Combining the best of two worlds: highly flexible chimeric antigen receptor adaptor molecules (CAR-adaptors) for the recruitment of chimeric antigen receptor T cells

Figures & data

Table 1. Comprehensive overview of current CARs and their respective adaptor molecules.

	Target (clone)	CAR-adaptor-tag	CAR-construct	Reference
Antibody derivative CAR-adaptors
scFv-based CAR-adaptors
	CD33 & CD123 (bispecific)	5B9	anti-La 5B9-CD28TM/CD28/CD3ζ	^Citation39,Citation76
	Sialyl-Tn (based on mAb L2A5)	E5B9	anti-La E5B9-CD28TM/CD28/CD3ζ	^Citation40
Fab-based CAR-adaptors
	CD19 (FMC63) CD20 (Ofatumumab)	GCN4	anti-GCN4 (52SR4)-CD8TM/4–1BB/CD3ζ	^Citation27
	CD19 (FMC63) CD22 (M971)	FITC FITC	anti-FITC-E2-CD8TM/4-1BB/CD3ζ	^Citation25,Citation27
	Her2 (Trastuzumab, 4D5)	FITC	anti-FITC-E2-CD8TM/4-1BB/CD3ζ	^Citation26
	Murine CD19 (1D3)	GCN4	anti-GCN4 (52SR4)-mCD28TM/mCD28TM/CD3ζ anti-GCN4 (52SR4)-IgG4mTM/m4-1BB/CD3ζ anti-GCN4 (52SR4)-IgG4mTM/mCD28/CD3ζ anti-GCN4 (52SR4)-IgG4mTM/mCD28/m4-1BB/CD3ζ anti-GCN4 (52SR4)-mCD8TM/m4-1BB/CD3ζ anti-GCN4 (52SR4)-mCD8TM/mCD28/CD3ζ anti-GCN4 (52SR4)-mCD8TM/mCD28/m4-1BB/CD3ζ	^Citation44
mAb-based CAR-adaptors
	EGFR (Cetuximab), HER2 (Trastuzumab), CD20 (Rituximab)	FITC	anti-FITC-CD28 TM/4-1BB/CD3ζ	^Citation77
Nanobody-based CAR-adaptors
	EGFR (camelid nanobody 7C12^Citation36)	E5B9	anti-E5B9-CD28/CD3ζ	^Citation37
	EGFR (bivalent (camelid nanobody 7C12^Citation36)	E5B9	anti-E5B9-CD28/CD3ζ	^Citation24
Small molecule-based CAR-adaptors
	FolR1 (EC-17^Citation78)	FITC	anti-FITC 4M5.3–4-1BB/CD3ζ	^Citation31
	FolR1 (EC-17)	FITC	anti-FITC^Citation25-4-1BB/CD3ζ	^Citation32
Substituted ECD + CAR-adaptors
	MSLN, EpCAM	scFv-biotinylated IgG-biotinylated	anti-biotin-dimeric avidin-CD28/CD3ζ	^Citation23
	CD19 (Rituximab), CD20 (Cetuximab)	IgG-biotinylated IgG -biotinylated	anti-biotin (mSA2 affinity enhanced) CD28TM/CD28-CD3ζ anti-biotin-CD28TM/4-1-BB/CD3ζ	^Citation58
	CD20 (Rituximab), Her2 (Trasutuzumab), CCR4 (Mogamulizumab)		CD16 (FCGR3A) 158(V/V)(FCGR3A)-CD3TM/CD3ζ	^Citation55,Citation56
	CD20 (Rituximab), Her2 (Trasutuzumab), G2D (hu14.18K322A)		CD16 (FCGR3A) V158 variant-CD8ɑ/41BB/CD3ζ	^Citation53
	CD20 (Obinutuzumab), CD20 (Rituximab), EGFR (Cetuximab), EGFR (Panitumumab), MCSP (LC007 M4-3 ML2), MCSP (LC007 M4-3 ML2 ML2-g2)		CD16 (FCGR3A) 48H and V158 variant-CD28TM/CD28ICD/CD3ζ CD16 (FCGR3A) 48L and F158 variant-CD28TM/CD28ICD/CD3ζ CD16 (FCGR3A) 48L and V158 variant-CD28TM/CD28ICD/CD3ζ	^Citation57
	CD20 (Rituximab), ErbB2 (Herceptin)		CD16A (V158) CD8TM/CD28-CD3ζ	^Citation63
	CD20 (Rituximab), SEA-BCMA^Citation65		CD16A-41BB/CD3ζ	ClinicalTrials.gov Identifier:, NCT02776813 ClinicalTrials.gov Identifier: NCT03266692
	MSLN Her2 AxI	scFv-leucine zipper (SYN5, SYN 3, and EE)	Leucine zipper-CD28/41BB/CD3ζ	^Citation59

Figure 1. Schematic representation of a second generation, conventional CAR-T cell. CAR-T cells target surface antigens directly in a major histocompatibility class-independent manner. They consist of an extracellular antigen recognition domain (ECD), conventionally a single chain variable fragment (scFv), which is linked via hinge region (hinge) to a transmembrane domain (TM), followed by an intracellular costimulatory domain (ICD) and a CD3ζ signaling domain.

Figure 2. Indirect and flexible antigen targeting through CAR-adaptor molecules (CAR-adaptors). Depicted are the different designs of antigen-targeting CAR-adaptors that are used with a modular CAR (modCAR)-engineered effector cell. (a) IgG-tag-based CAR-adaptor | (b) IgG CAR-adaptor | (c) small molecule-based CAR-adaptor | (d) Fab-based CAR-adaptor | (e) scFv-based CAR-adaptor | (f) bispecific-based CAR-adaptor | (g) nanobody-based CAR-adaptor | (h) bivalent nanobody-based CAR-adaptor.

Figure 3. Depicted is a modular CAR (modCAR) engineered effector cell with diverse ECDs able to target a CAR adaptor molecule (CAR-adaptor), here represented by an IgG. (i) scFv-ECD | (j): FcR-ECD | (k) and (l) monomeric and dimeric avidin-ECD require a biotinylated CAR-AM to enable antigen targeting | (m) leucine zipper.

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