Genetically engineered T cells expressing a chimeric antigen receptor (CAR) are rapidly emerging a promising new treatment for haematological and non-haematological malignancies. CAR-T therapy can induce rapid and durable clinical responses but is associated with unique acute toxicities. Moreover, CAR-T cells are vulnerable to immunosuppressive mechanisms. Here, we report that CAR-T cells release extracellular vesicles, mostly in the form of exosomes that carry CAR on their surface. The CAR-containing exosomes express a high level of cytotoxic molecules and inhibit tumour growth. Compared with CAR-T cells, CAR exosomes do not express Programmed cell Death protein 1 (PD1), and their antitumour effect cannot be weakened by recombinant PD-L1 treatment. In a preclinical in vivo model of cytokine release syndrome, the administration of CAR exosomes is relatively safe compared with CAR-T therapy. This study supports the use of exosomes as biomimetic nanovesicles that may be useful in future therapeutic approaches against tumours.

The TIGIT (T cell immunoreceptor with Ig and ITIM domains) protein is a co-inhibitory receptor that has been reported to suppress autoreactive T and B cells to trigger immunological tolerance. We generated a new recombinant protein by connecting the extracellular domain of murine TIGIT to the Fc region of the rat immunoglobulin IgG2a. The fusion protein was then characterized. The results suggested that among mice with lupus that were treated with the TIGIT-Ig fusion protein, the onset of proteinuria was delayed, serum concentrations of autoantibodies, such as antinuclear antibodies, were reduced without a decrease in the total IgG concentrations, and the survival rate was significantly increased compared to those of the controls.In conclusion, TIGIT-Ig administration showed promising results for both the prevention and treatment of autoimmune diseases in mice. This indicates that treatment with recombinant human TIGIT-Ig shows promise as an effective way to treat human autoimmune diseases.

Hepatocellular carcinoma (HCC) is the global leading cause of cancer-related deaths due to the deficiency of targets for precision therapy. A new modality of epigenetic regulation has emerged involving RNA–RNA crosstalk networks where two or more competing endogenous RNAs (ceRNAs) bind to the same microRNAs. However, the contribution of such mechanisms in HCC has not been well studied. Herein, potential HMGB1-driven RNA–RNA crosstalk networks were evaluated at different HCC stages, identifying the mTORC2 component RICTOR as a potential HMGB1 ceRNA in HBV+ early stage HCC. Indeed, elevated HMGB1 mRNA was found to promote the expression of RICTOR mRNA through competitively binding with the miR-200 family, especially miR-429. Functional assays employing overexpression or interference strategies demonstrated that the HMGB1 and RICTOR 3′untranslated regions (UTR) epigenetically promoted the malignant proliferation, self-renewal, and tumorigenesis in HCC cells. Intriguingly, interference against HMGB1 and RICTOR in HCC cells promoted a stronger anti-PD-L1 immunotherapy response, which appeared to associate with the production of PD-L1+ exosomes. Mechanistically, the HMGB1-driven RNA-RNA crosstalk network facilitated HCC cell glutamine metabolism via dual mechanisms, activating a positive feedback loop involving mTORC2-AKT-C-MYC to upregulate glutamine synthetase (GS) expression, and inducing mTORC1 signaling to derepress SIRT4 on glutamate dehydrogenase (GDH). Meanwhile, this crosstalk network could impede the efficacy of immunotherapy through mTORC1-P70S6K dependent PD-L1 production and PD-L1+ exosomes activity. In conclusion, our study highlights the non-coding regulatory role of HMGB1 with implications for RNA-based therapeutic targeting together with a prediction of anti-PD-L1 immunotherapy in HCC.

Purpose: Both EGFR and PI3K-Akt signaling pathways have been used as therapeutically actionable targets, but resistance is frequently reported. In this report, we show that enrichment of the cancer stem cell (CSC) subsets and dysregulation of Notch signaling underlie the challenges to therapy and describe the development of bispecific antibodies targeting both HER and Notch signaling.Experimental Design: We utilized cell-based models to study Notch signaling in drug-induced CSC expansion. Both cancer cell line models and patient-derived xenograft tumors were used to evaluate the antitumor effects of bispecific antibodies. Cell assays, flow cytometry, qPCR, and in vivo serial transplantation assays were employed to investigate the mechanisms of action and pharmacodynamic readouts.Results: We found that EGFR/Notch targeting bispecific antibodies exhibited a notable antistem cell effect in both in vitro and in vivo assays. Bispecific antibodies delayed the occurrence of acquired resistance to EGFR inhibitors in triplenegative breast cancer cell line-based models and showed efficacy in patient-derived xenografts. Moreover, the EGFR/ Notch bispecific antibody PTG12 in combination with GDC-0941 exerted a stronger antitumor effect than the combined therapy of PI3K inhibitor with EGFR inhibitors or tarextumab in a broad spectrum of epithelial tumors. Mechanistically, bispecific antibody treatment inhibits the stem cell-like subpopulation, reduces tumor-initiating cell frequency, and downregulates the mesenchymal gene expression.Conclusions: These findings suggest that the coblockade of EGFR and Notch signaling has the potential to increase the response to PI3K inhibition, and PTG12 may gain clinical efficacy when combined with PI3K blockage in cancer treatment.

Chronic PKA phosphorylation of ryanodine receptor 2 (RyR2) has been shown to increase diastolic sarcoplasmic reticulum (SR) Ca2+ leakage and lead to cardiac dysfunction. We hypothesize that intracellular gene delivery of an RyR2‐targeting phosphorylation site—specific nanobody could preserve the contractility of the failing myocardium. In the present study, we acquired RyR2‐specific nanobodies from a phage display library that were variable domains of Camelidae heavy chain—only antibodies. One of the nanobodies, AR185, inhibited RyR2 phosphorylation in vitro and was chosen for further investigation. We investigated the potential of adeno‐associated virus (AAV)9—mediated cardiac expression of AR185 to combat postischemic heart failure (HF). AAV gene delivery elevated the intracellular expression of the AR185 protein in a rat model of ischemic HF, and this treatment normalized the systolic and diastolic dysfunction of the failing myocardium in vivo by reversing myocardial Ca2+ handling. Furthermore, AR185 gene transfer to failing cardiomyocytes reduced the frequency of SR calcium leaks, thereby restoring the attenuated intracellular calcium transients and SR calcium load. Moreover, AR185 gene transfer inhibited the PKA‐mediated phosphorylation of RyR2 in failing cardiomyocytes. Our results provide preclinical experimental evidence that the cardiac expression of RyR2 nanobodies with AAV9 vectors is a promising therapeutic strategy for HF.—Li, T., Shen, Y., Lin, F., Fu, W., Liu, S., Wang, C, Liang, J., Fan, X., Ye, X., Tang, Y., Ding, M., Yang, Y., Lei, C., Hu, S. Targeting RyR2 with a phosphorylation site–specific nanobody reverses dysfunction of failing cardiomyocytes in rats. FASEB J. 33, 7467–7478 (2019). http://www.fasebj.org

Background: Gallbladder carcinoma (GBC) remains a highly lethal disease worldwide. MiR-552 family members promote the malignant progression of a variety of digestive system tumors, but the role of miR-552-3p in GBC has not been elucidated. miR-552-3p was predicted to target the 3'-untranslated region (3'UTR) of the mRNA for the tumor suppressor gene "repulsive guidance molecule BMP co-receptor a" (RGMA). The aim of the present study was to clarify the roles and mechanisms of miR-552-3p targeting RGMA in the malignant progression of GBC. Methods: In vitro: expression of miR-552-3p was detected by real-time quantitative PCR (qRT-PCR) in tumor and non-tumor adjacent tissues (NATs). Lentivirus-miR-552-3p was employed to knockdown this miRNA in GBC cell lines. Stem cell-related transcription factors and markers were assessed by qRT-PCR.Cell Counting Kit-8 (CCK-8), sphere formation and transwell assays were used to determine the malignant phenotypes of GBC cells. Targeting the 3'UTR of RGMA by miR-552-3p was verified by integrated analysis including bioinformatics prediction, luciferase assays, measures of changes of gene expression and rescue experiments. In vivo: mouse models of subcutaneous tumors and lung metastases were established to observe the effect of miR-552-3p on tumorigenesis and organ metastasis, respectively.Results: MiR-552-3p was abnormally highly expressed in GBC tissues and cancer stem cells. Interference with miR-552-3p in SGC-996 and GBC-SD cells significantly inhibited GBC stem cell expansion.Reciprocally, miR-552-3p promoted GBC cell proliferation, migration and invasion both in vitro and in vivo; hence, interference with this miRNA impeded the malignant progression of GBC. Furthermore, the important tumor suppressor gene RGMA was identified as a target of miR-552-3p. The effects of miR-552-3p on cell proliferation and metastasis were abrogated or enhanced by gain or loss of RGMA function, respectively. Mechanistically, miR-552-3p promoted GBC progression by reactivating the Akt/β-catenin Original ArticleSong et al. MiR-552-3p promotes progression of GBC by targeting RGMA

<p>Supplementary methods, Supplementary figures and tables</p>

<p>Supplementary methods, Supplementary figures and tables</p>