Biotechnological Requirements for Patient Monitoring in Gene Therapies

Gene therapies are advancing rapidly as a novel therapeutic modality to provide a functional cure for patients with monogenic diseases. Several gene therapies have already been approved in the past several years. This article provides a roadmap for the biotechnological advances required for monitoring safety and efficacy endpoints, as well as functional biomarkers relevant for gene therapies targeting the liver as a biofactory of defective or missing proteins, with hemophilia A as a representative disease.

Hemophilia is a group of X-linked inherited bleeding disorders caused by defects in, or the absence of, critical factors in the coagulation cascade. The prevalence of hemophilia A has been reported at 17 cases per 100,000 males for all severities and 6 cases per 100,000 males for severe hemophilia A.

Hemophilia A is caused by various mutations in a gene located in the X chromosome resulting in deficient production of the FVIII coagulation protein. Alterations in this gene include point mutations, deletions, and insertions resulting in absent, unfunctional or low levels of FVIII. Variations in the mutations causing the disorder result in different levels of the affected protein, which influences tendency to bleeding. Patients with plasma FVIII level less than 1% of normal are classified as having severe hemophilia, those with levels of 1 to 5% of normal as moderate hemophilia, and those with levels over 5 to 40% of normal as mild hemophilia (Pipe 2022).

The current standard of care treatment for hemophilia A involves regular infusion (prophylaxis) of concentrates of exogenously derived FVIII. The aim of prophylaxis is to raise FVIII activity above a level that is detectable (>1%) to prevent bleeding and reduce or delay the incidence of joint disease and other serious complications. The relatively short half-life of FVIII in the bloodstream (range 10 to 14 hours) requires regular infusions of factor concentrates (every 2 to 3 days). Newer modified recombinant products with enhanced pharmacokinetic behavior allow less frequent infusions.

Hemophilia is an ideal target for gene therapy, especially as the genetics and pathophysiology of the disease are well understood. The goal of gene therapy is long term expression of the transferred gene at levels high enough to be therapeutic, a so-called functional cure. A growing body of clinical experience supports the use of liver-directed recombinant adeno-associated virus (rAAV) as a gene therapy transporter of the replacement gene for the treatment of hemophilia.

ASC Therapeutics has developed ASC618, a synthetic gene, B-domain deleted, liver specific codon-optimized, bioengineered chimeric human-porcine factor VIII with a gene promoter, packaged within a capsid of rAAV8 serotype. ASC618 is designed to express the bioengineered FVIII protein for the treatment of patients with severe and moderately severe hemophilia A. Following a single administration of ASC618, FVIII is expected to be expressed by liver cells indefinitely and potentially eliminate or reduce the need for prophylactic or replacement FVIII therapy (Pipe 2022).

ASC Therapeutics is conducting a first-in-human, open-label, dose-finding study designed to assess the safety and preliminary efficacy of a single infusion of ASC618 in study participants with severe and moderately severe hemophilia A (FVIII activity ≤ 2 International Units/Deciliter) (NCT04676048).

The main safety monitoring parameters are:

1. Physical examination, including assessments of general appearance; head, eyes, ears, nose, and throat; the cardiovascular, dermatologic, lymphatic, respiratory, gastrointestinal, genitourinary, musculoskeletal, and neurologic systems. Height and weight will also be measured and recorded.
2. Vital signs, including oral temperature, pulse rate, respiratory rate, and blood pressure),
3. Viral shedding, assessing the evidence of potential viral transmission will be tested on samples of blood, saliva, urine, stool, and semen
4. Liver ultrasound

The main efficacy monitoring assessments are:

1. FVIII Activity, determined by validated assays, one-stage activated partial thromboplastin time (aPTT) and chromogenic FXa. FVIII levels should be taken at a trough or close to trough levels, meaning after a minimum of 72 hours has elapsed since the last infusion of FVIII protein concentrates.
2. Bleeding Episodes and FVIII Replacement Therapy will be captured on the participant’s diary. In addition, number of bleeding episodes requiring treatment, and number of bleedings that do not require treatment, following the administration of ASC618 infusion, will be recorded throughout the study.

The main exploratory assessments are:

1. Hemophilia A quality of life (Haem-A-QoL) questionnaire wcompleted by participants during the study.
2. Blood samples will be collected for the purpose of potential future research in the hemophilia A disease area to assess: a) Liver structure, including Fibroscan and abdominal ultrasound; b) Liver function, including Liver enzymes, GRP78/BiP, Enhanced Liver Fibrosis (ELF™, https://www.siemens-healthineers.com/laboratory-diagnostics/assays-by-diseases-conditions/liver-disease/elf-test) and microRNA exploration analysis; c) Liver Tumorigenesis, including GALAD score (serum biomarker-based model that predicts the probability of having HCC* incorporating gender, age, AFP-L3, AFP, and DCP), VirScan (comprehensively analyzing antiviral antibodies; https://cdi.bio/antygen-virscan-phip-seq-service/) and cell-free DNA assessment.

In order to complete these assessments, extensive monitoring including on-site and remote visits will be implemented. The levels of FVIII and liver function tests will be evaluated twice a week in the first 16 weeks and then slowly decreased as these measurements will reach a plateau.

ASC Therapeutics is conducting this investigational study after obtaining an IND, Investigational New Drug, clearance by the U.S. FDA, Food and Drug Administration. The goal of this study is to confirm pre-clinical data supporting the potential of ASC618 as a second-generation gene therapy that can be safe and effective while providing the advantage of lower cost and longer durability vs. first-generation gene therapies for hemophilia A (Pipe 2022).

Reference:

Pipe SW, Gonen-Yaacovi G, Segurado OG. Hemophilia A gene therapy: current and next-generation approaches. Expert Opin Biol Ther. 2022 Sep;22(9):1099-1115.