Development of sensitive and unbiased integration site analyses to comprehensivly assess biosafety and efficiency of innovative vectors

• Antragsteller: Dr. Manfred Schmidt, Ph.D.
• Fachliche Zuordnung: Hämatologie, Onkologie
• Förderung: Förderung von 2006 bis 2013
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 22811434

Gene therapy with insertional vector systems has been coming of age, achieving unprecedented levels of efficiency, success, but unfortunately also side effects. Our recent preclinical and clinical gene therapy studies have revealed that insertional mutagenesis is not rare, but may lead to frequent gene activation in hematopoiesis, producing insertion-activated expansion of cell clones, and in its worst cases has resulted in oncogenesis. We and others have demonstrated that redesigning integrating vectors might diminish or avoid unwanted insertional gene activation, but even with such vectors, any integration by definition remains mutagenic, and poses a finite biosafety risk. Therefore, the quest for cellular gene delivery has rendered integrase deficient vectors particularly attractive, because they maintain the ease of use, effectiveness and lack of acute toxicity of retrovirus and lentivirus vectors but potentially minimize the likelihood of severe side effects. Transduction by integrase deficient lentiviral vectors has been attributed to episomal DNA circles and was able to correct clinically relevant rodent in vivo disease models. Towards therapeutic applications in quiescent or less frequently dividing cells in retina, heart and muscle, we aim to characterize and quantify nuclear episomal vector forms, and characterize residual integrations by statistically relevant large-scale mapping analysis. Integrase deficient vectors dilute out1 transgene expression in dividing cells, thus becoming an ideal system to study the influence of particular transcription factors such as MDS1/EVI1on the proliferation and differentiation of early repopulating bone marrow cells. To combine non-toxic nucleic acid transfer of lentivirus particles with the preferable insertion and capacity of transposases, we will generate and characterize the potential biological advantages of a stably integrating combined vector system.

• DFG-Verfahren: Schwerpunktprogramme
• Teilprojekt zu: SPP 1230: Mechanisms of gene vector entry and persistence
• Beteiligte Personen: Professor Dr. Hanno Glimm; Professor Dr. Christof von Kalle