Gene Therapy for Beta Thalassemia

Virology

Mic A

Gonçalo Baptista

All the ___ words referer directly as answer to “Selected Gene”,”Pathology in study”, “Target
Tissue” and “Viral Vector”

Introduction

Beta Thalassemia is a blood disorder caused by defective hemoglobin production. Because the
disease was first described in communities living near the Mediterranean Sea, its name is
taken from the Greek word "thalassa”, which means "sea." However, the sickness is also found
in Africa, the Middle East, and Asia.

Thalassemia is a collection of illnesses that can range from a barely noticeable blood
abnormalities to life-threatening anemia. Two alpha and two beta polypeptide chains make up
adult hemoglobin. The hemoglobin alpha gene (HBA1) has two copies (HBA1 and HBA2), each
of which encodes a alpha-chain, and both genes are found on chromosome 16. The beta-chain
is encoded by the hemoglobin beta gene (HBB), which is found on chromosome 11.

The synthesis of alpha-chains is impaired in alpha-thalassemia. The extra beta-chains that arise
bind oxygen weakly, resulting in a low oxygen concentration in tissues (hypoxemia). Excess
alpha-chains, on the other hand, can create intractable clumps inside red blood cells. The
destruction of red blood cells and their progenitors is caused by these aggregates, resulting in
severe anemia. The spleen enlarges as damaged red blood cells are removed from
circulation.Most cases of alpha-thalassemia are caused by HBA1 and/or HBA2 gene deletions.
The severity of symptoms is determined by the number of these genes deleted. The loss of one
or two genes normally causes no symptoms, but the deletion of all four genes is lethal to the
unborn child.

Genetic Therapy

Beta-thalassemia gene therapy is based on the transfer of a human beta-globin gene into
autologous HSCs, which reduces the risk of GVHD and graft failure associated with allogeneic
BMT and resolves the lack of compatible donors. In beta-thalassemia, autologous HSC gene
therapy aims to restore normal beta-globin protein expression. The beta-globin gene and its
regulatory components were previously transferred using gamma-retroviral vectors. However,
oncoretroviral vectors including LCR sequences and the beta-globin regulatory elements were
difficult to manufacture at large viral vector titers and were exceedingly unstable (Novak et al.,
1990; Gelinas et al., 1992). The goal was to increase transduction efficiency by identifying and
deleting the DNA sequences that cause vector instability and poor titers.

Lentiviral vectors are a type of sophisticated retrovirus that inserts its viral genome into the
chromosomes of differentiated lymphocytes and macrophages in a stable manner. The human
immunodeficiency virus-type 1 (HIV-1) is the prototype of this viral family. HIV-1 is an immune
system pathogen with cytopathic effects that can be transduced into non-dividing cells at the
G1–S cell cycle boundary (Naldini et al., 1996a; Naldini et al., 1996b). Due to the existence of a
robust RNA export element (RRE) that binds the RNA-binding viral protein, lentiviral vectors
can accept the insertion of long and complicated DNA sequences (Kumar et al., 2001).
The capacity of lentiviral vectors to transduce human umbilical CB cells in NOD-scid IL2R null
immunodeficient mice was first tested in human umbilical CB cells transplanted into NOD-scid
IL2R null animals (NSG mice). Around 50% of human progenitors were genetically changed six
months after lentiviral beta-globin vector transduction and transplantation (Imren et al.,
2004), showing a high transduction efficacy of CB-HSCs by the lentiviral beta-globin vectors.

In cultivated erythroid cells generated from beta-thalassemia patients, the ability of lentiviral
vectors expressing the beta-globin gene to repair the beta-thalassemia major phenotype was
investigated (Centis et al., 2000). Normalized beta-globin levels were obtained, resulting in
successful cell expansion, normal erythroid cell differentiation, and apoptosis reduction
(Puthenveetil et al., 2004; Malik et al., 2005). NSG mice were given the gene-corrected human
thalassemia CD34+ cells. The erythroid progenitor cells generated from human HSCs had
normal amounts of human beta-globin and efficient erythropoiesis. Importantly, the
expression of the beta-globin protein in erythroid colonies produced from normal control
patients was comparable (Puthenveetil et al., 2004). Furthermore, in vitro transduction was
demonstrated in a number of samples from -thalassemia patients of various geographic and
ethnic origins, as well as from a variety of genotypes.