoplasia that acts as a physical barrier to limit infiltration by activated CD8+ T-cells117. Even osmotic perfusion can be limited by a dense desmoplastic stroma. TAFs can also stimulate anergy of APCs and activated CD8+ T-cells by expressing high levels of PDL-1/2. Finally, TAFs express high levels of cytokines that inactivate pro-immunogenic Th1 T-regs and M1 TAMs in addition to polarization towards their immunosuppressive counterparts. Several strategies have been undertaken to limit or reverse the immunosuppressive actions of TAFs in the TME using targeted NPs. They primarily involve either depleting TAFs in the TME or preventing cross-talk between TAFs and other cell populations. Similar to other inhibitory cell populations in the TME, targeted NPs offer a promising approach to selectively modulating TAF-mediated immunoresistance. One hallmark that distinguishes TAFs from untransformed mesenchymal fibroblasts is the overexpression of α-SMA which is also correlated with increased surface expression of sigma receptors. Anisamides, including aminoethylanisamide, are sigma receptor agonists that can be conjugated to the surface of cationic, polymeric, or liposomal NPs for targeted intracellular uptake. TAFs also tend to be enriched around the vasculature of desmoplastic tumors and relatively accessible to NP delivery vectors. Huang and colleagues have developed several preclinical strategies to target TAFs using engineered NPs. In one study, they used anisamide-decorated lipid calcium phosphate NPs to deliver plasmid DNA to TAFs and disrupt tumoral paracrine signaling necessary for the maintenance of the TAF phenotype in a desmoplastic models of bladder (UMUC3) and pancreatic (BXPC3) cancers118. The targeted NPs induced high expression of the TNF-related inducing secretory ligand sTRAIL in TAFs which in turn stimulated apoptosis of adjacent tumor cells. Significant levels of apoptosis were not detected in α-SMA+ TAFs. However, apoptosis of adjacent tumor cells was associated with a reversion of TAFs to a quiescent state and significant reductions in α-SMA and FAPα (markers of fibroblast activation).