Generation and Validation of an IL-12 Fusion Protein Based on a Novel Anti-human FAP Monoclonal Antibody

Overview

Journal J Immunother Cancer

Specialties Allergy & Immunology
Oncology
Pharmacology

Date 2022 Sep 14

PMID 36104101

Authors

Lisa Nadal

Frederik Peissert

Abdullah Elsayed

Tobias Weiss

Thomas Look

Michael Weller

Geny Piro

Carmine Carbone

Giampaolo Tortora

Mattia Matasci

Nicholas Favalli

Riccardo Corbellari

Cesare Di Nitto

Eleonora Prodi

Chiara Libbra

Simone Galeazzi

Claudiopietro Carotenuto

Cornelia Halin

Emanuele Puca

Dario Neri

Roberto De Luca

Affiliations

Soon will be listed here.

Abstract

Background: In this study, we describe the generation of a fully human monoclonal antibody (named '7NP2') targeting human fibroblast activation protein (FAP), an antigen expressed in the microenvironment of different types of solid neoplasms.

Methods: 7NP2 was isolated from a synthetic antibody phage display library and was improved by one round of mutagenesis-based affinity maturation. The tumor recognition properties of the antibody were validated by immunofluorescence procedures performed on cancer biopsies from human patients. A fusion protein consisting of the 7NP2 antibody linked to interleukin (IL)-12 was generated and the anticancer activity of the murine surrogate product (named mIL12-7NP2) was evaluated in mouse models. Furthermore, the safety of the fully human product (named IL12-7NP2) was evaluated in monkeys.

Results: Biodistribution analysis in tumor-bearing mice confirmed the ability of the product to selectively localize to solid tumors while sparing healthy organs. Encouraged by these results, therapy studies were conducted , showing a potent antitumor activity in immunocompetent and immunodeficient mouse models of cancer, both as single agent and in combination with immune checkpoint inhibitors. The fully human product was tolerated when administered to non-human primates.

Conclusions: The results obtained in this work provided a rationale for future clinical translation activities using IL12-7NP2.

Citing Articles

Taming interleukin-12: Engineering of bispecific antibody-based IL-12 mimetics with biased agonism capacities.

Lipinski B, Unmuth L, Arras P, Endruszeit R, Becker S, Rodel J Protein Sci. 2025; 34(3):e70072.

PMID: 39981925 PMC: 11843475. DOI: 10.1002/pro.70072.

4-1BB agonist targeted to fibroblast activation protein α synergizes with radiotherapy to treat murine breast tumor models.

Garate-Soraluze E, Serrano-Mendioroz I, Fernandez-Rubio L, de Andrea C, Barrio-Alonso C, Herrero C J Immunother Cancer. 2025; 13(2).

PMID: 39933836 PMC: 11815443. DOI: 10.1136/jitc-2024-009852.

Antibody-based delivery of interleukin-2 modulates the immunosuppressive tumor microenvironment and achieves cure in pancreatic ductal adenocarcinoma syngeneic mice.

Carbone C, De Luca R, Puca E, Agostini A, Caggiano A, Priori L J Exp Clin Cancer Res. 2025; 44(1):7.

PMID: 39773310 PMC: 11705946. DOI: 10.1186/s13046-024-03238-x.

Augmentation of Solid Tumor Immunotherapy With IL-12.

Geils C, Kathrein K J Gene Med. 2024; 26(12):e70000.

PMID: 39618102 PMC: 11609498. DOI: 10.1002/jgm.70000.

Targeting PD-1 T cells with small-format immunocytokines enhances IL-12 antitumor activity.

Silva-Pilipich N, Beloki U, Apaolaza P, Igea A, Salaberry L, Prats-Mari L Mol Ther. 2024; 33(1):297-316.

PMID: 39563030 PMC: 11764877. DOI: 10.1016/j.ymthe.2024.11.027.

References

Pure E, Blomberg R . Pro-tumorigenic roles of fibroblast activation protein in cancer: back to the basics. Oncogene. 2018; 37(32):4343-4357. PMC: 6092565. DOI: 10.1038/s41388-018-0275-3. View

Fallon J, Tighe R, Kradjian G, Guzman W, Bernhardt A, Neuteboom B . The immunocytokine NHS-IL12 as a potential cancer therapeutic. Oncotarget. 2014; 5(7):1869-84. PMC: 4039112. DOI: 10.18632/oncotarget.1853. View

Corbellari R, Stringhini M, Mock J, Ongaro T, Villa A, Neri D . A Novel Antibody-IL15 Fusion Protein Selectively Localizes to Tumors, Synergizes with TNF-based Immunocytokine, and Inhibits Metastasis. Mol Cancer Ther. 2021; 20(5):859-871. PMC: 7611336. DOI: 10.1158/1535-7163.MCT-20-0853. View

Little R, Pluda J, Wyvill K, Rodriguez-Chavez I, Tosato G, Catanzaro A . Activity of subcutaneous interleukin-12 in AIDS-related Kaposi sarcoma. Blood. 2006; 107(12):4650-7. PMC: 1475826. DOI: 10.1182/blood-2005-11-4455. View

Sahai E, Astsaturov I, Cukierman E, DeNardo D, Egeblad M, Evans R . A framework for advancing our understanding of cancer-associated fibroblasts. Nat Rev Cancer. 2020; 20(3):174-186. PMC: 7046529. DOI: 10.1038/s41568-019-0238-1. View

Millul J, Bassi G, Mock J, Elsayed A, Pellegrino C, Zana A . An ultra-high-affinity small organic ligand of fibroblast activation protein for tumor-targeting applications. Proc Natl Acad Sci U S A. 2021; 118(16). PMC: 8072232. DOI: 10.1073/pnas.2101852118. View

Lyerly H, Osada T, Hartman Z . Right Time and Place for IL12: Targeted Delivery Stimulates Immune Therapy. Clin Cancer Res. 2018; 25(1):9-11. DOI: 10.1158/1078-0432.CCR-18-2819. View

Claus C, Ferrara C, Xu W, Sam J, Lang S, Uhlenbrock F . Tumor-targeted 4-1BB agonists for combination with T cell bispecific antibodies as off-the-shelf therapy. Sci Transl Med. 2019; 11(496). PMC: 7181714. DOI: 10.1126/scitranslmed.aav5989. View

Busek P, Vanickova Z, Hrabal P, Brabec M, Fric P, Zavoral M . Increased tissue and circulating levels of dipeptidyl peptidase-IV enzymatic activity in patients with pancreatic ductal adenocarcinoma. Pancreatology. 2016; 16(5):829-38. DOI: 10.1016/j.pan.2016.06.001. View

10.

Spitaleri G, Berardi R, Pierantoni C, De Pas T, Noberasco C, Libbra C . Phase I/II study of the tumour-targeting human monoclonal antibody-cytokine fusion protein L19-TNF in patients with advanced solid tumours. J Cancer Res Clin Oncol. 2012; 139(3):447-55. DOI: 10.1007/s00432-012-1327-7. View

11.

De Luca R, Soltermann A, Pretto F, Pemberton-Ross C, Pellegrini G, Wulhfard S . Potency-matched Dual Cytokine-Antibody Fusion Proteins for Cancer Therapy. Mol Cancer Ther. 2017; 16(11):2442-2451. PMC: 5844457. DOI: 10.1158/1535-7163.MCT-17-0211. View

12.

Rosenberg S . IL-2: the first effective immunotherapy for human cancer. J Immunol. 2014; 192(12):5451-8. PMC: 6293462. DOI: 10.4049/jimmunol.1490019. View

13.

Calais J . FAP: The Next Billion Dollar Nuclear Theranostics Target?. J Nucl Med. 2020; 61(2):163-165. DOI: 10.2967/jnumed.119.241232. View

14.

Gerosa F, Baldani-Guerra B, Lyakh L, Batoni G, Esin S, Winkler-Pickett R . Differential regulation of interleukin 12 and interleukin 23 production in human dendritic cells. J Exp Med. 2008; 205(6):1447-61. PMC: 2413040. DOI: 10.1084/jem.20071450. View

15.

Nguyen K, Vrabel M, Mantooth S, Hopkins J, Wagner E, Gabaldon T . Localized Interleukin-12 for Cancer Immunotherapy. Front Immunol. 2020; 11:575597. PMC: 7593768. DOI: 10.3389/fimmu.2020.575597. View

16.

Trinchieri G, Pflanz S, Kastelein R . The IL-12 family of heterodimeric cytokines: new players in the regulation of T cell responses. Immunity. 2003; 19(5):641-4. DOI: 10.1016/s1074-7613(03)00296-6. View

17.

Guida M, Casamassima A, Monticelli G, Quaranta M, Colucci G . Basal cytokines profile in metastatic renal cell carcinoma patients treated with subcutaneous IL-2-based therapy compared with that of healthy donors. J Transl Med. 2007; 5:51. PMC: 2169206. DOI: 10.1186/1479-5876-5-51. View

18.

Puca E, Probst P, Stringhini M, Murer P, Pellegrini G, Cazzamalli S . The antibody-based delivery of interleukin-12 to solid tumors boosts NK and CD8 T cell activity and synergizes with immune checkpoint inhibitors. Int J Cancer. 2019; 146(9):2518-2530. DOI: 10.1002/ijc.32603. View

19.

Liu R, Li H, Liu L, Yu J, Ren X . Fibroblast activation protein: A potential therapeutic target in cancer. Cancer Biol Ther. 2012; 13(3):123-9. DOI: 10.4161/cbt.13.3.18696. View

20.

Micke P, Ostman A . Tumour-stroma interaction: cancer-associated fibroblasts as novel targets in anti-cancer therapy?. Lung Cancer. 2004; 45 Suppl 2:S163-75. DOI: 10.1016/j.lungcan.2004.07.977. View