Sensitive aptasensing of ATP based on a PAM site-regulated CRISPR/Cas12a activation.

The combination of CRISPR/Cas12a and functional DNA provides the possibility of constructing biosensors for detecting non-nucleic-acid targets. In the current study, the duplex protospacer adjacent motif (PAM) in the activator of CRISPR/Cas12a was used as a molecular switch, and a sensitive adenosine triphosphate (ATP) detection biosensor was constructed using an allosteric probe-conjugated PAM site formation in hybridization chain reaction (HCR) integrated with the CRISPR/Cas12a system (APF-CRISPR). In the absence of ATP, an aptamer-containing probe (AP) is in a stem-loop structure, which blocks the initiation of HCR. In the presence of ATP, the structure of AP is changed upon ATP binding, resulting in the release of the HCR trigger strand and the production of long duplex DNA with many PAM sites. Since the presence of a duplex PAM site is crucial for triggering the cleavage activity of CRISPR/Cas12a, the ATP-dependent formation of the PAM site in HCR products can initiate the FQ-reporter cleavage, allowing ATP quantification by measuring the fluorescent signals. By optimizing the sequence elements and detection conditions, the aptasensor demonstrated superior detection performance. The limit of detection (LOD) of the assay was estimated to be 1.16 nM, where the standard deviation of the blank was calculated based on six repeated measurements. The dynamic range of the detection was 25-750 nM, and the whole workflow of the assay was approximately 60 min. In addition, the reliability and practicability of the aptasensor were validated by comparing it with a commercially available chemiluminescence kit for ATP detection in serum. Due to its high sensitivity, specificity, and reliable performance, the APF-CRISPR holds great potential in bioanalytical studies for ATP detection. In addition, we have provided a proof-of-principle for constructing a CRISPR/Cas12a-based aptasensor, in which the PAM is utilized to regulate Cas12a cleavage activity.

Glycyrrhizae radix et rhizoma processing ameliorates adverse reactions of polygalae radix in zebra fish and rabbit models.

ETHNOPHARMACOLOGICAL RELEVANCE: Polygala tenuifilia Willd (Polygalaceae), a traditional Chinese medicine, has been used for a long time to treat various illnesses with serious adverse reactions. Glycyrrhizae radix et rhizoma processing is generally used to reduce the adverse reactions. AIM OF THE STUDY: The aim of this study was to validate the irritation caused by raw Polygalaceae (RPA), to investigate whether processed Polygalaceae (PGA) was less irritating, and to screen and validate irritant properties of virgaureagenin G (polygala acid, PA), 3,6'-disinapoylsucrose (DSS), Tenuifolia (TEN) and polygalaxanthone III (POL), which had pharmacologically active in Polygalaceae. Zebrafish model, Draize test and High-Performance Liquid Chromatography (HPLC) were utilized to achieve the aim. MATERIALS AND METHODS: Scanning Electron Microscopy (SEM) and optical microscope were used to determine the presence of calcium oxalate needle crystal in RPA and PGA. Zebrafish egg spinning changes and zebrafish embryo behavior were used for irritation validation, irritation comparison and irritant screening. For additional evidence, the Draize test, HE staining of rabbit eyes and ELISA kit were used. Finally, changes in the composition of RPA and PGA were investigated using HPLC. RESULTS: SEM and optical microscopy revealed no calcium oxalate needle crystals in Polygalaceae. RPA, PGA, PA and DSS were able to accelerate the spinning of zebrafish eggs and the movement of embryos, while TEN and POL were not. RPA, PGA, DSS and PA may cause rabbit eyes to become hyperemic and swollen, resulting in damage to the iris, cornea and conjunctiva and increased levels of interleukin-6 (IL-6) and interleukin-10 (IL-10). Comparatively, the effects caused by PGA were less severe than those caused by RPA. In addition, compared to RPA, PGA had lower levels of DSS and PA. CONCLUSIONS: RPA, PGA, DSS, and PA were irritating. However, processing and curing could reduce the irritation by reducing the levels of DSS and PA. DSS and PA could be two potential irritants of Polygalaceae.

An optimized microRNA detection platform based on PAM formation-regulated CRISPR/Cas12a activation.

MicroRNAs (miRNAs) have emerged as biomarkers for the diagnosis and prognosis of various diseases, such as cancer. Recent advancements in CRISPR/Cas12a-based biosensors in combination with hybridization chain reaction (HCR) make it a promising approach for miRNA detection. To increase the compatibility of HCR and CRISPR/Cas12a, we compared two design strategies of hairpin DNA in HCR. The results showed that different arrangements of the protospacer sequence and protospacer adjacent motif (PAM) in the hairpin DNA could affect the sensing performance. The "PAM Formation" strategy, by which the duplex PAM sites are absent in the hairpin DNA and present in the long duplex DNA after HCR, exhibited advantages in detection sensitivity. By optimizing the probe sequences and reaction conditions, we developed a miRNA detection platform. With the same crRNA, this platform enables the identification of different miRNAs by simply replacing the loop region of the target recognition probe. In addition, the proposed platform can detect single-stranded DNA and distinguishing single or multiple base mutations in the target strand. The application of discriminating the target miRNA expression levels from different cell lines validated the reliability and practicability of the sensor platform, indicating its potential applications in early clinical accurate diagnosis of cancers.

An ultrasensitive Cd2+ detection biosensor based on DNAzyme and CRISPR/Cas12a coupled with hybridization chain reaction.

The detection of cadmium is essential because it poses a significant threat to human health and the environment. Recent advancements in biosensors that detect nonnucleic-acid targets using CRISPR/Cas12a in combination with aptamers or DNAzymes show promising performance. Herein, we integrated DNAzyme, hybridization chain reaction (HCR) and CRISPR/Cas12a into a single biosensor for the first time and realized the ultrasensitive detection of Cd2+. A single phosphorothioate ribonucleobase (rA)-containing oligonucleotide (PS substrate) and a Cd2+-specific DNAzyme (Cdzyme) are used for Cd2+ recognition, releasing short single-stranded DNA. Then, the HCR is triggered by the cleavage products for signal transduction and amplification. Next, the trans-cleavage activity of Cas12a is activated due to the presence of crRNA complementary strands and PAM sites in the HCR products. As a result, FQ-reporters are cleaved, and the fluorescence values can be easily read using a fluorometer, allowing Cd2+ quantification by measuring the fluorescent signal. The Cd2+ detection biosensor is ultrasensitive with a detection limit of 1.25 pM. Moreover, the biosensor shows great stability under different pH and various anion conditions. The proposed sensor was utilized for environmental water sample detection, demonstrating the dependability of the detection system. Considering the high sensitivity and reliable performance of the assay, it could be further used in environmental monitoring. In addition, the design strategy reported in this study could extend the application of CRISPR/Cas12a in heavy metal detection.

Application of Foley balloon catheter in palliative surgery for pulmonary atresia with an intact ventricular septem, with additional cases of pulmonary atresia with ventricular septal defect and tetralogy of Fallot.

BACKGROUND: Pulmonary atresia and tetralogy of Fallot can require palliative surgery in the neonatal period due to severe hypoxia; however, limitations of established techniques include high failure rate and need for cardiopulmonary bypass. Herein, right ventricular outflow tract reconstruction on a beating heart using a Foley balloon catheter is described. METHODS: A retrospective review of patients who underwent right ventricular outflow tract reconstruction on a beating heart using a Foley balloon catheter at our institution between September 2018 and March 2022 was completed. During the procedure, a Foley balloon catheter was used to occlude the blood from the right ventricular inflow tract. RESULTS: Eight patients with pulmonary atresia and intact ventricular septum underwent an off-pump right ventricular outflow tract reconstruction. One patient with pulmonary atresia and ventricular septal defect, and two patients with tetralogy of Fallot underwent an on-pump right ventricular outflow tract reconstruction on a beating heart. The procedures were successful in all patients. Patent ductus arteriosus ligation without modified Blalock-Taussig shunt placement was performed in three patients with pulmonary atresia with intact ventricular septum and two patients with tetralogy of Fallot, ductus arteriosus was left open in four patients with pulmonary atresia with intact ventricular septum. All patients remained clinically well without serious complications. CONCLUSIONS: Right ventricular outflow tract reconstruction on a beating heart using a Foley balloon catheter for pulmonary atresia and tetralogy of Fallot is a feasible alternative to catheter-based interventions or traditional surgical treatment, especially in patients with muscular infundibular stenosis or hypoplastic pulmonary annulus. Further studies with more cases are needed to verify feasibility and superiority of this approach.

Sutureless technique combined with vertical vein incision and pulmonary veins unroofed for correction of infracardiac total anomalous pulmonary venous connection.

Objective: We report a surgical method (sutureless technique), combined with vertical vein incision and pulmonary veins unroofed (semisutureless technique), to correct infracardiac total anomalous pulmonary venous connection (TAPVC). Materials and methods: The clinical characteristics of 21 patients, who were diagnosed with infracardiac TAPVS between February 2017 and March 2022, were retrospectively analyzed. These patients were divided into three groups according to different surgical methods: conventional surgery group, sutureless technique group, and semisutureless technique group. The conventional surgery group enrolled five patients with a median age of 16 days (interquartile range, 9-27 days) and a median weight of 3.25 kg (interquartile range, 3.1-3.42 kg). In this group, no preoperative pulmonary vein obstruction (PVO), preoperative ventilator support, or emergency surgery were reported. The sutureless technique group enrolled seven patients with a median age of 12 days (interquartile range, 5-16 days) and a median weight of 3.04 kg (interquartile range, 2.76-3.20 kg). In this group, two patients with preoperative PVO, four patients with preoperative ventilator support, and seven patients requiring emergency operation were found. The semisutureless technique group enrolled nine patients with a median age of 14 days (interquartile range, 7-24 days) and a median weight of 3.22 kg (interquartile range, 3.15-3.50 kg). In this group, four patients with preoperative PVO, two patients with preoperative ventilator support, and seven patients requiring emergency operation were noted. Results: In the conventional surgery group, two patients with postoperative supraventricular tachycardia, one patient with postoperative low cardiac output syndrome, one patient with PVO, and no case of postoperative death were reported. In the sutureless technique group, two patients with postoperative low cardiac output syndrome, one patient with postoperative supraventricular tachycardia, one patient with postoperative PVO, and no postoperative deaths were determined. In the semisutureless technique group, three patients had low cardiac output syndrome, two patients had supraventricular tachycardia after the operation, and one patient, who had been admitted to the hospital after cardiopulmonary resuscitation in the emergency room, died early after the operation. No case of death or PVO was noted after the operation. Conclusion: The semisutureless technique has positive effects. This surgery method can enlarge the anastomotic stoma, increase the volume of the left atrium, reduce the tension of the anastomotic stoma, fix the pulmonary vein to avoid distortion, and prevent postoperative hemorrhage.

G-quadruplex-containing oligodeoxynucleotides as DNA topoisomerase I inhibitors.

DNA topoisomerase I was found to be highly abundant in fast-proliferating tumor cells and is a potential target for anticancer therapy. A series of G-quadruplex-containing oligodeoxynucleotides (ODNs) were designed and used as inhibitors of DNA topoisomerase I. It was demonstrated that ODNs with G-quadruplexes can efficiently inhibit the supercoiled DNA relaxation reaction catalyzed by DNA topoisomerase I. Compared with the other conformations, the parallel propeller-type G-quadruplex was the most efficient DNA topoisomerase I inhibitor. Further studies revealed that integrating G-quadruplexes with duplexes to form quadruplex-duplex hybrids could significantly improve the inhibition efficiency. In addition, a circular ODN that consists of a G-quadruplex motif and DNA topoisomerase I binding site was synthesized and used as a DNA topoisomerase I inhibitor. The results showed that the particularly designed circular ODN displayed high inhibitory efficiency on the activity of DNA topoisomerase I with an IC50 value of 54.8 nM. Moreover, the circular ODN exhibited excellent thermal stability and nuclease resistance. Considering the low cytotoxicity of DNA-based biopharmaceuticals, the design strategy and results reported in this study may shed new light on nucleic acid-based DNA topoisomerase I inhibitor construction for potential anticancer drugs.

Construction of a tumor immune infiltration macrophage signature for predicting prognosis and immunotherapy response in liver cancer.

Liver cancer is an extraordinarily heterogeneous malignant disease. The tumor microenvironment (TME) and tumor-associated macrophages (TAMs) are the major drivers of liver cancer initiation and progression. It is critical to have a better understanding of the complicated interactions between liver cancer and the immune system for the development of cancer immunotherapy. Based on the gene expression profiles of tumor immune infiltration cells (TIICs), upregulated genes in TAMs and downregulated genes in other types of immune cells were identified as macrophage-specific genes (MSG). In this study, we combined MSG, immune subtypes, and clinical information on liver cancer to develop a tumor immune infiltration macrophage signature (TIMSig). A four-gene signature (S100A9, SLC22A15, TRIM54, and PPARGC1A) was identified as the TAM-related prognostic genes for liver cancer, independent of multiple clinicopathological parameters. Survival analyses showed that patients with low TIMSig had a superior survival rate than those with high TIMSig. Additionally, clinical immunotherapy response and TIMSig was observed as highly relevant. In addition, TIMSig could predict the response to chemotherapy. Collectively, the TIMSig could be a potential tool for risk-stratification, clinical decision making, treatment planning, and oncology immunotherapeutic drug development.

Sensitive fluorescent aptasensing of tobramycin on graphene oxide coupling strand displacement amplification and hybridization chain reaction.

An ultrasensitive biosensor was designed and constructed for tobramycin detection. As a target recognition component, the DNA probe consists of an aptamer region for tobramycin binding and a template for amplification. In the absence of tobramycin, the probe was locked to form a stem-loop structure. In the presence of the target, the binding of tobramycin led to a conformational change in the probe. The released 3' end was used as a primer for the strand displacement amplification (SDA) to produce a large amount of single-stranded trigger DNA, which then efficiently initiated the following hybridization chain reaction (HCR) to produce a long duplex DNA with many fluorophores. The signals were detected after the addition of graphene oxide (GO) to quench the fluorescence from excess hairpin DNA. Through sequence and reaction condition optimization, the biosensor exhibited high selectivity for tobramycin. The linearity range and limit of detection (LOD) were 0.5-30 nM and 0.06 nM, respectively. Moreover, the application of detecting tobramycin in milk and lake water samples showed that this method is reliable and could be further used in food safety control and environmental monitoring.

Development of a DNAzyme-based colorimetric biosensor assay for dual detection of Cd2+ and Hg2.

A colorimetric biosensor assay has been developed for Cd2+ and Hg2+ detection based on Cd2+-dependent DNAzyme cleavage and Hg2+-binding-induced conformational switching of the G-quadruplex fragment. Two types of multifunctional magnetic beads (Cd-MBs and Hg-MBs) were synthesized by immobilizing two functionalized DNA sequences on magnetic beads via avidin-biotin chemistry. For Cd2+ detection, Cd-MBs are used as recognition probes, which are modified with a single phosphorothioate ribonucleobase (rA) substrate (PS substrate) and a Cd2+-specific DNAzyme (Cdzyme). In the presence of Cd2+, the PS substrate is cleaved by Cdzyme, and single-stranded DNA is released as the signal transduction sequence. After molecular assembly with the other two oligonucleotides, duplex DNA is produced, and it can be recognized and cleaved by FokI endonuclease. Thus, a signal output component consisting of a G-quadruplex fragment is released, which catalyzes the oxidation of ABTS with the addition of hemin and H2O2, inducing a remarkably amplified colorimetric signal. To rule out false-positive results and reduce interference signals, Hg-MBs modified with poly-T fragments were used as Hg2+ accumulation probes during the course of Cd2+ detection. On the other hand, Hg-MBs can perform their second function in Hg2+ detection by changing the catalytic activity of the G-quadruplex/hemin DNAzyme. In the presence of Hg2+, the G-quadruplex structure in Hg-MBs is disrupted upon Hg2+ binding. In the absence of Hg2+, an intensified color change can be observed by the naked eye for the formation of intact G-quadruplex/hemin DNAzymes. The biosensor assay exhibits excellent selectivity and high sensitivity. The detection limits for Cd2+ and Hg2+ are 1.9 nM and 19.5 nM, respectively. Moreover, the constructed sensors were used to detect environmental water samples, and the results indicate that the detection system is reliable and could be further used in environmental monitoring. The design strategy reported in this study could broadly extend the application of metal ion-specific DNAzyme-based biosensors.

A practical screening strategy for Lynch syndrome and Lynch syndrome mimics in colorectal cancer.

OBJECTIVES: The objective of the study is to provide an efficient and practical screening strategy to distinguish a broader spectrum of Lynch syndrome (LS) and LS mimics-associated colorectal cancer (CRC), including Lynch-like syndrome (LLS), constitutional mismatch repair-deficiency, familial CRC type X (FCCTX), and polymerase proofreading-associated polyposis syndrome. MATERIALS AND METHODS: 1294 cases of CRC samples were detected mismatch repair (MMR) status using immunohistochemistry (IHC) staining, in which the cases with MLH1-deficient CRC underwent BRAF mutation analysis by IHC. Following the personal and/or family history survey, next-generation sequencing (NGS) was used to detect gene variants. RESULTS: 1294 CRC patients were dichotomized into tumors caused by a deficient MMR (dMMR) system and a proficient MMR (pMMR) system after MMR status analysis. 45 patients with suspected sporadic dMMR CRC were then separated from MLH1-deficient CRC though BRAF mutation status analysis by IHC. Following the personal and/or family history survey for 1294 patients, as well as germline genetic testing by NGS, 34 patients were diagnosed as LS (8 cases), SLS (13 cases), LLS ( 6 cases), FCCTX (3 cases), and sporadic CRC (4 cases). CONCLUSIONS: Our screening strategy, which consists of clinical and molecular analyses, is expected to improve the screening efficiency and management for the LS and LS mimics.

Protein convertase subtilisin/Kexin type 9 inhibits hepatocellular carcinoma growth by interacting with GSTP1 and suppressing the JNK signaling pathway.

OBJECTIVE: Protein convertase subtilisin/Kexin type 9 (PCSK9) has been found to be closely associated with the occurrence and development of numerous tumors. However, the precise role of PCSK9 and its relationship to the development of hepatocellular carcinoma (HCC) remain largely unknown. This study aimed to clarify these issues. METHODS: The expression levels of PCSK9 in HCC tissues and HCC cell lines were determined by the quantitative reverse transcription polymerase chain reaction, Western blot, and immunohistochemical analyses, and the effects of PCSK9 expression on HCC cell biological traits were investigated by overexpressing and downregulating PCSK9 expression in vivo and in vitro. Additionally, the mechanism by which PCSK9 mediated dissociation of glutathione S-transferase Pi 1 (GSTP1) dimers and phosphorylation of the Jun N-terminal kinase (JNK) pathway components were investigated. RESULTS: PCSK9 expression levels were significantly lower in HCC tissues than in adjacent non-tumor samples. In vivo and in vitro experiments suggested that PCSK9 inhibited HCC cell proliferation and metastasis. Further analysis showed that PCSK9 interacted with GSTP1 and promoted GSTP1 dimer dissociation and JNK signaling pathway inactivation in HCC cells. Moreover, the relationships between PCSK9 protein expressions and clinical outcomes were investigated. The PCSK9-lo group displayed a significantly shorter overall survival (OS; median OS: 64.2 months vs. 83.2 months; log-rank statistic: 4.237; P = 0.04) and recurrence-free survival (RFS; median RFS: 26.5 months vs. 46.6 months; log-rank statistic: 10.498; P = 0.001) time than the PCSK9-hi group. CONCLUSIONS: PCSK9 inhibited HCC cell proliferation, cell cycle progression, and apoptosis by interacting with GSTP1 and suppressing JNK signaling, suggesting that PCSK9 might act as a tumor suppressor and be a therapeutic target in HCC patients.

Succinate dehydrogenase deficient gastrointestinal stromal tumor in a three month old boy with a fatal clinical course: a case report and review of literature.

BACKGROUND: Succinate dehydrogenase deficient gastrointestinal stromal tumors (SDH-deficient GISTs), which lack KIT or PDGFRA mutations demonstrate unique clinical and pathological features, and they respond poorly to standard targeted therapy. We herein present a novel case of SDH-deficient GIST in a three-month-old infant's colon mesentery, and he is the youngest patientto date. CASE PRESENTATION: The infantpresented with complaints of blood in the stool. CT showed a 6.3 × 4.6 cm mass in the left lower retroperitoneal. Complete resection of tumor and segmental bowel resection was performed without regional lymphadenectomy. Histologically, tumor cells were distinctive in their multinodular colon wall involvement with interspersed tracts of colon wall smooth muscle. The tumor was composed mainly of epithelioid cells. Immunohistochemically, the tumor cells were positive for Vim, CD117, PDGFR, while negative for SDHB. Mutational analysis showed a synonymous mutation for SDHB and wild-type for KIT and PDGFRA. Two months after surgery, metastases were found and Imatinib was administered. Unfortunately, the disease continued to progress, and the infant died 5 months after surgery. CONCLUSIONS: SDH-deficient GISTs comprise a subgroup of a relatively rare tumor type and show a number of clinically and biologically unique features, especially for infants. It is of great importance to developing new therapeutic targets and novel specific drugs.

Clinicopathological characteristics and surgical outcomes of sarcomatoid hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the sixth most common type of cancer and the fourth leading cause of cancer-related death worldwide. Sarcomatoid HCC, which contains poorly differentiated carcinomatous and sarcomatous components, is a rare histological subtype of HCC that differs from conventional HCC. It is highly aggressive and has a poor prognosis. Its clinicopathological characteristics, surgical outcomes and underlying mechanisms of its highly aggressive nature have not been fully elucidated. AIM: To examine the clinicopathological characteristics and surgical outcomes of sarcomatoid HCC and explore the histogenesis of sarcomatoid HCC. METHODS: In total, 196 patients [41 sarcomatoid HCC and 155 high-grade (Edmondson-Steiner grade III or IV) HCC] who underwent surgical resection between 2007 and 2017 were retrospectively reviewed. The characteristics and surgical outcomes of sarcomatoid HCC were compared with those of patients with high-grade HCC. The histological composition of invasive and metastatic sarcomatoid HCCs was evaluated. RESULTS: Sarcomatoid HCC was more frequently diagnosed at an advanced stage with a larger tumor and higher rates of nonspecific symptom, adjacent organ invasion and lymph node metastasis than high-grade HCC (all P < 0.05). Compared with high-grade HCC patients, sarcomatoid HCC patients are less likely to have typical dynamic imaging features of HCC (44.4% vs 72.7%, P = 0.001) and elevated serum alpha-fetoprotein levels (> 20 ng/mL; 36.6% vs 78.7%, P < 0.001). The sarcomatoid group had a significantly shorter median recurrence-free survival (5.6 mo vs 16.4 mo, log-rank P < 0.0001) and overall survival (10.5 mo vs 48.1 mo, log-rank P < 0.0001) than the high-grade group. After controlling for confounding factors, the sarcomatoid subtype was identified as an independent predictor of poor prognosis. Pathological analyses indicated that invasive and metastatic lesions were mainly composed of carcinomatous components. CONCLUSION: Sarcomatoid HCC was associated with a more advanced stage, atypical dynamic imaging, lower serum alpha-fetoprotein levels and a worse prognosis. The highly aggressive nature of sarcomatoid HCC is perhaps mediated by carcinomatous components.

E2F1-activated SPIN1 promotes tumor growth via a MDM2-p21-E2F1 feedback loop in gastric cancer.

Gastric cancer (GC) is one of the most common cancers around the world. Searching for specific gene expression changes during the development of GC could help identify potential therapy targets. We previously showed that the histone code reader SPIN1 may act as an oncogene in breast cancer. At present, the biological function and regulation of SPIN1 in GC remain unclear. Here, we demonstrate that SPIN1 is upregulated in GC tissues, compared with nontumorous gastric tissues. Increased expression of SPIN1 is closely associated with poor prognosis for patients with GC. Increased SPIN1 expression enhances GC cell proliferation, migration, and invasion and promotes cell cycle progression. Mechanically, SPIN1 sustains GC cell proliferation via activation of the MDM2-p21-E2F1 signaling pathway by binding to H3K4me3 of the MDM2 promoter region. Interestingly, E2F1 could directly bind to the SPIN1 promoter and activate its transcription, thus forming a positive feedback loop. Our data suggest that SPIN1 plays an important role in the development of GC and could be used as a promising prognostic biomarker and therapeutic target for GC.

Two novel fan-shaped trinuclear Pt(ii) complexes act as G-quadruplex binders and telomerase inhibitors.

Two new trinuclear Pt(ii) complexes {[Pt(dien)]3(tib)}(NO3)6 (1) and {[Pt(dpa)]3(tib)}(NO3)6 (2) (dien: diethylenetriamine, dpa: bis-(2-pyridylmethyl)amine, tib: 1,3,5-tris(1H-imidazol-1-yl)benzene) have been designed, synthesized, characterized and applied to a series of biochemical studies. We found that both of the Pt(ii) complexes exhibited much better selectivity for human telomeric G-quadruplex sequence than promoter G-quadruplexes (c-kit, c-myc, and bcl2) or duplex DNA. Both complexes displayed comparative stability and affinity towards human telomeric G-quadruplex by the studies from surface plasmon resonance, fluorescence resonance energy transfer and polymerase chain reaction stop assays. The circular dichroism indicated that both complexes could induce and stabilize anti-parallel G-quadruplex structures. Molecule docking presented that Pt(ii) complex intercalated into the large groove of human telomeric G-quadruplex (PDB ID: 143D). Furthermore, telomeric repeat amplification protocol assays quantitatively evaluated the inhibition of telomerase activity caused by the Pt(ii) complexes. The obtained IC50 values of 6.41 ± 0.042 µM and 2.67 ± 0.035 µM for 1 and 2, respectively, exhibited strong telomerase inhibitions. All results suggest that such fan-shaped trinuclear Pt(ii) complexes are effective and selective G-quadruplex binders, as well as strong telomerase inhibitors. This study provides insight into the development of human telomeric G-quadruplex targeted anticancer drugs based on the metal complex.

LncRNA-HNF1A-AS1 functions as a competing endogenous RNA to activate PI3K/AKT signalling pathway by sponging miR-30b-3p in gastric cancer.

BACKGROUND: Accumulating evidence demonstrated that long noncoding RNAs (lncRNAs) played important regulatory roles in many cancer types. However, the role of lncRNAs in gastric cancer (GC) progression remains unclear. METHODS: RT-qPCR assay was performed to detect the expression of HNF1A-AS1 in gastric cancer tissues and the non-tumourous gastric mucosa. Overexpression and RNA interference approaches were used to investigate the effects of HNF1A-AS1 on GC cells. Insight into competitive endogenous RNA (ceRNA) mechanisms was gained via bioinformatics analysis, luciferase assays and an RNA-binding protein immunoprecipitation (RIP) assay, RNA-FISH co-localisation analysis combined with microRNA (miRNA)-pulldown assay. RESULTS: This study displayed that revealed expression of HNF1A-AS1 was associated with positive lymph node metastasis in GC. Moreover, HNF1A-AS1 significantly promoted gastric cancer invasion, metastasis, angiogenesis and lymphangiogenesis in vitro and in vivo. In addition, HNF1A-AS1 was demonstrated to function as a ceRNA for miR-30b-3p. HNF1A-AS1 abolished the function of the miRNA-30b-3p and resulted in the derepression of its target, PIK3CD, which is a core oncogene involved in the progression of GC. CONCLUSION: This study demonstrated that HNF1A-AS1 worked as a ceRNA and promoted PI3K/AKT signalling pathway-mediated GC metastasis by sponging miR-30b-3p, offering novel insights of the metastasis mechanism in GC.

TREM2 acts as a tumor suppressor in hepatocellular carcinoma by targeting the PI3K/Akt/ß-catenin pathway.

Triggering receptor expressed on myeloid cells 2 (TREM2) is involved in nonmalignant pathological processes. However, TREM2's function in malignant diseases, especially in hepatocellular carcinoma (HCC) remains unknown. In the present study, we report that TREM2 is a novel tumor suppressor in HCC. TREM2 expression was obviously decreased in hepatoma cells (especially metastatic HCC cells), and in most human HCC tissues (especially extrahepatic metastatic tumors). Reduced tumor TREM2 expression was correlated with poor prognosis of HCC patients, and with aggressive pathological features (BCLC stage, tumor size, tumor encapsulation, vascular invasion, and tumor differentiation). TREM2 knockdown substantially promoted cell growth, migration, and invasion in vitro and in vivo, while TREM2 overexpression produced the opposite effect. TREM2 suppressed HCC metastasis by inhibiting epithelial-mesenchymal transition, accompanied by abnormal expression of epithelial and mesenchymal markers. Further study revealed that downregulation of TREM2 in HCC was regulated by miR-31-5p. Moreover, by directly interacting with ß-catenin, TREM2 attenuated oncogenic and metastatic behaviors by inhibiting Akt and GSK3ß phosphorylation, and activating ß-catenin. TREM2 suppressed carcinogenesis and metastasis in HCC by targeting the PI3K/Akt/ß-catenin pathway. Thus, we propose that TREM2 may be a candidate prognostic biomarker in malignant diseases and TREM2 restoration might be a prospective strategy for HCC therapy.