In vivo evaluation of scaffolds compatible for colonoid engraftments onto injured mouse colon epithelium.

Colon organoids (colonoids) are known to be similar to colon tissue in structure and function, which makes them useful in the treatment of intestinal de-epithelialized disease. Matrigel, which is used as a transplantation scaffold for colonoids, cannot be used in clinical applications because of its undefined composition and tumorigenicity. This study identifies clinically available scaffolds that are effective for colonoid transplantation in damaged intestinal mucosa. The colon crypt was isolated and cultured from C57BL/6-Tg[CAG enhanced green fluorescent protein (EGFP)131Osb/LeySopJ mice into EGFP + colonoids and subsequently transplanted into the EDTA colitis mouse model using gelatin, collagen, or fibrin glue scaffolds. To identify scaffolds suitable for colonoid engraftment in injured colon mucosa, the success rates of transplantation and secondary EGFP colonoid formation were measured, and the scaffolds' mediated toxicity in vitro and in vivo was observed in recipient mice. When colonoids were transplanted with gelatin, collagen, and fibrin glue into the EDTA colitis mouse model, all groups were found to be successfully engrafted. Fibrin glue, especially, showed significant increase in the engrafted area compared with Matrigel after 4 wk. The scaffolds used in the study did not induce colonic toxicity after transplantation into the recipients' colons and were thus deemed safe when locally administrated. This study suggests new methods for and provides evidence of the safety and utility of the clinical application of colonoid-based therapeutics. Furthermore, the methods introduced in this study will be helpful in developing cell treatment using the esophagus or a stomach organoid for various digestive-system diseases.-Jee, J., Jeong, S. Y., Kim, H. K., Choi, S. Y., Jeong, S., Lee, J., Ko, J. S., Kim, M. S., Kwon, M.-S., Yoo, J. In vivo evaluation of scaffolds compatible for colonoid engraftments onto injured mouse colon epithelium.

Celastrol, which targets IL-2/CD25 binding inhibition, induces T cell-mediated antitumor activity in melanoma.

Interleukin-2 (IL-2) induces contrasting immune responses depending on its binding receptor subunit; thus, selective receptor binding is considered a key challenge in cancer therapeutic strategies. In this study, we aimed to investigate the inhibition of IL-2 action and antitumor activity of celastrol (CEL), a compound identified in a screen for IL-2/CD25 binding inhibitors, and to elucidate the underlying role of CEL in immune cells. We found that CEL selectively impairs the binding of IL-2 and CD25 and directly binds to IL-2 but not to CD25. CEL significantly suppressed the proliferation and signaling of IL-2-dependent murine T cells and interfered with IL-2-responsive STAT5 phosphorylation in IL-2 reporter cells and human PBMCs. After confirming the impact of CEL on IL-2, we evaluated its antitumor activity in C57BL/6 mice bearing B16F10 tumors and found that CEL significantly inhibited tumor growth by increasing CD8+ T cells. We also found that CEL did not inhibit tumor growth in T cell-deficient BALB/c nude mice, suggesting that its activity was mediated by the T-cell response. Moreover, combination therapy with low-dose CEL and a TNFR2 antagonist synergistically improved the therapeutic efficacy of the individual monotherapies by increasing the ratio of intratumoral CD8/Treg cells and suppressing Foxp3 expression. These findings suggest that CEL, which inhibits CD25 binding by targeting IL-2, exerts antitumor activity by mediating the T-cell response and could be a promising candidate for combination therapy in cancer immunotherapy against melanoma.

A derivative of 3-(1,3-diarylallylidene)oxindoles inhibits dextran sulfate sodium-induced colitis in mice.

BACKGROUND: IA-0130 is a derivative of 3-(1,3-diarylallylidene)oxindoles, which is a selective estrogen receptor modulator (SERM). A previous study demonstrated that SERM exhibits anti-inflammatory effects on colitis by promoting the anti-inflammatory phenotype of monocytes in murine colitis. However, the therapeutic effects of oxindole on colitis remain unknown. Therefore, we evaluated the efficacy of IA-0130 on dextran sulfate sodium (DSS)-induced mouse colitis. METHODS: The DSS-induced colitis mouse model was established by administration of 2.5% DSS for 5 days. Mice were orally administered with IA-0130 (0.01 mg/kg or 0.1 mg/kg) or cyclosporin A (CsA; 30 mg/kg). Body weight, disease activity index score and colon length of mice were calculated and histological features of mouse colonic tissues were analyzed using hematoxylin and eosin staining. The expression of inflammatory cytokines and tight junction (TJ) proteins were analyzed using quantitative real-time PCR and enzyme-linked immunosorbent assay. The expression of interleukin-6 (IL-6) signaling molecules in colonic tissues were investigated using Western blotting and immunohistochemistry (IHC). RESULTS: IA-0130 (0.1 mg/kg) and CsA (30 mg/kg) prevented colitis symptom, including weight loss, bleeding, colon shortening, and expression of pro-inflammatory cytokines in colon tissues. IA-0130 treatment regulated the mouse intestinal barrier permeability and inhibited abnormal TJ protein expression. IA-0130 down-regulated IL-6 expression and prevented the phosphorylation of signaling molecules in colonic tissues. CONCLUSIONS: This study demonstrated that IA-0130 suppressed colitis progression by inhibiting the gp130 signaling pathway and expression of pro-inflammatory cytokines, and maintaining TJ integrity.

Chelerythrine, a novel small molecule targeting IL-2, inhibits melanoma progression by blocking the interaction between IL-2 and its receptor.

AIMS: In this study, we investigated the inhibition of IL-2 activity and anticancer efficacy of chelerythrine (CHE), a natural small molecule that targets IL-2 and inhibits CD25 binding, and elucidated the mechanism underlying the action of CHE on immune cells. MAIN METHODS: CHE was discovered by competitive binding ELISA and SPR analysis. The effect of CHE on IL-2 activity was evaluated in CTLL-2, HEK-Blue reporter and immune cells, and in ex vivo generation of regulatory T cells (Treg cells). The antitumor activity of CHE was evaluated in B16F10 tumor-bearing C57BL/6 or BALB/c nude mice. KEY FINDINGS: We identified that CHE, which acts as an IL-2 inhibitor, selectively inhibits the interaction between IL-2 and IL-2Rα and directly binds to IL-2. CHE inhibited the proliferation and signaling of CTLL-2 cells and suppressed IL-2 activity in HEK-Blue reporter and immune cells. CHE prevented the conversion of naive CD4+ T cells into CD4+CD25+Foxp3+ Treg cells in response to IL-2. CHE reduced tumor growth in C57BL/6 mice but not in T-cell-deficient mice, upregulated the expression of IFN-γ and cytotoxic molecules, and limited Foxp3 expression. Furthermore, the combination of CHE and a PD-1 inhibitor synergistically increased antitumor activity in melanoma-bearing mice and almost completely regressed the implanted tumors. SIGNIFICANCE: We found that CHE, which targets IL-2 and inhibits its binding to CD25, exhibits T cell-mediated antitumor activity and that combination therapy with CHE and PD-1 inhibitor induced synergistic antitumor effects, suggesting that CHE may be a promising anticancer agent for melanoma monotherapy and combination therapy.

Tannic acid, an IL-1ß-direct binding compound, ameliorates IL-1ß-induced inflammation and cartilage degradation by hindering IL-1ß-IL-1R1 interaction.

Interleukin-1ß (IL-1ß) is one of the most potent pro-inflammatory cytokines implicated in a wide range of autoinflammatory, autoimmune, infectious, and degenerative diseases. Therefore, many researchers have focused on developing therapeutic molecules that inhibit IL-1ß-IL-1 receptor 1 (IL-1R1) interaction for the treatment of IL-1-related diseases. Among IL-1-related diseases, osteoarthritis (OA), is characterized by progressive cartilage destruction, chondrocyte inflammation, and extracellular matrix (ECM) degradation. Tannic acid (TA) has been proposed to have multiple beneficial effects, including anti-inflammatory, anti-oxidant, and anti-tumor activities. However, it is unclear whether TA plays a role in anti-IL-1ß activity by blocking IL-1ß-IL-1R1 interaction in OA. In this study, we report the anti-IL-1ß activity of TA in the progression of OA in both in vitro human OA chondrocytes and in vivo rat OA models. Herein, using-ELISA-based screening, natural compound candidates capable of inhibiting the IL-1ß-IL-1R1 interaction were identified. Among selected candidates, TA showed hindering IL-1ß-IL-1R1 interaction by direct binding to IL-1ß using surface plasmon resonance (SPR) assay. In addition, TA inhibited IL-1ß bioactivity in HEK-Blue IL-1-dependent reporter cell line. TA also inhibited IL-1ß-induced expression of inducible nitric oxide synthase (NOS2), cyclooxygenase-2 (COX-2), IL-6, tumor necrosis factor-alpha (TNF-α), nitric oxide (NO), and prostaglandin E2 (PGE2) in human OA chondrocytes. Moreover, TA downregulated IL-1ß-stimulated matrix metalloproteinase (MMP)3, MMP13, ADAM metallopeptidase with thrombospondin type 1 motif (ADAMTS)4, and ADAMTS5, while upregulating collagen type II (COL2A1) and aggrecan (ACAN). Mechanistically, we confirmed that TA suppressed IL-1ß-induced MAPK and NF-κB activation. The protective effects of TA were also observed in a monosodium iodoacetamide (MIA)-induced rat OA model by reducing pain and cartilage degradation and inhibiting IL-1ß-mediated inflammation. Collectively, our results provide evidence that TA plays a potential role in OA and IL-1ß-related diseases by hindering IL-1ß-IL-1R1 interaction and suppressing IL-1ß bioactivity.