LOXL1-AS1 contributes to metastasis in sonic-hedgehog medulloblastoma by promoting cancer stem-like phenotypes.

BACKGROUND: Medulloblastomas (MBs) are one of the most common malignant brain tumor types in children. MB prognosis, despite improvement in recent years, still depends on clinical and biological risk factors. Metastasis is the leading cause of MB-related deaths, which highlights an unmet need for risk stratification and targeted therapy to improve clinical outcomes. Among the four molecular subgroups, sonic-hedgehog (SHH)-MB harbors clinical and genetic heterogeneity with a subset of high-risk cases. Recently, long non-coding (lnc)RNAs were implied to contribute to cancer malignant progression, but their role in MB remains unclear. This study aimed to identify pro-malignant lncRNAs that have prognostic and therapeutic significance in SHH-MB. METHODS: The Daoy SHH-MB cell line was engineered for ectopic expression of MYCN, a genetic signature of SHH-MB. MYCN-associated lncRNA genes were identified using RNA-sequencing data and were validated in SHH-MB cell lines, MB tissue samples, and patient cohort datasets. SHH-MB cells with genetic manipulation of the candidate lncRNA were evaluated for metastatic phenotypes in vitro, including cell migration, invasion, sphere formation, and expressions of stemness markers. An orthotopic xenograft mouse model was used to evaluate metastasis occurrence and survival. Finally, bioinformatic screening and in vitro assays were performed to explore downstream mechanisms. RESULTS: Elevated lncRNA LOXL1-AS1 expression was identified in MYCN-expressing Daoy cells and MYCN-amplified SHH-MB tumors, and was significantly associated with lower survival in SHH-MB patients. Functionally, LOXL1-AS1 promoted SHH-MB cell migration and cancer stemness in vitro. In mice, MYCN-expressing Daoy cells exhibited a high metastatic rate and adverse effects on survival, both of which were suppressed under LOLX1-AS1 perturbation. Integrative bioinformatic analyses revealed associations of LOXL1-AS1 with processes of cancer stemness, cell differentiation, and the epithelial-mesenchymal transition. LOXL1-AS1 positively regulated the expression of transforming growth factor (TGF)-ß2. Knockdown of TGF-ß2 in SHH-MB cells significantly abrogated their LOXL1-AS1-mediated prometastatic functions. CONCLUSIONS: This study proved the functional significance of LOXL1-AS1 in SHH-MB metastasis by its promotion of TGF-ß2-mediated cancer stem-like phenotypes, providing both prognostic and therapeutic potentials for targeting SHH-MB metastasis.

Coffee Husk By-Product as Novel Ingredients for Cascara Kombucha Production.

Kombucha, a fermented beverage, is gaining popularity due to its numerous beneficial health effects. Various substrates such as herbs, fruits, flowers, and vegetables, have been used for kombucha fermentation in order to enhance the flavor, aroma, and nutritional composition. This study aims to investigate the potential suitability of cascara as a novel ingredient for kombucha production. Our findings suggested that cascara is a suitable substrate for kombucha production. Fermentation elevated the total phenolic and flavonoid content in cascara, which enhanced the antioxidant, antibacterial, and prebiotic characteristics of the product. Furthermore, the accumulation of acetic acid-induced the pH lowering reached 2.7 after 14 days of fermentation, which achieved the microbiological safety of the product. Moreover, 14 days of fermentation resulted in a balanced amalgamation of acidity, sweetness, and fragrance according to sensory evaluation. Our findings not only highlight the potential of cascara kombucha as a novel substrate for kombucha production but also contribute to repurposing coffee by-products, promoting environmentally friendly and sustainable agricultural development.

Targeting of RRM2 suppresses DNA damage response and activates apoptosis in atypical teratoid rhabdoid tumor.

BACKGROUND: Atypical teratoid rhabdoid tumors (ATRT) is a rare but aggressive malignancy in the central nervous system, predominantly occurring in early childhood. Despite aggressive treatment, the prognosis of ATRT patients remains poor. RRM2, a subunit of ribonucleotide reductase, has been reported as a biomarker for aggressiveness and poor prognostic conditions in several cancers. However, little is known about the role of RRM2 in ATRT. Uncovering the role of RRM2 in ATRT will further promote the development of feasible strategies and effective drugs to treat ATRT. METHODS: Expression of RRM2 was evaluated by molecular profiling analysis and was confirmed by IHC in both ATRT patients and PDX tissues. Follow-up in vitro studies used shRNA knockdown RRM2 in three different ATRT cells to elucidate the oncogenic role of RRM2. The efficacy of COH29, an RRM2 inhibitor, was assessed in vitro and in vivo. Western blot and RNA-sequencing were used to determine the mechanisms of RRM2 transcriptional activation in ATRT. RESULTS: RRM2 was found to be significantly overexpressed in multiple independent ATRT clinical cohorts through comprehensive bioinformatics and clinical data analysis in this study. The expression level of RRM2 was strongly correlated with poor survival rates in patients. In addition, we employed shRNAs to silence RRM2, which led to significantly decrease in ATRT colony formation, cell proliferation, and migration. In vitro experiments showed that treatment with COH29 resulted in similar but more pronounced inhibitory effect. Therefore, ATRT orthotopic mouse model was utilized to validate this finding, and COH29 treatment showed significant tumor growth suppression and prolong overall survival. Moreover, we provide evidence that COH29 treatment led to genomic instability, suppressed homologous recombinant DNA damage repair, and subsequently induced ATRT cell death through apoptosis in ATRT cells. CONCLUSIONS: Collectively, our study uncovers the oncogenic functions of RRM2 in ATRT cell lines, and highlights the therapeutic potential of targeting RRM2 in ATRT. The promising effect of COH29 on ATRT suggests its potential suitability for clinical trials as a novel therapeutic approach for ATRT.

Multi-Strain Probiotics Enhance the Bioactivity of Cascara Kombucha during Microbial Composition-Controlled Fermentation.

Kombucha is a widely consumed fermented tea beverage with diverse health benefits. In a previous study, we demonstrated that the use of cascara as a substrate results in a special kombucha beverage with high bioactivity. Traditional kombucha fermentation using a symbiotic culture of bacteria and yeast (SCOBY) can lead to inconsistent product quality because of the lack of control over microbial composition. We successfully isolated and identified yeast and bacteria, including Saccharomyces cerevisiae, Komagataeibacter rhaeticus, and Lactobacillus brevis that are appropriate starter cultures for cascara kombucha fermentation. We also demonstrated that a supplementation with lactic acid bacteria (LAB) and a mixture of S. cerevisiae and K. rhaeticus resulted in higher total polyphenol and flavonoid content of cascara kombucha compared with the traditionally fermented product using SCOBY as the inoculum. The free radical scavenging activity, inhibitory effects on α-amylase, tyrosinase activity, and antibacterial properties of cascara kombucha were also enhanced as a result of LAB supplement. These findings provide valuable insights into the controlled microbiological composition required for the fermentation of cascara kombucha, thereby ensuring consistent quality and enhanced bioactivity of the product. Further, the use of cascara as a substrate for kombucha production not only offers various health benefits and biological effects, but also repurposes by-products from the coffee industry, which contributes to sustainable development and is eco-friendly.

Targeting Castration-Resistant Prostate Cancer Using Mesenchymal Stem Cell Exosomes for Therapeutic MicroRNA-let-7c Delivery.

BACKGROUND: Castration-resistant prostate cancer (PCa; CRPC) has a poor response to androgen deprivation therapy and is considered an incurable disease. MicroRNA (miR)-lethal 7c (let-7c) was implied to be a tumor suppressor in PCa, and treatment with exogenous let-7c targets both cancer cells and their associated mesenchymal stem cells (MSCs) to prevent CRPC progression and metastasis. Exosomes are nanometer-sized membrane-bound vesicles which have an absolute predominance in biocompatibility for drug delivery and gene therapy by mediating cell-to-cell communication. By utilizing the intrinsic tumor-targeting property of MSCs, this study aimed to investigate the feasibility of MSC-derived exosomes as an exogenous miR delivery system to target CRPC, using miR let-7c as an example. METHODS: Bioinformatics analysis was performed to observe miR-let-7c expression in clinical samples by utilizing the GEO database. MSC-derived exosomes were collected from a human bone marrow-derived MSC cell line after cell transfection with either a pre-miR negative control or pre-miR-let-7c, and further characterized through nanoparticle tracking analysis and Western blotting. miR-let-7c expression was determined using RT-qPCR, and the phenotypic effects of both naked and MSC-exosome-encapsulated let-7c on CRPC cells (PC3 and CWR22Rv1) were determined by WST-1 cell proliferation assay and wound healing migration assay. RESULTS: miR-let-7c was downregulated in metastatic PCa and high grade group patients. miR-let-7c expression was confirmed to be downregulated in PCa cell lines, with massively decreased in most metastatic CRPC-like cells. Exogenous miR-let-7c can be successfully packaged into MSC exosomes. Treatment with either naked or MSC-exosome-encapsulated miR-let-7c resulted in significant reductions in cell proliferation and migration in CRPC-like PC3 and CWR22Rv1 cells. CONCLUSIONS: MSC-derived exosomes could serve as a therapeutic let-7c delivery system to target CRPC.

The ADAM9/UBN2/AKR1C3 axis promotes resistance to androgen-deprivation in prostate cancer.

Metastatic and castration-resistant disease is a fatal manifestation of prostate cancer (PCa). The mechanism through which resistance to androgen deprivation in PCa is developed remains largely unknown. Our understanding of the tumor microenvironment (TME) and key signaling pathways between tumors and their TME is currently changing in light of the generation of new knowledge with regard to cancer progression. A disintegrin and metalloproteinase domain-containing protein 9 (ADAM9) is a membranous bridge forming cell-cell and cell-matrix connections that regulate tumor aggressiveness and metastasis. However, it is not known whether ADAM9 expressed in the TME contributes to the CRPC phenotype. In this study, we aimed to investigate the expression patterns of ADAM9 in prostate cancer-associated fibroblasts (CAFs). We also intended to elucidate the effects of both stromal cell- and cancer cell-derived ADAM9 on the progression of CRPC and the implicated molecular pathways. By using both clinical specimens and cell lines, we herein showed that unlike the membrane anchored ADAM9 overexpressed by both PCa cells and prostate CAFs, the secreted isoform of ADAM9 (sADAM9) was strongly detected in CAFs, but rarely in tumor cells, and that could be a serum marker for PCa patients. We demonstrated that functionally sADAM9 are characterized as chemoattractant for the directed movement of androgen-independent PCa cells through integrin downstream FAK/AKT pathway, supporting that elevated sADAM9 by prostate CAFs could be responsible for the promotion of CRPC metastasis. Moreover, by stimulating PCa cells with sADAM9, we found that ubinuclein-2 (UBN2) expression was increased. A positive correlation of ADAM9 and UBN2 expression was observed in androgen receptor-expressing PCa cell lines and further confirmed in clinical PCa specimens. Using a genetic modification approach, we identified UBN2 as a downstream target gene of ADAM9 that is critical for the survival of androgen-dependent PCa cells in response to androgen deprivation, through the induction and effect of the aldo-keto reductase family 1 member C3 (AKR1C3). Collectively, our results reveal a novel action of ADAM9 on the transition of androgen-dependent PCa cells into an androgen-independent manner through the UBN2/AKR1C3 axis; the aforementioned action could contribute to the clinically-observed acquired androgen-deprivation therapy resistance.

Application of Mesenchymal Stem Cells in Targeted Delivery to the Brain: Potential and Challenges of the Extracellular Vesicle-Based Approach for Brain Tumor Treatment.

Treating brain tumors presents enormous challenges, and there are still poor prognoses in both adults and children. Application of novel targets and potential drugs is hindered by the function of the blood-brain barrier, which significantly restricts therapeutic access to the tumor. Mesenchymal stem cells (MSCs) can cross biological barriers, migrate to sites of injuries to exert many healing effects, and be engineered to incorporate different types of cargo, making them an ideal vehicle to transport anti-tumor agents to the central nervous system. Extracellular vesicles (EVs) produced by MSCs (MSC-EVs) have valuable innate properties from parent cells, and are being exploited as cell-free treatments for many neurological diseases. Compared to using MSCs, targeted delivery via MSC-EVs has a better pharmacokinetic profile, yet avoids many critical issues of cell-based systems. As the field of MSC therapeutic applications is quickly expanding, this article aims to give an overall picture for one direction of EV-based targeting of brain tumors, with updates on available techniques, outcomes of experimental models, and critical challenges of this concept.

Lactobacillus rhamnosus JB3 inhibits Helicobacter pylori infection through multiple molecular actions.

BACKGROUND: Eradication of Helicobacter pylori infection is the most direct and effective way for preventing gastric cancer. Lactic acid bacteria are considered as alternative therapeutic agents against H. pylori infection. METHODS: Effects of Lactobacillus rhamnosus JB3 (LR-JB3) on the virulence gene expression of H. pylori and infection-induced cellular responses of AGS cells were investigated by co-cultivating infected AGS cells with different multiplicity of infections (MOIs) of LR-JB3. RESULTS: LR-JB3, specifically at a MOI of 25, suppressed the association ability of H. pylori and its induced IL-8 levels, as well as the mRNA levels of vacA, sabA, and fucT of H. pylori, infection-induced Lewis (Le)x antigen and Toll-like receptor 4 (TLR4) expressions in AGS cells. However, the apoptosis mediated by infection was inhibited by LR-JB3 in a dose-dependent manner. In addition, autoinducer (AI)-2 was observed to have increased the association ability and fucT expression of H. pylori, and Lex antigen and TLR4 expression of AGS cells. Interestingly, an unknown bioactive cue was hypothesized to have been secreted from LR-JB3 at a MOI of 25 to act as an antagonist of AI-2. CONCLUSIONS: LR-JB3 possesses various means to interfere with H. pylori pathogenesis and infection-induced cellular responses of AGS cells to fight against infection.

Antagonistic Activities of Lactobacillus rhamnosus JB3 Against Helicobacter pylori Infection Through Lipid Raft Formation.

Helicobacter pylori is a Gram-negative pathogen that can increase the risk of stomach cancer in infected patients. H. pylori exploits lipid rafts to infect host cells. Infection triggers clustering of Lewis x antigen (Lex) and integrins in lipid rafts to facilitate H. pylori adherence to the gastric epithelium. H. pylori infection can be treated with probiotics containing lactic acid bacteria that offer numerous benefits to the host while lacking the side effects associated with antibiotic therapy. Previously, we showed that the cell-free supernatant (CFS) derived from Lactobacillus rhamnosus JB3 (LR-JB3) at a multiplicity of infection (MOI) of 25 attenuated the pathogenicity of H. pylori. In this study, we established a mucin model to simulate the gastric environment and to further understand the influence of mucin on the pathogenesis of H. pylori. Porcine stomach mucin dramatically upregulated H. pylori virulence gene expression, including that of babA, sabA, fucT, vacA, hp0499, cagA, and cagL, as well as the adhesion and invasion ability of H. pylori and induced increased levels of IL-8 in infected-AGS cells. The CFS derived from LR-JB3 at a MOI of 25 reduced the expression of H. pylori sabA, fucT, and hp0499 in mucin, as well as that of the Lex antigen and the α5ß1 integrin in AGS cells during co-cultivation. These inhibitory effects of LR-JB3 also suppressed lipid raft clustering and attenuated Lewis antigen-dependent adherence, type IV secretion system-mediated cell contact, and lipid raft-mediated entry of VacA to host cells. In conclusion, LR-JB3 could affect H. pylori infection through mediating lipid raft formation of the host cells. The currently unknown cues secreted from LR-JB3 are valuable not only for treating H. pylori infection, but also for treating diseases that are also mediated by lipid raft signaling, such as cancer and aging-associated and neurodegenerative conditions.

Risk factors for cutaneous reactions to allopurinol in Kinh Vietnamese: results from a case-control study.

OBJECTIVE: The aim of this study was to investigate risk factors for cutaneous adverse reactions (CARs) in Kinh Vietnamese. METHODS: All patients were prospectively recruited in Ho Chi Minh City. Presence of the HLA-B*58:01 allele was determined by real-time PCR-sequence-specific amplification by using the PG5801 Detection Kit (Pharmigene, Taipei). Patients with severe (SCARs) and mild (MCARs) CARs and controls were compared for differences in features prospectively collected, and odds ratios (ORs) with 95% confidence intervals (CIs) were estimated. RESULTS: On comparing 32 patients with SCARs and 395 tolerant controls, we identified eight strong risk factors: increased age (OR 15.1 [95% CI 5.8-40.1], P < 0.0001), female sex (OR 333 [40-43,453], P < 0.0001), allopurinol for asymptomatic hyperuricemia (OR 955 [120-125,847], P < 0.0001), allopurinol starting dose > 150 mg (OR 316 [101-122], P < 0.0001), diuretics intake (OR 304 [35-40,018], P < 0.0001), eGFR < 60 ml/min/1.73 m2 (OR 100 [32-353], P < 0.0001), history of allopurinol-induced skin reaction (OR 78 [6-10,808], P = 0.004), and HLA-B*58:01 carriage (OR 147 [45-746], P < 0.0001). HLA-B*58:01 allele frequency in controls was 7.3%. For MCARs (n = 74), risk factors were eGFR < 60 ml/min/1.73 m2 (OR 4.9 [1.61-14.6], P = 0.006), history of allopurinol-induced skin reaction (OR 27 [2-3777], P = 0.01), and asymptomatic hyperuricemia (OR 27 [2-3777], P = 0.01). CONCLUSION: This study confirmed 8 risk factors, including HLA-B*58:01, for SCARs and identified 3 risk factors for MCARs in Kinh Vietnamese. HLA-B*58:01 genotyping could guide the indication for allopurinol in Kinh Vietnamese patients with gout.