INTRODUCTION TO NONCLINICAL SAFETY EVALUATIONS OF GENE THERAPIES
The concept of Delivering genes as a means of therapeutic intent has been slowly progressing to reality since the Early 1980. Initially researchers used method of introducing genes into hematopoietic stem cells by extacting HSCs from patients, transducing them in tissue culture and then returning the genetically modified cells back to the patient. This is a process that is now referred to as ex vivo gene therapy, As the machinery used to Introduce the genetic change occurs outside of the patient. as further development of viral vectors progressed through the 1990s, the other Approaches involved direct administration of the gene and the delivery System to the patient, now referred to as in vivo gene therapy. Each approach comes with some common and unique nonclinical Features & strategies
the Concept of delivering DNA directly to an individual with the intent of expressing a therapeutic gene has been ongoing in both preclinical and clinical experimentation since the early 1990s with the advent of the use of adenoviruses as a delivery mechanism. Attempts to treat a wide variety of medical conditions including haemophilia, monogenic rare diseases, respiratory aliments and, most notable cancer, however have been met with several issues related to delivery of a viral vector into the circulation. in several cases, pre-existing or evolving immunity of the vector Significantly limited sufficient exposure, thus resulting in ineffective levels of transgene expression; however, modifications to routes of administration and vector design have kept the field mowing forward.
potential adversities during gene therapy development
in 1999, a clinical trial designed to treat a rare Genetic disease resulting from OTC deficiency was initiated with an adenovirus intended to transduce the liver and express OTC. A Patient enrolled in the third cohort of the study was The second patient to be infused at a total dose of 3-10*3 total particles. Shortly after infusion, an apparent massive systemic inflammatory Response occurred that began with liver Dysfunction and concluded with multi-organ failure and death. other enrolled patients showed transient limited increases in liver enzymes. other, viral vectors such as the adeno-associated virus (AAVs) have also shown some propensity for inducting immune-based toxicities. A clinical trial Evaluating the efficacy of an AAV2 vector expressing the coagulation protein factor IX found that several weeks after Administering the gene therapy product via the hepatic artery, was followed by an elimination of transgene expression. Follow-up bioanalytic work indicated that the patient generated a CTL response agains AAV2. A subsequent clinical trial using the same therapeutical strategy, except With an AAV8 serotype found no CTL response, no liver injury and longer term FIX expression without any immunosuppressive therapy.
role of the toxicological pathologist
the Pathologist should be aware of the potential for the immune system to play a significant role in The tolerability of gene therapies Delivered in vivo. Such reactions have the potential to occur with vectors that are Administered via routes other than the bloodstream. While several organs systems are classically Characterised as ‘immune Privileged’ once the vector is Introduced via the needle or some other Apparatus to that locations, barriers can be compromised, leaving the Location susceptible to infiltrates from the immune system. Toxicological Programs Utilising AAV vectors delivered to the brain and the Eye have found that:
antibodies measured in the Term against the vector capsid protein can sometimes have a negative effect on efficiency of cellular transduction.
inflammatory infiltrates can occur following injection of viral particles. In some cases, this has resulted in significant local vascular and perivascular inflammatory processes in organs from preclinical species.
the pathologist may decide that histopathology can be complemented by immunophenotyping cellular infiltrates in lesions or more Commonly by IHC demonstration of Immune complexes where cases of vasculitis or glomerulonephritis have occurred Preclinical or clinically with such therapies.
the importance of the injection sites during preclinical development
As the in vivo gene therapy field has evolved, local injection to the Anatomie site of Interest has become an inceasinfly utilized approach With the intent of maximizing Exposure and chances of efficacy. While several organs systems have been investigated in nonclinical and clinical trials; the most notable Attempt have been delivery to the brain and eye.
Treatment of neurodegenerative diseases such as Parkinson’s disease and Spinal Muscular atrophy (O’Connor 2015) have employed the expression of a variety of factors that need to be locally expressed in particular areas of the brain such as putamen, striatum and subthalmic nucleus, or the cerebrospinal fluid.
Considering local injections are at least initially confined to enclosed tissue or fluid-filled spaces, the efficiency of distribution of a vector within that space is a key component to obtaining maximal effect. in this regard, the pathologist is likely to be consulted at early stages of such programs to lend expertise in target Localisation techniques and his or her ability to identify expressing cell populations.
The gene therapy product is also Freely modified to enhance cellular uptake by Changing the shell of proteins on outside of the virus called the capsids or also y Optimising cellular expression by testing different promoters of One expression (Castle, 2016, Zinn 2014). The Pathologist should be a key figure in all of these early investigations and can serve a critical role in selecting the optimal product to move into animal safety and efficacy experiments