OVERVIEW

The course will provide an overview of the most advanced gene therapies, starting from their molecular bases to their current translational applications. The course will deal with the fundamental biological and morphological characteristics, with the genetic content and organization and with the integrative competence of several viral families that makes them suitable as effective vectors for gene therapy, as well as a few “non-viral” based- and “genome editing”-approaches. Implication and relevance for human health ameliorations due to gene-therapy based approaches (current and envisageable) will also be taken into account. 

AIMS AND CONTENT

LEARNING OUTCOMES

To allow the student to acquire a detailed knowledge of the current and up-to date gene transfer and therapy approaches, along with their evolution in clinical settings, and to correlate them with the current knowledge on physiology, molecular biology and genetic engineering technologies; to allow the student to envisage possible future applications of gene therapy based on the current knowledge of monogenic diseases with a relevant clinical prevalence.

AIMS AND LEARNING OUTCOMES

At the end of the course the student must retain: a) the general applicative principles, methodologies and technical requirements for the preparation of gene therapy products; b) the biological, morphological, genetic and functional main requirements of the viral family types used gene therapy approaches; c) the basic concept of genome-editing. Students should be able to evaluate pros and cons and associated risks with gene therapies, particularly in confrontation with current gold-standard therapeutic approaches, as well as their implications for human health and its ameliorations, critically supporting their observations with an appropriate scientific language and terminology.

During the course students will implement their own alphabetical-functional advanced competences, developing comprehension of written texts in a different language, autonomously searching the existing current literature related to the subject of their presentation, critically organising the entire body of information and publically expressing their own conclusions in convincing and appropriate way.

PREREQUISITES

For the best comprehension of the contents of the course, acquired competences in subjects such as molecular and cellular biology, biochemistry and physiology are strongly recommended – although not mandatory.

TEACHING METHODS

The course lasts 20 hours of front lessons, during which course items are presented through PowerPoint presentations. Didactic material is available in AulaWeb. Interactive discussion during lessons is encouraged. Students will be offered to give a power-point presentation on a scientific paper provided by the teacher. The presentation will be a critical revision of the contents of the paper in which students will have to summarize, discuss and evaluate the paper, eventually also comparing it to additional literature. Papers will be made available by the teacher in the second half of the course. Students must be aware that the English language will be used for lessons and exams. However didactic/studying material is provided both in English and in Italian.

Students' alphabetical-functional advanced competences shall be implemented through Inquiry-based-learning approaches and  the debates that will follow each student's presentation. A peer-evaluation process will also be adopted to force criticism and analysis of each presentation by all the other students in the audience. If feasible -depending on the number of students enrolled for the course - debate activities may also be based on a variation of the think-pair-share approach, in which couples of students will hold-up -in a public confrontation- one the positive aspects and the other the negative aspects of the same scientific report/study.  

Students with certified Specific Learning Disorders (DSA), with disabilities, or other educational needs are invited to contact the teacher and the Scool of Medical Sciences representative teacher (Prof. Nicola Girtler; nicolagirtler@unige.it) at the beginning of the course to agree upon specific individual  teaching/evaluation methods.

SYLLABUS/CONTENT

Clinical trials based on gene therapy: geographical distribution and socio-economic aspects. General principles of gene therapy and choice of the proper target pathology. Side effects: consequences and complications. Examples of therapeutic approaches by means of gene therapy (for ex. in hematopoietic cells, in muscular dystrophy, in tumor treatment or in neuropathological disorders) will be presented along the description of the viral vectors used (Adenosine deaminase deficiency, SCID and X-SCID, Junctional Epidermolysis bullosa).Viral vectors used in gene therapy approaches: structure, characteristics, pros and cons.

Herpesviridae (HSV):-Structural characteristics of HSV capsid, matrix and envelope; structural organization of the viral genome; immediate-early (IE), early (E) and late (L) genes and corresponding proteins; HSV lifecycle (lytic and lysogenic cycles); viral tropism; packaging systems based on helper virus and helper virus-free systems; transgene insertion in HSV; optimization for targeting: gB and gD functions. -Sindbin Virus e Semliki Forest Viruses (SFV) as suicidal vectors.

Adenoviridae (AV):-Viral particle structure; genome structure; E1, E2, E3 and E4 genes; ITR and psi (ψ) region of the viral genome; effects of AVs and specific functions of their proteins against host defenses. AV vectors and their potentiation (promoter-specific activities, pro-drug enzymes, vector-spread molecules, anti-tumor molecules). Pre-clinical aspects: positive and negative results on murine experiments. Adeno-associated viruses (AAV) and applications of AV/AAV approaches.

Retroviridae(RV)-Viral particle structure of retroviruses; the RV genome and its integrated provirus; main relevant characteristics of the RV genome: gene products, LTRs, cPPT, TAR, RRE. Replication of the viral genome; generation of helper viruses for packaging cell lines; minigene vectors; Packaging cell lines bi- and tri-sected; -Accessory proteins in Lentiviridae (Tet, Rev, Vif, Vpi, Vpx, Vpu and Nef); SMAR sequences. Self-inactivating vectors (SIN). Current methods for the recovery transgene-carrying functional viral particles: ultracentrifugation, poly-lysine precipitation, ionic-exchange chromatography. Comparison between HSV, AV, AAV, RV and LV. Use of LV for shRNA transfer.

Non viral-based gene therapy approaches and principles of “genome editing”: Naked DNA inoculation, jet-gas injection, gene-gun, electroporation, sonoporation, hydrodynamic transfer, chemical vectors. Characteristics of target tissues for non-viral based gene transfer.

Frontiers of genome editing: Characteristics of the CRISPR-Cas 9 system; discovery and functions of the CRISPR locus; role of CAS proteins; potentiality of the CRISPR-Cas9 approach in Gene Therapy.

RECOMMENDED READING/BIBLIOGRAPHY

Suggested textbooks:

M. Giacca (2011) - Terapia Genica. Springer Biomed Ed.

A.J. Dunn - Elementi di virologia molecolare. CEA Ed.

B. Alberts et al. - Biologia molecolare della cellula. Zanichelli Ed.

J.D. Watson et al. - DNA Ricombinante. Zanichelli Ed.

TEACHERS AND EXAM BOARD

LESSONS

LESSONS START

The course is planned for the second semester. Location, date and timetable of the lessons will be provided by the competent office.

Class schedule

The timetable for this course is available here: Portale EasyAcademy

EXAMS

EXAM DESCRIPTION

Before the formal final exam the student is invited – if willing - to give a PowerPoint presentation during which he/she will expose the scientific results of a scientific publication dealing with the subjects of the course, in particular dealing with therapeutic approaches based on the technologies described during lessons. A critical overview, both relating methodology and results, is suggested for the presentation; the presentation and its issues will be discussed publically among all the students and the teacher. The presentation, however, is not mandatory.

The student's performance is evaluated during an oral examination (average duration: 30 min), according to the exam calendar and timetable set by the competent office or through direct contacts with the teacher.

ASSESSMENT METHODS

Questions of raising difficulty will be posed, relating major issues of the course In particular we will evaluate:

a) exactness of the student's response;

b) the student's ability to frame the topic in a wider context;

c) the student's ability to correlate the topic with other topics, not necessarily related to the course;

d) the terminology and language appropriateness;

e) the student's criticism.

For the final evaluation of the candidate the commission may also take into consideration the performance provided during the non-mandatory presentation of a scientific paper, selected within the most recent literature.

Exam schedule

FURTHER INFORMATION

The scientific articles for the students' critical presentations, as well as all the material presented during the course and including the access keys to the aulaweb site, will be provided directly to the students during lessons.