Safety of gene therapy and insertional mutagenesis
Retroviral and Lentiviral Vectors allow the stable and long term expression of the therapeutic transgenes in therapeutically relevant cell types thanks to their ability to insert their genetic cargo in the genome of target cells. However, enhancer sequences and/or other genetic elements present in the integrated vector may interact with regulatory elements of the host cell genome and cause unwanted side effects.
The interest of our group are:
- uncovering the interactions occurring between integrated viral vector and the surrounding host cell genome and understand how these interactions may affect the expression levels or the mRNA structure of cellular genes or induce changes in chromatin topology;
- addressing the impact of vector insertions on the physiology of cells, tissues and, ultimately, of the whole organism.
To reach these aims, we have developed sensitive genetic mouse models for testing the genotoxic potential of different vector types and backbone designs, state of the art high throughput retrieval and next generation sequencing of vector insertion sites and a solid bioinformatics and statistical framework to analyze and integrate genomic and phenotypic data.
Currently we are studying the phenotype and fate of transplanted human and mouse hematopoietic stem and progenitor cells marked with specific vector constructs and evaluating their impact on the hematopoietic reconstitution in different mouse models.
Finally, we are using chromatin conformation capture technology to study the physical interactions between integrated lentiviral vector constructs and the surrounding chromatin. This approach provides the opportunity to observe and compare chromatin conformation changes occurring in the genome after the semi-random insertion of vector constructs containing different genetic elements.
Cesana D*, Santoni de Sio FR, Rudilosso L, Gallina P, Calabria A, Beretta S, Merelli I, Bruzzesi E, Passerini L, Nozza S, Vicenzi E, Poli G, Gregori S, Tambussi G, Montini E*. *co-corresponding. HIV-1-mediated insertional activation of STAT5B and BACH2 trigger viral reservoir in T regulatory cells. Nat Commun. 2017 Sep 8;8(1):498.
Calabria A, Leo S, Benedicenti F, Cesana D, Spinozzi G, Orsini M, Merella S, Stupka E, Zanetti G, Montini E. VISPA: a computational pipeline for the identification and analysis of genomic vector integration sites. Genome Med. 2014 Sep 3;6(9):67.
Ranzani M, Annunziato S, Calabria A, Brasca S, Benedicenti F, Gallina P, Naldini L, Montini E. Lentiviral vector-based insertional mutagenesis identifies genes involved in the resistance to targeted anticancer therapies. Mol Ther. 2014 Dec;22(12):2056-68.
Cesana D, Ranzani M, Volpin M, Bartholomae C, Duros C, Artus A, Merella S, Benedicenti F, Sergi Sergi L, Sanvito F, Brombin C, Nonis A, Serio CD, Doglioni C, von Kalle C, Schmidt M, Cohen-Haguenauer O, Naldini L, Montini E. Uncovering and dissecting the genotoxicity of self-inactivating lentiviral vectors in vivo. Mol Ther. 2014 Apr;22(4):774-85.
Biffi A*, Montini E*, Lorioli L, Cesani M, Fumagalli F, Plati T, Baldoli C, Martino S, Calabria A, Canale S, Benedicenti F, Vallanti G, Biasco L, Leo S, Kabbara N, Zanetti G, Rizzo WB, Mehta NA, Cicalese MP, Casiraghi M, Boelens JJ, Del Carro U, Dow DJ, Schmidt M, Assanelli A, Neduva V, Di Serio C, Stupka E, Gardner J, von Kalle C, Bordignon C, Ciceri F, Rovelli A, Roncarolo MG, Aiuti A, Sessa M, Naldini L. *equal contribution. Lentiviral hematopoietic stem cell gene therapy benefits metachromatic leukodystrophy. Science. 2013 Aug 23;341(6148):1233158.
Ranzani M, Cesana D, Bartholomae CC, Sanvito F, Pala M, Benedicenti F, Gallina P, Sergi LS, Merella S, Bulfone A, Doglioni C, von Kalle C, Kim YJ, Schmidt M, Tonon G, Naldini L, Montini E. Lentiviral vector-based insertional mutagenesis identifies genes associated with liver cancer. Nat Methods. 2013 Feb;10(2):155-61.
Cesana D, Sgualdino J, Rudilosso L, Merella S, Naldini L, Montini E. Whole transcriptome characterization of aberrant splicing events induced by lentiviral vector integrations. J Clin Invest. 2012 May;122(5):1667-76.
Biffi A, Bartolomae CC, Cesana D, Cartier N, Aubourg P, Ranzani M, Cesani M, Benedicenti F, Plati T, Rubagotti E, Merella S, Capotondo A, Sgualdino J, Zanetti G, von Kalle C, Schmidt M, Naldini L, Montini E. Lentiviral vector common integration sites in preclinical models and a clinical trial reflect a benign integration bias and not oncogenic selection. Blood. 2011 May 19;117(20):5332-9.
Montini E*, Cesana D*, Schmidt M, Sanvito F, Bartholomae CC, Ranzani M, Benedicenti F, Sergi LS, Ambrosi A, Ponzoni M, Doglioni C, Di Serio C, von Kalle C, Naldini L. *equal contribution. The genotoxic potential of retroviral vectors is strongly modulated by vector design and integration site selection in a mouse model of HSC gene therapy. J Clin Invest. 2009 Apr;119(4):964-75.
Montini E, Cesana D, Schmidt M, Sanvito F, Ponzoni M, Bartholomae C, Sergi Sergi L, Benedicenti F, Ambrosi A, Di Serio C, Doglioni C, von Kalle C, Naldini L. Hematopoietic stem cell gene transfer in a tumor-prone mouse model uncovers low genotoxicity of lentiviral vector integration. Nat Biotechnol. 2006 Jun;24(6):687-96.
Senior postdoc fellow
Senior postdoc fellow
Adriano De Marino
Luca Del Core