Multi-dimensional mismatch and barriers for promoting PrEP among men who have sex with men in China: a cross sectional survey from the Demand-side.

BACKGROUND: Men who have sex with men (MSM) is a key population for preventing HIV in China, yet pre-exposure prophylaxis (PrEP) is not widely accepted in this population. The objective of this manuscript was to assessed the barriers in the acknowledgement and uptake focusing the demand side. METHODS: An online questionnaire survey was conducted from December 2018 to January 2019. All participants were required to scan two-dimensional code which was the online crowdsourcing survey platform to complete the electronic questionnaire anonymously. RESULTS: Among 1915 MSM from thirty-four cities of China, 512 (26.7%) versus 1617 (84.4%) had an objective or subjective need of PrEP, respectively. One hundred and six (5.5%) reported affordability and only 23 (1.2%) had ever taken it. Age, living alone and occupation were associated with the objective needs. Age, income, sexual behavior were associated with actual usage. The participants who they had objective need to use PrEP are the population which we should focus on. CONCLUSION: A wide disconnect exists among the objective need, willingness, affordability and uptake of PrEP. Cost was the most prevalent barrier, accounting for 78.22% of individuals who needed and wished for PrEP but finally failed to receive it. The findings might facilitate optimizing future allocation of resources to better promote PrEP in Chinese MSM.

Correction to: Identification of miRNA-mRNA crosstalk in CD4+ T cells during HIV-1 infection by integrating transcriptome analyses.

An amendment to this paper has been published and can be accessed via the original article.

MicroRNA miR-126-5p Enhances the Inflammatory Responses of Monocytes to Lipopolysaccharide Stimulation by Suppressing Cylindromatosis in Chronic HIV-1 Infection.

Persistent immune activation during chronic human immunodeficiency virus type 1 (HIV-1) infection facilitates immune dysfunction and thereby fuels disease progression. The translocation of bacterial derivatives into blood and the hyperinflammatory responsiveness of monocytes have been considered important causative factors for persistent immune activation. Whether microRNAs (miRNAs) are involved in regulating monocyte-mediated inflammatory responses during chronic HIV-1 infection remains elusive. In this study, we show that miR-126-5p functions as a positive regulator of monocyte-mediated inflammatory responses. Significantly increased miRNA miR-126-5p and decreased cylindromatosis (CYLD) were observed in primary monocytes from chronic HIV-1 patients. Inhibition of miR-126-5p in monocytes from chronic HIV-1 patients attenuated the responsiveness of these cells to lipopolysaccharide (LPS) stimulation. Gain-of-function assays confirmed that miR-126-5p could downregulate CYLD, which in turn caused an upregulation of phosphorylation of JNK protein (pJNK) and enhanced inflammatory responses of monocytes to LPS stimulation. Overall, miR-126-5p upregulates the responsiveness of monocytes to LPS stimulation in chronic HIV-1 infection, and the suppression of miR-126-5p and the promotion of CYLD expression in primary monocytes may represent a practical immune intervention strategy to contain persistent inflammation in chronic HIV-1 infection.IMPORTANCE Monocyte-mediated hyperinflammatory responses during chronic HIV-1 infection are important causative factors driving AIDS progression; however, the underlying mechanism has not been fully addressed. We demonstrated that miR-126-5p, one of the most upregulated miRNAs during chronic HIV-1 infection, could enhance the inflammatory responses of monocytes to LPS by suppressing the inhibitory protein CYLD and thereby unleashing the expression of pJNK in the LPS/Toll-like receptor 4/mitogen-activated protein kinase pathway. This observation reveals a new mechanism for HIV-1 pathogenesis, which could be targeted by immune intervention.

Identification of miRNA-mRNA crosstalk in CD4+ T cells during HIV-1 infection by integrating transcriptome analyses.

BACKGROUND: HIV-1-infected long-term nonprogressors (LTNPs) are characterized by infection with HIV-1 more than 7-10 years, but keeping high CD4+ T cell counts and low viral load in the absence of antiretroviral treatment, while loss of CD4+ T cells and high viral load were observed in the most of HIV-1-infected individuals with chronic progressors (CPs) However, the mechanisms of different clinical outcomes in HIV-1 infection needs to be further resolved. METHODS: To identify microRNAs (miRNAs) and their target genes related to distinct clinical outcomes in HIV-1 infection, we performed the integrative transcriptome analyses in two series GSE24022 and GSE6740 by GEO2R, R, TargetScan, miRDB, and Cytoscape softwares. The functional pathways of these differentially expressed miRNAs (DEMs) targeting genes were further analyzed with DAVID. RESULTS: We identified that 7 and 19 DEMs in CD4+ T cells of LTNPs and CPs, respectively, compared with uninfected controls (UCs), but only miR-630 was higher in CPs than that in LTNPs. Further, 478 and 799 differentially expressed genes (DEGs) were identified in the group of LTNPs and CPs, respectively, compared with UCs. Compared to CPs, four hundred and twenty-four DEGs were identified in LTNPs. Functional pathway analyses revealed that a close connection with miRNA-mRNA in HIV-1 infection that DEGs were involved in response to virus and immune system process, and RIG-I-like receptor signaling pathway, whose DEMs or DEGs will be novel biomarkers for prediction of clinical outcomes and therapeutic targets for HIV-1. CONCLUSIONS: Integrative transcriptome analyses showed that distinct transcriptional profiles in CD4+ T cells are associated with different clinical outcomes during HIV-1 infection, and we identified a circulating miR-630 with potential to predict disease progression, which is necessary to further confirm our findings in the future.

Morphologic classification of the right auricule on 256-slice computed tomography.

PURPOSE: To investigate the shape of right auricule on 256-slice computed tomography (CT). MATERIALS AND METHODS: Five hundred people (250 men, age range 16-84 years) who had cardiac multidetector CT angiography were recruited in this study. All patients had normal sinus rhythm with normal blood pressure (<140/90 mmHg for systolic/diastolic pressure). The morphology of the right auricule was studied and compared after reconstruction of the raw images. RESULTS: All patients successfully had cardiac CT angiography (100%), and the right auricule morphology was divided into five types and nine subtypes, including Type I of triangular shape (Ia and Ib), Type II of M shape (IIa and IIb), Type III of L shape (IIIa and IIIb), Type IV of reverse L shape (IVa and IVb), and Type V of balanced shape. The most common type of right auricule is Type IV (28.4%) followed by Type II (24.0%), whereas the least common is Type V (11.0%). Type Ia was present significantly (P < 0.0001) more frequently in females than in males, whereas Type IIa significantly (P = 0.042) more frequently in males than females. No other significant (P > 0.05) sex difference existed in the constitution ratio of the types. The normal angle was greater in Type Ib than in Ia. The greater the normal angle in Type I, the greater the deviation of the right auricule tip towards the left. CONCLUSION: A good understanding of the right auricule anatomical morphology can better guide atrial pacing, radiofrequency ablation and other surgical procedures while preventing possible intra-procedural complications.

[Preparation and purification of polyclonal antibody against Toxoplasma gondii rhoptry protein 2 (ROP2) and its application in immunofluorescence localization].

OBJECTIVE: To prepare and purify polyclonal antibody against Toxoplasma gondii rhoptry protein 2 (ROP2) and apply it to immunofluorescence localization. METHODS: The constructed recombinant plasmid pET32a-ROP2 was transformed into E. coli BL21 (DE3) and the protein was expressed under the condition of 0.5 mmol/L IPTG induction. Cells were lysed by multiple rounds of sonication to obtain supernatant and inclusion body, respectively. The washed inclusion bodies were dissolved in urea. New Zealand White rabbits were immunized with 200 microg purified recombinant ROP2 mixing with the same volume of Freund's adjuvant for 3 times at interval of 14 days and 19 days, respectively. Rabbit serum was collected at 10 days after the last immunization. Polyclonal antibody in rabbit serum was purified with HiTrap Protein G HP affinity purification column. Indirect ELISA and Western blotting were used to detect antibody titer and specificity of polyclonal antibody against the recombinant ROP2. The polyclonal antibody was used to the localization of ROP2 on the parasitophorous vacuole membrane in human foreskin fibroblasts infected by Toxoplasma tachyzoites by the immunofluorescence method. RESULTS: The recombinant ROP2 protein was obtained and specific rabbit-derived polyclonal antibody was prepared. Indirect ELISA confirmed that the rabbit-derived polyclonal antibody titer reached 1:102400, and the recombinant ROP2 protein was recognized by specific polyclonal antibody. Immunofluorescence localization test showed that the ROP2 protein was located on the parasitophorous vacuole membrane. CONCLUSION: The rabbit-derived polyclonal antibody against ROP2 is prepared, and used in immunofluorescence localization of ROP2 on parasitophorous vacuole membrane.

Generation and Functional Verification of Hypoxia-sensitive Chimeric Antigen Receptor-T Cells.

Extensive studies have proven the promise of chimeric antigen receptor T (CAR-T) cell therapy in treating hematological malignancies. However, treating solid tumors remains challenging, as exemplified by the safety concerns that arise when CAR-T cells attack normal cells expressing the target antigens. Researchers have explored various approaches to enhance the tumor selectivity of CAR-T cell therapy. One representative strategy along this line is the construction of hypoxia-sensitive CAR-T cells, which are designed by fusing an oxygen-dependent degradation domain to the CAR moiety and are strategized to attain high CAR expression only in a hypoxic environment-the tumor microenvironment (TME). This paper presents a protocol for the generation of such CAR-T cells and their functional characterization, including methods to analyze the changes in CAR expression and killing capacity in response to different oxygen levels established by a mobile incubator chamber. The constructed CAR-T cells are anticipated to demonstrate CAR expression and cytotoxicity in an oxygen-sensitive manner, thus supporting their capability to distinguish between hypoxic TME and normoxic normal tissues for selective activation.

Efficacy and safety of novel multifunctional M10 CAR-T cells in HIV-1-infected patients: a phase I, multicenter, single-arm, open-label study.

Chimeric antigen receptor T (CAR-T) cells have been proposed for HIV-1 treatment but have not yet demonstrated desirable therapeutic efficacy. Here, we report newly developed anti-HIV-1 CAR-T cells armed with endogenic broadly neutralizing antibodies (bNAbs) and the follicle-homing receptor CXCR5, termed M10 cells. M10 cells were designed to exercise three-fold biological functions, including broad cytotoxic effects on HIV-infected cells, neutralization of cell-free viruses produced after latency reversal, and B-cell follicle homing. After demonstrating the three-fold biological activities, M10 cells were administered to treat 18 HIV-1 patients via a regimen of two allogenic M10 cell infusions with an interval of 30 days, with each M10 cell infusion followed by two chidamide stimulations for HIV-1 reservoir activation. Consequently, 74.3% of M10 cell infusions resulted in significant suppression of viral rebound, with viral loads declining by an average of 67.1%, and 10 patients showed persistently reduced cell-associated HIV-1 RNA levels (average decrease of 1.15 log10) over the 150-day observation period. M10 cells were also found to impose selective pressure on the latent viral reservoir. No significant treatment-related adverse effects were observed. Overall, our study supported the potential of M10 CAR-T cells as a novel, safe, and effective therapeutic option for the functional cure of HIV-1/AIDS.

Rapid generation of a mouse model for evaluating on-target normal tissue toxicity of human CAR-T cells using replication-defective recombinant adenovirus.

INTRODUCTION: The on-target off-tumor toxicity of chimeric antigen receptor-engineered T cells (CAR-T) might lead to fatal side effects in cancer patients, which remains as a major obstacle to the clinical application of CAR-T immunotherapy. The off-tumor on-target normal tissue toxicity of CAR-T cells needs to be evaluated in preclinical studies using rational animal models. OBJECTIVES: We aim to develop a rational animal model for assessing the off-tumor on-target normal tissue toxicity of various CAR-T cell designs quickly. METHODS: We used a recombinant adenovirus type 5 carrying human HER2/ERBB2 (Ad5-HER2) or CD47 gene (Ad5-CD47) to rapidly generate a mouse model with tunable human antigen expression on normal liver tissue to determine immunotoxicity of traditional CAR-T and hypoxia-response CAR-T cells in vivo. RESULTS: The obvious liver damage and lymphocyte infiltration were not observed in mice with human antigen-high livers 8 days post-infection. Interestingly, the lethal liver damage, systemic cytokine release and CAR-T cells infiltration in liver were only observed in mice that received traditional CAR-T cells, but not in hypoxia-response CAR-T cells. CONCLUSION: Adenovirus-based expression of target antigen in normal mouse tissue may be a useful method for assessing on-target CAR-T cell toxicity in normal tissues, especially various CAR-T cell designs that have the potency of conditional regulation in tumor microenvironment (TME).

Epicardial fat volume evaluated with multidetector computed tomography and other risk factors for prevalence of three-vessel coronary lesions.

PURPOSE: To retrospectively investigate the epicardial fat volume with multidetector computed tomography (MDCT) and other risk factors for the prevalence of three-vessel coronary lesion. MATERIALS AND METHODS: MDCT was performed on 424 subjects with or without three-vessel coronary lesion. Blood was tested for triglyceride, high-density lipoprotein (HDL), low-density lipoprotein (LDL), apolipoprotein A (ApoA), apolipoprotein B (ApoB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), lipoprotein a, and fasting blood glucose. RESULTS: Among all the subjects, a significant (P < 0.05) negative linear correlation existed between age and ALT or ALT/AST. The epicardial fat had a significant (P < 0.05) negative linear correlation with HDL and Apo A but a positive correlation with age and ApoB/ApoA. The epicardial fat volume and the fasting blood glucose were significantly (P = 0.001) greater in the patients than in the control group, whereas HDL and Apo A were both significantly (P < 0.0001) smaller in the patients than in the control groups. A significant prediction value (P < 0.05) existed in age increase, male gender, epicardial fat increase, low HDL, high LDL, and elevated fasting blood glucose. CONCLUSION: Three-vessel coronary lesions are more prevalent in subjects with greater volume of epicardial fat and in male gender.

Pathologically complete remission to combination of invariant NK T cells and anti-CD20 antibody in a refractory HIV+ diffuse large B-cell lymphoma patient.

Although there is a high curability rate with rituximab chemotherapy, approximately 40% of patients with diffuse large B-cell lymphoma (DLBCL) develop disease relapse or primary-refractory lymphoma. The prognosis of HIV+ DLBCL patients is even worse with limited therapeutic options. The case is presented of a 28-year-old man who was diagnosed with HIV-DLBCL, refractory to rituximab-based chemo-immunotherapies and radiotherapy before and maintained a pathologically complete regression with the infusion of haplotype-matched invariant NK T cells and anti-CD20 antibody. His abdominal mass kept shrinking during the period of follow-up without relapse to date. A combination of haplotype-matched invariant NK T cells was likely to reinvigorate the efficacy of anti-CD20 antibody and may offer a viable treatment option for refractory DLBCL patients.

Diffuse large B-cell lymphoma (DLBCL) is one of the most frequent types of lymphoid cancer. The prognosis of relapsed/refractory DLBCL patients has been dismal with resistance to rituximab-based chemo-immunotherapy. The combination of effective cells with rituximab might improve clinical response by enhancing antibody-dependent cytotoxicity. The authors reported the case of a 28-year-old man diagnosed with HIV-DLBCL, refractory to 3 lines of rituximab-based chemo-immunotherapies and radiotherapy before, and who received 6 experimental infusions of haplotype-matched invariant NK T cells combined with anti-CD20 antibody. The lesion on radiological examination was partially regressed after cycle 3 and cycle 6. The pathological examination, performed 4 weeks after cycle 6, suggested pathologically complete regression. The mass was completely regressed on CT scanning without relapse to date. A combination of invariant NK T cells and anti-CD20 antibody may offer a viable treatment option for relapsed/refractory DLBCL patients for whom rituximab chemotherapy was not effective.

Long-term sequelae of different COVID-19 variants: The original strain versus the Omicron variant.

Although Omicron appears to cause less severe acute illness than the original strain, the potential for large numbers of patients to experience long COVID is a major concern. Little is known about the recovery phase in cases of Omicron, highlighting the importance of dynamically monitor long COVID in those patients. Subjects of the current study were patients available for a three-month follow-up who were admitted from January 13 to May 22, 2020 (period of the original strain) and from January 1 to May 30, 2022 (period of Omicron). Twenty-eight-point-four percent of patients infected with the original strain had long-term symptoms of COVID-19 and 5.63% of those infected with the Omicron strain had such symptoms. The most common symptom was a cough (18.5%), followed by tightness in the chest (6.5%), in patients infected with the original strain. Fatigue (2.4%) and dyspnea (1.7%) were the most commonly reported symptoms in patients infected with the Omicron strain. The respiratory system is the primary target of SARSCoV-2. Supportive treatment is the basis for the treatment of respiratory symptoms in patients with COVID-19. Quality sleep and good nutrition may alleviate fatigue and mental issues. Further knowledge about a long-term syndrome due to Omicron needs to be discussed and assembled so that healthcare and workforce planners can rapidly obtain information to appropriately allocate resources.

Freeze-Drying Formulations Increased the Adenovirus and Poxvirus Vaccine Storage Times and Antigen Stabilities.

Successful vaccines induce specific immune responses and protect against various viral and bacterial infections. Noninactivated vaccines, especially viral vector vaccines such as adenovirus and poxvirus vaccines, dominate the vaccine market because their viral particles are able to replicate and proliferate in vivo and produce lasting immunity in a manner similar to natural infection. One challenge of human and livestock vaccination is vaccine stability related to the antigenicity and infectivity. Freeze-drying is the typical method to maintain virus vaccine stability, while cold chain transportation is required for temperatures about 2 °C-8 °C. The financial and technological resource requirements hinder vaccine distribution in underdeveloped areas. In this study, we developed a freeze-drying formula consisting of bovine serum albumin (BSA), L-glutamic acid (L-Glu), polyethylene glycol (PEG), and dextran (DEX) to improve the thermal stability and activity of viral vaccines, including vaccinia recombinant vaccine (rTTV-OVA) and adenovirus vaccine (Ad5-ENV). We compared a panel of five different formulations (PEG: DEX: BSA: L-GLU = 50:9:0:0(#1), 50:5:4:0(#2), 50:10:9:0(#3), 50:0:0:9(#4), and 50:1:0:8(#5), respectively) and optimized the freeze-drying formula for rTTV-OVA and Ad5-ENV. We found that the freeze-drying formulations #2 and #3 could maintain rTTV-OVA infectivity at temperatures of 4 °C and 25 °C and that rTTV-OVA immunogenicity was retained during lyophilization. However, formulations #4 and #5 maintained Ad5-ENV infectivity under the same conditions, and Ad5-ENV immunogenicity had maximum retention with freeze-drying formulation #4. In summary, we developed new freeze-drying formulations that increased virus vaccine storage times and retained immunogenicity at an ambient temperature.

IL-21 arming potentiates the anti-tumor activity of an oncolytic vaccinia virus in monotherapy and combination therapy.

BACKGROUND: Oncolytic viruses (OVs) have shown promise in containing cancer progression in both animal models and clinical trials. How to further improve the efficacy of OVs are intensively explored. Arming OVs with immunoregulatory molecules has emerged as an important means to enhance their oncolytic activities majorly based on the mechanism of reverting the immunosuppressive nature of tumor environment. In this study, we aimed to identify the optimal combination of different OVs and immunomodulatory molecules for solid tumor treatment as well as the underlying mechanism, and subsequently evaluated its potential synergy with other immunotherapies. METHODS: Panels of oncolytic viruses and cells stably expressing immunoregulatory molecules were separately evaluated for treating solid tumors in mouse model. A tumor-targeted replicating vaccinia virus Tian Tan strain with deletion of TK gene (TTVΔTK) was armed rationally with IL-21 to create rTTVΔTK-IL21 through recombination. CAR-T cells and iNKT cells were generated from human peripheral blood mononuclear cells. The impact of rTTVΔTK-IL21 on tumor-infiltrating lymphocytes was assessed by flow cytometry, and its therapeutic efficacy as monotherapy or in combination with CAR-T and iNKT therapy was assessed in mouse tumor models. RESULTS: IL-21 and TTV was respectively identified as most potent immunomodulatory molecule and oncolytic virus for solid tumor suppression in mouse models. A novel recombinant oncolytic virus that resulted from their combination, namely rTTVΔTK-mIL21, led to significant tumor regression in mice, even for noninjected distant tumor. Mechanistically, rTTV∆TK-mIL21 induced a selective enrichment of immune effector cells over Treg cells and engage a systemic response of therapeutic effect. Moreover, its human form showed a notable synergy with CAR-T or iNKT therapy for tumor treatment when coupled in humanized mice. CONCLUSION: With a strong potency of shaping tumor microenvironment toward favoring TIL activities, rTTVΔTK-IL21 represents a new opportunity worthy of further exploration in clinical settings for solid tumor control, particularly in combinatorial strategies with other immunotherapies. ONE SENTENCE SUMMARY: IL21-armed recombinant oncolytic vaccinia virus has potent anti-tumor activities as monotherapy and in combination with other immunotherapies.

Conditioned CAR-T cells by hypoxia-inducible transcription amplification (HiTA) system significantly enhances systemic safety and retains antitumor efficacy.

BACKGROUND: Hypoxia is a striking feature of most solid tumors and could be used to discriminate tumors from normoxic tissues. Therefore, the design of hypoxia-conditioned Chimeric Antigen Receptor (CAR) T cells is a promising strategy to reduce on-target off-tumor toxicity in adoptive cell therapy. However, existing hypoxia-conditioned CAR-T designs have been only partially successful in enhancing safety profile but accompanied with reduced cytotoxic efficacy. Our goal is to further improve safety profile with retained excellent antitumor efficacy. METHODS: In this study, we designed and constructed a hypoxia-inducible transcription amplification system (HiTA-system) to control the expression of CAR in T (HiTA-CAR-T) cells. CAR expression was determined by Flow cytometry, and the activation and cytotoxicity of HiTA-CAR-T cells in vitro were evaluated in response to antigenic stimulations under hypoxic or normoxic conditions. The safety of HiTA-CAR-T cells was profiled in a mouse model for its on-target toxicity to normal liver and other tissues, and antitumor efficacy in vivo was monitored in murine xenograft models. RESULTS: Our results showed that HiTA-CAR-T cells are highly restricted to hypoxia for their CAR expression, activation and cytotoxicity to tumor cells in vitro. In a mouse model in vivo, HiTA-CAR-T cells targeting Her2 antigen showed undetectable CAR expression in all different normoxic tissues including human Her2-expresing liver, accordingly, no liver and systemic toxicity were observed; In contrast, regular CAR-T cells targeting Her2 displayed significant toxicity on human Her2-expression liver. Importantly, HiTA-CAR-T cells were able to achieve significant tumor suppression in murine xenograft models. CONCLUSION: Our HiTA system showed a remarkable improvement in hypoxia-restricted transgene expression in comparison with currently available systems. HiTA-CAR-T cells presented significant antitumor activities in absence of any significant liver or systemic toxicity in vivo. This approach could be also applied to design CAR-T cell targeting other tumor antigens.

Engineering T cells with hypoxia-inducible chimeric antigen receptor (HiCAR) for selective tumor killing.

Chimeric antigen receptor-modified T cells (CAR-T cells) have shown good effects in the treatment of hematologic cancers; however, they may cause on-target off-tumor toxicity because of minimal expression of tumor-associated antigens (TAAs) on normal tissues, particularly in the context of treating solid tumors. Hypoxia is a common hallmark of solid tumors because of the Warburg effect. To minimize side effects, we designed a hypoxia-inducible CAR (HiCAR), which is driven by a hypoxia response element (HRE), and consists of a conventional CAR and an oxygen-dependent degradation domain (ODD) that is actively degraded under normoxia but stabilized under hypoxia. HiCAR-T cells showed enhanced cytotoxicity against tumor cells under hypoxia compared to normoxia in vitro and antitumor efficacy comparable to that of conventional CAR-T cells in vivo. Overall, our study demonstrates the potential of the HiCAR for improving the safety of CAR-T cells to promote the clinical application of CAR-T immunotherapy.

PD-L1 chimeric costimulatory receptor improves the efficacy of CAR-T cells for PD-L1-positive solid tumors and reduces toxicity in vivo.

BACKGROUND: On-target off-tumor toxicity impedes the clinical application of chimeric antigen receptor-modified T cells (CAR-T cells) in the treatment of solid tumors. Previous reports proved that the combinatorial antigen recognition strategy could improve the safety profile of CAR-T cells by targeting two different tumor-associated antigens (TAAs), one as a CAR-T targeted antigen and the other as a chimeric costimulatory receptor (CCR) ligand. The programmed death-ligand 1 (PD-L1, also known as B7-H1) is preferentially overexpressed on multiple tumors, it will be highly interesting to explore the potential of PD-L1 as a universal target for designing CCR. METHODS: A novel dual-targeted CAR, which is composed of first-generation CD19/HER2 CAR with CD3ζ signaling domain and PD-L1 CCR containing the CD28 costimulatory domain, was constructed and delivered into T cells by pseudotyped lentivirus. The cytokine release, cytotoxicity and proliferation of dual-targeted CAR-T cells were tested in vitro, and their safety and therapeutic efficacy were evaluated using a human tumor xenograft mouse model in vivo. RESULTS: The dual-targeted CAR-T cells exerted a similar cytotoxic activity against CD19/HER2+ tumor cells with or without PD-L1 in vitro, however, enhanced cytokine releases and improved proliferative capacity were only observed in the presence of both CD19/HER2 and PD-L1. Importantly, the dual-targeted CAR-T cells displayed no cytotoxicity against PD-L1+ cells alone in the absence of tumor antigen CD19/HER2. In addition, the dual-targeted CAR-T cells preferably destroyed tumor xenografts bearing both CD19/HER2 and PD-L1, but spared only antigen-positive tumor xenografts without PD-L1 in vivo. Furthermore, PD-L1 CCR also improved the antitumor efficacy of the low-affinity HER2 CAR-T cells against PD-L1+ tumors expressing high levels of HER2. CONCLUSION: Our observations demonstrated that PD-L1 could be used as a universal target antigen for designing CCR, and the dual-targeted CAR-T cells equipped with PD-L1 CCR could be used to reduce the risk of on-target off-tumor toxicity while retaining their potent antitumor efficacy in the treatment of PD-L1+ solid tumors.

TSC1 and DEPDC5 regulate HIV-1 latency through the mTOR signaling pathway.

The latent reservoir of HIV-1 presents a major barrier to viral eradication. The mechanism of the establishment and maintenance of the latent viral reservoir is not yet fully understood, which hinders the development of effective curative strategies. In this study, we identified two inhibitory genes, TSC1 and DEPDC5, that maintained HIV-1 latency by suppressing the mTORC1 pathway. We first adapted a genome-wide CRISPR screening approach to identify host factors required for HIV latency in a T-cell-based latency model and discovered two inhibitory genes, TSC1 and DEPDC5, which are potentially involved in HIV-1 latency. Knockout of either TSC1 or DEPDC5 led to enhanced HIV-1 reactivation in both a T-cell line (C11) and a monocyte cell line (U1), and this enhancement could be antagonized by the mTORC1 inhibitor rapamycin. Further evaluation of the mechanism revealed that TSC1 suppresses AKT-mTORC1-S6 via downregulation of Rheb, whereas DEPDC5 inhibits AKT-mTORC1-S6 through RagA. Overall, both TSC1 and DEPDC5 negatively regulate the AKT-mTORC1 pathway, and thus their agonists could be used in the development of new therapeutic approaches for activating HIV-1 latency.

Spatiotemporal Analysis of the Malaria Epidemic in Mainland China, 2004-2014.

The purpose of this study is to characterize spatiotemporal heterogeneities in malaria distribution at a provincial level and investigate the association between malaria incidence and climate factors from 2004 to 2014 in China to inform current malaria control efforts. National malaria incidence peaked (4.6/100,000) in 2006 and decreased to a very low level (0.21/100,000) in 2014, and the proportion of imported cases increased from 16.2% in 2004 to 98.2% in 2014. Statistical analyses of global and local spatial autocorrelations and purely spatial scan statistics revealed that malaria was localized in Hainan, Anhui, and Yunnan during 2004-2009 and then gradually shifted and clustered in Yunnan after 2010. Purely temporal clusters shortened to less than 5 months during 2012-2014. The two most likely clusters detected using spatiotemporal analysis occurred in Anhui between July 2005 and November 2007 and Yunnan between January 2010 and June 2012. Correlation coefficients for the association between malaria incidence and climate factors sharply decreased after 2010, and there were zero-month lag effects for climate factors during 2010-2014. Overall, the spatiotemporal distribution of malaria in China changed from relatively scattered (2004-2009) to relatively clustered (2010-2014). As the proportion of imported cases increased, the effect of climate factors on malaria incidence has gradually become weaker since 2011. Therefore, new warning systems should be applied to monitor resurgence and outbreaks of malaria in mainland China, and quarantine at borders should be reinforced to control the increasingly trend of imported malaria cases.

Potential Role of Circulating microRNA-21 for Hepatocellular Carcinoma Diagnosis: A Meta-Analysis.

BACKGROUND: Circulating microRNA-21 (miR-21) is known to be aberrantly expressed in hepatocellular carcinoma (HCC) patients, and this implies that microRNA-21 is a promising and novel indicator of HCC. However, a systematic evaluation of the performance of microRNA-21 as a diagnostic marker for HCC has yet to be conducted. Therefore, the test performance of circulating miR-21 for HCC was assessed in this study. METHODS: Three common international databases and a Chinese electronic database were used to search for literature on the diagnostic accuracy of microRNA-21 for HCC. The pooled results included the sensitivity and specificity of microRNA-21 for HCC detection and were analyzed with a random effect model. The area under summary receiver operating characteristic curve (AUC) was used to estimate overall test performance. RESULTS: A total of 339 HCC patients and 338 controls without HCC from four published studies were eligible for the meta-analysis and included in our study. The test performance of circulating miR-21 in HCC detection was assessed with the summary estimates of sensitivity and specificity, which were 81.2% (95% CI: 70.8% to 88.4%) and 84.8% (95% CI: 75.1% to 91.2%), respectively. The value of AUC was 0.90 (95% CI: 0.87 to 0.92). Significant inter-study heterogeneity was detected by our analysis, and sub-group analyses suggested that the type of control group was probably a source of heterogeneity. CONCLUSIONS: Our current findings suggested that circulating miR-21 can serve as a potential co-biomarker for early-stage HCC diagnosis. Thorough large-scale studies are needed to confirm the generalizability of our findings.