Development of novel platforms for PEGylation of proteins with therapeutic potential using microfluidics

• Grant number: 18/25994-2
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: September 01, 2019
• End date: June 21, 2021
• Field of knowledge: Engineering - Chemical Engineering - Chemical Process Industries
• Principal Investigator: Carlota de Oliveira Rangel Yagui
• Grantee: João Henrique Picado Madalena Santos
• Host Institution: Faculdade de Ciências Farmacêuticas (FCF). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 13/08617-7 - Production of extracellular L-asparaginase: from bioprospecting to the engineering of an antileukemic biopharmaceutical, AP.TEM

Abstract

PEGylation of therapeutic proteins has been employed to improve biopharmaceuticals clinical performance (considering immunogenicity and plasma half-life). Despite the advantages of PEGylated biopharmaceuticals, the parallel reactions between reactive PEG and the functional amino acids of proteins result in heterogeneity of PEGylated species. This problem associated with the hydrolysis of reactive PEG is limitations of this technique. In addition, the purification of the PEGylated proteins is complex and inefficient, compromising the economical viability of the process. In this project the study of more efficient PEGylation reactions for the model proteins L-asparaginase and catalase using microfluidic technology is proposed. The continuous PEGylation reaction process will be developed in micro-reactors and aqueous biphasic systems (ABS) will be investigated as purification technology, both by microfluidics and by the centrifugal partition chromatography (CPC) platforms. As expected results, it is intended to develop a site-specific PEGylation reaction in micro-reactors for the two therapeutic proteins in a continuous flow reactive media, with higher yield, shorter reaction time and applying less amount of reactive PEG than is currently practiced on a laboratory scale (batch process). Another expected result will be the development of a continuous purification process with ABS associated with the microfluidic reaction stage. Optimization of the continuous purification technique using CPC technology will also be implemented, allowing the efficient separation of PEGylated protein forms with high purity.

Scientific publications

(13)

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

SILVA, AMANDA R. P.; GUIMARAES, MARINA S.; RABELO, JHENIFFER; HERRERA BELEN, LISANDRA; JOSE PERECIN, CAIO; FARIAS, JORGE G.; SANTOS, JOAO H. P. M.; RANGEL-YAGUI, CARLOTA O.. Recent advances in the design of antimicrobial peptide conjugates. JOURNAL OF MATERIALS CHEMISTRY B, v. 10, n. 19, p. 15-pg., 2022-02-22. (18/25994-2)

D'ANGELO, NATALIA A.; NORONHA, MARIANA A.; CAMARA, MAYRA C. C.; KURNIK, ISABELLE S.; FENG, CHUYING; ARAUJO, VICTOR H. S.; SANTOS, JOAO H. P. M.; FEITOSA, VALKER; V.D. MOLINO, JOAO; RANGEL-YAGUI, CARLOTA O.; et al. Doxorubicin nanoformulations on therapy against cancer: An overview from the last 10 years. BIOMATERIALS ADVANCES, v. 133, p. 23-pg., 2022-02-01. (18/10799-0, 18/25994-2, 17/10789-1, 20/03727-2, 19/08549-8)

SANTOS, JOAO H. P. M.; FEITOSA, VALKER A.; MENEGUETTI, GIOVANNA P.; CARRETERO, GUSTAVO; COUTINHO, JOAO A. P.; VENTURA, SONIA P. M.; RANGEL-YAGUI, CARLOTA O.. Lysine-PEGylated Cytochrome C with Enhanced Shelf-Life Stability. BIOSENSORS-BASEL, v. 12, n. 2, p. 13-pg., 2022-02-01. (16/22065-5, 18/25994-2)

SANTOS, JOAO H. P. M.; MENDONCA, CARLOS M. N.; SILVA, AMANDA R. P.; OLIVEIRA, RICARDO P. S.; PESSOA JR, ADALBERTO; COUTINHO, JOAO A. P.; VENTURA, SONIA P. M.; RANGEL-YAGUI, CARLOTA O.. An integrated process combining the reaction and purification of PEGylated proteins. GREEN CHEMISTRY, v. 21, n. 23, p. 6407-6418, DEC 7 2019. (16/22065-5, 13/08617-7, 18/25994-2)

SANTOS, JOAO H. P. M.; MARTINS, MARGARIDA; SILVA, AMANDA R. P.; CUNHA, JHENIFFER R.; RANGEL-YAGUI, CARLOTA O.; VENTURA, SONIA P. M.. Imidazolium-based Ionic Liquids as Adjuvants to Form Polyethylene Glycol with Salt Buffer Aqueous Biphasic Systems. JOURNAL OF CHEMICAL AND ENGINEERING DATA, v. 65, n. 8, p. 3794-3801, AUG 13 2020. (16/22065-5, 18/25994-2)

MAGRI, AGNES; PIMENTA, MARCELA V.; SANTOS, JOAO H. P. M.; COUTINHO, JOAO A. P.; VENTURA, SONIA P. M.; MONTEIRO, GISELE; RANGEL-YAGUI, CARLOTA O.; PEREIRA, JORGE F. B.. Controlling the l-asparaginase extraction and purification by the appropriate selection of polymer/salt-based aqueous biphasic systems. JOURNAL OF CHEMICAL TECHNOLOGY AND BIOTECHNOLOGY, v. 95, n. 4, p. 1016-1027, APR 2020. (13/08617-7, 18/15104-0, 14/16424-7, 18/25994-2, 14/19793-3, 15/07749-2)

TORRES-OBREQUE, KARIN M.; MENEGUETTI, GIOVANNA P.; MUSO-CACHUMBA, JORGE J.; FEITOSA, VALKER A.; SANTOS, JOAO H. P. M.; VENTURA, SONIA P. M.; RANGEL-YAGUI, CARLOTA O.. Building better biobetters: From fundamentals to industrial application. DRUG DISCOVERY TODAY, v. 27, n. 1, p. 65-81, JAN 2022. (16/22065-5, 18/25994-2)

SOARES, BRUNA P.; SANTOS, JOAO H. P. M.; MARTINS, MARGARIDA; ALMEIDA, MAFALDA R.; SANTOS, V, NATHALIA; FREIRE, MARA G.; SANTOS-EBINUMA, VALERIA C.; COUTINHO, JOAO A. P.; PEREIRA, JORGE F. B.; VENTURA, SONIA P. M.. Purification of green fluorescent protein using fast centrifugal partition chromatography. Separation and Purification Technology, v. 257, FEB 15 2021. (14/16424-7, 18/25994-2, 16/07529-5)

PILLACA-PULLO, OMAR S.; INTIQUILLA, ARTURO; SANTOS, JOAO H. P. M.; SANCHEZ-MOGUEL, IGNACIO; BRANDELLI, ADRIANO; ZAVALETA, I, AMPARO. urification of Pseudomonas sp. proteases through aqueous biphasic systems as an alternative source to obtain bioactive protein hydrolysate. BIOTECHNOLOGY PROGRESS, v. 37, n. 3, APR 2020. (18/25994-2)

FREIRE, ROMINNE KARLA BARROS; MENDONCA, CARLOS MIGUEL NOBREGA; FERRARO, RAFAEL BERTELLI; MOGUEL, IGNACIO SANCHEZ; TONSO, ALDO; LOURENCO, FELIPE REBELLO; SANTOS, JOAO HENRIQUE PICADO MADALENA; SETTE, LARA DURAES; PESSOA JUNIOR, ADALBERTO. Glutaminase-free L-asparaginase production byLeucosporidium muscorumisolated from Antarctic marine-sediment. PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY, v. 51, n. 3, SEP 2020. (18/25994-2, 13/08617-7)

SANTOS, JOAO H. P. M.; BRUMANO, LARISSA P.; PESSOA, JR., ADALBERTO; RANGEL-YAGUI, CARLOTA O.. Tailoring Protein PEGylation Reaction: An Undergraduate Laboratory Experiment. Journal of Chemical Education, v. 97, n. 5, p. 1443-1447, MAY 12 2020. (17/21819-9, 16/22065-5, 13/08617-7, 18/25994-2)

TORRES-OBREQUE, KARIN; KLEINGESINDS, EDUARDO KREBS; SANTOS, JOAO H. P. M.; CARRETERO, GUSTAVO; RABELO, JHENIFFER; CONVERTI, ATTILIO; MONTEIRO, GISELE; PESSOA JR, ADALBERTO; RANGEL-YAGUI, CARLOTA O.. PEGylation versus glycosylation: effect on the thermodynamics and thermostability of crisantaspase. PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY, v. N/A, p. 11-pg., 2023-09-11. (16/22065-5, 17/20384-9, 13/08617-7, 18/25994-2)

MARTINS, MARGARIDA; SOARES, BRUNA P.; SANTOS, JOAO H. P. M.; BHARMORIA, PANKAJ; ACOSTA, MARIO A. TORRES; DIAS, V, ANA C. R.; COUTINHO, JOAO A. P.; VENTURA, SONIA P. M.. Sustainable Strategy Based on Induced Precipitation for the Purification of Phycobiliproteins. ACS SUSTAINABLE CHEMISTRY & ENGINEERING, v. 9, n. 10, p. 3942-3954, MAR 15 2021. (18/25994-2)