CD38-targeted attenuated interferon alpha immunocytokine activates both innate and adaptive immune cells to drive anti-tumor activity
James F. Sampson, Hong Zhang, Dongmei Zhang, Mingying Bi, Adam Hinthorne, Sakeena Syed, Yuhong Zhang, Nibedita Chattopadhyay, Sabrina Collins, Sarah Pogue, Pia Björck, Michael Curley
Abstract
Recombinant interferon alpha (IFNα) has been used to treat cancer patients for over 30 years; however, its clinical utility has been limited by a narrow therapeutic index. Given the recognized anti-tumor and immunomodulatory impacts of IFNα, the development of novel strategies to harness these attributes while minimizing associated toxicity could provide significant benefit for patients. The concept of attenuating IFNα binding affinity for its receptor was conceived to address this challenge and led to the development of CD38-targeted Attenukine™, a CD38-targeted antibody attenuated IFNα immunocytokine. In this study, we sought to delineate the effects of targeting AttenukineTM specifically to tumor cells and/or immune cells using an antibody to CD38, a cell surface glycoprotein expressed on certain tumor and immune cells, using different mouse models and anti-human or anti-mouse CD38-targeted Attenukine™. Our results demonstrate that an anti-human CD38 AttenukineTM inhibits tumor growth through direct anti-proliferative effects of IFNα on CD38 + tumor cells as well as by indirectly modulating the anti-tumor immune response.
Introduction
Interferon alpha (IFNα) is a pleiotropic cytokine that, through binding to the widely expressed IFNα receptor (IFNAR), mediates broad-spectrum signal activation with diverse outcomes including immune activation, direct tumor cell cytotoxicity, and inhibition of tumor cell proliferation [1]. Despite its potent anti-tumor and pro-inflammatory attributes and some promising signs of activity in certain tumor indications, the clinical utility of recombinant IFNα has been limited due to a narrow therapeutic index, driven by significant IFNα-related toxicities including nausea, severe flu-like symptoms, vasculopathic complications (e.g., thrombocytopenia and leukopenia), and negative impacts on the nervous system, resulting in depression and anxiety [2–5].
Materials and method
Generation of treatment agents
Anti-human CD38-human AttenukineTM (hCD38-hAtt) was generated as previously described [11]. Briefly, reference anti-CD38 antibody variable regions were generated by polymerase chain reaction (PCR) from published V region sequences (WO 2013/059885). The human IFNα2b gene was isolated from HEK293 genomic DNA, and one attenuating point mutation was introduced by PCR at a residue which interacts with the high-affinity IFNα receptor chain, IFNAR2 [12]. Anti-CD38 and IFNα2b gene fragments were cloned into the pTT5 mammalian expression vector [13] containing human immunoglobulin (Ig)G4 and kappa Ig constant region genes.
Results
hCD38-hAtt demonstrates anti-tumor activity in xenograft models of multiple myeloma
Anti-human CD38-hAtt (hCD38-hAtt) has been shown to reduce the proliferation of leukemia and lymphoma cell lines in vitro, and to inhibit the growth of CD38-expressing xenograft tumors in immunodeficient mice [11]. Given the historical use of recombinant IFNα as a treatment option for patients with multiple myeloma, the impact of hCD38-hAtt on the growth of multiple myeloma xenograft tumors was evaluated. In studies evaluating the anti-tumor activity of hCD38-hAtt against ANBL-6, LP-1, MM1.S, and JJN-3 multiple myeloma tumors in vivo, CRs were observed in most hCD38-hAtt-treated mice (Fig 1 A–D). Anti-tumor activity of hCD38-hAtt in these xenograft tumor models did not directly correlate with the levels of expression of CD38 or IFNAR2 of these tumor cell lines in vitro (Fig 1E).
Discussion
In this study, we sought to delineate the effects of a CD38-targeted attenuated IFNα immunocytokine (CD38-AttenukineTM) on tumor cells and/or immune cells, through leveraging different immunocompromised and immunocompetent mouse models. In immunocompromised, CD38-expressing tumor model settings, robust anti-tumor activity of hCD38-hAtt was demonstrated. The depth of response somewhat aligned, but was not directly correlated, with tumor CD38 expression levels in vitro (Fig 1), though we do acknowledge that levels of target expression on cell lines may not necessarily remain consistent when evaluated in in vitro and in vivo settings.
Acknowledgments
Authors would like to sincerely thank James Garnsey and Adrienne Xenos for in vitro data presented in supplementary figures, Marjan Fatholahi for assay design support, Tetsuya Taura for reagent generation, and David Wilson for scientific advice.
Citation: Sampson JF, Zhang H, Zhang D, Bi M, Hinthorne A, Syed S, et al. (2025) CD38-targeted attenuated interferon alpha immunocytokine activates both innate and adaptive immune cells to drive anti-tumor activity. PLoS One 20(5): e0321622. https://doi.org/10.1371/journal.pone.0321622
Editor: Abhinava Kumar Mishra, University of California Santa Barbara, UNITED STATES OF AMERICA
Received: July 25, 2024; Accepted: March 10, 2025; Published: May 2, 2025
Copyright: © 2025 Sampson et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: Takeda’s policy on data sharing is that each data request needs to be done formally and assessed individually by Takeda, therefore we are unable to deposit the data on a database. The datasets generated and/or analyzed during the current study are available from the corresponding author upon request. In addition to the corresponding author, data requests can be sent to the Global Head of Oncology Drug Discovery, Kathy Seidl ([email protected]) or the Head of Scientific Communications, Caroline Ojaimi ([email protected]). There are no third-party data provided in the submitted manuscript. All data were generated by authors, and could be made available upon request. The authors did not have any special access privileges that others would not have
Funding: The funder provided support in the form of salaries for authors [JFS, HZ, DZ, AH, SS, YZ, NC, SC, and MC], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.
Competing interests: J.F.S. reports employment with Takeda. H.Z. reports employment with Takeda at the time of the study. D.Z. reports employment with Takeda. M.B. reports employment with Teva. A.H. reports employment with Takeda. S.S. reports employment with Takeda at the time of the study. Y.Z. reports employment with and ownership of stock/shares in Takeda. N.C. reports employment with Takeda at the time of the study. S.C. reports employment with and ownership of stock/shares in Takeda. S.P. and P.B. report employment with and ownership of stock/shares in Teva. M.C. reports employment with and ownership of stock/shares in Takeda, and ownership of stock/shares in Merrimack Pharmaceuticals.
