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mAbexperts commits to the future of immunotherapy: Complete VHH-CAR-T design

 

Chimeric antigen receptors (CARs) are recombinant receptors consisting of an extracellular antigen-recognition domain—usually a single-chain variable fragment (scFv) from a monoclonal antibody (mAb)—linked to intracellular signaling and costimulatory domains from CD3ζ and CD28/4-1BB, respectively.

 

The scFvs are composed of a heavy-chain variable fragment connected to a light-chain variable fragment by a flexible linker. They are typically reformatted from a full-length Ig, with the linker optimized to preserve heavy- and light-chain variable region pairing.

 

CAR-T are the result of a transduced T lymphocyte with a ScFv fragment antibody able to recognize specific cancer antigens.

 

However, scFvs can present several drawbacks:

 

ü they do not always fold efficiently and can be prone to aggregation,

ü murine ScFv may induce immunogenicity,

ü they can’t access all the different epitopes, since scFv present flat or concave topologies paratopes.

 

In contrast, the variable regions of camelid-derived single-domain antibody fragments (VHHs or nanobodies):

 

ü are small, stable, with affinities comparable to traditional scFvs,

ü are less immunogenic than murine scFv,

ü can access epitopes different from those seen by scFvs, since they offer convex topology associated with smaller size,

ü do not require the additional folding and assembly steps that come with V-region pairing (in scFv libraries, natural VH–VL pairings are usually lost).

 

VHHs could therefore serve as suitable antigen recognition domains in CAR T cells, and several potentially interesting VHHs have been tested since the beginning of 2019.

 

These new constructs are called CAR-VHH-T and may be a potential future of CAR adoptive cell therapy.

 

mAbexperts is now able to provide a one one-stop-shop toolbox for cancer treatment and proceed as follow :

 

1. RD Biotech (https://www.rd-biotech.com) starts with the antigen coding sequence and the expression of recombinant cancer antigen in order to specifically generate the adequate camelid heavy chain-only antibodies (HCAbs) against it.

 

2.  After immunization in llama and/or alpaca with this specific antigen, QVQ (http://www.qvquality.com) generates phage-display libraries from the immunized camelids. They generate a lead panel of monoclonal sdAbs binding with high affinity to the tumor associated antigens.

 

3.   Following the collection of a patient's T cells by leukapheresis at Blood Transfusion Center in Besançon, the cells are genetically engineered by RD-Biotech through inactivated lentivirus particles containing the coding sequence for the VHH.

 

4.    To ensure that the CAR-VHH-T is well activated, Diaclone (https://www.diaclone.com) launches its specific interferon gamma ELISpot test assay attesting that the CAR-VHH-T combines the specificity of an antibody for CD proteins, with the cytotoxic and memory functions of T cells.

 

5.    Drawing on its expertise in secondary monoclonal antibodies, SYnAbs (https://www.synabs.be) has already generated an universal rat monoclonal antibody against the VHH construct. Thanks to this new available tool, mAbexperts’ partners are now able to detect and select specific transfected CAR-VHH-T populations on flow cytometry platform !

 

We could add that VHHs are also well suited in the generation of bi- and multispecific antibodies. Bispecific antibodies provide a possible solution in which they could bind simultaneously two or more tumor epitopes (bi-paratopic) or antigens simultaneously. Can we conclude that we should expect new developments in the near future?

 

Business to be continued...    

 

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