Isolation, partial characterization, and molecular cloning of a human colon adenocarcinoma cell-surface glycoprotein recognized by the C215 mouse monoclonal antibody.

Björk, Per; Jönsson, Ulf; Svedberg, Helena; Larsson, Kerstin; Lind, Peter; Dillner, Joakim; Hedlund, Gunnar; Dohlsten, Mikael; Kalland, Terje

doi:10.1016/s0021-9258(20)80515-8

Cited by 38 publications

(5 citation statements)

References 27 publications

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“…Trop-1/EpCAM and Trop-2 were previously shown to trigger Ca 2+ signals and to drive cancer cell growth 1,2,5,6,8,14,15 . We noticed that Trop-1 and Trop-2, despite distinct intra-cellular distribution patterns (Figure S1), were recruited at overlapping sites at the cell membrane (Figure 1).…”

Section: Resultsmentioning

confidence: 99%

“…The transmembrane glycoproteins Trop-1/EpCAM and Trop-2 have been shown to drive tumor growth [1][2][3] and metastatic diffusion 4,5 . Trops drive tumor progression upon overexpression as wildtype molecules 1,2,6,7 , which are activated by ADAM10 cleavage at a conserved position in the thyroglobulin domain 4,[8][9][10][11] . Trop proteolytic cleavage triggers a downstream proteolytic cascade, carried out by the TNF-a-converting enzyme (TACE) followed by g-secretase cleavage within the transmembrane domain, which leads to nuclear signaling and transcription factor activation 4,10,12,13 .…”

Section: Introductionmentioning

confidence: 99%

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Membrane cliffs are giant, recursive platforms that drive calcium and protein kinase signaling for cell growth

Trerotola,

Relli,

Tripaldi

et al. 2024

Preprint

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The transmembrane glycoproteins Trop-1/EpCAM and Trop-2 independently trigger Ca2+and kinase signals for cell growth and tumor progression. We discovered that Trop-1 and Trop-2 are recruited at overlapping sites at free cell edges. Z-stack analysis and three-dimensional reconstruction of these sites revealed previously unrecognized, protruding membrane regions ≥20 µm-long, up to 1.5 µm high, then named ‘cliffs’. Cliffs appeared confined to essentially immobile sites of the cell membrane, where they recursively assembled over hundreds of seconds. Cliffs were shown to recruit growth-driving kinases and downstream cytoplasmic effectors. Trop-2 stimulates cell growth through a membrane super-complex that comprises CD9 and PKCα. Our findings indicated that the growth-driving Trop-2 super-complex assembles at cliffs. Cliffs acted as sites of phosphorylation/activation of growth-driving kinases and as origins of Ca2+signaling waves, indicating cliffs as novel signaling platforms for drivers of cell growth. Cliffs were induced by growth factors and disappeared upon growth factor deprivation, suggesting cliffs as pivotal platforms for signaling for cell growth.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Membrane cliffs are giant, recursive platforms that drive calcium and protein kinase signaling for cell growth

Trerotola,

Relli,

Tripaldi

et al. 2024

Preprint

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“…3 ). In addition to the parental UBS54 ( K D = 0.88 μM) and affinity-tuned Y6V variant ( K D = 10.8 μM) CARs, we included a nanomolar affinity CAR ( K D = 3.95 nM 28 ) using scFv derived from the anti-EpCAM antibody C215 29 . After co-incubation with target cells with varying antigen density, i.e., 293T low , SW-1990 mid or HT-29 high cells (superscript indicates EpCAM expression levels), the levels of GFP induction in C215 CAR-T cells were similar to GFP expression obtained after stimulation with phorbol 12-myristate 13-acetate (PMA) and ionomycin (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…CAR constructs were synthesized and cloned by GenScript (Piscataway, New Jersey, USA) into a lentiviral plasmid backbone (VectorBuilder Inc., Vector Design Studio) under the regulation of a human elongation factor 1α (EF1α) promoter. EpCAM-specific CAR constructs contain from the 5′-LTR end: c-Myc tag, scFv derived from anti-EpCAM monoclonal antibody UBS54 39 or its affinity-tuned scFv variants (D1A, F3A, L4A, Y6A, Y6F, Y6L and Y6F), or scFv derived from C215 monoclonal antibody 29 , the CD8 α hinge, the transmembrane and cytoplasmic domains of CD28, and the cytoplasmic domain of CD3ζ. ICAM-1-specific CAR was composed of c-Myc tag, LFA-1 I domain (F292A), the CD8 α hinge and transmembrane domain, 4-1BB co-stimulatory domain and CD3ζ intracellular domain.…”

Section: Methodsmentioning

confidence: 99%

Inducible expression of interleukin-12 augments the efficacy of affinity-tuned chimeric antigen receptors in murine solid tumor models

Yang

Alcaina

et al. 2023

Nat Commun

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The limited number of targetable tumor-specific antigens and the immunosuppressive nature of the microenvironment within solid malignancies represent major barriers to the success of chimeric antigen receptor (CAR)-T cell therapies. Here, using epithelial cell adhesion molecule (EpCAM) as a model antigen, we used alanine scanning of the complementarity-determining region to fine-tune CAR affinity. This allowed us to identify CARs that could spare primary epithelial cells while still effectively targeting EpCAMhigh tumors. Although affinity-tuned CARs showed suboptimal antitumor activity in vivo, we found that inducible secretion of interleukin-12 (IL-12), under the control of the NFAT promoter, can restore CAR activity to levels close to that of the parental CAR. This strategy was further validated with another affinity-tuned CAR specific for intercellular adhesion molecule-1 (ICAM-1). Only in affinity-tuned CAR-T cells was NFAT activity stringently controlled and restricted to tumors expressing the antigen of interest at high levels. Our study demonstrates the feasibility of specifically gearing CAR-T cells towards recognition of solid tumors by combining inducible IL-12 expression and affinity-tuned CAR.

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“…Primary hepatocytes were obtained from HTCR Services GmbH (Munich, Germany). The following primary antibodies were used: EpCAM (clone C215 20 ; a kind gift of Dr. H. Lindhofer, Munich), SRRM2 (Abcam; clone 122719, Biozol; catalog no. 9206; Invitrogen; clone PA5-6682, and Sigma Aldrich; clone SC-35).…”

Section: Methodsmentioning

confidence: 99%

The nuclear speckles protein SRRM2 is a new therapeutic target molecule on the surface of cancer cells

Kellner,

Hörmann,

Fackler

et al. 2024

Preprint

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The membrane composition of extracellular vesicles (EVs) largely reflects that of the plasma membrane of the cell of origin. We therefore hypothesized that EVs are a source for the detection of hitherto unknown tumor-associated druggable target molecules. For this, we used EVs derived from cancer cell lines for an immunization of a rat. From this immunization, we obtained a monoclonal antibody specific for SRRM2, a protein involved in splicing a major component of nuclear speckles. Here, we used this antibody to demonstrate that SRRM2 is exposed at the surface of most cancer cell lines from various entities and, even more important, on cancer cells in vivo. Moreover, we demonstrate that SRRM2-specific CAR-T cells are killing SRRM2-positive cancer cells effectively. Collectively, we identified SRRM2 as a promising new target molecule exposed on the cancer cell surface and show that our SRRM2-specific antibody can be used as a basis for the development of new targeted cancer therapies.

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Isolation, partial characterization, and molecular cloning of a human colon adenocarcinoma cell-surface glycoprotein recognized by the C215 mouse monoclonal antibody.

Cited by 38 publications

References 27 publications

Membrane cliffs are giant, recursive platforms that drive calcium and protein kinase signaling for cell growth

Membrane cliffs are giant, recursive platforms that drive calcium and protein kinase signaling for cell growth

Inducible expression of interleukin-12 augments the efficacy of affinity-tuned chimeric antigen receptors in murine solid tumor models

The nuclear speckles protein SRRM2 is a new therapeutic target molecule on the surface of cancer cells

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