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Enhanced ADCC and NK Cell Activation of an Anticarcinoma Bispecific Antibody by Genetic Insertion of a Modified IL-15 Cross-linker.

ABSTRACT: Previously, we constructed a bispecific NK-cell-engager (BiKE) bearing single-chain variable fragments (scFv) against CD16 on NK cells and EpCAM on tumor cells. This BiKE facilitated antigen-specific antibody-dependent cell-mediated cytotoxicity (ADCC) but did not induce NK cell expansion. We incorporated a modified interleukin-15 cross-linker to create a trispecific construct (TriKE) in order to improve activation, proliferation, and survival of NK cells. Synthesis and assembly of hybrid genes encoding the TriKE was accomplished using DNA-shuffling and DNA-ligation techniques. The TriKE was tested for specificity, efficacy, proliferative capability, and cytokine profile using functional assays. The molecular modifications improved yield without compromising binding to EpCAM(+) HT-29 colorectal carcinoma cells. (51)Chromium-release and degranulation assays showed better killing rates with TriKE compared to BiKE. TriKE was more active in a variety of different carcinoma cell lines. TriKE showed the ability to stimulate expansion of CD56(+)CD3(-) NK cells. BiKE and TriKE showed enhanced but not supraphysiologic levels of cytokine secretion. 1615EpCAM TriKE drives enhanced ADCC while significantly improving proliferation, activation, and survival of NK cell effectors. The TriKE provides a selectively delivered self-sustaining signal at the NK/tumor cell synapse. Targeted cytokine stimulation, rather than systemic cytokine administration, may impact toxicity in patients rendering the TriKE a promising new off-the-shelf carcinoma therapy.

SUBMITTER: Schmohl JU

PROVIDER: S-EPMC5088765 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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Enhanced ADCC and NK Cell Activation of an Anticarcinoma Bispecific Antibody by Genetic Insertion of a Modified IL-15 Cross-linker.

Schmohl Joerg U JU Felices Martin M Taras Elizabeth E Miller Jeff S JS Vallera Daniel A DA

Molecular therapy : the journal of the American Society of Gene Therapy 20160509 7

Previously, we constructed a bispecific NK-cell-engager (BiKE) bearing single-chain variable fragments (scFv) against CD16 on NK cells and EpCAM on tumor cells. This BiKE facilitated antigen-specific antibody-dependent cell-mediated cytotoxicity (ADCC) but did not induce NK cell expansion. We incorporated a modified interleukin-15 cross-linker to create a trispecific construct (TriKE) in order to improve activation, proliferation, and survival of NK cells. Synthesis and assembly of hybrid genes ...[more]

PMID: 27157665

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Project description:NK cells have potent antitumor capacity. They are enriched in the human liver, with a large subset specialized for tissue-residence. The potential for liver-resident versus liver-infiltrating NK cells to populate, and exert antitumor functions in, human liver tumors has not been studied. We examined liver-resident and liver-infiltrating NK cells directly ex vivo from human hepatocellular carcinomas (HCCs) and liver colorectal (CRC) metastases, compared with matched uninvolved liver tissue. We found that NK cells were highly prevalent in both HCC and liver CRC metastases, although at lower frequencies than unaffected liver. Up to 79% of intratumoral NK cells had the CXCR6+CD69+ liver-resident phenotype. Direct ex vivo staining showed that liver-resident NK cells had increased NKG2D expression compared to their non-resident counterparts, but both subsets had NKG2D downregulation within liver tumors compared to uninvolved liver. Proliferation of intratumoral NK cells (identified by Ki67) was selectively impaired in those with the most marked NKG2D downregulation. Human liver tumor NK cells were functionally impaired, with reduced capacity for cytotoxicity and production of cytokines, even when compared to the hypo-functional tissue-resident NK cells in unaffected liver. Coculture of human liver NK cells with the human hepatoma cell line PLC/PRF/5, or with autologous HCC, recapitulated the defects observed in NK cells extracted from tumors, with downmodulation of NKG2D, cytokine production, and target cell cytotoxicity. Transwells and conditioned media confirmed a requirement for cell contact with PLC/PRF/5 to impose NK cell inhibition. IL-15 was able to recover antitumor functionality in NK cells inhibited by in vitro exposure to HCC cell lines or extracted directly from HCC. In summary, our data suggest that the impaired antitumor function of local NK cells reflects a combination of the tolerogenic features inherent to liver-resident NK cells together with additional contact-dependent inhibition imposed by HCC itself. The demonstration that IL-15 can recover hepatic NK cell function following tumor exposure supports its inclusion in immunotherapy strategies.

Dataset Information

Enhanced ADCC and NK Cell Activation of an Anticarcinoma Bispecific Antibody by Genetic Insertion of a Modified IL-15 Cross-linker.

Publications

Enhanced ADCC and NK Cell Activation of an Anticarcinoma Bispecific Antibody by Genetic Insertion of a Modified IL-15 Cross-linker.

Similar Datasets

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