Recent Patents on Biotechnology - Volume 15, Issue 4, 2021

Background: L-asparaginase (L-ASNase, L-asparagine amidohydrolase, E.C.3.5.1.1) is an enzyme with wide therapeutic applicability. Currently, the commercialized L-ASNase comes from mesophilic organisms, presenting low specificity to the substrate and limitations regarding thermostability and active pH range. Such factors prevent the maximum performance of the enzyme in different applications. Therefore, extremophilic organisms may represent important candidates for obtaining amidohydrolases with particular characteristics desired by the biotechnological market. Objectives: The present study aims to carry out a technological prospecting of patents related to the L-asparaginases derived from extremophilic organisms, contributing to pave the way for further rational investigation and application of such enzymes. Methods: This patent literature review used six patents databases: The LENS, WIPO, EPO, USPTO, Patent Inspiration, and INPI. Results: It was analyzed 2860 patents, and 14 were selected according to combinations of descriptors and study criteria. Approximately 57.14% of the patents refer to enzymes obtained from archaea, especially from the speciesPyrococcus yayanosii (35.71% of the totality). Conclusion: The present prospective study has singular relevance since there are no recent patent reviews for L-asparaginases, especially produced by extremophilic microorganisms. Although such enzymes have well-defined applications, corroborated by the patents compiled in this review, the most recent studies allude to new uses, such as the treatment of infections. The characterization of the catalytic profiles allows us to infer that there are potential sources still unexplored. Hence, the search for new L-ASNases with different characteristics will continue to grow in the coming years and, possibly, ramifications of the technological routes will be witnessed.

Background: Hutchinson-Gilford progeria syndrome (HGPS), also known as progeria of childhood or progeria is a rare, rapid, autosomal dominant genetic disorder characterized by premature aging which occurs shortly after birth. HGPS occurs as a result of de novo point mutation in the gene recognized as LMNA gene that encodes two proteins, Lamin A protein and Lamin C protein which are the structural components of the nuclear envelope. Mutations in the gene trigger abnormal splicing and induce internal deletion of 50 amino acids leading to the development of a truncated form of Lamin A protein known as Progerin. Progerin generation can be considered the crucial step in HGPS since the protein is highly toxic to human cells, permanently farnesylated, and exhibits variation in several biochemical and structural properties within the individual. HGPS also produces complications such as skin alterations, growth failure, atherosclerosis, hair and fat loss, and bone and joint diseases. We have also revised all relevant patents relating to Hutchinson-Gilford progeria syndrome and its therapy in the current article. Methods: The goal of the present review article is to provide information about Hutchinson- Gilford progeria syndrome (HGPS) and the use of CRISPR/Cas technology as a promising treatment approach in the treatment of the disease. The review also discusses about different pharmacological and non-pharmacological methods of treatment currently used for HGPS. Results: The main limitation associated with progeria is the lack of a definitive cure. The existing treatment modality provides only symptomatic relief. Therefore, it is high time to develop a therapeutic method that hastens premature aging in such patients. Conclusion: CRISPR/Cas technology is a novel gene-editing tool that allows genome editing at specific loci and is found to be a promising therapeutic approach for the treatment of genetic disorders such as HGPS where dominant-negative mutations take place.

Background: L-Methioninase (EC 4.4.1.11; MGL) is a pyridoxal phosphate (PLP)-dependent enzyme that is produced by a variety of bacteria, fungi, and plants. L-methioninase, especially from Pseudomonas and Citrobacter sp., is considered as the efficient therapeutic enzyme, particularly in cancers such as glioblastomas, medulloblastoma, and neuroblastoma that are more sensitive to methionine starvation. Objective: The low stability is one of the main drawbacks of the enzyme; in this regard, in the current study, different features of the enzyme, including phylogenetic, functional, and structural from Pseudomonas, Escherichia, Clostridium, and Citrobacter strains were evaluated to find the best bacterial L-Methioninase. Methods: After the initial screening of L-Methioninase sequences from the above-mentioned bacterial strains, the three-dimensional structures of enzymes from Escherichia fergusonii, Pseudomonas fluorescens, and Clostridium homopropionicum were determined through homology modeling via GalaxyTBM server and refined by GalaxyRefine server. Results and Conclusion: Afterwards, PROCHECK, verify 3D, and ERRAT servers were used for verification of the obtained models. Moreover, antigenicity, allergenicity, and physico-chemical analysis of enzymes were also carried out. In order to get insight into the interaction of the enzyme with other proteins, the STRING server was used. The secondary structure of the enzyme is mainly composed of random coils and alpha-helices. However, these outcomes should further be validated by wet-lab investigations.

Background: Methotrexate (MTX) is a general chemotherapeutic agent utilized to treat a variety of malignancies, woefully, its high doses can cause nephrotoxicity and subsequent defect in the process of MTX excretion. The recombinant form of glucarpidase is produced by engineered E. coli and is a confirmed choice to overcoming this problem. Objective: In the present study, in silico analyses were performed to select suitable SPs for the secretion of recombinant glucarpidase in E. coli. Methods: The signal peptide website and UniProt database were employed to collect the SPs and protein sequences. In the next step, SignalP-5.0 helped us to predict the SPs and the position of cleavage sites. Moreover, physicochemical properties and solubility were evaluated using Prot- Param and Protein-sol online software, and finally, ProtCompB was used to predict the final subcellular localization. Results: Luckily, all SPs could form soluble fusion proteins. At last, it was found that PPB and TIBA could translocate the glucarpidase into the extracellular compartment. Conclusion: This study showed that there are only 2 applicable SPs for the extracellular translocation of glucarpidase. Although the findings were remarkable with high degrees of accuracy and precision based on the utilization of bioinformatics analyses, additional experimental assessments are required to confirm and validate it. Recent patents revealed several inventions related to the clinical aspects of vaccine peptides against human disorders.

Background: All parts of Momordica charantia L. have potential hypoglycemic properties in reversing the metabolic disorder of diabetes mellitus. However, there exists a need for preparing an effective and safer formulation of active phytochemicals. We have also reviewed and analyzed certain patents on such preparatory methods for Momordica charantia L. formulations. Objective: This study aimed to isolate essential oil from the seeds of Momordica charantia L., analyze its phytochemicals, and study their anti-diabetic effects. Methods: The essential oil was isolated by the hydrodistillation method and analyzed for phytochemicals by GC-MS. Furthermore, its acute toxicity was tested in rats. Anti-diabetic effects were evaluated in Streptozotocin-induced diabetic rats with 17.5 and 55 mg/kg b.wt of essential oil by evaluating blood glucose, serum lipid profile, liver glycogen, protein, and other serum markers such as ALT, AST, ALP, urea, and creatinine. The histologic changes in the liver, pancreas, and kidney were evaluated using Haematoxylin and Eosin staining. Results: The phytochemicals having hypoglycaemic and insulin induction potency were identified in the GC-MS analysis. A highly significant (p≤0.01; p≤0.001) reduction in blood glucose was observed from 17.5 mg/kg and 55 mg/kg essential oil treatments, respectively. Diabetes-associated metabolic alterations (p≤0.001) observed in diabetic control rats such as lipid profile, enzymes, glycogen, protein, urea, and creatinine were normalized upon treatment with essential oil. Moreover, the histologic changes in vital organs reversed in treated rats. Conclusion: The essential oil of Momordica charantia L. seed has promising potency to normalize the metabolic changes of type II diabetes mellitus.