Oligonucleotides: Current Trends and Innovative Applications in the Synthesis, Characterization, and Purification.

biopharmaceuticals chromatography continuous chromatography drugs gene therapy multicolumn solvent gradient process oligonucleotides solid-phase

Journal

Biotechnology journal
ISSN: 1860-7314
Titre abrégé: Biotechnol J
Pays: Germany
ID NLM: 101265833

Informations de publication

Date de publication:
Aug 2020
Historique:
received: 22 01 2020
revised: 17 03 2020
pubmed: 17 4 2020
medline: 27 2 2021
entrez: 17 4 2020
Statut: ppublish

Résumé

Oligonucleotides (ONs) are gaining increasing importance as a promising novel class of biopharmaceuticals. Thanks to their fundamental role in gene regulation, they can be used to develop custom-made drugs (also called N-to-1) able to act on the gene expression at pre-translational level. With recent approvals of ON-based therapeutics by the Food and Drug Administration (FDA), a growing demand for high-quality chemically modified ONs is emerging and their market is expected to impressively prosper in the near future. To satisfy this growing market demand, a scalable and economically sustainable ON production is needed. In this paper, the state of the art of the whole ON production process is illustrated with the aim of highlighting the most promising routes toward the auspicated market-size production. In particular, the most recent advancements in both the upstream stage, mainly based on solid-phase synthesis and recombinant technology, and the downstream one, focusing on chromatographic techniques, are reviewed. Since ON production is projected to expand to the large scale, automatized multicolumn countercurrent technologies will reasonably be required soon to replace the current ones based on batch single-column operations. This consideration is supported by a recent cutting-edge application of continuous chromatography for the ON purification.

Identifiants

pubmed: 32298041
doi: 10.1002/biot.201900226
doi:

Substances chimiques

Biological Products 0
Oligonucleotides 0

Types de publication

Journal Article Review

Langues

eng

Sous-ensembles de citation

IM

Pagination

e1900226

Subventions

Organisme : Italian University and Scientific Research Ministry
ID : PRIN 2017Y2PAB8 003

Informations de copyright

© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Références

B. Ravi, A. Antonellis, C. J. Sumner, A. P. Lieberman, Hum. Mol. Genet. 2019, 28, R55.
D. M. Ozata, I. Gainetdinov, A. Zoch, D. O'Carroll, P. D. Zamore, Nat. Rev. Genet. 2019, 20, 89.
J. Li, F. Yu, Y. Chen, D. Oupický, J. Controlled Release 2015, 219, 369.
F. Atyabi, F. Zahir, F. Khonsari, A. Shafiee, F. Mottaghitalab, in Nanostructures for Cancer Therapy, Elsevier, New York 2017, pp. 541-561.
M. J. Glennie, P. W. Johnson, Immunol. Today 2000, 21, 403.
P. Chames, M. Van Regenmortel, E. Weiss, D. Baty, Br. J. Pharmacol. 2009, 157, 220.
A. L. Nelson, E. Dhimolea, J. M. Reichert, Nat. Rev. Drug Discovery 2010, 9, 767.
S. L. Ginn, I. E. Alexander, M. L. Edelstein, M. R. Abedi, J. Wixon, J. Gene Med. 2013, 15, 65.
J. A. Gilbert, M. J. Blaser, J. G. Caporaso, J. K. Jansson, S. V. Lynch, R. Knight, Nat. Med. 2018, 24, 392.
S. Goodwin, J. D. McPherson, W. R. McCombie, Nat. Rev. Genet. 2016, 17, 333.
P. M. Visscher, N. R. Wray, Q. Zhang, P. Sklar, M. I. McCarthy, M. A. Brown, J. Yang, Am. J. Hum. Genet. 2017, 101, 5.
V. G. LeBlanc, M. A. Marra, Cancers 2015, 7, 1925.
T. Meißner, K. M. Fisch, L. Gioia, A. I. Su, BMC Med. Genomics 2015, 8, 24.
L. Gold, J. Biol. Chem. 1995, 270, 13581.
P. Y. Ho, A.-M. Yu, Wiley Interdiscip. Rev.: RNA 2016, 7, 186.
S. M. Hoy, Drugs 2018, 78, 1625.
S. J. Keam, Drugs 2018, 78, 1371.
J. Kim, C. Hu, C. Moufawad El Achkar, L. E. Black, J. Douville, A. Larson, M. K. Pendergast, S. F. Goldkind, E. A. Lee, A. Kuniholm, A. Soucy, J. Vaze, N. R. Belur, K. Fredriksen, I. Stojkovska, A. Tsytsykova, M. Armant, R. L. DiDonato, J. Choi, L. Cornelissen, L. M. Pereira, E. F. Augustine, C. A. Genetti, K. Dies, B. Barton, L. Williams, B. D. Goodlett, B. L. Riley, A. Pasternak, E. R. Berry, K. A. Pflock, S. Chu, C. Reed, K. Tyndall, P. B. Agrawal, A. H. Beggs, P. E. Grant, D. K. Urion, R. O. Snyder, S. E. Waisbren, A. Poduri, P. J. Park, A. Patterson, A. Biffi, J. R. Mazzulli, O. Bodamer, C. B. Berde, T. W. Yu, N. Engl. J. Med. 2019, 381, 1644.
C. A. Stein, D. Castanotto, Mol. Ther. 2017, 25, 1069.
R. L. Juliano, Nucleic Acids Res. 2016, 44, 6518.
S. T. Crooke, S. Wang, T. A. Vickers, S. Wen, X.-H. Liang, Nat. Biotechnol. 2017, 35, 230.
A. Khvorova, J. K. Watts, Nat. Biotechnol. 2017, 35, 238.
F. Eckstein, Nucleic Acid Ther. 2014, 24, 374.
R. Z. Yu, J. S. Grundy, R. S. Geary, Expert Opin. Drug Metab. Toxicol. 2013, 9, 169.
C. Godfrey, L. Desviat, B. Smedsrød, F. Piétri-Rouxel, M. Denti, P. Disterer, S. Lorain, G. Nogales-Gadea, V. Sardone, R. Anwar, S. El Andaloussi, T. Lehto, B. Khoo, C. Brolin, W. van Roon-Mom, A. Goyenvalle, A. Aartsma-Rus, V. Arechavala-Gomeza, EMBO Mol. Med. 2017, 9, 545.
P. Rimessi, P. Sabatelli, M. Fabris, P. Braghetta, E. Bassi, P. Spitali, G. Vattemi, G. Tomelleri, L. Mari, D. Perrone, A. Medici, M. Neri, M. Bovolenta, E. Martoni, N. M. Maraldi, F. Gualandi, L Merlini, M Ballestri, L Tondelli, K Sparnacci, P Bonaldo, A Caputo, M Laus, A Ferlini, Mol. Ther. 2009, 17, 820.
K. E. Lundin, O. Gissberg, C. I. E. Smith, R. Zain, Methods Mol. Biol. 2019, 2036, 3.
S. T. Crooke, S. Wang, T. A. Vickers, W. Shen, X. H. Liang, Nat. Biotechnol. 2017, 35, 230.
S. Wang, N. Allen, T. A. Vickers, A. S. Revenko, H. Sun, X. H. Liang, S. T. Crooke, Nucleic Acids Res. 2018, 46, 3279.
P. Martin, Helv. Chim. Acta 1995, 78, 486.
T. P. Prakash, Chem. Biodiversity 2011, 8, 1616.
P. S. G. E. M. Pallan, P. A. Jicman, R. K. Pandey, M. Manoharan, E. Rozners, M. Egli, Nucleic Acids Res. 2011, 39, 3482.
S. M. Gryaznov, Chem. Biodiversity 2010, 7, 477.
X. Wang, C. S. Hu, B. Petersen, J. Qiu, F. Ye, J. Houldsworth, K. Eng, F. Huang, R. Hoffmann, Blood Adv. 2018, 2, 2378.
A. Vater, S. Klussmann, Drug Discovery Today 2015, 20, 147.
A. Soler-Bisté, A. Zorreguieta, M. E. Tolmasky, Molecules 2019, 24, 2297.
S. K. Singh, P. Nielsen, A. A. Koshkin, J. Wengel, Chem. Commun. 1998, 455.
S. K. Singh, J. Wengel, Chem. Commun. 1998, 1247.
C. Thorpe, S. Epple, B. Woods, A. H. El-Sagheera, T. Brown, Org. Biomol. Chem. 2019, 17, 5341.
D. Ittig, S. Liu, D. Rennember, D. Schumperli, C. J. Leumann, Nucleic Acids Res. 2004, 32, 346.
K. Relizani, G. Griffith, L. Echevarrìa, F. Zarrouki, P. Facchinetti, C. Vaillend, C. Laumann, L. Garcia, A. Goyenvalle, Mol. Ther.-Nucleic Acids 2017, 8, 144.
K. R. Lim, R. Mauyama, T. Yokota, Drug Des., Dev. Ther. 2017, 11, 533.
S. Montazersaheb, M. S. Hejazi, C. H. Nozad, Adv. Pharm. Bull. 2018, 8, 551.
D. Pfister, L. Nicoud, M. Morbidelli, Continuous Biopharmaceutical Processes: Chromatography, Bioconjugation, and Protein Stability, Cambridge University Press, Cambridge 2018.
D. M. Ecker, S. D. Jones, H. L. Levine, mAbs 2015, 7, 9.
R. K. Kumar, A. P. Guzaev, C. Rentel, V. T. Ravikumar, Tetrahedron 2006, 62, 4528.
M. Septak, Nucleic Acids Res. 1996, 24, 3053.
D. Chen, Z. Yan, D. L. Cole, G. S. Srivatsa, Nucleic Acids Res. 1999, 27, 389.
K. L. Fearon, J. T. Stults, B. J. Bergot, L. M. Christensen, A. M. Raible, Nucleic Acids Res. 1995, 23, 2754.
J. Temsamami, M. Kubert, S. Agrawal, Nucleic Acids Res. 1995, 23, 1841.
X. Wei, Tetrahedron 2013, 69, 3615.
D. Capaldi, H. J. Gaus, R. L. Carty, M. N. Moore, B. J. Turney, S. D. Decottignies, J. V. McArdle, A. N. Scozzari, V. T. Ravikumar, A. H. Krotz, Bioorg. Med. Chem. Lett. 2004, 14, 4683.
A. A. Rodriguez, I. Cedillo, A. K. McPherson, Bioorg. Med. Chem. Lett. 2016, 26, 3468.
A. Rodriguez, I. Cedillo, B. P. Mowery, H. J. Gaus, S. S. Krishnamoorty, A. McPherson, Bioorg. Med. Chem. Lett. 2014, 24, 3243.
D. C. Capaldi, H. Gaus, A. H. Krotz, J. Arnold, R. L. Carty, N. M. Moore, A. N. Scozzari, K. Lowery, D. L. R. V. T. Cole, Org. Process Res. Dev. 2003, 7, 832.
W. B. Wan, M. T. Migawa, G. Vasquez, H. M. Murray, J. G. Nichols, H. Gaus, A. Berdeja, S. Lee, C. E. Hart, W. F. Lima, E. E. Swayze, P. P. Seth, Nucleic Acids Res. 2014, 42, 13456.
N. Iwamoto, D. C. D. Butler, N. Svrzikapa, S. Mohapatra, I. S. D. W. Y. Zlatev, Meena, S. M. S, G. Lu, L. H. Apponi, M. Frank-Kamenetsky, J. J. Zhang, C. Vargeese, G. L. Verdine, Nat. Biotechnol. 2017, 35, 845.
J. Summerton, D. Weller, Antisense Nucleic Acid Drug Dev. 1997, 7, 187.
L. Christensen, R. Fitzpatrick, B. Gildea, K. H. Petersen, H. F. Hansen, T. Koch, M. B. O. Egholm, P. E. Nielsen, J. Coull, R. H. Berg, J. Pept. Sci. 1995, 1, 175.
K. L. Fearon, J. S. Nelson, Curr. Protoc. Nucleic Acid Chem. 2000, 3, 4.7.1.
E. Paredes, V. Aduda, K. Ackley, H. Cramer, Compr. Med. Chem. III 2017, 233.
T. R. Shepherd, R. R. Du, H. Huang, E.-C. Wamhoff, M. Bathe, Sci. Rep. 2019, 9, 6121.
H. Lönnberg, Beilstein J. Org. Chem. 2017, 13, 1368.
A.-M. Yu, C. Jian, A. H. Yu, M.-J. Tu, Pharmacol. Ther. 2019, 196, 91.
L. Ponchon, G. Beauvais, S. Nonin-Lecomte, F. Dardel, Nat. Protoc. 2009, 4, 947.
L. Ponchon, F. Dardel, Nat. Methods 2007, 4, 571.
L. Huang, J. Jin, P. Deighan, E. Kiner, L. McReynolds, J. Lieberman, Nat. Biotechnol. 2013, 31, 350.
Y. Liu, V. G. Stepanov, U. Strych, R. C. Willson, G. W. Jackson, G. E. Fox, BMC Biotechnol. 2010, 10, 85.
P. Y. Ho, Z. Duan, N. Batra, J. L. Jilek, M.-J. Tu, J.-X. Qiu, Z. Hu, T. Wun, P. N. Lara, R. W. {DeVere White}, H.-W. Chen, A.-M. Yu, J. Pharmacol. Exp. Ther. 2018, 365, 494.
P. Pereira, A. Q. Pedro, J. Tomás, C. J. Maia, J. A. Queiroz, A. Figueiras, F. Sousa, Appl. Microbiol. Biotechnol. 2016, 100, 3723.
S. Simoens, I. Huys, Gene Ther. 2017, 24, 539.
X. Yang, in Manual of Industrial Microbiology and Biotechnology, ASM Press, Washington, DC 2010, pp. 669-675.
S. Xu, H. Chen, J. Biotechnol. 2016, 231, 149.
D. J. Karst, F. Steinebach, M. Morbidelli, Curr. Opin. Biotechnol. 2018, 53, 76.
N. M. El Hazar, N. Magdy, A. M. El-Kosasy, M. G. Bartlett, Biomed. Chromatogr. 2017, 32, 4088.
A. K. Palm, G. Marko-Varga, J. Pharm. Biomed. Anal. 2004, 35, 415.
J. Carmody, B. Noll, in Handbook of Analysis of Oligonucleotides and Related Products (Eds: J. V. Bonilla, G. S. Srivatsa), CRC Press, Boca Raton, FL 2011, pp. 243-264.
D. Wegman, F. Ghasemi, A. F. Stasheuski, A. Khorshidi, B. B. Yang, S. K. Liu, G. M. I. Yousef, S. G. Krylov, Anal. Chem. 2016, 88, 2472.
L. Greenough, K. M. Schermerhorn, L. Mazzola, J. Bybee, D. Rivizzigno, E. Cantin, B. E. Slatko, A. F. Gardner, Nucleic Acids Res. 2016, 44, e15.
J. Wang, Y. Qin, S. Jiang, L. Liu, Y. Lu, J. Li, l. Qiu, P. Jiang, Sens. Actuators, B 2016, 237, 106.
V. Datinská, K. Klepárník, B. Belšánová, M. Minárik, F. Foret, J. Sep. Sci. 2018, 41, 2961.
V. Bosi, E. Sarti, N. M. L., D. P. L. Perrone, A. Cavazzini, M. L. Capobianco, Anal. Bioanal. Chem. 2015, 407, 5405.
V. Dolnik, J. Liu, J. F. Banks Jr., M. V. Novotny, P. Boček, J. Chromatogr. A 1989, 480, 321.
A. T. H. Le, S. M. Krylova, M. Kanoatov, S. Desai, S. N. Krylov, Angew. Chem., Int. Ed. 2019, 58, 2739.
A. T. H. Le, S. M. Krylova, S. M. Krylov, Anal. Chem. 2019, 91, 8532.
A. T. H. Le, S. M. Krylova, S. N. Krylov, Electrophoresis 2019, 40, 2553.
C. Fu, S. Smith, S. G. Simkins, P. F. Agris, Anal. Biochem. 2002, 306, 135.
V. D'Atri, T. Causon, O. Hernandez-Alba, A. Mutabazi, J.-L. Veuthey, S. Cianferani, D. Guillarme, J. Sep. Sci. 2018, 41, 20.
A. Goyon, P. Yehl, K. Zhang, J. Pharm. Biomed. Anal. 2020, 182, 113105.
M. Biba, J. P. Foley, C. J. Welch, in Liquid Chromatography (Eds: S. Fanali, P. R. Haddad, C. F. Poole, M. L. Riekkola), Elsevier, Amsterdam 2017, pp. 159-182.
L. Wang, C. Li, Bioanalysis 2016, 8, 143.
Q. Tian, J. Rogness, M. Meng, Z. Li, Bioanalysis 2017, 9, 861.
S. Studzińska, Talanta 2018, 176, 329.
A. Kaczmarkiewicz, L. Nuckowski, S. Studzinska, B. Buszewski, Crit. Rev. Anal. Chem. 2019, 49, 256.
A. C. McGinnis, B. Chen, M. G. Bartlett, J. Chromatogr. B 2012, 883-884, 76.
C. G. Huber, P. J. Oefner, G. K. Bonn, Anal. Biochem. 1993, 212, 351.
M. Gilar, K. J. Fountain, Y. Budman, U. D. Neue, K. R. Yardley, P. D. Rainville, R. J. Russell II, J. C. Gebler, J. Chromatogr. A 2002, 958, 167.
C. Huber, P. Oefner, G. Bonn, J. Chromatogr. A 1992, 599, 113.
L. Li, T. Leone, J. P. Foley, C. J. Welch, J. Chromatogr. A 2017, 1500, 84.
M. Enmark, M. Rova, J. Samuelsson, E. Örnskov, F. Schweikart, T. Fornstedt, Anal. Bioanal. Chem. 2019, 411, 3383.
A. Apffel, J. A. Chakel, S. Fischer, K. Lichtenwalter, W. S. Hancock, Anal. Chem. 1997, 69, 1320.
A. Kaczmarkiewicz, L. Nuckowski, S. Studzińska, Talanta 2019, 196, 54.
S. Studzińska, P. Cywoniuk, K. Sobczak, Analyst 2019, 144, 622.
N. Li, N. M. El Hazar, J. G. Saad, E. R. E. van der Hage, M. G. Bartlett, J. Chromatogr. A 2018, 1580, 110.
M. G. Roussis, M. Pearce, C. Rentel, J. Chromatogr. A 2019, 1594, 105.
B. Basiri, H. van Hattum, W. D. van Dongen, M. M. Murph, M. G. Bartlett, J. Am. Soc. Mass Spectrom. 2017, 28, 190.
R. Liu, Y. Ruan, Z. Liu, L. Gong, Rapid Commun. Mass Spectrom. 2019, 33, 697.
S. Studzinska, R. Rola, B. Buszewski, J. Pharm. Biomed. Anal. 2017, 138, 146.
V. K. Sharma, J. Glick, P. Vouros, J. Chromatogr. A 2012, 1245, 65.
S. Studzińska, S. Bocian, L. Siecińska, B. Buszewski, J. Chromatogr. B 2017, 1060, 36.
A. O. Nwokeoji, M. E. Earll, P. M. Kilby, D. E. Portwood, M. J. Dickman, J. Chromatogr. B 2019, 1104, 212.
J. Qiao, C. Liang, Z. Zhu, Z. Cao, W. Zheng, H. Lian, J. Chromatogr. A 2018, 1569, 168.
K. Kawamura, K. Ikoma, Y. Maruoka, H. Hisamoto, Chromatographia 2015, 78, 487.
A. Sabarudin, J. Huang, S. Shu, S. Sakagawa, T. Umemura, Anal. Chim. Acta 2012, 736, 108.
A. Alpert, J. Chromatogr. A 1990, 499, 177.
P. Holdšvendová, J. Suchánková, M. Bunček, V. Bačkovská, P. Coufal, J. Biochem. Biophys. Methods 2007, 70, 23.
L. Gong, J. McCullagh, J. Chromatogr. A 2011, 1218, 5480.
R. Easter, K. Kröning, J. Caruso, P. Limbach, Analyst 2010, 135, 2560.
R. Easter, C. Barry, J. Caruso, P. Limbach, Anal. Methods 2013, 5, 2657.
Q. Li, F. Lynen, J. Wang, H. Li, G. Xu, P. Sandra, J. Chromatogr. A 2012, 1255, 237.
S. Studzińska, F. Łobodziński, B. Buszewski, J. Chromatogr. B 2017, 1040, 282.
L. Gong, Rapid Commun. Mass Spectrom. 2017, 31, 2125.
P. Lobue, M. Jora, B. Addepalli, P. Limbach, J. Chromatogr. A 2019, 39-48, 1595.
A. Zimmermann, R. Greco, I. Walker, J. Horak, A. Cavazzini, M. Lämmerhofer, J. Chromatogr. A 2014, 1354, 43.
M. Biba, E. Jiang, B. Mao, D. Zewge, J. P. Foley, J. Chromatogr. A 2013, 1304, 69.
B. Buszewski, Z. Safaei, S. Studzińska, Open Chemistry. 2015, 13, 1286.
S. Bäurer, A. Zimmermann, U. Woiwode, O. Sánchez-Muñoz, M. Kramer, J. Horak, W. Lindner, W. Bicker, M. Lämmerhofer, J. Chromatogr. A 2019, 1593, 110.
F. Kanwal, C. Lu, J. Chromatogr. B 2019, 1120, 71.
O. C. Uhlenbeck, RNA 1995, 1, 4.
P. J. Lukavsky, J. D. Puglisi, RNA 2004, 10, 889.
I. Kim, S. A. McKenna, E. V. Puglisi, J. D. Puglisi, RNA 2007, 13, 289.
C. Anacleto, R. Ouye, N. Schoenbrunner, J. Chromatogr. A 2014, 1329, 78.
W. Ausserer, M. Biros, BioTechniques 1995, 19, 136.
L. E. Easton, Y. Shibata, P. J. Lukavsky, RNA 2010, 16, 647.
J. Koubek, K. Lin, Y. Chen, R. Cheng, J. Huang, RNA 2013, 19, 1449.
C. Lin, Z. Huang, W. Jaremko, L. Niu, Anal. Biochem. 2014, 449, 106.
M. Biba, B. Mao, C. J. Welch, J. P. Foley, LCGC North Am. 2014, 32, 42.
Q. Zhang, H. Lv, L. Wang, M. Chen, F. Li, C. Liang, Y. Yu, F. Jiang, A. Lu, G. Zhang, Int. J. Mol. Sci. 2016, 17, 2134.
K. J. Fountain, M. Gilar, Y. G. C. J. Budman, J. Chromatogr. B 2003, 783, 61.
L. Aumann, M. Morbidelli, Biotechnol. Bioeng. 2007, 98, 1043.
T. Mueller-Spath, G. Strohlein, O. Lyngberg, D. Maclean, Chemistry Today 2013, 31, 56.
F. Steinebach, N. Ulmer, L. Decker, L. Aumann, M. Morbidelli, J. Chromatogr. A 2017, 1492, 19.
S. Vogg, N. Ulmer, J. Souquet, H. Broly, M. Morbidelli, Biotechnol. J. 2019, 14, 1800732.
Purification of a therapeutic oligonucleotide using twin-column chromatography (MCSGP), https://www.chromacon.com/resources/public/lava3/media/kcfinder/files/Oligonucleotide_MCSGP_application_note.pdf (accessed: December 2019).

Auteurs

Martina Catani (M)

Department of Chemistry and Pharmaceutical Sciences, University of Ferrara, via L. Borsari 46, Ferrara, 44121, Italy.

Chiara De Luca (C)

Department of Chemistry and Pharmaceutical Sciences, University of Ferrara, via L. Borsari 46, Ferrara, 44121, Italy.

João Medeiros Garcia Alcântara (J)

Department of Chemistry, Materials and Chemical Engineering "Giulio Natta,", Politecnico di Milano, via Mancinelli 7, Milano, 20131, Italy.

Nicolò Manfredini (N)

Department of Chemistry, Materials and Chemical Engineering "Giulio Natta,", Politecnico di Milano, via Mancinelli 7, Milano, 20131, Italy.

Daniela Perrone (D)

Department of Chemistry and Pharmaceutical Sciences, University of Ferrara, via L. Borsari 46, Ferrara, 44121, Italy.

Elena Marchesi (E)

Department of Chemistry and Pharmaceutical Sciences, University of Ferrara, via L. Borsari 46, Ferrara, 44121, Italy.

Richard Weldon (R)

ChromaCon AG, Technoparkstrasse 1, Zürich, 8005, Switzerland.

Thomas Müller-Späth (T)

ChromaCon AG, Technoparkstrasse 1, Zürich, 8005, Switzerland.

Alberto Cavazzini (A)

Department of Chemistry and Pharmaceutical Sciences, University of Ferrara, via L. Borsari 46, Ferrara, 44121, Italy.

Massimo Morbidelli (M)

Department of Chemistry, Materials and Chemical Engineering "Giulio Natta,", Politecnico di Milano, via Mancinelli 7, Milano, 20131, Italy.

Mattia Sponchioni (M)

Department of Chemistry, Materials and Chemical Engineering "Giulio Natta,", Politecnico di Milano, via Mancinelli 7, Milano, 20131, Italy.

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