Skip to main content

Advertisement

SpringerLink
Log in

CD20/TNFR1 dual-targeting antibody enhances lysosome rupture-mediated cell death in B cell lymphoma

  • Research
  • Published:
Cancer Immunology, Immunotherapy Aims and scope Submit manuscript

Abstract

Obinutuzumab is a therapeutic antibody for B cell non-Hodgkin’s Lymphoma (BNHL), which is a glyco-engineered anti-CD20 antibody with enhanced antibody-dependent cellular cytotoxicity (ADCC) and causes binding-induced direct cell death (DCD) through lysosome membrane permeabilization (LMP). Tumour necrosis factor receptor 1 (TNFR1), a pro-inflammatory death receptor, also evokes cell death, partly through lysosomal rupture. As both obinutuzumab- and TNFR1-induced cell deaths are mediated by LMP and combining TNFR1 and obinutuzumab can amplify LMP-mediated cell death, we made dual-targeting antibody for CD20 and TNFR1 to enhance DCD of obinutuzumab.

Obinutuzumab treatment-induced CD20 and TNFR1 colocalisation, and TNFR1-overexpressing cells showed increased obinutuzumab-induced DCD. Two targeting modes, anti-CD20/TNFR1 bispecific antibodies (bsAbs), and obinutuzumab-TNFα fusion proteins (OBI-TNFαWT and OBI-TNFαMUT), were designed to cluster CD20 and TNFR1 on the plasma membrane. OBI-TNFαWT and OBI-TNFαMUT showed significantly enhanced LMP, DCD, and ADCC compared with that induced by obinutuzumab. TNFR1 expression is upregulated in many BNHL subtypes compared to that in normal B cells; OBI-TNFαMUT specifically increased DCD and ADCC in a B cell lymphoma cell line overexpressing TNFR1. Further, OBI-TNFαMUT blocked NF-κB activation in the presence of TNF-α, implying that it can antagonise the proliferative role of TNF-α in cancers.

Our study suggests that dual targeting of CD20 and TNFR1 can be a new therapeutic strategy for improving BNHL treatment. The OBI-TNFαMUT fusion protein enhances DCD and ADCC and prevents the proliferating effect of TNFα signalling; therefore, it may provide precision treatment for patients with BNHL, especially those with upregulated TNFR1 expression.

Graphical Abstract

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

ADCC:

Antibody-dependent cellular cytotoxicity

BNHL:

B cell non-Hodgkin lymphoma

bsAb:

Bispecific antibody

CLL:

Chronic lymphocytic leukaemia

DCD:

Direct cell death

DLBCL:

Diffuse large B cell lymphoma

EGFR:

Epidermal growth factor receptor

EV:

Empty vector

FL:

Follicular lymphoma

LMP:

Lysosomal membrane permeabilization

MMML:

Molecular mechanism in malignant lymphoma

NHL:

Non-Hodgkin lymphoma

OBI-TNFαWT :

Obinutuzumab-tumour necrosis factor alpha wild-type fusion protein

OBI-TNFαMUT :

Obinutuzumab-tumour necrosis factor alpha mutant fusion protein

TNFR1:

Tumour necrosis factor receptor 1

References

  1. Shaffer AL III, Young RM, Staudt LM (2012) Pathogenesis of human B cell lymphomas. Annu Rev Immunol 30:565–610. https://doi.org/10.1146/annurev-immunol-020711-075027

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Crisci S, Di Francia R, Mele S, Vitale P, Ronga G, De Filippi R, Berretta M, Rossi P, Pinto A (2019) Overview of targeted drugs for mature B-cell non-hodgkin lymphomas. Front Oncol 9:443. https://doi.org/10.3389/fonc.2019.00443

    Article  PubMed  PubMed Central  Google Scholar 

  3. Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, Bray F (2021) Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 71:209–249. https://doi.org/10.3322/caac.21660

    Article  PubMed  Google Scholar 

  4. Plosker GL, Figgitt DP (2003) Rituximab: a review of its use in non-Hodgkin’s lymphoma and chronic lymphocytic leukaemia. Drugs 63:803–843. https://doi.org/10.2165/00003495-200363080-00005

    Article  CAS  PubMed  Google Scholar 

  5. Coiffier B, Lepage E, Brière J et al (2002) CHOP chemotherapy plus rituximab compared with CHOP alone in elderly patients with diffuse large-B-cell lymphoma. N Engl J Med 346:235–242. https://doi.org/10.1056/NEJMoa011795

    Article  CAS  PubMed  Google Scholar 

  6. Cartron G, Trappe RU, Solal-Céligny P, Hallek M (2011) Interindividual variability of response to rituximab: from biological origins to individualized therapies. Clin Cancer Res 17:19–30. https://doi.org/10.1158/1078-0432.Ccr-10-1292

    Article  CAS  PubMed  Google Scholar 

  7. Czuczman MS, Olejniczak S, Gowda A et al (2008) Acquirement of rituximab resistance in lymphoma cell lines is associated with both global CD20 gene and protein down-regulation regulated at the pretranscriptional and posttranscriptional levels. Clin Cancer Res 14:1561–1570. https://doi.org/10.1158/1078-0432.Ccr-07-1254

    Article  CAS  PubMed  Google Scholar 

  8. Bonavida B (2014) Postulated mechanisms of resistance of B-cell non-Hodgkin lymphoma to rituximab treatment regimens: strategies to overcome resistance. Semin Oncol 41:667–677. https://doi.org/10.1053/j.seminoncol.2014.08.006

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Shanafelt TD, Wang XV, Kay NE et al (2019) Ibrutinib-rituximab or chemoimmunotherapy for chronic lymphocytic leukemia. N Engl J Med 381:432–443. https://doi.org/10.1056/NEJMoa1817073

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Tobinai K, Klein C, Oya N, Fingerle-Rowson G (2017) A review of obinutuzumab (GA101), a novel type II Anti-CD20 monoclonal antibody, for the treatment of patients with B-cell malignancies. Adv Ther 34:324–356. https://doi.org/10.1007/s12325-016-0451-1

    Article  CAS  PubMed  Google Scholar 

  11. Freeman CL, Sehn LH (2018) A tale of two antibodies: obinutuzumab versus rituximab. Br J Haematol 182:29–45. https://doi.org/10.1111/bjh.15232

    Article  PubMed  Google Scholar 

  12. Mössner E, Brünker P, Moser S et al (2010) Increasing the efficacy of CD20 antibody therapy through the engineering of a new type II anti-CD20 antibody with enhanced direct and immune effector cell-mediated B-cell cytotoxicity. Blood 115:4393–4402. https://doi.org/10.1182/blood-2009-06-225979

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Niederfellner G, Lammens A, Mundigl O et al (2011) Epitope characterization and crystal structure of GA101 provide insights into the molecular basis for type I/II distinction of CD20 antibodies. Blood 118:358–367. https://doi.org/10.1182/blood-2010-09-305847

    Article  CAS  PubMed  Google Scholar 

  14. Kumar A, Planchais C, Fronzes R, Mouquet H, Reyes N (2020) Binding mechanisms of therapeutic antibodies to human CD20. Science 369:793–799. https://doi.org/10.1126/science.abb8008

    Article  CAS  PubMed  Google Scholar 

  15. Alduaij W, Ivanov A, Honeychurch J et al (2011) Novel type II anti-CD20 monoclonal antibody (GA101) evokes homotypic adhesion and actin-dependent, lysosome-mediated cell death in B-cell malignancies. Blood 117:4519–4529. https://doi.org/10.1182/blood-2010-07-296913

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Honeychurch J, Alduaij W, Azizyan M, Cheadle EJ, Pelicano H, Ivanov A, Huang P, Cragg MS, Illidge TM (2012) Antibody-induced nonapoptotic cell death in human lymphoma and leukemia cells is mediated through a novel reactive oxygen species-dependent pathway. Blood 119:3523–3533. https://doi.org/10.1182/blood-2011-12-395541

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Boya P, Kroemer G (2008) Lysosomal membrane permeabilization in cell death. Oncogene 27:6434–6451. https://doi.org/10.1038/onc.2008.310

    Article  CAS  PubMed  Google Scholar 

  18. Jak M, van Bochove GG, Reits EA et al (2011) CD40 stimulation sensitizes CLL cells to lysosomal cell death induction by type II anti-CD20 mAb GA101. Blood 118:5178–5188. https://doi.org/10.1182/blood-2011-01-331702

    Article  CAS  PubMed  Google Scholar 

  19. Domagala A, Bobrowicz M, Stachura J, Siernicka M, Mrowka P, Dwojak M, Pyrzynska B, Firczuk M, Winiarska M (2016) Lysosomal disruption augments obinutuzumab-induced direct cell death. Blood 128:2766. https://doi.org/10.1182/blood.V128.22.2766.2766

    Article  Google Scholar 

  20. van Horssen R, ten Hagen TLM, Eggermont AMM (2006) TNF-α in cancer treatment: molecular insights, antitumor effects, and clinical utility. Oncologist 11:397–408. https://doi.org/10.1634/theoncologist.11-4-397

    Article  PubMed  Google Scholar 

  21. Montfort A, Colacios C, Levade T, Andrieu-Abadie N, Meyer N, Ségui B (2019) The TNF paradox in cancer progression and immunotherapy. Front Immunol. https://doi.org/10.3389/fimmu.2019.01818

    Article  PubMed  PubMed Central  Google Scholar 

  22. Dürr C, Hanna BS, Schulz A et al (2018) Tumor necrosis factor receptor signaling is a driver of chronic lymphocytic leukemia that can be therapeutically targeted by the flavonoid wogonin. Haematologica 103:688–697. https://doi.org/10.3324/haematol.2017.177808

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Nakayama S, Yokote T, Hirata Y et al (2014) TNF-α expression in tumor cells as a novel prognostic marker for diffuse large B-cell lymphoma, not otherwise specified. Am J Surg Pathol 38:228–234. https://doi.org/10.1097/pas.0000000000000094

    Article  PubMed  Google Scholar 

  24. Nakayama S, Yokote T, Tsuji M et al (2014) TNF-α receptor 1 expression predicts poor prognosis of diffuse large B-cell lymphoma, not otherwise specified. Am J Surg Pathol 38:1138–1146. https://doi.org/10.1097/pas.0000000000000219

    Article  PubMed  Google Scholar 

  25. Schneider-Brachert W, Tchikov V, Neumeyer J et al (2004) Compartmentalization of TNF receptor 1 signaling: internalized TNF receptosomes as death signaling vesicles. Immunity 21:415–428. https://doi.org/10.1016/j.immuni.2004.08.017

    Article  CAS  PubMed  Google Scholar 

  26. Schütze S, Tchikov V, Schneider-Brachert W (2008) Regulation of TNFR1 and CD95 signalling by receptor compartmentalization. Nat Rev Mol Cell Biol 9:655–662. https://doi.org/10.1038/nrm2430

    Article  CAS  PubMed  Google Scholar 

  27. Ullio C, Casas J, Brunk UT, Sala G, Fabriàs G, Ghidoni R, Bonelli G, Baccino FM, Autelli R (2012) Sphingosine mediates TNFα-induced lysosomal membrane permeabilization and ensuing programmed cell death in hepatoma cells. J Lipid Res 53:1134–1143. https://doi.org/10.1194/jlr.M022384

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Werneburg NW, Guicciardi ME, Bronk SF, Gores GJ (2002) Tumor necrosis factor-alpha-associated lysosomal permeabilization is cathepsin B dependent. Am J Physiol Gastrointest Liver Physiol 283:G947–G956. https://doi.org/10.1152/ajpgi.00151.2002

    Article  CAS  PubMed  Google Scholar 

  29. Ono K, Kim SO, Han J (2003) Susceptibility of lysosomes to rupture is a determinant for plasma membrane disruption in tumor necrosis factor alpha-induced cell death. Mol Cell Biol 23:665–676. https://doi.org/10.1128/MCB.23.2.665-676.2003

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Zhang F, Yang J, Li H et al (2016) Combating rituximab resistance by inducing ceramide/lysosome-involved cell death through initiation of CD20-TNFR1 co-localization. Oncoimmunology 5:e1143995. https://doi.org/10.1080/2162402x.2016.1143995

    Article  PubMed  PubMed Central  Google Scholar 

  31. Chan S, Belmar N, Ho S et al (2022) An anti-PD-1–GITR-L bispecific agonist induces GITR clustering-mediated T cell activation for cancer immunotherapy. Nature Cancer 3:337–354. https://doi.org/10.1038/s43018-022-00334-9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Janda CY, Dang LT, You C et al (2017) Surrogate Wnt agonists that phenocopy canonical Wnt and β-catenin signalling. Nature 545:234–237. https://doi.org/10.1038/nature22306

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Yen M, Ren J, Liu Q et al (2022) Facile discovery of surrogate cytokine agonists. Cell 185:1414–30.e19. https://doi.org/10.1016/j.cell.2022.02.025

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Bajic D, Chester K, Neri D (2020) An antibody-tumor necrosis factor fusion protein that synergizes with oxaliplatin for treatment of colorectal cancer. Mol Cancer Ther 19:2554–2563. https://doi.org/10.1158/1535-7163.Mct-19-0729

    Article  CAS  PubMed  Google Scholar 

  35. Shibata H, Yoshioka Y, Ohkawa A et al (2008) Creation and X-ray structure analysis of the tumor necrosis factor receptor-1-selective mutant of a tumor necrosis factor-alpha antagonist. J Biol Chem 283:998–1007. https://doi.org/10.1074/jbc.M707933200

    Article  CAS  PubMed  Google Scholar 

  36. Ridgway JB, Presta LG, Carter P (1996) ‘Knobs-into-holes’ engineering of antibody CH3 domains for heavy chain heterodimerization. Protein Eng 9:617–621. https://doi.org/10.1093/protein/9.7.617

    Article  CAS  PubMed  Google Scholar 

  37. Lewis SM, Wu X, Pustilnik A et al (2014) Generation of bispecific IgG antibodies by structure-based design of an orthogonal Fab interface. Nat Biotechnol 32:191–198. https://doi.org/10.1038/nbt.2797

    Article  CAS  PubMed  Google Scholar 

  38. Santich BH, Park JA, Tran H, Guo HF, Huse M, Cheung NV (2020) Interdomain spacing and spatial configuration drive the potency of IgG-[L]-scFv T cell bispecific antibodies. Sci Transl Med. https://doi.org/10.1126/scitranslmed.aax1315

    Article  PubMed  PubMed Central  Google Scholar 

  39. Loeffler-Wirth H, Kreuz M, Hopp L et al (2019) A modular transcriptome map of mature B cell lymphomas. Genome Med 11:27. https://doi.org/10.1186/s13073-019-0637-7

    Article  PubMed  PubMed Central  Google Scholar 

  40. Kapoor P, Greipp PT, Morice WG, Rajkumar SV, Witzig TE, Greipp PR (2008) Anti-CD20 monoclonal antibody therapy in multiple myeloma. Br J Haematol 141:135–148. https://doi.org/10.1111/j.1365-2141.2008.07024.x

    Article  CAS  PubMed  Google Scholar 

  41. Bradley JR (2008) TNF-mediated inflammatory disease. J Pathol 214:149–160. https://doi.org/10.1002/path.2287

    Article  CAS  PubMed  Google Scholar 

  42. Lech-Maranda E, Bienvenu J, Broussais-Guillaumot F, Warzocha K, Michallet AS, Robak T, Coiffier B, Salles G (2010) Plasma TNF-alpha and IL-10 level-based prognostic model predicts outcome of patients with diffuse large B-cell lymphoma in different risk groups defined by the International Prognostic Index. Arch Immunol Ther Exp (Warsz) 58:131–141. https://doi.org/10.1007/s00005-010-0066-1

    Article  CAS  PubMed  Google Scholar 

  43. Zhang H, Yan D, Shi X et al (2008) Transmembrane TNF-alpha mediates “forward” and “reverse” signaling, inducing cell death or survival via the NF-kappaB pathway in Raji Burkitt lymphoma cells. J Leukoc Biol 84:789–797. https://doi.org/10.1189/jlb.0208078

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Goede V, Fischer K, Busch R et al (2014) Obinutuzumab plus chlorambucil in patients with CLL and coexisting conditions. N Engl J Med 370:1101–1110. https://doi.org/10.1056/NEJMoa1313984

    Article  CAS  PubMed  Google Scholar 

  45. Lee JM, Lee SH, Hwang JW et al (2016) Novel strategy for a bispecific antibody: induction of dual target internalization and degradation. Oncogene 35:4437–4446. https://doi.org/10.1038/onc.2015.514

    Article  CAS  PubMed  Google Scholar 

  46. Lu D, Zhang H, Koo H et al (2005) A fully human recombinant IgG-like bispecific antibody to both the epidermal growth factor receptor and the insulin-like growth factor receptor for enhanced antitumor activity. J Biol Chem 280:19665–19672. https://doi.org/10.1074/jbc.M500815200

    Article  CAS  PubMed  Google Scholar 

  47. Gabandé-Rodríguez E, Boya P, Labrador V, Dotti CG, Ledesma MD (2014) High sphingomyelin levels induce lysosomal damage and autophagy dysfunction in Niemann Pick disease type A. Cell Death Differ 21:864–875. https://doi.org/10.1038/cdd.2014.4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Reddy V, Dahal LN, Cragg MS, Leandro M (2016) Optimising B-cell depletion in autoimmune disease: is obinutuzumab the answer? Drug Discov Today 21:1330–1338. https://doi.org/10.1016/j.drudis.2016.06.009

    Article  CAS  PubMed  Google Scholar 

  49. Reddy V, Klein C, Isenberg DA, Glennie MJ, Cambridge G, Cragg MS, Leandro MJ (2017) Obinutuzumab induces superior B-cell cytotoxicity to rituximab in rheumatoid arthritis and systemic lupus erythematosus patient samples. Rheumatology (Oxford) 56:1227–1237. https://doi.org/10.1093/rheumatology/kex067

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors are very grateful to Dr. Henry Löffler-Wirth and Dr. Hans of the Interdisciplinary Center for Bioinformatics at the University of Leipzig for making the whole expression data set of mature B cell lymphomas available to the public in the repository ‘The Leipzig Health Atlas’ (https://www.health-atlas.de/index.php/en/lha/7VT47TM4GV-1). We would also thank Professor Joon Yong Ahn of the Department of Biosystem and Biomedical Science, Korea University, for his advise on big data analysis.

Funding

This work was supported by grants from the National Research Foundation of Korea, Project Nos. NRF-2021R1I1A1A01059867 to D.H.L and Project Nos. NRF-2019R1A2C1086348 to J.Y.K.

Author information

Authors and Affiliations

  1. Department of Pharmacology and Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, 50-1 Yonsei-Ro, Seodaemun-Gu, Seoul, 03080, Republic of Korea

    Jeong Ryeol Kim, Donghyuk Lee, Yerim Kim & Joo Young Kim

Authors
  1. Jeong Ryeol Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Donghyuk Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Yerim Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Joo Young Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

JRK performed and analyzed most of the experiments. JYK organized and supervised the whole project. YK helped and reproduced Lysotracker assays, DL produced obinutuzumab-TNFα fusion protein and provided technical advice. JRK and JYK wrote the manuscript and YK and DL edited the English.

Corresponding author

Correspondence to Joo Young Kim.

Ethics declarations

Conflict of interest

All authors have declared that no competing interest exists.

Consent to participate

Patient-derived PBMC samples were isolated with approval of Yonsei University Institutional Review Committee after obtaining informed consent, under IRP procedure (#4-2016-0600).

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (PDF 828 kb)

Supplementary file2 (PDF 406 kb)

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kim, J.R., Lee, D., Kim, Y. et al. CD20/TNFR1 dual-targeting antibody enhances lysosome rupture-mediated cell death in B cell lymphoma. Cancer Immunol Immunother 72, 1567–1580 (2023). https://doi.org/10.1007/s00262-022-03344-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00262-022-03344-9

Keywords

Search

Navigation