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Smart responsive hybrid bioplatforms obtained by laser methods with tailored antibacterial and antitumor activity

Project code: PN-III-P2-2.1-PED-2019-2695

Contract number: 529PED

Supported by Romanian Ministry of Education and Research,

UEFISCDI - Executive Agency for Higher Education, Research, Development and Innovation Funding

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to About

    About    

The project scope is to develop an experimental prototype of a new generation of responsive hybrid smart platforms engineered by laser method, with tailored antibacterial and antitumor dual activity, addressing the current needs for research studies and medical applications (e.g. controlled drug delivery, development of next-generation precision medications testing platform, tunable interfaces for basic cell research).

The platform envisaged by this project consists in a pH and Temperature responsive copolymeric matrix (i.e. Poly(N-isopropylacrylamide-co-bu tylacrylate (polyNIPAM-BA)) in which bioactive compounds such as antimicrobial peptides AMPs, Hydrophobines and Hydrophobines–AMPs conjugates) are embedded by the use of Matrix Assisted Pulsed Laser Evaporation (MAPLE).

Based on the fact that tumor and bacteria cells exhibit more acidic medium and higher T compared to normal cells, this platform is designed for tumor and bacteria microenvironment-active /triggered drugs/bioactive compounds release.

To achieve its  goal, the project will follow 4 specific objectives:

1-Designing the new smart hybrid platform based on innovative and complex hybrid coatings (i.e. pNIPAM-co-BA with/without single active bio-compound);

2. Developing the laboratory MAPLE technology for obtaining hybrid coating with enhanced sensitivity, antitumor and antibacterial potential by tailoring deposition parameters (laser, fluence, target system and composition) and by correlating it with the hybrid coating characteristics (thickness, porosity, composition: various ratio of pNIPAM-co-BA, Hydrophobines, AMPs).

3. Optimizing the smart dual responsive engineered hybrid polymeric platforms with different compositions via laser methods and to demonstrate the bio-functionality using a tumor and bacteria microenvironment-active/triggere d release of bioactive compounds;

4. To validate the efficacity of the hybrid platform at laboratory level (TRL 4) for various tumoral cells and bacterial lines.

 

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Project duration: 09.11.2020 - 09.11.2022

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    Team   

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INFLPR team

 IBAR Team

Dr. Laurentiu Nicolae Rusen - CS III  / Project Manager

http://ssll.inflpr.ro/rusen/

Dr. Valentina Carmen Dinca -  CS I /  Member

Drd. Luminita Nicoleta Dumitrescu - Doctorand / Member

Drd. Anca Bonciu - Doctorand / Member

Dr. Iulia Anghel -  CS /  Member

Dr. Petronela Gheorghe -  CS /  Member

Dr. Antoniu Moldovan -  CS III /  Member

Dr. Adrian Petris -  CS I /  Member

Dr. Maria Dinescu -  CS I /  Member

Dr. Anca Roseanu - CS I / Partner manager

Dr. Mihaela Trif - CS I / Member

Dr. Paula Florian - CS / Member

Dr. Madalina Icriverzi - ACS / PostDoc Member

to Work

    Work    

First Phase 09.11.2020 - 31.12.2020

Preliminary coating design technology based on MAPLE and the target system.

Activities:

1.1.1-1.2 Measuring / characterizing the spatial, temporal and energetic the parameters of the laser beam. Optimization of the MAPLE experimental device. Design, development and implementation of a new multi-target support for MAPLE experiments to obtain coverage based on target system adaptation. - INFLPR

1.1.2 Preliminary tests to evaluate the cytotoxicity of substrates obtained by MAPLE technique. - IBAR

 

 

Results: In this stage, experiments were performed to measure / characterize the real values ​​of the parameters of the spatial, temporal and energetic beam. The MAPLE experimental device was optimized according to the project requirements. A new multi-target support for MAPLE experiments has been designed, developed and preliminarily tested to obtain the necessary coverage for the project. Preliminary p-films (NIPAM-co-BA) were made using different solvents. Preliminary tests were performed to evaluate the cytotoxicity of the substrates obtained by the MAPLE technique.

Second Phase 01.01.2021 - 31.12.2021

Activities:

2.1. Determination of laser processing parameters (fluence, number of pulses) and specifications of the target composition (concentration, solvent) for each component element. Adapting the MAPLE device to the project requirements. - INFLPR

2.2. Morphological and structural characterization of the platforms obtained from each component by analysis: AFM, SEM, FTIIR, contact angle . - INFLPR

2.3 Analysis of the double thermal and pH response of polyNIPAM-BA coatings. - INFLPR

2.4 Optimization of single component layers for the dual responsive platform. - INFLPR

2.5 Adaptation of the MAPLE target system for the deposition of hybrid layers based on polyNIPAM-BA_AMP-M, AMP-M_Hydrophobin, polyNIPAM-BA _Hydrophobin, polyNIPAM-BA_AMP-M_Hydrophobin. 2.6 Submission of hybrid coatings (polyNIPAM-BA_AMP, AMP-M_Hydrophobin, polyNIPAM-BA - INFLPR _Hydrophobin, polyNIPAM-BA_AMP-M_Hydrophobin) through MAPLE and the multiple target system. Morphological and structural characterization of the obtained deposits. Preliminary selection of the optimal platform. - INFLPR

2.7 In vitro evaluation of the viability and proliferation of murine melanoma cells cultured on single-element films (polyNIPAM-BA, Hydrophobin, AMP-M) and new complex hybrid coatings (ie pNIPAM-BA_Hydrophobin, polyNIPAM-BA_AMP-M, Hydrophob -M and polyNIPAM-BA_Hydrophobin_AMP-M).

2.8 Adhesion and morphology of murine melanoma cells cultured on single-element films (polyNIPAM-BA, hydrophobin, AMP) and new complex hybrid coatings (ie polyNIPAM-BA_Hydrophobin, polyNIPAM-BA_AMP-M, Hydrophobin_AMP-MP and poly -M). - IBAR

2.9 Preliminary studies on the effect of platforms on cell proliferation and cell morphology of metastatic murine melanoma. - IBAR

Results:

             In this stage, the laser processing parameters (fluency, number of pulses) and the specifications of the target composition (concentration, solvent) for each component element were determined. The MAPLE system has been adapted to the experimental requirements. Hybrid layers based on polyNIPAM-BA_AMP-M, AMP-M_Hydrophobin, polyNIPAM-BA_Hydrophobin, polyNIPAM-BA_AMP-M_Hydrophobin were obtained by the optimized MAPLE technique. The platforms obtained from each component were characterized morphologically and structurally by analysis: Atomic Force Microscopy (AFM), SEM, FTIR, contact angle. The double thermal and pH response of polyNIPAM-BA coatings was analyzed. During this stage, the research activity of the IBAR research group involved a study on the in vitro biological evaluation of simple (1 single element) and complex / hybrid coatings based on Poly (NIPAM-co-BA), Hydrophobin and AMP -M. To this end, the effect of these materials on the viability, proliferation, morphology and adhesion of normal human embryonic epithelial kidney cells HEK-293T and murine melanoma B16-F1 and B16-F10 was investigated.

             The project web page has been updated.

             The results obtained were disseminated through participation in international / national conferences.

             A scientific article is being prepared to be submitted to an ISI Journal.

Third Phase 01.01.2022 - 09.11.2022

Activities:

3.1 Analysis of the double thermal and pH response of the coatings based on polyNIPAM. - INFLPR

3.2 Selection and producing an optimal platform. Optimization and selection of the new intelligent hybrid polymer platform with double reaction. - INFLPR

3.3 Biological evaluation of simple and hybrid platforms. Studies on the effect of platforms on cell proliferation and the morphology of metastatic murine melanoma cells. - IBAR

3.4.1 Investigating the in vitro bio-functionality of the integrated intelligent platform. The capacity of the intelligent hybrid platform that contains antitumor and antibacterial compounds on the cell. - INFLPR

3.4.2 Evaluation and correlation of the chemical effects of the platform surface with the antibacterial capacity and the corresponding optimization. Defining the optimal composition of the platform to be validated according to bio-tests. - IBAR

 

Results:

            A new polymer platform based on pNIPAM-co-BA and the incorporation of 2 bioactive factors with antitumor and antibacterial action was obtained. The selection and optimization of the bioactive factors was done according to the surface characteristics. The stability of the films was subjected to conditions that mimicked normal and tumor or bacterial cell cultures, as well as based on the biological response given by the normal and tumor lines.

           Validation of the biological functionality of a new generation of hybrid polymer platforms was achieved by corroborating the results obtained by evaluating cell viability/proliferation (MTS assay, flow cytometry for apoptosis/necrosis and cell cycle), morphological analysis (SEM, fluorescence microscopy) highlighted the antitumor capacity of simple coatings with AMP and AMP-MG complexes (Magainin), property dependent on the type of cell line studied. The effect is due to the action of AMP and is modulated by the presence of MG in the case of complex coatings.

             The project web page has been updated.

             The results obtained were disseminated through participation in international / national conferences (1 oral presentation and 2 poster presentations) as well in 2 ISI articles.

             A scientific article is being prepared to be submitted to an ISI Journal.

 

Concluzii proiect 529PED:

           În cadrul proiectului 529PED-BioSMARTplat a fost dezvoltat un prototip experimental al unei noi generații de platforme polimerice hibride, bazate pe un polimer susceptibil la actiunea unor stimuli de temperatura şi pH şi care poate  încorpora  compuși bioactivi multifuncţionali (de ex. peptide cu acţiune antimicrobiană și antitumorală).

           Aceste platforme inteligente hibride au fost realizate prin metoda laser MAPLE (depunere laser pulsată asistată de o matrice - Matrix Assisted Pulse Laser Evaporation) şi sunt dedicate nevoilor actuale apărute în studiile de cercetare și domeniul aplicațiilor medicale.

             Un câștig important al implementarii proiectului îl reprezintă  impactul major pe care îl poate avea asupra pacienţilor  utilizarea unei astfel de matrice polimerice conţinând compuși biologici activi pentru reducerea contaminării bacteriene a dispozitivelor medicale și implicit al riscurilor de infecţie.  Totodată se extind perspectivele aplicaţiilor practice în direcţia dezvoltării de vehicule pentru livrarea controlată de medicamente, designul platformelor de testare a medicamentelor de ultimă generație, interfețe reglabile pentru cercetarea celulară de bază, dispozitive biomedicale performante.

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Papers

Results

-Olivia L. Rose, Anca Bonciu, Valentina Marascu, Andreea Matei, Qian Liu, Laurentiu Rusen, Valentina Dinca* and Cerasela Zoica Dinu / Thin Films of Metal-Organic Framework Interfaces Obtained by Laser Evaporation / Nanomaterials 2021, 11(6), 1367; https://doi.org/10.3390/nano11061367

-Dumitrescu, NLIcriverzi, MBonciu, AFlorian, PMoldovan, ARoseanu, ARusen, LDinca, V; Grama, F / New Poly(N-isopropylacrylamide-butylacrylate) Copolymer Biointerfaces and Their Characteristic Influence on Cell Behavior In Vitro / INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Volume 23 Issue7Article Number 3988 APR 2022 DOI 10.3390/ijms23073988

- Satulu, V; Dinca, V; Bacalum, M; Mustaciosu, C; Mitu, B; Dinescu, G  / Chemistry-Induced Effects on Cell Behavior upon Plasma Treatment of pNIPAAM / POLYMERS Volume 14 Issue 6 Article Number1081 MAR 2022 DOI 10.3390/polym14061081

Conferences

- N. Dumitrescu, A. Bonciu, A. Moldovan, V. Dinca and L. Rusen   / Preparation and Properties of P(N-Isopropylacrylamide-co-butylacrylate) Coatings using Matrix Assisted Pulsed Laser Evaporation / Oral presentation  at 13th International Conference on Physics of Advanced Materials (ICPAM-13) - 2021.

- M. Icriverzi, P. Florian, N. Dumitrescu, A. Bonciu, V. Dinca, L. Rusen, Anca Roseanu / CHARACTERIZATION AND IN VITRO BIOLOGICAL EVALUATION OF NOVEL OBTAINED PNIPAM-BASED COATINGS / Poster presenatation at The International Symposium "Priorities of Chemistry for a Sustainable Development" - PRIOCHEM XVIIth Edition 2021.

- V. Dinca, M. Icriverzi, A. Bonciu, L.N. Dumitrescu, A. Roseanu, L. Rusen, L.E. Sima, S. Nistorescu / Hierarchical biointerfaces obtained by laser processing and surface functionalization/ Poster presentation  at 13th International Conference on Physics of Advanced Materials (ICPAM-13) - 2021.

- P. Florian, M. Icriverzi, L.N. Dumitrescu, A. Bonciu, L. Rusen, V. Dinca, A. Roseanu / Novel pNIPAM-based coatings for medical applications: physical, chemical and biological characteristics / Oral presentations at International Conference Materials, Methods &Technologies 2022.

- L.N. Dumitrescu, A. Bonciu, M. Icriverzi, P. Florian, A. Moldovan, S. Nistorescu, A. Roseanu,  L. Rusen, V. Dinca / Biointerfaces based on Poly(N-isopropylacrylamide-butylacrylate) Copolymer obtained by MAPLE for bioengineering applications / Poster presentation at International Conference on Lasers, Plasma, and Radiation – Science and Technology ICLPR-ST 2022.

- L.N. Dumitrescu, L. Rusen, A. Bonciu, A. Moldovan, A. Roseanu, M. Icriverzi, V. Dinca, F. Grama / New poly(N-isopropylacrylamide-butylacrylate) copolymer biointerfaces and their characteristics influence on cells behavior in vitro / Poster presentations at 5th International Conference on Applied Surface Science ICASS-2022.

November 2022

    Contact    

Atomistilor 409 Street

Magurele, ILFOV,  077125

 

http://ssll.inflpr.ro/rusen/

Tel: to be added

Fax: 0214574243

Dr. Rusen Laurentiu Nicolae

email : laurentiu.rusen@inflpr.ro

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