Natural evidence of coronaviral 2'-O-methyltransferase activity affecting viral pathogenesis via improved substrate RNA binding.

Previous studies through targeted mutagenesis of K-D-K-E motif have demonstrated that 2'-O-MTase activity is essential for efficient viral replication and immune evasion. However, the K-D-K-E catalytic motif of 2'-O-MTase is highly conserved across numerous viruses, including flaviviruses, vaccinia viruses, coronaviruses, and extends even to mammals. Here, we observed a stronger 2'-O-MTase activity in SARS-CoV-2 compared to SARS-CoV, despite the presence of a consistently active catalytic center. We further identified critical residues (Leu-36, Asn-138 and Ile-153) which served as determinants of discrepancy in 2'-O-MTase activity between SARS-CoV-2 and SARS-CoV. These residues significantly enhanced the RNA binding affinity of 2'-O-MTase and boosted its versatility toward RNA substrates. Of interest, a triple substitution (Leu36 → Ile36, Asn138 → His138, Ile153 → Leu153, from SARS-CoV-2 to SARS-CoV) within nsp16 resulted in a proportional reduction in viral 2'-O-methylation and impaired viral replication. Furthermore, it led to a significant upregulation of type I interferon (IFN-I) and proinflammatory cytokines both in vitro and vivo, relying on the cooperative sensing of melanoma differentiation-associated protein 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2). In conclusion, our findings demonstrated that alterations in residues other than K-D-K-E of 2'-O-MTase may affect viral replication and subsequently influence pathogenesis. Monitoring changes in nsp16 residues is crucial as it may aid in identifying and assessing future alteration in viral pathogenicity resulting from natural mutations occurring in nsp16.

Predictive factors that influence the clinical efficacy of umbilical cord-derived mesenchymal stromal cells in the treatment of type 2 diabetes mellitus.

BACKGROUND: Our previous single-center, randomized, double-blinded, placebo-controlled phase 2 study evaluated the safety and effectiveness of human umbilical cord mesenchymal stromal cell (UC-MSC) transfusion for treating patients with type 2 diabetes mellitus (T2DM). Indeed, this potential treatment strategy was able to reduce insulin use by half in a considerable number of patients. However, many other patients' responses to UC-MSC transfusion were insignificant. The selection of patients who might benefit from UC-MSC treatment is crucial from a clinical standpoint. METHODS: In this post hoc analysis, 37 patients who received UC-MSC transfusions were divided into two groups based on whether their glycated hemoglobin (hemoglobin A1c, or HbA1c) level was less than 7% after receiving UC-MSC treatment. The baseline differences between the two groups were summarized, and potential factors influencing efficacy of UC-MSCs for T2DM were analyzed by univariate and multivariate logistic regression. The correlations between the relevant hormone levels and the treatment effect were further analyzed. RESULTS: At the 9-week follow-up, 59.5% of patients achieved their targeted HbA1c level. Male patients with lower baseline HbA1c and greater C-peptide area under the curve (AUCC-pep) values responded favorably to UC-MSC transfusion, according to multivariate analysis. The effectiveness of UC-MSCs transfusion was predicted by AUCC-pep (cutoff value: 14.22 ng/h/mL). Further investigation revealed that AUCC-pep was increased in male patients with greater baseline testosterone levels. CONCLUSIONS: Male patients with T2DM with greater AUCC-pep may be more likely to respond clinically to UC-MSC therapy, and further large-scale multi-ethnic clinical studies should be performed to confirm the conclusion.

NSUN2-mediated m5C modification of HBV RNA positively regulates HBV replication.

Chronic hepatitis B virus (HBV) infection is a major cause of liver cirrhosis and liver cancer, despite strong prevention and treatment efforts. The study of the epigenetic modification of HBV has become a research hotspot, including the N6-methyladenosine (m6A) modification of HBV RNA, which plays complex roles in the HBV life cycle. In addition to m6A modification, 5-methylcytosine (m5C) is another major modification of eukaryotic mRNA. In this study, we explored the roles of m5C methyltransferase and demethyltransferase in the HBV life cycle. The results showed that m5C methyltransferase NSUN2 deficiency could negatively regulate the expression of HBV while m5C demethyltransferase TET2 deficiency positively regulates the expression of HBV. Subsequently, we combined both in vitro bisulfite sequencing and high-throughput bisulfite sequencing methods to determine the distribution and stoichiometry of m5C modification in HBV RNA. Two sites: C2017 and C131 with the highest-ranking methylation rates were identified, and mutations at these two sites could lead to the decreased expression and replication of HBV, while the mutation of the "fake" m5C site had no effect. Mechanistically, NSUN2-mediated m5C modification promotes the stability of HBV RNA. In addition, compared with wild-type HepG2-NTCP cells and primary human hepatocytes, the replication level of HBV after NSUN2 knockdown decreased, and the ability of the mutant virus to infect and replicate in wild-type HepG2-NTCP cells and PHHs was substantially impaired. Similar results were found in the experiments using C57BL/6JGpt-Nsun2+/- mice. Interestingly, we also found that HBV expression and core protein promoted the endogenous expression of NSUN2, which implied a positive feedback loop. In summary, our study provides an accurate and high-resolution m5C profile of HBV RNA and reveals that NSUN2-mediated m5C modification of HBV RNA positively regulates HBV replication by maintaining RNA stability.

Efficacy of Umbilical Cord-Derived Mesenchymal Stem Cells in the Treatment of Type 2 Diabetes Assessed by Retrospective Continuous Glucose Monitoring.

Umbilical cord-derived mesenchymal stem cells (UC-MSCs) have been proved a promising clinical strategy for the treatment of diabetes, and time in range (TIR) has been demonstrated a new metric of glycemic control links to diabetes complications. To further assess the therapeutic effect of UC-MSCs on TIR, a phase II study investigating the efficacy of UC-MSCs in Chinese adults with type 2 diabetes (T2D) assessed by retrospective continuous glucose monitoring (CGM) was conducted. In this randomized and placebo-controlled trial, a total of 73 patients were randomly assigned to receive intravenous infusion of UC-MSCs (n = 37) or placebo (n = 36) 3 times at 4-week intervals and followed up for 48 weeks. The primary endpoint was the changes in TIR and glycosylated hemoglobin (HbA1c). TIR and HbA1c were both significantly improved in UC-MSCs and placebo groups after 48 weeks of therapy compared with baseline. Compared with placebo group, UC-MSCs group exhibited more pronounced changes at 9 and 48 weeks from baseline in TIR (26.54 vs. 15.84 and 21.36 vs. 6.32) and HbA1c (-1.79 vs. -0.96 and -1.36 vs. -0.51). More patients in UC-MSCs group achieved the glycemic control target of TIR ≥ 70% and HbA1c < 7% at 9 and 48 weeks than in placebo group (59.5% vs. 27.8% and 43.2% vs. 11.1%). The C-peptide area under the curve (AUCC-pep) was an independent risk factor associated with efficacy in T2D undergoing UC-MSCs intervention. These results illustrate that UC-MSCs administration via intravenous infusion is an effective approach for ameliorating TIR.

NETosis promotes chronic inflammation and fibrosis in systemic lupus erythematosus and COVID-19.

Pulmonary fibrosis, a serious complication of systemic lupus erythematosus (SLE) and coronavirus disease 2019 (COVID-19), leads to irreversible lung damage. However, the underlying mechanism of this condition remains unclear. In this study, we revealed the landscape of transcriptional changes in lung biopsies from individuals with SLE, COVID-19-induced pulmonary fibrosis, and idiopathic pulmonary fibrosis (IPF) using histopathology and RNA sequencing, respectively. Despite the diverse etiologies of these diseases, lung expression of matrix metalloproteinase genes in these diseases showed similar patterns. Particularly, the differentially expressed genes were significantly enriched in the pathway of neutrophil extracellular trap formation, showing similar enrichment signature between SLE and COVID-19. The abundance of Neutrophil extracellular traps (NETs) was much higher in the lungs of individuals with SLE and COVID-19 compared to those with IPF. In-depth transcriptome analyses revealed that NETs formation pathway promotes epithelial-mesenchymal transition (EMT). Furthermore, stimulation with NETs significantly up-regulated α-SMA, Twist, Snail protein expression, while decreasing the expression of E-cadherin protein in vitro. This indicates that NETosis promotes EMT in lung epithelial cells. Given drugs that are efficacious in degrading damaged NETs or inhibiting NETs production, we identified a few drug targets that were aberrantly expressed in both SLE and COVID-19. Among these targets, the JAK2 inhibitor Tofacitinib could effectively disrupted the process of NETs and reversed NET-induced EMT in lung epithelial cells. These findings support that the NETs/EMT axis, activated by SLE and COVID-19, contributes to the progression of pulmonary fibrosis. Our study also highlights that JAK2 as a potential target for the treatment of fibrosis in these diseases.

Epigenetic reprogramming of Runx3 reinforces CD8 + T-cell function and improves the clinical response to immunotherapy.

BACKGROUND: Checkpoint blockade immunotherapy, represented by PD-1 or PD-L1 antibody treatment, has been of tremendous success in clinical practice. However, the low clinical response rate and lack of biomarkers for prediction of the immune response limit the clinical application of anti-PD-1 immunotherapy. Our recent work showed that a combination of low-dose decitabine and PD-1-ab significantly improved the complete response (CR) rate of cHL patients from 32 to 71%, which indicates that there is a significant correlation between epigenetic regulation and the clinical response to immunotherapy. METHODS: We recruited two groups of Hodgkin lymphoma patients who were treated with anti-PD-1 and DAC+anti-PD-1. CD8+ T cells were isolated from the patients' peripheral blood, DNA methylation was analyzed by EPIC, the expression profile was analyzed by RNA-seq, and multigroup analysis was performed with IPA and GSEA functional annotations. We explored the effect of DAC on the function of CD8+ T cells in the blood, spleen, tumor and lymph nodes using a mouse model. Furthermore, we explored the function of Tils in the tumor microenvironment. Then, we constructed Runx3-knockout mice to confirm the T-cell-specific function of Runx3 in CD8+ T cells and analyzed various subtypes of T cells and cytokines using mass cytometry (CyTOF). RESULTS: Multiomics analysis identified that DNA methylation reprogramming of Runx3 was a crucial mediator of CD8+ T-cell function. Multiomics data showed that reversal of methylation of the Runx3 promoter promoted the infiltration of CD8+ TILs and mitigated the exhaustion of CD8+ T cells. Furthermore, experiments on tissue-specific Runx3-knockout mice showed that Runx3 deficiency reduced CD8+ T infiltration and the differentiation of effector T and memory T cells. Furthermore, Runx3 deficiency significantly decreased CCR3 and CCR5 levels. Immunotherapy experiments in Runx3 conditional knockout mice showed that DAC could not reverse the resistance of anti-PD-1 in the absence of Runx3. Moreover, both our clinical data and data from TISIDB showed that Runx3 could be a potential biomarker for immunotherapy to predict the clinical response rate. CONCLUSION: We demonstrate that the DNA methylation of Runx3 plays a critical role in CD8+ T-cell infiltration and differentiation during decitabine-primed PD-1-ab immunotherapy, which provides a supporting mechanism for the essential role of epiregulation in immunotherapy.

NSUN2-mediated M5c methylation of IRF3 mRNA negatively regulates type I interferon responses during various viral infections.

5-Methylcytosine (m5C) is a widespread post-transcriptional RNA modification and is reported to be involved in manifold cellular responses and biological processes through regulating RNA metabolism. However, its regulatory role in antiviral innate immunity has not yet been elucidated. Here, we report that NSUN2, a typical m5C methyltransferase, negatively regulates type I interferon responses during various viral infections, including SARS-CoV-2. NSUN2 specifically mediates m5C methylation of IRF3 mRNA and accelerates its degradation, resulting in low levels of IRF3 and downstream IFN-ß production. Knockout or knockdown of NSUN2 enhanced type I interferon and downstream ISGs during various viral infection in vitro. And in vivo, the antiviral innate response is more dramatically enhanced in Nsun2+/- mice than in Nsun2+/+ mice. The highly m5C methylated cytosines in IRF3 mRNA were identified, and their mutation enhanced cellular IRF3 mRNA levels. Moreover, infection with Sendai virus (SeV), vesicular stomatitis virus (VSV), herpes simplex virus 1 (HSV-1), or Zika virus (ZIKV) resulted in a reduction of endogenous NSUN2 levels. Especially, SARS-CoV-2 infection (WT strain and BA.1 omicron variant) also decreased endogenous levels of NSUN2 in COVID-19 patients and K18-hACE2 KI mice, further increasing type I interferon and downstream ISGs. Together, our findings reveal that NSUN2 serves as a negative regulator of interferon response by accelerating the fast turnover of IRF3 mRNA, while endogenous NSUN2 levels decrease during SARS-CoV-2 and various viral infections to boost antiviral responses for effective elimination of viruses.

Efficacy and safety of umbilical cord-derived mesenchymal stem cells in Chinese adults with type 2 diabetes: a single-center, double-blinded, randomized, placebo-controlled phase II trial.

BACKGROUND: To determine the efficacy and safety of umbilical cord-derived mesenchymal stem cells (UC-MSCs) in Chinese adults with type 2 diabetes mellitus (T2DM). METHODS: In this single-center, double-blinded, randomized, placebo-controlled phase II trial, 91 patients were randomly assigned to receive intravenous infusion of UC-MSCs (n = 45) or placebo (n = 46) three times with 4-week intervals and followed up for 48 weeks from October 2015 to December 2018. The primary endpoint was the percentage of patients with glycated hemoglobin (HbA1c) levels of < 7.0% and daily insulin reduction of ≥ 50% at 48 weeks. Additional endpoints were changes of metabolic control, islet ß-cell function, insulin resistance, and safety. RESULTS: At 48 weeks, 20% of the patients in the UC-MSCs group and 4.55% in the placebo group reached the primary endpoint (p < 0.05, 95% confidence interval (CI) 2.25-28.66%). The percentage of insulin reduction of the UC-MSCs group was significantly higher than that of the placebo group (27.78% versus 15.62%, p < 0.05). The levels of HbA1c decreased 1.31% (9.02 ± 1.27% to 7.52 ± 1.07%, p < 0.01) in the UC-MSCs group, and only 0.63% in the placebo group (8.89 ± 1.11% to 8.19 ± 1.02%, p˃0.05; p = 0.0081 between both groups). The glucose infusion rate (GIR) increased significantly in the UC-MSCs group (from 3.12 to 4.76 mg/min/kg, p < 0.01), whereas no significant change was observed in the placebo group (from 3.26 to 3.60 mg/min/kg, p ˃ 0.05; p < 0.01 between both groups). There was no improvement in islet ß-cell function in both groups. No major UC-MSCs transplantation-related adverse events occurred. CONCLUSIONS: UC-MSCs transplantation could be a potential therapeutic approach for Chinese adults with T2DM. Trial registration This study was registered on ClinicalTrials.gov (identifier: NCT02302599).

Nucleolar RNA in action: Ultrastructure revealed during protein translation through a terpyridyl manganese(II) complex.

Nucleolar RNA stored within the interphase nucleus offers a targetable site for processing ribosomal RNAs that can significantly impact the synthesis of cytosolic protein. Nevertheless, information regarding nucleolus ultrastructure is still largely limited to observations from electron microscopy, while its interior information including the correlation between nucleolar RNA and ribosomes remains to be elucidated. In this work, we designed a highly charged terpyridyl manganese(II) complex which could rapidly penetrate into live cell nucleus and selectively 'light up' nucleolar RNA. Its superior optical properties allow the observation nucleolar ultrastructures under super-resolution nanoscopy including RNA rich dense fibrillar components and fibrillar centers. Moreover, this complex allowed us to explain the nucleolar RNA dynamics and its relationship with ribosomes under the translation of protein synthesis.

Human adenoviruses: A suspect behind the outbreak of acute hepatitis in children amid the COVID-19 pandemic.

As of 10 May 2022, at least 450 cases of pediatric patients with acute hepatitis of unknown cause have been reported worldwide. Human adenoviruses (HAdVs) have been detected in at least 74 cases, including the F type HAdV41 in 18 cases, which indicates that adenoviruses may be associated with this mysterious childhood hepatitis, although other infectious agents or environmental factors cannot be excluded. In this review, we provide a brief introduction of the basic features of HAdVs and describe diseases caused by different HAdVs in humans, aiming to help understand the biology and potential risk of HAdVs and cope with the outbreak of acute child hepatitis.

Revealing Sulfur Dioxide Regulation to Nucleophagy in Embryo Development by an Adaptive Coloration Probe.

Understanding signaling molecules in regulating organelles dynamics and programmed cell death is critical for embryo development but is also challenging because current imaging probes are incapable of simultaneously imaging the signaling molecules and the intracellular organelles they interact with. Here, we report a chemically and environmentally dual-responsive imaging probe that can react with gasotransmitters and label cell nuclei in distinctive fluorescent colors, similar to the adaptive coloration of chameleons. Using this intracellular chameleon-like probe in three-dimensional (3D) super-resolution dynamic imaging of live cells, we discovered SO2 as a critical upstream signaling molecule that activates nucleophagy in programmed cell death. An elevated level of SO2 prompts kiss fusion between the lysosomal and nuclear membranes and nucleus shrinkage and rupture. Significantly, we revealed that the gasotransmitter SO2 is majorly generated in the yolk, induces autophagy there at the initial stage of embryo development, and is highly related to the development of the auditory nervous system.

Live attenuated coronavirus vaccines deficient in N7-Methyltransferase activity induce both humoral and cellular immune responses in mice.

Coronaviruses (CoVs) can infect a variety of hosts, including humans, livestock and companion animals, and pose a serious threat to human health and the economy. The current COVID-19 pandemic, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has killed millions of people. Unfortunately, effective treatments for CoVs infection are still lacking, suggesting the importance of coronavirus vaccines. Our previous work showed that CoV nonstuctural protein 14 (nsp14) functions as (guanine-N7)-methyltransferase (N7-MTase), which is involved in RNA cap formation. Moreover, we found that N7-MTase is well conserved among different CoVs and is a universal target for developing antivirals against CoVs. Here, we show that N7-MTase of CoVs can be an ideal target for designing live attenuated vaccines. Using murine hepatitis virus strain A59 (MHV-A59), a representative and well-studied model of coronaviruses, we constructed N7-MTase-deficient recombinant MHV D330A and Y414A. These two mutants are highly attenuated in mice and exhibit similar replication efficiency to the wild-type (WT) virus in the cell culture. Furthermore, a single dose immunization of D330A or Y414A can induce long-term humoral immune responses and robust CD4+ and CD8+ T cell responses, which can provide full protection against the challenge of a lethal-dose of MHV-A59. Collectively, this study provides an ideal strategy to design live attenuated vaccines for coronavirus by abolishing viral RNA N7-MTase activity. This approach may apply to other RNA viruses that encode their own conservative viral N7-methyltransferase.

Low-Dose Decitabine Augments the Activation and Anti-Tumor Immune Response of IFN-γ+ CD4+ T Cells Through Enhancing IκBα Degradation and NF-κB Activation.

BACKGROUND: CD4+ T cells play multiple roles in controlling tumor growth and increasing IFN-γ+ T-helper 1 cell population could promote cell-mediated anti-tumor immune response. We have previously showed that low-dose DNA demethylating agent decitabine therapy promotes CD3+ T-cell proliferation and cytotoxicity; however, direct regulation of purified CD4+ T cells and the underlying mechanisms remain unclear. METHODS: The effects of low-dose decitabine on sorted CD4+ T cells were detected both in vitro and in vivo. The activation, proliferation, intracellular cytokine production and cytolysis activity of CD4+ T cells were analyzed by FACS and DELFIA time-resolved fluorescence assays. In vivo ubiquitination assay was performed to assess protein degradation. Moreover, phosphor-p65 and IκBα levels were detected in sorted CD4+ T cells from solid tumor patients with decitabine-based therapy. RESULTS: Low-dose decitabine treatment promoted the proliferation and activation of sorted CD4+ T cells, with increased frequency of IFN-γ+ Th1 subset and enhanced cytolytic activity in vitro and in vivo. NF-κB inhibitor, BAY 11-7082, suppressed decitabine-induced CD4+ T cell proliferation and IFN-γ production. In terms of mechanism, low-dose decitabine augmented the expression of E3 ligase ß-TrCP, promoted the ubiquitination and degradation of IκBα and resulted in NF-κB activation. Notably, we observed that in vitro low-dose decitabine treatment induced NF-κB activation in CD4+ T cells from patients with a response to decitabine-primed chemotherapy rather than those without a response. CONCLUSION: These data suggest that low-dose decitabine potentiates CD4+ T cell anti-tumor immunity through enhancing IκBα degradation and therefore NF-κB activation and IFN-γ production.

Improved clinical outcome in a randomized phase II study of anti-PD-1 camrelizumab plus decitabine in relapsed/refractory Hodgkin lymphoma.

BACKGROUND: Programmed death-1 (PD-1) blockade monotherapy induced durable remission in a subset of patients with relapsed/refractory classical Hodgkin lymphoma (cHL). We asked whether the anti-PD-1 agent, camrelizumab, combined with the DNA demethylating agent, decitabine, improves progression-free survival (PFS) in patients with relapsed/refractory cHL over camrelizumab alone. METHODS: This extended follow-up of an ongoing randomized phase II trial analyzed PFS among patients enrolled from January 2017 through July 2018. Sixty-one patients with relapsed/refractory cHL who were clinically naïve to PD-1 blockade and had received ≥2 previous therapies were randomized 1:2 to receive either camrelizumab (200 mg) monotherapy or camrelizumab (200 mg, day 8) combined with decitabine (10 mg/day, days 1-5) every 3 weeks. RESULTS: With a median follow-up of 34.5 months, complete remission was 79% (95% CI 63% to 90%) in the decitabine-plus-camrelizumab group versus 32% (95% CI 13% to 57%) in the camrelizumab group (p=0.001). Median duration of response was not reached in the decitabine-plus-camrelizumab group, with an estimated 63% (95% CI 46% to 75%) of patients maintaining a response at 24 months. Median PFS with decitabine-plus-camrelizumab therapy was 35.0 months (95% CI not reached) and 15.5 months (95% CI 8.4 to 22.7 months) with camrelizumab monotherapy (HR, 0.46; 95% CI 0.21 to 1.01; p=0.02). Female gender, lower tumor burden, and fewer previous therapies were favorable prognostic factors for durable remission with camrelizumab monotherapy. The PFS benefits of decitabine-plus-camrelizumab versus camrelizumab were observed in most subgroups, especially in patients with relative larger tumor burdens and those treated with ≥3 prior therapies. After decitabine-plus-camrelizumab treatment, the percentage increase of circulating peripheral central memory T-cells correlated with both improved clinical response and PFS, suggesting a putative biomarker of decitabine-plus-camrelizumab therapy for cHL. CONCLUSIONS: Decitabine-plus-camrelizumab results in longer PFS compared with camrelizumab alone in patients with relapsed/refractory cHL. TRIAL REGISTRATION NUMBERS: NCT02961101 and NCT03250962.

Efficacy of Decitabine plus Anti-PD-1 Camrelizumab in Patients with Hodgkin Lymphoma Who Progressed or Relapsed after PD-1 Blockade Monotherapy.

PURPOSE: Programmed death-1 (PD-1) blockade monotherapy is effective in relapsed/refractory classical Hodgkin lymphoma (cHL), but a subset of patients is recalcitrant to PD-1 inhibitors and only a minority of patients achieves durable remission. Effective treatment regimens for those with relapsed/progressive cHL after single-agent anti-PD-1 are urgently needed. Anti-PD-1 combination with the DNA-demethylating agent decitabine showed positive preliminary results in our test cohort patients who were resistant to anti-PD-1. Here, we assess the efficacy of decitabine plus anti-PD-1 therapy in an expansion cohort and after longer follow-up. PATIENTS AND METHODS: We present the response and progression-free survival rates from patients with relapsed/refractory cHL who relapsed/progressed after prior anti-PD-1 monotherapy, and who received decitabine (10 mg/day, days 1-5) plus the anti-PD-1 camrelizumab (200 mg, day 8), every 3 weeks in a phase II trial (ClinicalTrials.gov: NCT02961101 and NCT03250962). RESULTS: Overall, 51 patients (test cohort: 25, expansion cohort: 26) were treated and 50 evaluated for efficacy. The objective response rate was 52% [nine complete responses (CR); 36%] in the test cohort, and 68% (six CRs; 24%) in the expansion cohort. Median progression-free survival with decitabine plus camrelizumab was 20.0 and 21.6 months, respectively, which was significantly longer than that achieved with prior anti-PD-1 monotherapy. Durable response was observed in an estimated 78% of patients who achieved CR at 24 months. After decitabine plus camrelizumab, the ratio increase of circulating peripheral central memory T cells directly correlated with both clinical response and progression-free survival. CONCLUSIONS: Decitabine plus camrelizumab is associated with high response rates and long-term benefits in patients with relapsed/refractory cHL who failed PD-1 inhibitors.

Reversion of early- and late-stage ß-cell dedifferentiation by human umbilical cord-derived mesenchymal stem cells in type 2 diabetic mice.

BACKGROUND AIMS: The authors aimed to observe ß-cell dedifferentiation in type 2 diabetes mellitus (T2DM) and investigate the reversal effect of umbilical cord-derived mesenchymal stem cells (UC-MSCs) on early- and late-stage ß-cell dedifferentiation. METHODS: In high-fat diet (HFD)/streptozotocin (STZ)-induced T2DM mice, the authors examined the predominant role of ß-cell dedifferentiation over apoptosis in the development of T2DM and observed the reversion of ß-cell dedifferentiation by UC-MSCs. Next, the authors used db/db mice to observe the progress of ß-cell dedifferentiation from early to late stage, after which UC-MSC infusions of the same amount were performed in the early and late stages of dedifferentiation. Improvement in metabolic indices and restoration of ß-cell dedifferentiation markers were examined. RESULTS: In HFD/STZ-induced T2DM mice, the proportion of ß-cell dedifferentiation was much greater than that of apoptosis, demonstrating that ß-cell dedifferentiation was the predominant contributor to T2DM. UC-MSC infusions significantly improved glucose homeostasis and reversed ß-cell dedifferentiation. In db/db mice, UC-MSC infusions in the early stage significantly improved glucose homeostasis and reversed ß-cell dedifferentiation. In the late stage, UC-MSC infusions mildly improved glucose homeostasis and partially reversed ß-cell dedifferentiation. Combining with other studies, the authors found that the reversal effect of UC-MSCs on ß-cell dedifferentiation relied on the simultaneous relief of glucose and lipid metabolic disorders. CONCLUSIONS: UC-MSC therapy is a promising strategy for reversing ß-cell dedifferentiation in T2DM, and the reversal effect is greater in the early stage than in the late stage of ß-cell dedifferentiation.

Photodynamic Therapy Directed by Three-Photon Active Rigid Plane Organic Photosensitizer.

Multi-photon photosensitizers (PSs) could significantly improve the efficacy of photodynamic therapy due to the long-wavelength favorability for deeper tissue penetration and lower biological damage. However, most studies are limited to single-photon or two-photon PSs at a relatively short-wave excitation window. To overcome this barrier, we rationally design a series of rigid plane compounds with efficient reactive oxygen species (ROS) production in vitro under laser irradiation. Furthermore, the studies show that one of the compounds (U-TsO) could induce rapid multi-types of cell death under three-photon exposure, suggesting a promising clinical outcome in ex vivo 3D multicellular tumor spheroid. This work offers a novel strategy to construct functional materials with competitive multi-photon photodynamic therapy (PDT) outcome.

Manganese is critical for antitumor immune responses via cGAS-STING and improves the efficacy of clinical immunotherapy.

CD8+ T cell-mediated cancer clearance is often suppressed by the interaction between inhibitory molecules like PD-1 and PD-L1, an interaction acts like brakes to prevent T cell overreaction under normal conditions but is exploited by tumor cells to escape the immune surveillance. Immune checkpoint inhibitors have revolutionized cancer therapeutics by removing such brakes. Unfortunately, only a minority of cancer patients respond to immunotherapies presumably due to inadequate immunity. Antitumor immunity depends on the activation of the cGAS-STING pathway, as STING-deficient mice fail to stimulate tumor-infiltrating dendritic cells (DCs) to activate CD8+ T cells. STING agonists also enhance natural killer (NK) cells to mediate the clearance of CD8+ T cell-resistant tumors. Therefore STING agonists have been intensively sought after. We previously discovered that manganese (Mn) is indispensable for the host defense against cytosolic dsDNA by activating cGAS-STING. Here we report that Mn is also essential in innate immune sensing of tumors and enhances adaptive immune responses against tumors. Mn-insufficient mice had significantly enhanced tumor growth and metastasis, with greatly reduced tumor-infiltrating CD8+ T cells. Mechanically, Mn2+ promoted DC and macrophage maturation and tumor-specific antigen presentation, augmented CD8+ T cell differentiation, activation and NK cell activation, and increased memory CD8+ T cells. Combining Mn2+ with immune checkpoint inhibition synergistically boosted antitumor efficacies and reduced the anti-PD-1 antibody dosage required in mice. Importantly, a completed phase 1 clinical trial with the combined regimen of Mn2+ and anti-PD-1 antibody showed promising efficacy, exhibiting type I IFN induction, manageable safety and revived responses to immunotherapy in most patients with advanced metastatic solid tumors. We propose that this combination strategy warrants further clinical translation.

Efficacy and biomarker analysis of nivolumab plus gemcitabine and cisplatin in patients with unresectable or metastatic biliary tract cancers: results from a phase II study.

BACKGROUND: The prognosis of patients with unresectable or metastatic biliary tract cancer (BTC) is unacceptably low. This study aimed to determine the efficacy, safety and predictive biomarkers of the immune checkpoint inhibitor nivolumab in combination with chemotherapy in advanced BTCs. METHODS: In this open-label, single-arm, phase II trial, a chemotherapy and immunotherapy combination consisting of gemcitabine 1000 mg/m2, cisplatin 75 mg/m2 and nivolumab 3 mg/kg was administered every 3 weeks for up to six cycles. Maintenance treatment with gemcitabine plus nivolumab was administered to patients achieving disease control following the combination therapy. The primary outcome was the objective response rate. Secondary outcomes included safety, disease control rate (DCR), progression-free survival (PFS) and overall survival (OS). The exploratory objective was to assess biomarkers for predicting clinical response and prognosis. RESULTS: Thirty-two patients with a median age of 60 (range 27-69) years were enrolled. As of September 31, 2019, the median follow-up was 12.8 (95% CI 10.8 to 14.8) months. Twenty-seven response-evaluable patients received a median of 4 (IQR, 3-6) cycles of combination therapy, of whom 15 (55.6%) patients achieved an objective response, including 5 (18.6%) with a complete response (CR), and the DCR was 92.6%. Of the six patients in cohort A who were resistant to gemcitabine-based or cisplatin-based chemotherapy, one achieved CR and one achieved partial response. Thirteen of 21 chemotherapy-naive patients (61.9%) in cohort B achieved an objective response. The median PFS of all patients in cohorts A+B was 6.1 months. The median OS was 8.5 months, with a 33.3% 12-month OS rate. The most frequent grade 3 or higher adverse events were thrombocytopenia (56%) and neutropenia (22%). Fitness might be a biomarker for predicting clinical response. On-therapy changes in serum soluble FasL, MCP-1 and interferon-γ were correlated with prognosis. CONCLUSIONS: Nivolumab in combination with gemcitabine and cisplatin offers promising efficacy and a manageable safety profile for patients with advanced BTCs. TRIAL REGISTRATION NUMBER: NCT03311789.