By Jamshed Arslan, Pharm. D., PhD.
Colon cancer is responsible for over 600,000 deaths per year worldwide. Colon cancer can be classified into two categories: mismatch repair (MMR)-deficient and MMR-proficient cancers.1 MMR-deficient colon cancer shows high CD8+ cytotoxic T-lymphocyte (CTL) and low Th17 cells, while MMR-proficient colon cancer shows an opposite trend. MMR-proficient colon cancer is associated with poor prognosis and is resistant to anti-cancerous immune checkpoint inhibitors. In other words, outcome of immune therapy is dependent on the balance between Th17 and CTLs. In this regard, research institutes in Louisville and Albany, USA, developed a novel oral IL-10 formulation that could suppress colon cancer, possibly by acting on both Th17 and CTLs.2
To test their hypothesis, the scientists treated colon cancer model mice with IL-10. They also treated animals with a similar oral formulation of IL-12, which is a Th1 cytokine that can activate tumor-related CTLs. They expected a synergistic increase in CTLs when IL-10 and IL-12 were combined. As expected, combination therapy virtually eradicated the disease, more so than IL-10 alone, by enhancing the level of colon’s tight junction proteins. They noticed that IL-12 monotherapy exacerbated the disease by compromising gut barrier function. In sum, therapeutic outcome of immune therapy against colon cancer depends on gut barrier integrity.
Combination therapy (IL-10 with IL-12) reduces colon cancer by acting on T cell subsets
The team administered oral cytokines (IL-10 and/or IL-12) to the colon cancer mice and examined the tumors. Three weeks of IL-10 monotherapy reduced the tumor burden without affecting tumor histopathology. IL-12 monotherapy, on the other hand, increased the tumor burden. The combination of IL-10 and IL-12 eliminated tumors, improved colon histology, and enhanced animal survival.
Human peripheral blood mononuclear cells (PBMCs) either (A) unstimulated or (B) stimulated to induce Th17 cells were stained with Rabbit Anti-Human ROR gamma /RORC/NR1F3 Monoclonal Antibody (Catalog # MAB61093) followed by Phycoerythrin-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # F0110) and Mouse Anti-Human CD4 APC‑conjugated Monoclonal Antibody (Catalog # FAB3791A). Quadrant markers were set based on control antibody staining (Catalog # MAB1050). To facilitate intracellular staining, cells were fixed and permeabilized with FlowX FoxP3 Fixation & Permeabilization Buffer Kit (Catalog # FC012).
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To explore the cellular mechanism behind this anti-tumor synergy, the T cell subsets isolated from mesenteric lymph nodes were analyzed by flow cytometry. IL-10 monotherapy decreased Th17 cells positive for CD4, ROR gamma t and IL-17 without affecting CD8+ CTLs. IL-12 monotherapy distinctly enhanced CD8+ CTLs. Combination therapy reduced Th17 cells and enhanced CD8+ CTL activity similar to what is found in MMR-deficient colon cancer. In other words, IL-12 is beneficial against colorectal cancer only when given in combination with IL-10.
The next step was to explore if the paradoxical role of IL-12 was dependent on its downstream effector IFN-gamma.
Gut barrier integrity and IFN-gamma are required for successful immunotherapy against colon cancer
Treating tumor-bearing mice with anti-IFN-gamma antibody abrogated all effects of IL-12, indicating that both detrimental and advantageous effects of IL-12 depend on IFN-gamma. To see if gut integrity had any role in this phenomenon, mice were given oral FITC-labeled dextran and fluorescence was analyzed in their blood. IL-12 was found to enhance gut permeability, but neutralizing IFN-gamma abrogated this effect. Giving IL-10 in addition to IL-12 restored the steady state levels of FITC-labeled dextran.2 In light of a previous study,3 researchers hypothesized that this restoration of gut integrity could be due to IFN-gamma-induced expression of IL-10 receptor called IL-10 R alpha.2 Confocal microscopy revealed a robust IL-10 R alpha expression in the colon of mice receiving IL-12 or IL-12 with IL-10. This means that IL-12 enhances the responsiveness of gut epithelium to IL-10, thereby restoring the barrier integrity.
In sum, combined immunotherapy with IL-10 and IL-12 enhances tight junction integrity to treat colon cancer because IFN-gamma (generated by IL-12) sensitizes the gut epithelium to IL-10.
This study2 describes a unique synergy: IL-10 is anti-inflammatory while IL-12 is a pro-inflammatory cytokine, and when combined, they can ameliorate colon cancer. The paper proposes oral formulations of cytokines to change colon physiology, which can ensure patient compliance.
Jamshed Arslan, Pharm D., PhD.
Previously at the University of Alabama at Birmingham, School of Medicine.
Dr. Arslan studies cell signaling in mitochondrial defects in
C. elegans and transgenic mice
- Wirta, Erkki-Ville, et al. "Immunoscore in Mismatch Repair‐Proficient and ‐Deficient Colon Cancer." Journal of Pathology: Clinical Research, vol. 3, no. 3, 2017, pp. 203–213. https://doi.org/10.1002/cjp2.71
- Bhutiani, Neal, et al. "Enhanced Gut Barrier Integrity Sensitizes Colon Cancer to Immune Therapy." Oncoimmunology, vol. 7, no. 11, 2018, n.pag. https://doi.org/10.1080/2162402X.2018.1498438
- Kominsky, Douglas J., et al. "IFN-γ–Mediated Induction of an Apical IL-10 Receptor on Polarized Intestinal Epithelia." Journal of Immunology, vol. 192, no. 3, 2014, pp. 1267–1276. https://doi.org/10.4049/jimmunol.1301757