Regulatory T cells (Tregs) expressing FOXP3 are essential for the maintenance of self-tolerance and are deficient in many common autoimmune diseases. Immune tolerance is maintained in part by IL-2 and deficiencies in the IL-2 pathway cause reduced Treg function and an increased risk of autoimmunity. Recent studies expanding Tregs in vivo with low-dose IL-2 achieved major clinical successes highlighting the potential to optimize this pleiotropic cytokine for inflammatory and autoimmune disease indications. Here we compare the clinically approved IL-2 molecule, Proleukin, with two engineered IL-2 molecules with long half-lives owing to their fusion in monovalent and bivalent stoichiometry to a non-FcRγ binding human IgG1. Using nonhuman primates, we demonstrate that single ultra-low doses of IL-2 fusion proteins induce a prolonged state of in vivo activation that increases Tregs for an extended period of time similar to multiple-dose Proleukin. One of the common pleiotropic effects of high dose IL-2 treatment, eosinophilia, is eliminated at doses of the IL-2 fusion proteins that greatly expand Tregs. The long half-lives of the IL-2 fusion proteins facilitated a detailed characterization of an IL-2 dose response driving Treg expansion that correlates with increasingly sustained, suprathreshold pSTAT5a induction and subsequent sustained increases in the expression of CD25, FOXP3 and Ki-67 with retention of Treg-specific epigenetic signatures at FOXP3 and CTLA4.

Advancements in flow cytometers with capability to measure 15 or more parameters have enabled us to characterize cell populations at unprecedented levels of detail. Beyond discovery research, there is now a growing demand to dive deeper into evaluating the immune response in clinical trials for immune modulating compounds. However, for high-volume, complex flow cytometry data generated in clinical trials, conventional manual gating remains the standard of practice. Traditional manual gating is resource intense and becomes a bottleneck and an impractical method to complete high volumes of flow cytometry data analysis. Current efforts to automate "manual gating" have shown that computational algorithms can facilitate the analysis of daunting multi-parameter data; however, a greater degree of precision in comparison with traditional manual gating is needed for wide-scale adoption of automated gating methods. In an effort to more closely follow the manual gating process, our automated gating pipeline was created to include negative controls (Fluorescence Minus One [FMO]) to enhance the reliability of gate placement. We demonstrate that use of an automated pipeline, heavily relying on FMO controls for population discrimination, can analyze multi-parameter, large-scale clinical datasets with comparable precision and accuracy to traditional manual gating.

Serum interleukin (IL)‑17A level is associated with higher microvessel density and poor prognosis in liver cancer. However, the specific mechanism underlying the role of IL‑17A in liver cancer remains controversial. In the present study, the effect of IL‑17A on liver cancer cells was examined. IL‑17A had no evident impact on vascular endothelial growth factor A (VEGFA) production in HepG2 and Huh7.5 cells as determined by reverse transcription‑quantitative PCR and ELISA, but it did stimulate angiogenic CXC chemokine secretion, including chemokine (C‑X‑C motif) ligand 1 (CXCL1), CXCL2, CXCL3, CXCL5, CXCL6 and CXCL8 in Huh7.5 cells and CXCL2 in HepG2 cells. In addition, the production of angiostatic chemokines such as CXCL10 was not affected. The supernatant of Huh7.5‑IL17A cells promoted endothelial cell chemotaxis, which was attenuated by the C‑X‑C chemokine receptor type 2 (CXCR2) inhibitor SB225002. Although there was no role of IL‑17A in promoting in vitro cell proliferation, IL‑17A markedly increased the tumor growth of Huh7.5 cells in both subcutaneous and orthotopic xenograft models with increased vascularization. Taken together, these results demonstrated that IL‑17A may stimulate chemokine‑induced angiogenesis and promote tumor progression, independent of VEGF signaling. The CXCL‑CXCR2 axis may be a novel target for the anti‑angiogenesis treatment of liver cancer.

Although guidelines recommend extended surgical resection, radical resection and lymphadenectomy for patients with tumor stage (T)1b gallbladder cancer, these procedures are substantially underutilized. This population-based, retrospective cohort study aimed to evaluate treatment patterns and outcomes of 401 patients using the US Surveillance, Epidemiology, and End Results (SEER) database from 2004 to 2013. Results showed that median overall survival (OS) was 69 months for lymphadenectomy patients and 37 months for those without lymphadenectomy. Lymphadenectomy also tended to prolong cancer-specific survival (CSS), although the differences were not statistically significant. OS and CSS were similar for patients who received simple cholecystectomy and extended surgical resection. Cox proportional hazards regression models revealed survival advantages in patients with stage T1bN0 gallbladder cancer compared to those with stage T1bN1, and patients who received simple cholecystectomy plus lymphadenectomy compared to those who did not receive lymph node dissection. In further analyses, patients undergoing simple cholecystectomy who had five or more lymph nodes excised had better OS and CSS than those without lymph node dissection. In conclusion, survival advantages are shown for patients with T1b gallbladder cancer undergoing surgeries with lymphadenectomy. Future studies with longer follow-up and control of potential confounders are highly warranted.

Methods accurately predicting the responses of colorectal cancer (CRC) and colorectal cancer liver metastasis (CRLM) to personalized chemotherapy remain limited due to tumor heterogeneity. This study introduces an innovative patient-derived CRC and CRLM tumor model for preclinical investigation, utilizing 3d-bioprinting (3DP) technology. Efficient construction of homogeneous in vitro 3D models of CRC/CRLM is achieved through the application of patient-derived primary tumor cells and 3D bioprinting with bioink. Genomic and histological analyses affirm that the CRC/CRLM 3DP tumor models effectively retain parental tumor biomarkers and mutation profiles. In vitro tests evaluating chemotherapeutic drug sensitivities reveal substantial tumor heterogeneity in chemotherapy responses within the 3DP CRC/CRLM models. Furthermore, a robust correlation is evident between the drug response in the CRLM 3DP model and the clinical outcomes of neoadjuvant chemotherapy. These findings imply a significant potential for the application of patient-derived 3DP cancer models in precision chemotherapy prediction and preclinical research for CRC/CRLM.