molecular test 2 (therapies)

adeno-associated virus (AAV)

used in first FDA-approved in vivo gene therapy Luxturna to deliver a functional RPE65 gene into retinal cells

BCL11A

suppression reactivates fetal hemoglobin (HbF) production.

Beta-globin

a component of adult hemoglobin, mutation in the gene that codes for this causes SCA

Beta-thalassemia

an inherited blood disorder where the body does not make enough beta-globin

Hematopoietic stem cell

a stem cell that produces all the different types of blood cells

ex vivo

Cells are removed from the patient, genetically engineered outside the body, and reinfused. dominate strategy for gene therapy in SCA

Bone marrow

where formation of blood cells takes place

Erythrocyte

red blood cell, carries oxygen around the body

Fetal hemoglobin

type of hemoglobin that is mainly produced before birth.

It is made using gamma-globin (γ-globin) chains rather than the beta-globin (β-globin) chains that are mainly used in adult hemoglobin.

CRISPR

is used ex vivo to disrupt a regulatory region controlling the BCL11A gene

Gene therapy

a medical treatment that modifies a person's genes or genetic material to treat, prevent, or potentially cure a disease.

genetic disorder

inherited by parents

sickle cell anemia

a hereditary blood disorder caused by a mutation in the HBB gene encoding β-globin, a component of adult hemoglobin (HbA).

The disease is caused by a single amino-acid substitution:

glutamate (E) → valine (V) at position 6, commonly abbreviated E6V or Glu6Val.

This mutation creates a hydrophobic patch on hemoglobin molecules that promotes abnormal intermolecular interactions when oxygen levels decrease.

a natural process in cells that reduces or “silences” the expression of a specific gene. Can bind to a target mRNA and cause it to be destroyed or blocked, preventing the cell from making as much of the corresponding protein.

Zinc-finger nuclease

a gene-editing tool that uses a zinc finger protein to recognize a specific DNA sequence and cut the DNA at that location.

G-CSF

a cytokine used clinically to mobilize hematopoietic stem cells from bone marrow into the bloodstream. generally avoided in many sickle-cell patients because it can precipitate severe vaso-occlusive complications.

busulfan

a DNA-alkylating chemotherapeutic agent used in conditioning regimens prior to stem-cell transplantation. kills rapidly dividing hematopoietic stem cells, creating space for transplanted or engineered stem cells to engraft

Crizanlizumab (Adakveo)

a monoclonal antibody targeting P-selectin, an adhesion molecule expressed on activated endothelial cells and platelets. Blocking P-selectin reduces interactions between blood cells and vessel walls, helping decrease vaso-occlusive crises.

Voxelotor (Oxbryta)

a small molecule that stabilizes oxygenated hemoglobin, reducing HbS polymerization and sickling

ASO

Single-stranded nucleic acids. Function through complementary base pairing.

Can:

* block translation,

* alter splicing,

* induce RNA degradation,

* or interfere with RNA processing

RNA therapies targeting nucleic acids

Housekeeping RNAs

RNAs that are needed for basic, everyday functions of a cell

Lipid nanoparticle

where RNA can be packaged, These systems:

* protect RNA,

* improve delivery,

* and facilitate cellular uptake

miRNA

A small RNA molecule that regulates gene expression by binding to target mRNA and reducing protein production.

siRNA

A small RNA molecule that targets a specific mRNA and causes it to be degraded, preventing the protein from being made. RNA therapies targeting nucleic acids

RNA aptamer

RNA therapies targeting proteins, fold into highly specific three-dimensional structures and bind protein targets similarly to antibodies. binding depends on complementary shape

mRNA vaccines

These therapies use synthetic mRNAs that are translated by the patient’s own ribosomes

Fomivirsen

was the first FDA-approved antisense drug (1998).

It treated CMV retinitis, an opportunistic eye infection common in AIDS patients before modern HIV therapies.

Major features:

o single-stranded phosphorothioate DNA ASO,

o delivered by intravitreal injection,

o inhibited viral replication through antisense binding

Patisiran

was the first FDA-approved siRNA therapeutic (2018).

It treats hereditary transthyretin amyloidosis (hATTR).

Mechanism:

o double-stranded siRNA enters hepatocytes,

o activates the RNAi pathway,

o and reduces transthyretin mRNA levels.

This decreases production of the aggregation-prone transthyretin protein

Why are liver and eye diseases major RNA-therapy targets?

The liver is highly accessible because it naturally absorbs circulating molecules from blood.

Hepatocytes also express well-characterized uptake receptors that facilitate delivery.

The eye is also attractive because:

o local injection is straightforward,

o immune responses are relatively limited,

o and treatment effects are easy to monitor.

IgGs

abundant Y-shaped plasma proteins, and the most common class of antibodies in human blood. contains:

o two identical heavy chains,

o and two identical light chains, held together by disulfide bonds

Fc region

The “body” of the antibody

This region is relatively conserved among IgGs, and recruits immune effector mechanisms

Fab domain

The two tips of the antibody

These regions:

o differ enormously among antibodies,

o and specifically recognize molecular targets called antigens

What are the major modes of action of monospecific antibodies in cancer therapy?

Direct recruitment of immune effector cells and Immune checkpoint inhibition

Multiple myeloma

cancer of antibody-producing plasma cells.

CD38

a membrane glycoprotein expressed on many immune cells, myeloma cells often express extremely high levels of this.

This makes it an attractive therapeutic target

CD3

oa component of the T-cell receptor (TCR) complex,

and is found almost exclusively on T cells.

When the TCR recognizes this antigen it transmits intracellular activation signals

CD28

co-stimulatory receptor expressed on T cells.

T-cell activation normally requires:

1. antigen recognition through the TCR/CD3 complex,

2. and co-stimulation through receptors such as CD28.

What is the mode of action of anti-CD38/CD3 bispecific antibodies?

simultaneously bind:

o CD38 on myeloma cells,

o and CD3 on T cells.

This physically brings T cells into close contact with cancer cells, allowing redirected T-cell-mediated killing

What is the mode of action of anti-CD38/CD3/CD28 trispecific antibodies?

add CD28 co-stimulation to:

o tumor targeting (CD38),

o and T-cell activation (CD3).

This strategy aims to:

o enhance sustained T-cell activation,

o stimulate proliferation,

o prevent apoptosis,

o and increase tumor killing.

CRS (cytokine release syndrome)

a potentially life-threatening inflammatory condition caused by excessive immune activation.

Hybridomas

o grow indefinitely,

o and continuously secrete antibodies into culture medium