Dataset Information

The IgG4 hinge with CD28 transmembrane domain improves V_HH-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.

ABSTRACT: Heterogeneous antigen expression is a key barrier influencing the activity of chimeric antigen receptor (CAR) T cells in solid tumors. Here, we develop CAR T cells targeting glypican-1 (GPC1), an oncofetal antigen expressed in pancreatic cancer. We report the generation of dromedary camel V_HH nanobody (D4)-based CAR T cells targeting GPC1 and the optimization of the hinge (H) and transmembrane domain (TM) to improve activity. We find that a structurally rigid IgG4H and CD28TM domain brings the two D4 fragments in proximity, driving CAR dimerization and leading to enhanced T-cell signaling and tumor regression in pancreatic cancer models with low antigen density in female mice. Furthermore, single-cell-based proteomic and transcriptomic analysis of D4-IgG4H-CD28TM CAR T cells reveals specific genes (e.g., HMGB1) associated with high T-cell polyfunctionality. This study demonstrates the potential of V_HH-based CAR T for pancreatic cancer therapy and provides an engineering strategy for developing potent CAR T cells targeting membrane-distal epitopes.

SUBMITTER: Li N

PROVIDER: S-EPMC10082787 | biostudies-literature | 2023 Apr

REPOSITORIES: biostudies-literature

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The IgG4 hinge with CD28 transmembrane domain improves V<sub>H</sub>H-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.

Li Nan N Quan Alex A Li Dan D Pan Jiajia J Ren Hua H Hoeltzel Gerard G de Val Natalia N Ashworth Dana D Ni Weiming W Zhou Jing J Mackay Sean S Hewitt Stephen M SM Cachau Raul R Ho Mitchell M

Nature communications 20230408 1

Heterogeneous antigen expression is a key barrier influencing the activity of chimeric antigen receptor (CAR) T cells in solid tumors. Here, we develop CAR T cells targeting glypican-1 (GPC1), an oncofetal antigen expressed in pancreatic cancer. We report the generation of dromedary camel V<sub>H</sub>H nanobody (D4)-based CAR T cells targeting GPC1 and the optimization of the hinge (H) and transmembrane domain (TM) to improve activity. We find that a structurally rigid IgG4H and CD28TM domain b ...[more]

PMID: 37031249

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Project description:BackgroundPepsin agglutinators, discovered over 50 years ago, have been recently referred to as anti-hinge antibodies (AHAs) because of their reaction with the IgG hinge epitope. AHAs have different reactivity for each hinge epitope generated by each protease that cleaves the hinge region at different sites. Moreover, AHAs have different reactivity against different hinge epitopes derived from each IgG subclass even when the same protease is used. Since the expression of matrix metalloproteinase-3 (MMP-3) is enhanced in rheumatoid arthritis (RA), AHA production could also be increased. The purpose of this study was to determine whether the levels of AHAs against IgG hinge epitopes produced by MMP-3 are elevated in RA.MethodsThe serum levels of IgG or IgA AHAs against the IgG1/IgG4 F(ab')2 fragments, generated by either MMP-3 or pepsin, were measured using ELISA in 111 patients with RA and 81 healthy controls (HC). Receiver operating characteristic (ROC) analysis was used for obtaining optimal cutoff values and cutoff values indicating high specificity (> 95%) of the AHA. The targeted epitope of a specific AHA was investigated through inhibition ELISA.ResultsSeven AHAs were statistically higher in RA patients than in HC, except IgG AHA against IgG1 F(ab')2, which was generated by MMP-3 proteolytic cleavage. The areas under the ROC curve were 0.66-0.80, although the sensitivities at high specificity were low (5.4-24.3%). The cumulative number of positive AHAs in each individual was statistically higher in RA patients than in HC, suggesting the extreme extent of AHA repertoires in RA. Inhibition studies revealed that IgG AHAs against IgG4 F(ab')2 fragments generated by pepsin cross-reacted with IgG1 F(ab')2 fragments generated by pepsin. Multivariate logistic regression analysis identified the IgG AHA against IgG4 F(ab')2 fragments generated by pepsin as an independent variable for RA diagnosis, even in RA patients who were negative for both RF and ACPA (odds ratio 1.18, 95% CI 1.06-1.32; P = 0.003). Additional experiments using non-RA patients finally strengthened the diagnostic utility.ConclusionIn RA patients, we observed diversification and amplification of AHA repertoires and diagnostic utility of the specific AHA against IgG4 F(ab')2 fragments generated by pepsin but not MMP-3.

Dataset Information

The IgG4 hinge with CD28 transmembrane domain improves V_HH-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.

Publications

The IgG4 hinge with CD28 transmembrane domain improves V<sub>H</sub>H-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.

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Dataset Information

The IgG4 hinge with CD28 transmembrane domain improves VHH-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.

Publications

The IgG4 hinge with CD28 transmembrane domain improves V<sub>H</sub>H-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

The IgG4 hinge with CD28 transmembrane domain improves V_HH-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer.