Effector function of NK cells disrupted by lipid-rich environment: implications in tumor defense

It is estimated that in 2018 more than 18 million new cases of cancer will be diagnosed worldwide and an estimated 9.5 million individuals will die as a result of cancer (Bray et al. 2018). While there are many biological and environmental risk factors associated with cancer, increasing studies have placed an emphasis on the link between obesity and certain types of cancers (O’Rourke, 2016; Colditz and Peterson, 2018). A research by Renehan et al. (2008) showed that increased BMI was associated with an increased risk of developing cancers. In fact, 49% of cancer types were attributed to obesity (Renehan et al. 2008). Some of the mechanistic links between cancer and obesity include the overproduction of hormones and insulin which favors tumor growth and proliferation (Renehan et al. 2015). Other theories behind obesity and increased risk of cancer development involved the role of inflammation and the immune system and the production of cytokines (Divella et al. 2016; Deng et al. 2016; Renehan et al. 2015). Even though there are increasing publications on this front of cancer research, the mechanisms behind inflammation and the immune system on obesity-induced dysfunction and cancer risk is not fully understood.

A recently published article by Michelet et al. (2018) in Nature Immunology gave insights into the role of natural killer (NK) cells in fighting against tumor cells under normal (lean) and obese (high fat) conditions. The focus was placed on NK cells because of their important role in suppressing tumor development via the secretion of cytotoxic elements - lytic granules and perforin - to kill tumor cells (Mace et al. 2014). Additionally, NK cells have demonstrated metabolic reprogramming in order to meet the high energy demands required to carry out its effector function against tumor cells (Donnelly et al. 2014). The researchers therefore investigated whether a high fat diet interferes with NK cells’ metabolism to inhibit their effector function to target and destroy tumor cells. They investigated the function of NK cells in a mouse model of obesity and in NK cells isolated form obese and lean human subjects. In summary, their research found that in both mice and human, NK cells uptake lipid form their environment which disrupts their metabolism. This disruption lead to loss in tumor suppression ability by blocking the trafficking of cytotoxic elements to the synapse of NK and tumor cells.

Michelet et al. first investigate the function of NK cells in mice placed on a high fat diet over an 8 week period. They found that over 3000 genes, most of which were involved in lipid metabolism, were upregulated and a downregulation of genes that encode granzymes. The NK-mediated cytotoxicity pathway was the most downregulated pathway in obese mice. For comparison, they isolated NK cells from obese and lean humans and noticed that there was a decrease in the abundance of NK cells from obese individuals. NK cells from obese individuals also produced lower amounts of IFN-γ and killed fewer tumor cells. Similar to NK cells from obese mice, NK from obese human had higher intracellular lipid accumulation and lower expression of granzymes and perforin, suggesting that obesity limit effector function of NK cells in humans as well. Importantly, they demonstrated that the loss of effector function of NK cells were not due to increased amounts of glucose and insulin. The loss of effector function was only observed when NK cells from lean individuals were cultured in lipid.

Lean NK cells stained positive for lipids only
in the cell membrane, while NK cells from
obese individuals were positive for lipid in
the cytoplasm

In NK cells that were treated with oleate (lipid), there was in
increase accumulation of lipid within the cell and a decrease in
perforin 

To determine the mechanism behind effector functional loss of NK cells, they focused on the mTORC1 pathway because it has been demonstrated to be important for the expression of granzyme and production of IFN-γ (Donnelly et al. 2014). They found reduced activation of the mTOR pathway after NK cells were treated with lipid. Other downstream pathway activation of mTORC1 were significantly reduced when NK cells from lean individuals were pre-treated with lipid. This resulted in a reduction in NK effector function when stimulated with cytokines or when they encountered tumor cells.

mTOR is activated when phosphorylated. Lipid
treatment resulted in a decrease of phosphorylated mTOR (p-mTOR) 

Looking into the metabolism of lean NK cells pre-treated with lipid, the group found that the cells took up less glucose compared to the controls. As a result, lipid pre-treated NK cells displayed impairment in both glycolysis and oxidative phosphorylation, and the ultimate outcome was a significantly lower production of ATP. Similar observations were made from NK cells isolated from obese humans and NK cells from obese mice. The next question that they investigated was how does the disruption in metabolic pathway leads to the loss of cytotoxic function. To answer this question, they investigated different stages in NK cells targeting and killing of tumor cells. Although NK cells were able to form immunological synapse with tumor cells, cytotoxic elements such as lytic granules and perforin were not being trafficked to the synapse. The disruption in trafficking of cytotoxic machinery was due to the inability of microtubule-organizing centers to polarize to immunological synapse. Furthermore, when the metabolic deficits were reversed, NK cytotoxicity was restored.

Reversing metabolic deficits with etomoxir (right panel) allows
cytotoxic elements (green) to traffic to synapse similar to the untreated panel
on the left. When NK was treated with lipid (middle panel), cytotoxic
elements did not travel to synapse

To complete their model, they looked at how these functional deficits in obese NK contribute to tumor growth. Using a tumor bearing mouse model, they showed that mice transfected with stimulated control NK cells reduced the tumor burden of mice while mice transfected with lipid pre-treated NK cells had no effect on tumor burden. The lipid-treated NK cells were characterized by lower IFN-γ production and granzyme expression. To further investigate this pathway, they fed wild type mice a high fat diet and inject them with tumor cells. As expected, tumor growth was more aggressive in obese mice; NK cells were also significantly reduced in tumors of obese mice and produced less IFN-γ.

Mice that were obese (red) had a greater tumor burden and lower levels of
circulating NK cells 

The research conducted by Michelet et al. provided a mechanism by which NK cells in obese individuals lose their tumor suppressive ability by taking up lipids from their environment. The accumulation of lipids leads to disruption in metabolism and inhibition of cytotoxic proteins trafficking to the immunological synapse to facilitate killing of tumor cells. Targeting this metabolic pathway can provide a therapeutic approach to cancer treatment in obese individuals. However, this research also opens the possibility to investigate how other possible metabolic disorders may affect the effector function of NK cells towards tumor development. Further research can be done on ways to improve the function of NK cells under high lipid conditions which can also be a potential therapy. For instance, they demonstrate that blocking peroxisome proliferator-activated receptor (PPAR) reduced the accumulation of lipids in NK cells and prevented the loss of perforin. Further research on this pathway and similar pathways that can prevent lipid accumulation in NK cells would be beneficial in this field of study.

References:

Bray, Freddie, et al. "Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries." CA: a cancer journal for clinicians (2018).

Colditz, Graham A., and Lindsay L. Peterson. "Obesity and cancer: evidence, impact, and future directions." Clinical chemistry 64.1 (2018): 154-162.

Donnelly, Raymond P., et al. "mTORC1-dependent metabolic reprogramming is a prerequisite for NK cell effector function." The Journal of Immunology (2014): 1401558.

Mace, Emily M., et al. "Cell biological steps and checkpoints in accessing NK cell cytotoxicity." Immunology and cell biology92.3 (2014): 245-255.

Michelet, Xavier, et al. "Metabolic reprogramming of natural killer cells in obesity limits antitumor responses." Nature Immunology (2018): 1.

O’Rourke, Robert W. "Obesity and cancer." Metabolic Syndrome and Diabetes. Springer, New York, NY, 2016. 111-123.

Renehan, Andrew G., et al. "Body-mass index and incidence of cancer: a systematic review and meta-analysis of prospective observational studies." The Lancet 371.9612 (2008): 569-578.

Renehan, Andrew G., Marcel Zwahlen, and Matthias Egger. "Adiposity and cancer risk: new mechanistic insights from epidemiology." Nature Reviews Cancer 15.8 (2015): 484.