Tumor-targeted gypenoside nanodrug delivery system with double protective layers.

OBJECTIVES: Gypenoside (Gyp) is easily degraded in the gastrointestinal tract, resulting in its low bioavailability. We aimed to develop a tumor-targeted Gyp nanodrug delivery system and to investigate its antitumor effect in vitro. MATERIALS AND METHODS: We used Gyp as the therapeutic drug molecule, mesoporous silica (MSN) and liposome (Lipo) as the drug carrier and protective layers, and aptamer SYL3C as the targeting element to establish a tumor-targeted nanodrug delivery system (i.e., SYL3C-Lipo@Gyp-MSN). The characteristics of SYL3C-Lipo@Gyp-MSN were investigated, and its drug release performance, cell uptake, and antitumor activity in vitro were evaluated. RESULTS: A tumor-targeted Gyp nanodrug delivery system was successfully prepared. The SYL3C-Lipo@Gyp-MSN was spherical or ellipsoidal; had good dispersion, which enabled it to specifically target and kill the liver tumor cell HepG2; and effectively protected the early leakage of Gyp. CONCLUSIONS: We have established a tumor-targeted nanodrug delivery system that can target and kill liver cancer cells and may provide a strategy for preparing new nanodrug-loaded preparations of traditional Chinese medicine.

Visual analysis of hotspots and trends in long COVID research based on bibliometric.

After severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, a series of symptoms may persist for a long time, which is now called long COVID. It was found that long COVID can affect all patients with COVID-19. Therefore, long COVID has become a hot topic. In this study, we used the WOS database as a sample data source to conduct a bibliometric and visual analysis of 1765 long COVID articles over the past three years through VOSviewer and R package. The results show that countries/authors in Europe and The United States of America contribute most of the articles, and their cooperation is also the most active. Keyword co-occurrence identified four clusters, with important topics including the mechanism, clinical symptoms, epidemiological characteristics, and management/treatment of long COVID. Themes such as "cognitive impairment", "endothelial dysfunction", "diagnosis", and "biomarkers" are likely to be the focus of new attention in the coming period. In addition, we put forward the possible research opportunities on long COVID for researchers and practitioners to facilitate future research.

Generation of in situ CRISPR-mediated primary and metastatic cancer from monkey liver.

Non-human primates (NHPs) represent the most valuable animals for drug discovery. However, the current main challenge remains that the NHP has not yet been used to develop an efficient translational medicine platform simulating human diseases, such as cancer. This study generated an in situ gene-editing approach to induce efficient loss-of-function mutations of Pten and p53 genes for rapid modeling primary and metastatic liver tumors using the CRISPR/Cas9 in the adult cynomolgus monkey. Under ultrasound guidance, the CRISPR/Cas9 was injected into the cynomolgus monkey liver through the intrahepatic portal vein. The results showed that the ultrasound-guided CRISPR/Cas9 resulted in indels of the Pten and p53 genes in seven out of eight monkeys. The best mutation efficiencies for Pten and p53 were up to 74.71% and 74.68%, respectively. Furthermore, the morbidity of primary and extensively metastatic (lung, spleen, lymph nodes) hepatoma in CRISPR-treated monkeys was 87.5%. The ultrasound-guided CRISPR system could have great potential to successfully pursue the desired target genes, thereby reducing possible side effects associated with hitting non-specific off-target genes, and significantly increasing more efficiency as well as higher specificity of in situ gene editing in vivo, which holds promise as a powerful, yet feasible tool, to edit disease genes to build corresponding human disease models in adult NHPs and to greatly accelerate the discovery of new drugs and save economic costs.

An Efficient Cell-Targeting Drug Delivery System Based on Aptamer-Modified Mesoporous Silica Nanoparticles.

How to deliver chemotherapeutic drugs efficiently and selectively to tumor cells to improve therapeutic efficacy remains a difficult problem. We herein construct an efficient cell-targeting drug delivery system (Sgc8-MSN/Dox) based on aptamer-modified mesoporous silica nanoparticles that relies on the tumor-targeting ability of the aptamer Sgc8 to deliver doxorubicin (Dox) to leukemia cells in a targeted way, thereby improving therapeutic efficacy and reducing toxicity. In this work, Sgc8-MSN/Dox showed sustained Dox release, and they targeted and efficiently killed CCRF-CEM human acute T lymphocyte leukemia cells, suggesting potential as a cancer therapy.

Generation of Cancer-Specific Cytotoxic PD-1- T Cells Using Liposome-Encapsulated CRISPR/Cas System with Dendritic/Tumor Fusion Cells.

T-cell immunotherapy is showing great promise and therefore undergoing intensive developments for cancer treatment. In this study, we applied liposome-encapsulated Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated protein-9 nuclease (Cas9) (CRISPR/Cas9) genome editing tool to specifically knock out the programmed death-1 (PD-1) gene from T cells (PD-1- T cells). We then activated these cells by dendritic/tumor fusion cells (FCs) and examined their anti-cancer potential. Results showed that, following the antigen presentation and activation by DC/HepG2 FCs, PD-1- T cells showed a significantly higher ability than PD-1+ T cells to proliferate, secrete pro-inflammatory cytokine IFN-γ, and kill HepG2 cells in vitro. Consistently, in vitro activated PD-1- T cells inhibited proliferation and induced apoptosis in HepG2 xenografts in vivo, leading to significantly suppressed tumor growth and improved mouse survival. Liposome-encapsulated CRISPR/Cas9 genome editing technology effectively knocked out PD-1 gene in T cells, stimulating T cell activation in response to DC/tumor FCs and affording T cell-mediated cancer immunotherapy. Our study provides evidence to target checkpoint receptors in adoptively transfected T cells, as a novel therapeutic modality for adoptive T cell transfer.

Magnetic Endoglin Aptamer Nanoprobe for Targeted Diagnosis of Solid Tumor.

Based on molecular targeting, magnetic resonance imaging (MRI) is an ideal noninvasive approach for tumor diagnosis. Construction of targeting probes to enhance the MRI efficacy has become a research hotspot recently. In this study, magnetic endoglin aptamer (mEND) imaging nanoprobes based on mEND-modified magnetic carboxymethyl chitosan (CMCS) nanoparticles (denoted mEND-Fe3O4@CMCS) were developed. The mEND-Fe3O4@CMCS naoprobe was prepared using mEND as the recognition molecule and Fe3O4@CMCS as the carrier to enhance the MRI efficacy of hepatocellular carcinoma (HCC). On the one hand, the CMCS self-assembled on the surface of Fe3O4 improved the biocompatibility and nontoxicity of magnetic nanoparticles. On the other hand, chemical groups provided by CMCS contributed to the modification of more aptamers. More importantly, the assembled aptamers significantly improved the probe targeting ability, thus enhancing the diagnosis efficacy of MRI of HCC. As a result, the average diameter and zeta potential of the nanoprobe was 87.15±1.66 nm and -31.9±0.5 mV, respectively. The MRI imaging result indicated that this probe effectively targeted neovascularization of mouse HCC and improved the imaging contrast of subcutaneous tumor in mice. Cytotoxicity and histological tests confirmed that the constructed probe possessed low toxicity. In conclusion, the mEND-Fe3O4@CMCS nanoprobe showed high targeting affinity, enhanced MRI effect and good biocompatibility. This study provides new MRI probes to target CD105 positive cells and is a promising candidate for HCC early diagnosis.

TLS11a Aptamer/CD3 Antibody Anti-Tumor System for Liver Cancer.

New therapeutic approaches are needed for hepatocellular carcinoma (HCC), which is the most common primary malignancy of the liver. Bispecific T-cell engagers (BiTE) can effectively redirect T cells against tumors and show a strong anti-tumor effect. However, the potential immunogenicity, complexity, and high cost significantly limit their clinical application. In this paper, we used the hepatoma cells-specific aptamer TLS11a and anti-CD3 for to establish an aptamer/antibody bispecific system (AAbs), TLS11a/CD3, which showed advantages over BiTE and can specifically redirect T cells to lyse tumor cells. TLS11a-SH and anti-CD3-NH2 were crosslinked with sulfosuccinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (sulfo-SMCC). T cell activation, proliferation, and cytotoxicity of TLS11a/CD3 were analyzed by flow cytometry. Cytokine array was used to detect cytokine released from activated T cells. Hepatoma xenograft model was used to monitor the tumor volume and survival. TLS11a/CD3 could specifically bind hepatoma cells (H22) and T cells, activated T cells to mediate antigen-specific lysis of H22 cells in vitro, and effectively inhibited the growth of implanted H22 tumors as well as prolonged mice survival. TLS11a/CD3 could simultaneously target hepatoma cells and T cells, specifically guide T cells to kill tumor cells, and enhance the anti-tumor effect of T cells both in vitro and in vivo.

Correction to: A Graphene Oxide-Based Fluorescent Aptasensor for the Turn-on Detection of CCRF-CEM.

In the original publication of this article [1] the author Liping Zhong was omitted. In this correction article the author and the corresponding details are provided. The publisher apologizes to the readers and authors for the inconvenience.

A Graphene Oxide-Based Fluorescent Aptasensor for the Turn-on Detection of CCRF-CEM.

A convenient, low-cost, and highly sensitive fluorescent aptasensor for detection of leukemia has been developed based on graphene oxide-aptamer complex (GO-apt). Graphene oxide (GO) can absorb carboxyfluorescein-labeled Sgc8 aptamer (FAM-apt) by π-π stacking and quench the fluorescence through fluorescence resonance energy transfer (FRET). In the absence of Sgc8 target cell CCRF-CEM, the fluorescence is almost all quenched. Conversely, when the CCRF-CEM cells are added, the quenched fluorescence can be recovered rapidly and significantly. Therefore, based on the change of fluorescence signals, we can detect the number of CCRF-CEM cells in a wide range from 1 × 102 to 1 × 107 cells/mL with a limit of detection (LOD) of 10 cells/mL. Therefore, this strategy of graphene oxide-based fluorescent aptasensor may be promising for the detection of cancer.

Graphene-Based Multifunctional Nanomaterials in Cancer Detection and Therapeutics.

Nanotechnology for early diagnosis and treatment of malignant tumor is a forefront topic in the international field of biotechnology and medicine. In order to improve the effect of cancer therapy, the timely and accurate detection of the cancer is important and necessary. Graphene and its derivatives have various excellent characteristics. For example, biological sensors based on graphene are good at amplifying detection signals, and its derivatives play an important role in the early diagnosis and cancer therapy. In view of this, we discussed the biological sensor application based on graphene and its derivatives in the detection and therapy of cancer.

Recent Progress of Wnt Pathway Inhibitor Dickkopf-1 in Liver Cancer.

Hepatocellular carcinoma (HCC) is one of the most common cancers around the world. Multiple etiologic factors such as virus and environment can lead to HCC. It is a challenge for us to successfully detect early HCC due to the lack of effective characterized and specific biomarkers. However, if the early diagnosis is successfully realized, it provides crucial chance for HCC patients to receive effective treatment as early as possible. Dickkopf-1 (DKK-1) is a secretary glycoprotein, which negatively regulates Wnt pathway through binding to surface receptors LRP5/6 and Kremen 1/2. The expression of DKK-1 is regulated by p53, V-Myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog (MYCN), ß-catenin, etc. Ectopic expression of DKK-1 can inhibit cell proliferation, or induce apoptosis with apoptosis enhancing factors. DKK-1 is low-expressed in many tumors, but overexpressed in others. Growing evidences show that DKK-1 plays complex and different roles in tumorigenesis, tumor progression and metastasis of different cancers. We herein review the recent progress in the expression and function of DKK-1 in hepatocellular carcinoma.

Screening and antitumor effect of an anti­CTLA­4 nanobody.

Cytotoxic T­lymphocyte antigen­4 (CTLA­4) is a critical negative regulator of immune responses. CTLA­4 is rapidly upregulated following T­cell activation, and then binds to B7 molecules with a higher affinity than CD28. CTLA­4 may abolish the initiation of the responses of T cells by raising the threshold of signals required for full activation of T cells, and it also may terminate ongoing T-cell responses. This regulatory role has led to the development of monoclonal antibodies (mAbs) designed to block CTLA­4 activity for enhancing immune responses against cancer. mAbs have several disadvantages including high production cost and unstable behavior. Nanobodies (Nbs) are single­domain antigen­binding fragments derived from the camelid heavy­chain antibodies, which are highly attractive in cancer immunotherapy due to their small size, high specificity, and stability. We selected CTLA­4­specific Nbs from a high quality dromedary camel immune library by phage display technology. Four positive colonies were sequenced and classified based on the amino acids sequences in the CDR3 region. These Nbs recognized unique epitopes on CTLA­4 and displayed high binding rates when used on PHA­stimulated human T cells. Treatment of B16 melanoma­bearing C57BL/6 mice with anti­CTLA­4 nanobody 16 (Nb16) delayed melanoma growth and prolonged the survival time of mice. These data indicate that anti­CTLA­4 Nbs selected from a high quality phage display library may be effective for the treatment of patients with tumors.

Collagen I enhances the efficiency and anti-tumor activity of dendritic-tumor fusion cells.

Low fusion efficiency and nominal activity of fusion cells (FCs) restrict the clinical application of dendritic cell (DC)/tumor fusion cells. Collagen I (Col I) is an interstitial collagen with a closely-knit structure used to repair damaged cell membranes. This study evaluated whether Col I could improve the fusion efficiency of polyethylene glycol (PEG)-induction and enhance the immunogenicity of fusion vaccine. DC/B16 melanoma and controlled DC/H22 hepatoma cell fusions were induced by PEG with or without Col I. Col I/PEG treatment increased the levels of DC surface molecules and the secretion of lactate, pro- and anti-inflammatory cytokines in fusion cells. Col I/PEG-treated FCs enhanced T-cell proliferation and cytotoxic T lymphocyte activity. The Col I-prepared fusion vaccine obviously suppressed tumor growth and prolonged mice survival time. Thus Col I treatment could significantly improve the efficiency of PEG-induced DC/tumor fusion and enhance the anticancer activity of the fusion vaccine. This novel fusion strategy might promote the clinical application of DC/tumor fusion immunotherapy.

A New Theranostic System Based on Endoglin Aptamer Conjugated Fluorescent Silica Nanoparticles.

Background: Tumor vessels can potentially serve as diagnostic, prognostic and therapeutic targets for solid tumors. Fluorescent dyes are commonly used as biological indicators, while photobleaching seriously hinders their application. In this study, we aim to generate a fluorescent silica nanoparticles (FSiNPs) theranostic system marked by the mouse endgolin (mEND) aptamer, YQ26. Methods: A highly specific YQ26 was selected by using gene-modified cell line-based SELEX technique. FSiNPs were prepared via the reverse microemulsion method. The YQ26-FSiNPs theranostic system was developed by combining YQ26 with the FSiNPs for in vivo tumor imaging, treatment and monitoring. Results: Both in vitro experiments (i.e. cellular and tumor tissue targeting assays) and in vivo animal studies (i.e. in vivo imaging and antitumor efficacy of YQ26-FSiNPs) clearly demonstrated that YQ26-FSiNPs could achieve prominently high targeting efficiency and therapeutic effects via aptamer YQ26-mediated binding to endoglin (END) molecule. Conclusion: This simple, sensitive, and specific YQ26-FSiNPs theranostic system has a great potential for clinical tumor targeting imaging and treatment.

An 'activatable' aptamer-based fluorescence probe for the detection of HepG2 cells.

It is significant to develop a probe with sensitivity and specificity for the detection of cancer cells. The present study aimed to develop an 'activatable' aptamer-based fluorescence probe (AAFP) to detect cancer cells and frozen cancer tissue. This AAFP consisted of two fragments: aptamer TLS11a that targets HepG2 cells, and two short extending complementary DNA sequences with a 5'- and 3'-terminus that make the aptamer in hairpin structure a capable quencher to fluorophore. The ability of the AAFP to bind specifically to cancer cells was assessed using flow cytometry, fluorescence spectroscopy and fluorescence microscopy. Its ability to bind to frozen cancer tissue was assessed using fluorescence microscopy. As a result, in the absence of cancer cells, AAFP showed minimal fluorescence, reflecting auto-quenching. In the presence of cancer cells, however, AAFP showed a strong fluorescent signal. Therefore, this AAFP may be a promising tool for sensitive and specific detection of cancer.

Aptamer Combined with Fluorescent Silica Nanoparticles for Detection of Hepatoma Cells.

PURPOSE: The purpose of this study is to develop a simple, effective method to label hepatoma cells with aptamers and then detect them using fluorescent silica nanoparticles (FSNPs). METHOD: Streptavidin was conjugated to carboxyl-modified fluorescein isothiocyanate (FITC)-doped silica nanoparticles which were prepared by the reverse microemulsion method. The resulting streptavidin-conjugated fluorescent silica nanoparticles (SA-FSNPs) were mixed with hepatoma cells that had been labeled with biotin-conjugated aptamer TLS11a (Bio-TLS11a). The specificity and sensitivity of the nanoprobes were assessed using flow cytometry and fluorescence microscopy. Their toxicity was assessed in normal human liver cell cultures using the MTT assay, as well as in nude mice using immunohistochemistry. RESULTS: SA-FSNPs showed uniform size and shape, and fluorescence properties of them was similar to the free FITC dye. SA-FSNPs were able to detect aptamer-labeled hepatoma cells with excellent specificity and good sensitivity, and they emitted strong, photobleach-resistant fluorescent signal. SA-FSNPs showed no significant toxic effects in vitro or in vivo. CONCLUSION: The combination of biotin-conjugated aptamers and SA-FSNPs shows promise for sensitive detection of hepatoma cells, and potentially of other tumor cell types as well.

Aptamer-Functionalized Fluorescent Silica Nanoparticles for Highly Sensitive Detection of Leukemia Cells.

A simple, highly sensitive method to detect leukemia cells has been developed based on aptamer-modified fluorescent silica nanoparticles (FSNPs). In this strategy, the amine-labeled Sgc8 aptamer was conjugated to carboxyl-modified FSNPs via amide coupling between amino and carboxyl groups. Sensitivity and specificity of Sgc8-FSNPs were assessed using flow cytometry and fluorescence microscopy. These results showed that Sgc8-FSNPs detected leukemia cells with high sensitivity and specificity. Aptamer-modified FSNPs hold promise for sensitive and specific detection of leukemia cells. Changing the aptamer may allow the FSNPs to detect other types of cancer cells.