Optimization of Best Model for TanCAR T Cell Targeting CD123 and Folate Receptor Beta

Background and Aim: Acute myeloid leukemia (AML) is considered asa type of cancer with abnormal myeloblasts in the bone marrow. Totackle this disease, we capitalized on the chimeric antigen receptors(CARs), which are artificial molecules that change the specificity of Tcells to particular antigens. A CAR was consisted of two different antigens(CD123 and FRβ) recognition domains presented in tandem on a singletransgenic receptor that mediates bispecific activation and targeting ofT cells. We assumed that this tandem CAR (TanCAR) can exclusivelyrecognize each target molecule and facilitate a synergistic activation andfunctionality when both are used concurrently. While there exist somepossibilities in modeling the CAR, we capitalized on various methods inorder to optimize the physical properties of the final CAR protein.Methods: Different computational modeling tools were exploited topredict the functionality of CAR molecules that can mediate the bispecificactivation and targeting of T cells. To uncover whether the aforementionedmolecular arrangements were possible, we used TanCAR structuralmodels predicted by modeling web server, ModWeb. Patchdock andFireDock, software tools for docking and refining two structures. Theseanalyses were carried out based on the shape complementarity anddocking of FRβ- and CD123- scFvs to their respective target molecules.We also looked into some other important parameters in computationaldesigning of the chimeric protein, including stability, proper energylevel and etc. Finally, the best model was selected through comparisonbetween the predicted structures and physical properties, which mightsimulate the function of CAR molecule in the in vivo condition.Results: It was found that the potential interactions of the best TanCARmodel could accommodate the planned bispecificity, and as such,an initial model to explore the ability of TanCAR to interface with thetarget molecules individually predicted. The designed chimeric antigenreceptor demonstrated stability and same docking as the original scFvs.The modeled construct displayed a good energy level to making theproper conformation based on its sequences. Both paratope domainspossessed free physical configuration and were able to recognize eachtarget molecule solely.Conclusion: Various de novo CAR T cell therapy modalities have recentlybeen articulated. Among them, bi-specific CAR constructs are the mostattractive methods. They are able to recognize and bind to two differentantigens separately, and thus, result in an enhanced specific functionof recombinant T cells. Likewise, the TanCAR molecules provided bothrecognition domains in tandem, imposing T cells to facilitate a synergisticactivation and functionality. This study confirmed the potency of the bestTanCAR based on CD123 and FRβ as a new candidate for the treatmentof AML, which might simulate the function of CAR molecule in vivo.

Keywords:

Acute myeloid leukemia , CD123 , FRβ , TanCAR , Modeling