Recent trends in targeted delivery of smart nanocarrier-based microbial enzymes for therapeutic applications.

Smart carrier-based immobilization has widened the use of enzymes for the treatment of several disorders. Large surface areas, tunable morphology, and surface modification ability aid the targeted and controlled release of therapeutic enzymes from such formulations. Smart nanocarriers, such as polymeric carriers, liposomes, and silica have also increased the stability, half-life, and permeability of these enzymes. In this review, summarize recent advances in the smart immobilization of microbial enzymes and their development as precision nanomedicine for the treatment of cancer, thrombosis, phenylketonuria (PKU), and wound healing. We also discuss the challenges and measures to be adopted for the successful clinical translation of these formulations.

In silico and in vitro analysis of arginine deiminase from Pseudomonas furukawaii as a potential anticancer enzyme.

Arginine deiminase (ADI), a promising anticancer enzyme from Mycoplasma hominis, is currently in phase III of clinical trials for the treatment of arginine auxotrophic tumors. However, it has been associated with several drawbacks in terms of low stability at human physiological conditions, high immunogenicity, hypersensitivity and systemic toxicity. In our previous work, Pseudomonas furukawaii 24 was identified as a potent producer of ADI with optimum activity under physiological conditions. In the present study, phylogenetic analysis of microbial ADIs indicated P. furukawaii ADI (PfADI) to be closely related to experimentally characterized ADIs of Pseudomonas sp. with proven anticancer activity. Immunoinformatics analysis was performed indicating lower immunogenicity of PfADI than MhADI (M. hominis ADI) both in terms of number of linear and conformational B-cell epitopes and T-cell epitope density. Overall antigenicity and allergenicity of PfADI was also lower as compared to MhADI, suggesting the applicability of PfADI as an alternative anticancer biotherapeutic. Hence, in vitro experiments were performed in which the ADI coding arcA gene of P. furukawaii was cloned and expressed in E. coli BL21. Recombinant ADI of P. furukawaii was purified, characterized and its anticancer activity was assessed. The enzyme was stable at human physiological conditions (pH 7 and 37 °C) with Km of 1.90 mM. PfADI was found to effectively inhibit the HepG2 cells with an IC50 value of 0.1950 IU/ml. Therefore, the current in silico and in vitro studies establish PfADI as a potential anticancer drug candidate with improved efficacy and low immunogenicity. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03292-2.

Microbial arginine deiminase: A multifaceted green catalyst in biomedical sciences.

Arginine deiminase is a well-recognized guanidino-modifying hydrolase that catalyzes the conversion of L-arginine to citrulline and ammonia. Their biopotential to regress tumors via amino acid deprivation therapy (AADT) has been well established. PEGylated formulation of recombinant Mycoplasma ADI is in the last-phase clinical trials against various arginine-auxotrophic cancers like hepatocellular carcinoma, melanoma, and mesothelioma. Recently, ADIs have attained immense importance in several other biomedical applications, namely treatment of Alzheimer's, as an antiviral drug, bioproduction of nutraceutical L-citrulline and bio-analytics involving L-arginine detection. Considering the wide applications of this biodrug, the demand for ADI is expected to escalate several-fold in the coming years. However, the sustainable production aspects of the enzyme with improved pharmacokinetics is still limited, creating bottlenecks for effective biopharmaceutical development. To circumvent the lacunae in enzyme production with appropriate paradigms of 'quality-by-design' an explicit overview of its properties with 'biobetter' formulations strategies are required. Present review provides an insight into all the potential biomedical applications of ADI along with the improvements required for its reach to clinics. Recent research advances with special emphasis on the development of ADI as a 'biobetter' enzyme have also been comprehensively elaborated.

In-vitro and in-silico evaluation of the anti-chikungunya potential of Psidium guajava leaf extract and their synthesized silver nanoparticles.

Chikungunya is a notorious viral infection, which affects a large segment of world populations in absence of vaccines and antivirals. The current study evaluates of anti-chikungunya activities of Psidium guajava leaves extract and their green synthesized silver nanoparticles. Green synthesized nanoparticles were well characterized for their size and stability by dynamic light scattering (DLS), zeta potential, scanning electron microscopy (SEM) and their functional groups were analyzed by FTIR. Maximum non-toxic doses (MNTD) of extracts and nanoparticles were analysed by using Vero cell-lines. Anti-chikungunya activities of extracts and nano-particles were determined on Vero cells and their effects on cell viability were measured by MTT assay. The P. guajava nano-particles and extracts revealed the anti-chikungunya activities in the Vero cell. The cells viability was increased by 40% and 60% as compared to the virus control, when these cells were treated with MNTD of P. guajava nano-particles and extracts, respectively. To know the reason for antiviral activity, molecular docking of phytochemicals was done against a replication essential cysteine protease (nsP2) of Chikungunya. It was found that phytochemicals; Longifollen and Quercetin showed the minimum binding energy with nsP2. P. guajava extracts can be exploited to develop an effective anti-chikungunya agent. In the absence of CHIKV vaccines and antivirals, P. guajava may be used to develop rapid, responsive, specific, and cost-effective anti-chikungunya agents. Supplementary Information: The online version contains supplementary material available at 10.1007/s13337-021-00685-4.

Microbial Enzymes used in Prodrug Activation for Cancer Therapy: Insights and Future Perspectives.

Enzyme prodrug therapy has gained momentum in recent years due to its ability to improve therapeutic index (benefits versus toxic side-effects) and efficacy of chemotherapy in cancer treatment. Inactive prodrugs used in this system are converted into active anti-cancerous drugs by enzymes, specifically within the tumor cells. This therapy involves three components namely prodrug, enzyme and gene delivery vector. Past reports have clearly indicated that the choice of enzyme used is the major determinant for the success of this therapy. Generally, enzymes from nonhuman sources are employed to avoid off-target toxicity. Exogenous enzymes also give better control to the clinician regarding the calibration of treatment by site-specific initiation. Amongst these exo-enzymes, microbial enzymes are preferred due to their high productivity, stability and ease of manipulation. The present review focuses on the commonly used microbial enzymes, particularly cytosine deaminase, nitroreductase, carboxypeptidase, purine nucleoside phosphorylase in prodrug activation therapy. Various aspects viz. source of the enzymes, types of cancer targeted, mode of action and efficacy of the enzyme/prodrug system, efficient vectors used and recent research developments of each of these enzymes are comprehensively elaborated. Further, the results of the clinical trials and various strategies to improve their clinical applicability are also discussed.

Biomedical applications of microbial phenylalanine ammonia lyase: Current status and future prospects.

Phenylalanine ammonia lyase (PAL) has recently emerged as an important therapeutic enzyme with several biomedical applications. The enzyme catabolizes l-phenylalanine to trans-cinnamate and ammonia. PAL is widely distributed in higher plants, some algae, ferns, and microorganisms, but absent in animals. Although microbial PAL has been extensively exploited in the past for producing industrially important metabolites, its high substrate specificity and catalytic efficacy lately spurred interest in its biomedical applications. PEG-PAL drug named Palynziq™, isolated from Anabaena variabilis has been recently approved for the treatment of adult phenylketonuria (PKU) patients. Further, it has exhibited high potency in regressing tumors and treating tyrosine related metabolic abnormalities like tyrosinemia. Several therapeutically valuable metabolites have been biosynthesized via its catalytic action including dietary supplements, antimicrobial peptides, aspartame, amino-acids, and their derivatives. This review focuses on all the prospective biomedical applications of PAL. It also provides an overview of the structure, production parameters, and various strategies to improve the therapeutic potential of this enzyme. Engineered PAL with improved pharmacodynamic and pharmacokinetic properties will further establish this enzyme as a highly efficient biological drug.