Targeting a membrane-proximal epitope on mesothelin increases the tumoricidal activity of a bispecific antibody blocking CD47 on mesothelin-positive tumors

Our plain-English summary of Targeting a membrane-proximal epitope on mesothelin increases the tumoricidal activity of a bispecific antibody blocking CD47 on mesothelin-positive tumors, by E. Hatterer, X. Chauchet, F. Richard, et al (mAbs, 2020).

The study

• Authors: E. Hatterer, X. Chauchet, F. Richard, L. Barba, V. Moine, L. Chatel, L. Broyer, G. Pontini, T. Bautzova, F. Juan, S. Calloud, N. Bosson, M. Charreton, K. Masternak, V. Buatois, L. Shang
• Journal: mAbs
• Published: 2020;12(1):1739408
• Cited by: 39 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Mesothelin (MSLN) is a cell surface glycoprotein overexpressed in several solid malignancies, including gastric, lung, mesothelioma, pancreatic and ovarian cancers. While several MSLN-targeting therapeutic approaches are in development, only limited efficacy has been achieved in patients. A potential shortcoming of several described antibody-based approaches is that they target the membrane distal region of MSLN and, additionally, are known to be handicapped by the high levels of circulating soluble MSLN in patients. We show here, using monoclonal antibodies (mAbs) targeting different MSLN-spanning epitopes, that the membrane-proximal region resulted in more efficient killing of MSLN-positive tumor cells in antibody-dependent cell-mediated cytotoxicity (ADCC) assays. Surprisingly, no augmented killing was observed in antibody-dependent cellular phagocytosis (ADCP) by mAbs targeting this membrane-proximal region. To further increase the ADCP potential, we, therefore, generated bispecific antibodies (bsAbs) coupling a high-affinity MSLN binding arm to a blocking CD47 arm. Here, targeting the membrane-proximal domain of MSLN demonstrated enhanced ADCP activity compared to membrane-distal domains when the bsAbs were used in in vitro phagocytosis killing assays. Importantly, the superior anti-tumor activity was also translated in xenograft tumor models. Furthermore, we show that the bsAb approach targeting the membrane-proximal epitope of MSLN optimized ADCC activity by augmenting FcγR-IIIA activation and enhanced ADCP via a more efficient blockade of the CD47/SIRPα axis.

In plain English

This laboratory study tested whether epitope position on mesothelin (MSLN) changes how well antibodies kill MSLN-positive tumour cells. Using the gastric cancer line NCI-N87 as targets and human donor PBMCs or macrophages as effectors, the authors compared anti-MSLN antibodies binding a membrane-proximal region (mAb#1) or membrane-distal regions (mAb#2, amatuximab). The mAb#1 or mAb#2 arms were then coupled to a CD47-blocking arm as bispecific antibodies, also tested on the lung line NCI-H226. The membrane-proximal mAb#1 gave more antibody-dependent cell-mediated cytotoxicity, but not more phagocytosis. In a representative experiment, bsAb#1 reached a maximal phagocytosis index of 61 tumour cells engulfed per 100 macrophages versus 27 for bsAb#2; FcγR-IIIA reporter cells gave about a 13.4-fold maximal luciferase increase with bsAb#1 versus twofold with bsAb#2. In NOD scid mice bearing MSLN-transfected HepG2 tumours, bsAb#1 completely inhibited tumour growth while bsAb#2 only partially did. The authors concluded that a membrane-proximal MSLN epitope with a CD47-blocking arm improved ADCC and ADCP in vitro and tumour control in vivo. All co-authors were company employees.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Preclinical laboratory study: in vitro antibody binding and effector-function assays with human cancer cell lines and donor immune cells, plus mouse xenograft experiments
• Who was studied: In vitro: human cancer cell lines (gastric NCI-N87 for the monoclonal antibody comparisons, lung NCI-H226 for a bispecific antibody comparison) with human PBMCs and monocyte-derived macrophages from healthy donors as effector cells. In vivo: immunodeficient NOD scid female mice aged 7-9 weeks transplanted with HepG2 cells transformed to express human MSLN and luciferase
• Exposure: Anti-MSLN monoclonal antibodies binding membrane-proximal region-III (mAb#1) or membrane-distal region-I (mAb#2, amatuximab), control mAbs 22A10 and 7D9, and CD47xMSLN bispecific antibodies (bsAb#1, bsAb#2, bsAb#3) sharing the same CD47 arm; dosed in mice at 6 or 60 mg/kg intravenously once weekly
• Main results:
  • Binding to NCI-N87 cells: EC50 19.95 ng/mL for mAb#2, 49.14 ng/mL for mAb#1 and 19.98 ng/mL for amatuximab
  • In ADCC with NCI-N87 targets and human PBMC effectors, the membrane-proximal mAb#1 was more effective than the membrane-distal mAb#2 and amatuximab
  • In phagocytosis of NCI-N87 cells, membrane-proximal targeting was less potent: EC50 3.2 ng/mL (mAb#2), 1.6 ng/mL (amatuximab) and 65.8 ng/mL (mAb#1), with similar maximal killing across the mAbs
  • In a representative phagocytosis assay with NCI-N87 cells, the maximal phagocytosis index (target cells engulfed per 100 macrophages) was 61 for bsAb#1 (membrane-proximal) versus 27 for bsAb#2 (membrane-distal)
  • With the lung carcinoma line NCI-H226, bsAb#1 induced higher ADCC and ADCP at maximum concentration than bsAb#2
  • In engineered FcγR-IIIA reporter Jurkat cells, bsAb#1 gave about a 13.4-fold increase in maximal luciferase activity versus a twofold increase with bsAb#2; FcγR-I and FcγR-IIA signalling were similarly increased by both
  • bsAb#1 was about 4.3-fold more potent than the matched monovalent anti-MSLN antibody in phagocytosis (EC50 2515 ng/mL vs 10794 ng/mL), while bsAb#2 was similar to its monovalent variant (19.4 vs 34.5 ng/mL)
  • F(ab')2 fragments combined with a suboptimal dose of anti-MSLN mAb 7D9 raised the phagocytosis index to 28.1 (B6H12) and 29 (bsAb#1) versus 16.5 with 7D9 alone; bsAb#2 F(ab')2 did not increase it
  • In the HepG2-hMSLN xenograft in NOD scid mice, bsAb#1 completely inhibited tumour growth while bsAb#2 only partially inhibited it
  • Both anti-MSLN mAbs inhibited tumour growth compared with hIgG1 isotype control, mAb#1 slightly more than mAb#2, without prolonging mouse survival
• The authors' conclusion: The authors concluded that the targeting domain on a tumour-associated antigen must be chosen carefully, and that for MSLN-positive solid tumours a membrane-proximal epitope coupled to a CD47-blocking arm gave improved ADCC and ADCP that translated into greater efficacy in their in vivo model
• Limitations the authors state:
  • The authors noted that NK effector cells are deficient in the NOD scid model used, so additional models with functional NK cells would be needed to assess the in vivo contribution of ADCC
  • The authors stated that further investigation would be needed into the effects of the bispecific antibodies on the phagocytosis signalling cascades they describe
  • All co-authors were employees of the company whose antibodies were tested

Topics

Medical terms in this study, explained in our Lexicon: Mesothelin, Glycoprotein, Mesothelioma, Ovarian Cancer, Monoclonal, Phagocytosis.

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