Biotechnology Journal最新文献

Cancer continues to remain challenging due to heterogeneity and therapeutic resistance. Conventional therapeutic approaches, such as surgery, chemotherapy, and radiotherapy, often encounter critical limitations such as extensive tissue damage and ineffective elimination of metastatic malignancies. To resolve these issues, nanomaterial-based therapeutics have progressed to develop improved anticancer treatment outcomes. In this context, a CuS-R848-mediated therapeutic nanovaccine (NV) for synergizing photothermal therapy (PTT) with TLR7/8-driven immunity has been developed. To improve the stability and specificity, the NV was systematically coated with hyaluronic acid (HA) to effectively target and reduce systemic toxicity via CD44-targeted delivery. Furthermore, the NV under near-infrared (NIR) light exhibited 98% tumor ablation, including immunogenic cell death (ICD) induction via DAMPs, DC maturation, and significantly enhanced tumor eradication to strengthened immune activation. This dual-approach combining PTT with ICD demonstrated excellent therapeutic vaccine efficacy in preventing tumor recurrence and metastasis compared to monotherapy.

Vascular endothelial growth factor (VEGF) is a key pro-angiogenic glycoprotein used in both research and therapeutic applications. While Pichia pastoris (Komagataella phaffii) provides a cost-effective expression system for recombinant VEGF, its glycosylation patterns differ significantly from those in mammalian systems, and the impact of these modifications remains underexplored. Recombinant human VEGF165 was expressed in P. pastoris, purified via heparin affinity chromatography, and enzymatically deglycosylated using Endo H. Both glycosylated (yVEGF_g) and deglycosylated (yVEGF_dg) forms were characterized for their binding to an anti-VEGF antibody, bevacizumab, via ELISA and SPR and evaluated for biological activity using HUVEC proliferation assays. Comparative analyses were also conducted with mammalian-expressed VEGF. Deglycosylated VEGF exhibited reduced binding affinity to bevacizumab and a lower proliferative effect on endothelial cells compared to its glycosylated counterpart. Despite the N-glycosylation site being structurally distant from receptor/antibody epitopes, glycosylation enhanced VEGF's in vitro binding kinetics and angiogenic activity. Partially glycosylated VEGF from P. pastoris even surpassed mammalian-expressed VEGF in inducing endothelial proliferation. These findings demonstrate that glycosylation significantly enhances the functional properties of recombinant VEGF produced in P. pastoris, emphasizing the importance of glycosylation control for the development of bioactive VEGF.

This study investigated the biosynthesis of nanoparticles utilizing orange peel extract (OPE) to synthesize monometallic CuO and ZnO nanoparticles, in addition to bimetallic CuO–ZnO nanoparticles. A comparative examination was performed to assess their antibacterial and insecticidal properties. A range of characterization techniques was utilized to ascertain the shape, size, particle size distribution, crystallinity, surface charge, morphology, and optical properties of the biosynthesized CuO–ZnO nanoparticles. The bimetallic CuO–ZnO nanoparticles show markedly superior antibacterial activity relative to their monometallic equivalents. The minimum inhibitory concentrations (MICs) for CuO–ZnO nanoparticles against pathogenic bacteria, including Pseudomonas aeruginosa and Klebsiella pneumoniae, was (12.5 to 50 µg/mL). The insecticidal effect on Culex pipiens were evaluated. The LC₅₀ and LC₉₀ values for the bimetallic CuO–ZnO nanoparticles were significantly lower at (18.878, 66.447) µg/mL and (26.344, 82.458) µg/mL, respectively, demonstrating enhanced toxicity relative to both CuO and ZnO NPs. The biosynthesized bimetallic CuO–ZnO nanoparticles exhibited remarkable antibacterial and insecticidal characteristics, indicating their potential uses in medicinal and environmental domains.

Precise control of critical quality attributes, including titer and glycosylation, is essential in bioprocessing, yet conventional design‑of‑experiments methods are challenged by the high-dimensional, nonlinear design space for media and process parameters. We assemble a comprehensive glycan‑focused Chinese hamster ovary (CHO) fed‑batch dataset and develop a computational workflow (i) to train machine learning (ML) models to predict key CQAs, (ii) apply a hybrid ML + knowledge-based strategy to select potentially impactful features, and (iii) generate combinatorial media designs. The resulting models predict final titer (R² ≈ 0.93) and major glycan metrics—mannosylation, fucosylation, galactosylation (R² ≈ 0.79–0.95)—directly from initial media composition and process parameters without requiring spent media analysis. Feature selection shortlisted 20 features out of 76 for a second-tier validation, from which 15 were confirmed as actionable levers impacting titer and glycosylation, uncovering glycan effects independent of nucleotide sugar supplementation. Finally, we incorporated our workflow, utilizing a ML surrogate model coupled with simulated annealing, in a proof‑of‑concept active learning step, successfully proposing a media composition and process parameter combination that reduced mannosylation by 10% while increasing titer. Together, these results underscore how ML‑enhanced DOE can accelerate CHO process development and explore complex biomanufacturing spaces with greater efficiency.

With accelerating climate change and the urgent need to stack polygenic traits, multiplex CRISPR/Cas offers a scalable route to resilient crops—yet low editing efficiency and regeneration bottlenecks remain critical constraints. This review centers on multiplex strategies for polygenic trait engineering in plants, surveying compact nucleases (Cas9, Cas12, Cas13 and emerging ultra-compact variants), polycistronic gRNA platforms (tRNA–gRNA arrays, self-cleaving ribozymes, Csy4 processing), and delivery routes (Agrobacterium, biolistics, protoplast transfection, viral vectors). We highlight concrete outcomes—for example, targeted edits in PYL ABA-receptors increased rice grain yield by up to 31% in field tests—and applications from yield and disease resistance to abiotic-stress tolerance, nutrient biofortification and de novo domestication. Technical risks (off-targets, mosaicism, chromosomal rearrangements, transformability) are appraised alongside emerging fixes: compact/engineered nucleases, RNA-processing arrays, morphogenic regulators, and AI-driven sgRNA design integrated with multi-omics. By prioritizing multiplex approaches for polygenic trait stacking, the review argues that these tools are essential to accelerate precision breeding for climate-adapted agriculture.

Process conditions and cell culture media components significantly impact glycosylation in ways that are still poorly understood. Here, we evaluated the glycan distribution of CHO-K1 clones producing adalimumab, a biosimilar version of the anti-TNF antibody Humira. To modulate the glycan profile, the cell culture medium was supplemented with manganese and/or galactose. Both manganese and galactose increased galactosylation, and the effects were synergistic. Notably, the levels of Man5 were higher than in Humira, even when galactosylation was similar. qPCR analysis revealed few differences in glycosylation enzyme concentrations between different productivity clones or when 50 µM manganese was added. We modeled the glycosylation pathways using a dynamic mathematical model to elucidate the mechanisms by which high mannose glycans increased and to develop a more predictive approach to culture modulation. Modeling results suggest that the concentration of uridine triphosphate (UTP), a component of activated sugars, is limiting and that increased uridine diphosphate (UDP)-galactose results in decreased UDP-N-acetylglucosamine, limiting complex glycan synthesis.

Proteolytic degradation of recombinant proteins in Chinese hamster ovary (CHO) cells remains a major challenge in biopharmaceutical manufacturing, often reducing product yield and quality. Matriptase-1, a type II transmembrane serine protease, has been identified as a key contributor to unwanted proteolysis. This study investigates hepatocyte growth factor activator inhibitor-1 (HAI-1) overexpression as a strategy to mitigate matriptase-1-mediated degradation in CHO cell culture. Using an IL-12 IgG1 Fc fusion protein and a multi-specific antibody (M molecule) as model proteins, we employed genetic and biochemical approaches to assess the impact of Chinese hamster HAI-1 overexpression on protein quality and yield. Our results demonstrate that HAI-1 overexpression effectively inhibits matriptase-1 activity, achieving up to a 98% reduction in proteolytic clipping while maintaining cell growth, viability, and product quality. Compared to other protease control strategies, HAI-1 overexpression presents a practical and scalable solution that does not disrupt essential cellular functions. These findings establish HAI-1 as a key modulatory tool in CHO-based protein production, with implications for reducing proteolysis of therapeutic proteins and optimizing biopharmaceutical manufacturing processes.

Bacillus coagulans is a preferred producer for lactic acid, still facing the challenge of poor carbon metabolic flow ability. Here, an engineered B. coagulans strain P14 with improved carbon metabolic flow ability was developed for the first time through heavy ion mutagenesis combined with an enrichment strategy of acetic acid and sodium acetate, showing increased titer, productivity and yield of L-lactic acid compared to the parental strain. Notably, the activities of four enzymes involved in the glycolytic pathway of B. coagulans P14 were also higher. For batch fermentations, the engineered B. coagulans P14 strain showed improved production economy than the parental strain, which can produce 124.6 g/L L-lactic acid with a productivity of 7.1 g/L/h and a yield of 93% in a 5-L bioreactor under unsterile conditions, showing an increase of 6.0%, 24.3%, and 8.1% compared to the parental strain, respectively. This work showcases a method for rapidly breeding a robust lactic acid-producing strain through heavy ion mutagenesis combined with an enrichment strategy.