Nanoparticles targeting immune checkpoint protein VISTA induce potent antitumor immunity.

BACKGROUND: Immune checkpoint protein V-domain immunoglobulin suppressor of T cell activation (VISTA) controls antitumor immunity and is a valuable target for cancer immunotherapy. Previous mechanistic studies have indicated that VISTA impairs the toll-like receptor (TLR)-mediated activation of myeloid antigen-presenting cells, promoting the expansion of myeloid-derived suppressor cells, and suppressing tumor-reactive cytotoxic T cell function. METHODS: The aim of this study was to develop a dual-action lipid nanoparticle (dual-LNP) coloaded with VISTA-specific siRNA and TLR9 agonist CpG oligonucleotide. We used three murine preclinical tumor models, melanoma YUMM1.7, melanoma B16F10, and colon carcinoma MC38 to assess the functional synergy of the two cargoes of the dual LNP and therapeutic efficacy. RESULTS: The dual-LNP synergistically augmented antitumor immune responses and rejected large established tumors whereas LNPs containing VISTA siRNA or CpG alone were ineffective. In comparison with therapies using the soluble CpG and a VISTA-specific monoclonal antibody, the dual-LNP demonstrated superior therapeutic efficacy yet with reduced systemic inflammatory cytokine production. In three murine models, the dual-LNP treatment achieved a high cure rate. Tumor rejection was associated with influx of immune cells to tumor tissues, augmented dendritic cell activation, production of proinflammatory cytokines, and improved function of cytotoxic T cells. CONCLUSIONS: Our studies show the dual-LNP ensured codelivery of its synergistic cargoes to tumor-infiltrating myeloid cells, leading to simultaneous silencing of VISTA and stimulation of TLR9. As a result, the dual-LNP drove a highly potent antitumor immune response that rejected large aggressive tumors, thus may be a promising therapeutic platform for treating immune-cold tumors.

Can targeted nanoparticles distinguish cancer metastasis from inflammation?

Targeting ligands have been widely used to increase the intratumoral accumulation of nanoparticles and their uptake by cancer cells. However, these ligands aim at targets that are often also upregulated in inflamed tissues. Here, we assessed the ability of targeted nanoparticles to distinguish metastatic cancer from sites of inflammation. Using common targeting ligands and a 60-nm liposome as a representative nanoparticle, we generated three targeted nanoparticle (NP) variants that targeted either fibronectin, folate, or αvß3 integrin, whose deposition was compared against that of standard untargeted NP. Using fluorescently labeled NPs and ex vivo fluorescence imaging of organs, we assessed the deposition of the NPs into the lungs of mice modeling 4 different biological landscapes, including healthy lungs, aggressive metastasis in lungs, dormant/latent metastasis in lungs, and lungs with general pulmonary inflammation. Among the four NP variants, fibronectin-targeting NP and untargeted NP exhibited the highest deposition in lungs harboring aggressive metastases. However, the deposition of all targeted NP variants in lungs with metastasis was similar to the deposition in lungs with inflammation. Only the untargeted NP was able to exhibit higher deposition in metastasis than inflammation. Moreover, flow-cytometry analysis showed all NP variants accumulated predominantly in immune cells rather than cancer cells. For example, the number of NP+ macrophages and dendritic cells was 16-fold greater than NP+ cancer cells in the case of fibronectin-targeting NP. Overall, targeted NPs were unable to distinguish cancer metastasis from general inflammation, which may have clinical implications to the nanoparticle-mediated delivery of cancer drugs.

Intestinal mucosal triacylglycerol accumulation secondary to decreased lipid secretion in obese and high fat fed mice.

The ectopic deposition of fat in liver and muscle during obesity is well established, however surprisingly little is known about the intestine. We used the ob/ob mouse and C57BL6/J mice fed a high fat (HF) diet to examine the effects of obesity and the effects of HF feeding, respectively, on intestinal mucosal triacylglycerol (TG) accumulation. Male C57BL6/J (wild-type, WT) mice were fed low fat (LF; 10% kcal as fat) or HF (45%) diets, and ob/ob mice were fed the LF diet, for 3 weeks. In this time frame, the WT-HF mice did not become obese, enabling independent examination of effects of the HF diet and effects of obesity. Analysis of intestinal lipid extracts from fed and fasted animals demonstrated that the mucosa, like other tissues, accumulates excess lipid. In the fed state, mucosal triacylglycerol (TG) levels were threefold and fivefold higher in the WT-HF and ob/ob mice, respectively, relative to the WT-LF mice. In the fasted state, mucosa from ob/ob mice had threefold higher TG levels relative to WT-LF mucosa. q-PCR analysis of mucosal mRNA from fed state mice showed alterations in the expression of several genes related to both anabolic and catabolic lipid metabolism pathways in WT-HF and ob/ob mice relative to WT-LF controls. Fewer changes were found in mucosal samples from the fasted state animals. Remarkably, oral fat tolerance tests showed a striking reduction in the plasma appearance of an oral fat load in the ob/ob and WT-HF mice compared to WT-LF. Overall, the results demonstrate that the intestinal mucosa accumulates excess TG during obesity. Changes in the expression of lipid metabolic and transport genes, as well as reduced secretion of dietary lipid from the mucosal cells into the circulation, may contribute to the TG accumulation in intestinal mucosa during obesity. Moreover, even in the absence of frank obesity, HF feeding leads to a large decrease in the rate of intestinal lipid secretion.