Wangikar, P. P. | Search bioRxiv

bioRxiv2024

Metabolomic profiling reveals grade-specific niacinamide accumulation and its therapeutic Potential via SIRT1-CD38-EMT axis modulation in cervical cancer progression

Despite the availability of new therapies for cervical cancer, innovative strategies are essential to address challenges related to drug resistance and high toxicity. The present study focuses on the metabolic profiling of cervical carcinoma using non-targeted metabolomics approach using liquid chromatography-mass spectrometry. Our study identified over 100 metabolites in cervical tissue samples (both cancerous and adjacent normal) using HILIC and reversed-phase chromatography in the positive and negative ionization modes. The major metabolic alterations included changes in glycolysis, the citric acid cycle, amino acid metabolism, nucleotide metabolism, and nicotinamide metabolism in a grade-dependent manner. Interestingly, metabolic differences between HPV-positive and HPV-negative tumors included elevated arginine and proline metabolism, nicotinamide metabolism, and phosphatidylcholine biosynthesis. We further validated our findings by analyzing transcriptomics datasets from the Gene Expression Omnibus database to understand the expression patterns of the underlying genes involved in the dysregulated pathways. We observed that nicotinamide metabolism exhibits significant effects in lower-grade cervical cancers and specific HPV genotypes. We treated cervical cancer cell lines with niacinamide (NAM), an amide form of niacin, to evaluate its potential therapeutic efficacy. NAM treatment modulated NAD+ metabolism with key players such as CD38, PARP, NAMPT, and SIRT1, promoting apoptosis and inhibiting cell proliferation in cervical cancer cells. Importantly, the metabolic responses differed between the HPV-positive SiHa cells and HPV-negative C33A cells, reflecting distinct NAD+ metabolic adaptations. The study highlights the metabolic adaptation or shifts in cancer progression and provides insights into NAMs molecular mechanisms and therapeutic potential for precision medicine in cervical cancer.

cancer biology↗

bioRxiv2018

A plug-and-play system for enzyme production at commercially viable levels in fed-batch cultures of Escherichia coli BL21 (DE3)

Commercial exploitation of enzymes in biotransformation necessitates a robust method for enzyme production that yields high enzyme titer. Nitrilases are a family of hydrolases that can transform nitriles to enantiopure carboxylic acids, which are important pharmaceutical intermediates. Here, we report a fed-batch method that uses a defined medium and involves growth under carbon limiting conditions using DO-stat feeding approach combined with an optimized post-induction strategy, yielding high cell densities and maximum levels of active and soluble enzyme. This strategy affords strict control of nutrient feeding and growth rates, and ensures sustained protein synthesis over a longer period. The method was optimized for highest titer of nitrilase reported so far (247 kU/l) using recombinant E. coli expressing the Alcaligenes sp. ECU0401 nitrilase. The fed-batch protocol presented here can also be employed as template to produce a wide variety of enzymes with minimal modification, as demonstrated for alcohol dehydrogenase and formate dehydrogenase.

bioengineering↗