Activatable Nanoreporters for Real-Time Tracking of Macrophage Phenotypic States Associated with Disease Progression.

Diagnosis of inflammatory diseases is characterized by identifying symptoms, biomarkers, and imaging. However, conventional techniques lack the sensitivities and specificities to detect disease early. Here, it is demonstrated that the detection of macrophage phenotypes, from inflammatory M1 to alternatively activated M2 macrophages, corresponding to the disease state can be used to predict the prognosis of various diseases. Activatable nanoreporters that can longitudinally detect the presence of the enzyme Arginase 1, a hallmark of M2 macrophages, and nitric oxide, a hallmark of M1 macrophages are engineered, in real-time. Specifically, an M2 nanoreporter enables the early imaging of the progression of breast cancer as predicted by selectively detecting M2 macrophages in tumors. The M1 nanoreporter enables real-time imaging of the subcutaneous inflammatory response that rises from a local lipopolysccharide (LPS) administration. Finally, the M1-M2 dual nanoreporter is evaluated in a muscle injury model, where an initial inflammatory response is monitored by imaging M1 macrophages at the site of inflammation, followed by a resolution phase monitored by the imaging of infiltrated M2 macrophages involved in matrix regeneration and wound healing. It is anticipated that this set of macrophage nanoreporters may be utilized for early diagnosis and longitudinal monitoring of inflammatory responses in various disease models.

Supramolecular nanotherapeutics enable metabolic reprogramming of tumor-associated macrophages to inhibit tumor growth.

Tumor-associated macrophages (TAMs) exist in multiple phenotypes across the spectrum, defined by an M1 antitumorigenic phenotype and an M2 pro-tumorigenic phenotype on two ends of the spectrum. A largely immunosuppressive tumor-microenvironment aids the polarization of the infiltrating macrophages to a pro-tumorigenic M2 phenotype that promotes tumor progression and metastasis. Recent developments in macrophage immunotherapy have focused on strategies to re-educate TAMs from an M2 to M1 phenotype. Recent findings in the realm of immuno-metabolism have indicated that distinct metabolic signatures accompany macrophages based on their polarization states (M1-Glycolysis and M2-TCA cycle). These metabolites are important drivers of cellular signaling responsible for acquiring these polarization states, with evidence showing that metabolism is essential to facilitate the energy requirements of immune cells and regulate immune cell response. We hypothesized that TAMs could be reprogrammed metabolically by co-delivery of drugs using a supramolecular nanoparticle system that could effectively rewire macrophage metabolism by simultaneous inhibition of the TCA cycle and upregulation of the glycolytic metabolic pathway. TLR7/8 agonist and Fatty Acid Oxidation (FAO) inhibitor loaded metabolic supramolecular nanoparticles (MSNPs) were synthesized. In vitro assays showed macrophages treated with MSNPs were reprogrammed from an M2 phenotype to an M1 phenotype while significantly upregulating phagocytosis. When injected in 4T1 tumor-bearing mice, MSNPs treatment reduced tumor growth progression more than other treatments. Hence, the delivery of TLR7/8 agonist combined with an FAO inhibitor can enhance antitumor efficacy through metabolic reprogramming of tumor-associated macrophages.

Rational combination of an immune checkpoint inhibitor with CSF1R inhibitor-loaded nanoparticle enhances anticancer efficacy.

Since the advent of immune checkpoint inhibitors, rapid strides have been made in the realm of cancer immunotherapy. Of the abundance of infiltrating immune cells in the tumor microenvironment (TME), macrophages contribute a significant portion and make up to 50% of the tumor mass. In addition to this, the relative plasticity of macrophages makes it an attractive target to modulate macrophage functions to initiate an anti-tumor response. However, many challenges hinder this strategy. Macrophage colony-stimulating factor (MCSF) secreted by cancer cells binds to the colony-stimulating factor receptor present on macrophages and negatively influences macrophage functions. MCSF, along with a cocktail of immunosuppressive cytokines present in the TME, polarizes macrophages to an immunosuppressive pro-tumorigenic M2-like phenotype. M2-like macrophages dampen tumor response and are known to be associated with increased tumor progression and metastasis. Indeed, clinical interventions aimed to reprogram macrophage response from an M2-like tumor aiding phenotype to an M1-like tumor-killing phenotype using small-molecule inhibitors of the CSF1R axis have gathered much attention in the recent past. However, poor response and systemic toxicities observed in these therapies necessitate alternative therapeutic strategies. Furthermore, another key signaling pathway that has been recently implicated in aiding the CSF1R signaling in TAMs is the PDL1 signaling axis. Hence, in this study, we designed a self-assembled lipid nanoparticle system encompassing a potent small-molecule inhibitor of the CSF1R signaling axis, while the surface of the nanoparticle was tethered with anti-PDL1 mAb. The purpose of this is twofold; the nanoparticles can deliver the cargo in a targeted manner to PDL1 expressing M2-like macrophages while simultaneously blocking the receptor. The resulting nanoparticle system termed α-PDL1-CSF-LNP showed enhanced repolarization of M2 like macrophages in vitro while also upregulating the phagocytic index. Furthermore, suboptimal dose administration of α-PDL1-CSF-LNP in an aggressive melanoma mouse model resulted in superior anti-tumor efficacy with minimal toxicities. These results were validated by ex vivo mechanistic analysis showing that TAMs have successfully been repolarized to a predominantly M1-like phenotype. This, along with increased tumor infiltration of CD8+ T cells, worked in synergy to provide an effective anti-tumor strategy.