TOWARDS SMART BIOMANUFACTURING 4.0 OF RECOMBINANT PROTEIN PRODUCTION PROCESSES IN YEAST CELL FACTORIES

INDUSTRY 4.0 HAS BEEN GLOBALLY ACCEPTED BY GOVERNMENTS AND WORLDWIDE ECONOMIC ORGANIZATION AS A NEW INDUSTRIAL REVOLUTION WAVE LEVERAGED BY EMERGENT TECHNOLOGIES AS INTERNET OF THINGS (IOT), ARTIFICIAL INTELLIGENCE (AI), VIRTUAL AND AUGMENTED REALITY (AR) AND OTHER TECHNOLOGICAL INNOVATIONS DIRECTLY DRIVEN BY THE CAPABILITIES OF THE INTERNET AND POWER COMPUTING. THE FOURTH INDUSTRIAL REVOLUTION IS ALSO CALLED SMART INDUSTRY ACCORDING TO THE SMART CITIES MODEL, WHERE THE COMPETENCES BROUGHT BY IOT TECHNOLOGIES ARE INTEGRATED IN THE FACTORIES IN ORDER TO AUTOMATE INDUSTRIAL OPERATIONS. HOWEVER, THE INTRINSIC COMPLEXITY AND VARIABILITY OF BIOMANUFACTURING, MAKES ITS PATH TO DIGITALIZATION DIFFERENT AND MORE COMPLEX. UPSTREAM AND DOWNSTREAM PROCESSING MUST ADDRESS THE SINGULAR REQUIREMENTS OF LIVING CELLS AND RESPECTIVE PRODUCTS. THEREFORE, THE BIO-MANUFACTURING INDUSTRY, ESPECIALLY BIOPHARMA, IS STILL FAR FROM BEING FULLY AUTOMATED. IN FACT, MOST BIOPHARMA OPERATIONS ARE STILL ADAPTING FROM MANUAL TO AUTOMATED SYSTEMS (KNOWN AS INDUSTRY 3.0) BESIDES THE POTENTIAL OF INDUSTRY 4.0 OPERATIONS FOR IMPROVED PROCESS EFFICIENCY, MAXIMIZING PRODUCTIVITY WHILE MAINTAINING PRODUCT QUALITY, THE CURRENT BIOMANUFACTURING INDUSTRY ALSO REQUIRES ACCELERATING BIOPROCESS DEVELOPMENT, REDUCING TIME-TO-MARKET OF NEW BIOPRODUCTS, ESPECIALLY BIOPHARMACEUTICALS. AS A PARADIGMATIC EXAMPLE, THE COVID19 OUTBREAK HAS DEMONSTRATED THE NEED OF BIOTECHNOLOGICAL INDUSTRIES TO PRODUCE, IN A VERY SHORT TIME, VACCINES AND DIAGNOSTIC TESTS TO CONTROL AND MINIMIZE THE EFFECT OF THE PANDEMIC. THUS, IT IS A HIGH-PRIORITY OBJECTIVE THE DEVELOPMENT OF STANDARDISED AND PRE-OPTIMISED CELL FACTORIES PLATFORMS AND BIOPROCESSES ABLE TO DELIVER IN A COST-EFFECTIVE WAY ENOUGH AMOUNT OF TARGET BIOPRODUCTS UNDER SAFETY AND REPRODUCIBLE CONDITIONS. IN THIS CONTEXT, THIS PROJECT AIMS TO EMPOWER THE CURRENT BIOMANUFACTURING INDUSTRY, AND IN PARTICULAR, THE PRODUCTION OF BIOPHARMACEUTICALS, BY DEVELOPING AND INTEGRATING THE KEY TECHNOLOGIES THAT ENABLE THE TRANSITION TO SMART BIOMANUFACTURING ERA. AS A MODEL PRODUCTION PLATFORM, THE PROPOSAL FOCUSES ON THE PICHIA PASTORIS (KOMOGATAELLA PHAFFII) YEAST, A WIDELY USED CELL FACTORY. AS A MODEL PRODUCT, WE CONSIDER NANOBODIES (SINGLE DOMAIN ANTIBODIES, VHH), WHICH ARE ATTRACTIVE ALTERNATIVES TO CONVENTIONAL ANTIBODIES FOR MANY APPLICATIONS AS NEUTRALIZING DRUGS. TO THIS END, WE AIM TO UPGRADE THE CURRENT P. PASTORIS PLATFORM TO A “PICHIA4.0” SYSTEM. IT INCLUDES NOVEL COMPUTATIONALLY DESIGNED CHASSIS STRAINS, AN EXPANDED SYNTHETIC GENETIC BIOLOGY TOOLBOX, USE OF BIOSENSORS AND ADVANCED CULTURE STRATEGIES READY FOR FUTURE. IN THE PROJECT THE IMPLEMENTATION OF A DIGITAL TWIN (DT) FOR BIOPHARMACEUTICALS BIOMANUFACTURING WILL BE CARRIED OUT, WHERE THE APPLICATION OF ON-LINE SENSORS WILL SERVE AS A BRIDGE BETWEEN THE CELL FACTORY AND THE AI MODELS, DETECTING ANOMALIES AND PROVIDING PREDICTION FROM THE EDGE AND FROM THE CLOUD. IN ADDITION, THE AUTONOMOUS CONTROL IN REAL TIME OF THE BIOPROCESS ORCHESTRATED BY SEVERAL AI MODELS WILL PROVE THE 4.0 CAPABILITIES BROUGHT BY THE ADAPTIVE CONCEPT IN CONTINUED PROCESS VALIDATION (CPV). FINALLY, THE PLATFORM DEVELOPED IN A 5-L LAB BIOREACTOR WILL BE SCALE-UP TO A PILOT PLANT OF 60 L. IN SUMMARY, THE PROJECT IS FOCUSSED ON THE INTENSIFICATION AND TRANSITION TO SMART BIOMANUFACTURING OF MICROBIAL BIOPROCESSES USING CUTTING EDGE METHODOLOGIES IN THE FRAMEWORK OF THE BIOINDUSTRY 4.0 CONCEPT.