Immunogene therapy is a combination of two different treatment methods: gene therapy and immunotherapy. In immunogene therapy, a protein/antibody, gene, or nucleic acid molecule is delivered to patients by gene therapy methods with the intention of inducing targeted immunomodulatory effects. Recently, researchers have developed a novel cancer immunogene therapy method that overcomes the limitations of currently used CD19 chimeric antigen receptor T (CART)-cell therapy and bispecific antibody-based T-cell immunotherapy (BsAbs^{) 1} 

CART-cell immunogene therapy and limitations

The CART-cell therapy is an immunogene therapy currently used for treating cancer and autoimmune diseases. Utilizing a combination of cell therapy, gene therapy and immunotherapy, T-cells are modified with engineered T-cell receptor by transfection or adenoviral associated virus (AAV)-based gene therapy. These genetically modified T-cells are then introduced to patients for activating or inducing immunomodulatory effects.

One drawback of CAR-T Cell therapy is that it requires isolation and manufacturing of genetically modified CART-cells as well as continuous infusion due to its noticeably short half-life. Like most cancer therapy methods, CAR-T cells are administered intermittently which results in pharmacokinetic peaks and troughs.

Limitations of bispecific antibody-based T-cell therapies

Blinatumomab, a bispecific, T-cell engaging, antibody-based drug, is approved by FDA for treating Philadelphia chromosome-negative relapsed or refractory acute lymphoblastic leukaemia. It is made up of an anti-CD19 scFv (single chain) which is linked to an anti-CD3 scFv, where the anti-CD3 scFV binds to T-cells and the anti-CD19 binds to CD19 positive cancer cells. Therapy utilizing blinatumomab requires frequent administration as it has a short half-life and is rapidly cleared from the system.

Single-dose AAV-mediated immunogene therapy overcomes the limitations of currently used immunotherapies.

Researchers from the Nationwide Children’s Hospital (Ohio, USA) have developed a single-dose immunogene therapy method to overcome the current imitations of CART-cell and bispecific antibody-based T-cell immunotherapy. The single-dose immunogene therapy consists of introducing a gene encoding blinatumomab-mimic (αCD19-αCD3 dimert) into cells by AAV. The incorporation of signal peptide sequence facilitates the secretion of AAV expressed blinatumomab-mimic into the bloodstream and facilitates binding of T-cells to cancer cells (this method is named as CD19 TransJoin). Thus, AAV-mediated gene therapy has resulted in maintaining consistent level of blinatumomab-mimic expression in the bloodstream of mouse model of Burkitt’s lymphoma and did not show any toxicity for up to one year. Moreover, over a span of 64 days, lymphoma shrinkage was. When examined with bioluminescence, calliper measurement, and animal survival the single dose of rAAV injection can effectively reduce tumour size. Further, qPCR analysis of AAV genome revealed that the viral vectors were widely distributed throughout the body with its remarkably higher abundance in the heart, liver, and muscle, but none in the spleen and brain. This clearly indicates that CD19 TransJoin is an effective tool for systemic production of blinatumomab-mimic. The details of AAV vector constructs are shown in Fig. 1.

Single-dose immunogene therapy combined with on-demand expression of an immunotherapeutic

Continuous expression of immunotherapeutic (e.g., antibody) may not be warranted in all clinical needs. Often, short-term, or intermittent expression of an immunotherapeutic can be a preferable method of treatment. To achieve this goal, researchers have developed an AAV construct known as TransSkip. In the TransSkip, they inserted two introns flanking a defecting exon in the transgene (blinatumomab-mimic) coding sequence. The expression of functional transgene mRNA is induced by treatment with KTS morpholino, which binds to which exon-intron junction and induces exon skipping. CD19 TransSkip-treated mice (experimental model) were administered either a single dose or two doses of morpholino (Vivo-KTS) on two consecutive days. A robust expression of blinatumomab-mimic. was observed in the experimental set as expected. However, the TransSkip method is not as effective as CD19 TransJoin method. The details of AAV vector constructs are shown in Fig. 1.

Future considerations 

Conceptually, this modern technology opens a broad avenue of immunogen therapy and gene therapy which encompasses more than just single-gene disorder therapy. TransJoin, however, could pose the risk of potential toxicities arising both from the vector and the transgene. The researchers suggest that administration of lower dose of the drug can reduce the chances of toxicity that often arise from AAV vector. In addition to vector-mediated toxicities of TransJoin, blinatumomab-mimic poses the risk of evoking cytokine release syndrome (CRS), as well as immune effector cell-associated neurotoxicity syndrome (ICANS). CRS and ICANS can occur from CAR-T cell therapy in the similar vein. Nevertheless, the researchers anticipate that a slow and gradually accumulation of blinatumomab-mimic can minimize the risk, but they still believe that patient undergoing TransJoin therapy needs to rely on concomitant anti-interleukin-6 therapies to minimize the risk of inflammation.

Fig. 1: Sing-dose immunogene therapy methods.

Abbreviations:

• EFS: Elongation factor 1α short.
• K: Kozak sequence
• Sec1A: Signal peptide sequence
• WPRE: Woodchuck Hepatitis Virus (WHV) Posttranscriptional Regulatory Element.
• PA: Poly-A tail