CD70/CD27 Signaling Promotes Expansion of Clonal Plasma Cells in Multiple Myeloma and Is a Promising Therapeutic Target in Advanced Disease

Introduction: Multiple Myeloma (MM) is a hematological malignancy characterized by the clonal expansion of malignant plasma cells within the bone marrow (BM). In the last decade, substantial advances in the treatment of MM patients have been made resulting in improved progression-free and overall survival rates. However, to date, MM remains incurable. CD70 and CD27 belong to the TNF receptor family. CD70 is the only known ligand for its receptor CD27. Physiologically, the CD70/CD27 interaction is involved in the regulation of immune cell expansion. However, numerous solid and hematological cancer types overexpress CD70. Aside, CD70 and CD27 co-expression has been described on leukemic stem cells (LSC) in acute myeloid leukemia patients where an autonomous signaling activation drives LSC self-renewal and expansion. While CD70/CD27 signaling plays a key role in the differentiation process of B lymphocytes into plasma cells, only little is known about its impact on myelomagenesis and MM progression. In this study, we analyzed the role of CD70/CD27 signaling in the pathogenesis of MM and studied the possibility to target CD70 by monoclonal antibodies to treat the disease.

Materials and Methods: We evaluated CD70 and CD27 gene expression levels and its correlations with overall survival (OS), MM progression and high-risk molecular alterations using different databases (CoMMpass IA15, GSE6477). CD70 and CD27 protein expression patterns were analyzed in a newly established MM tissue microarray (TMA) and a collection of matched BM samples from initial and advanced disease stages (in collaboration with the Institutes of Pathology in Bern, Basel and Tübingen). To assess the functional role of CD70/CD27 signaling on MM initiation and expansion, we generated CD70 knock-out MM cell lines and newly established patient-derived xenografts (PDX) using CRISPR/Cas9 genome editing. CD70-expressing MM cell lines or patient-derived myeloma cells were grown in NOD/SCID (NSG) mice. Moreover, different monoclonal antibodies (Abs) and Ab-derivates were tested including a CD70 blocking antibody (αCD70) and an antibody-dependent cell-mediated cytotoxicity (ADCC) optimized antibody (cusatuzumab, ArgenX) that activates FcγR-expressing effector cells to assess the therapeutic potential of targeting CD70 in MM.

Results: High CD70 gene expression levels correlate with increased MKI67 gene expression and elevated lactate dehydrogenase serum levels indicating a more rapid proliferation of CD70-expressing myeloma cells. In addition, high CD70 expression in MM patients correlated with shorter OS. Longitudinal analyses of CD70 gene expression in the precursor stages monoclonal gammopathy of undetermined significance and smoldering myeloma as well as MM and relapsed MM stages showed highest CD70 gene expression levels in relapsed MM patients. Assessment of the molecular background further revealed increased frequencies of the prognostically unfavorable genomic translocations t(4;14) and t(14;16), as well as gain/amplification of the chromosomal region 1q21 (1q+) in CD70 high expressing MM patients. In line with our previous findings, a TMA analysis of over 200 MM core biopsies as well as a longitudinal analysis of matched biopsies from patients at initial diagnosis and MM relapse indicated that CD70 was overexpressed in advanced disease. Functionally, we found that CD70/CD27 signaling promotes plasma cell survival inducing the expansion of MM cell lines and primary malignant plasma cells. CD70 blockade (αCD70) or CD70 knock-out (CD70 KO) resulted in a significant inhibition or complete abrogation of growth of a CD70-expressing MM patient-derived xenograft (PDX) (EMM01 CD70+) compared to controls (Figure 1). Interestingly, in vivo limiting dilution assays revealed that CD70+CD27+ double-positive myeloma subpopulations showed significantly higher cancer stem cell frequencies compared to CD70 negative fractions suggesting that CD70 expression might be related to an immature or stem-cell like phenotype. The ADCC-optimized CD70 specific antibody custatuzumab efficiently eliminated CD70-expressing MM cells in the presence of PBMC-derived natural killer (NK) cells in vitro and in a PDX model in vivo.

Conclusion: Our results indicate that CD70-expressing MM cells have a stem-cell like phenotype and propagate the disease. Targeting CD70 is a promising new therapy especially in advanced disease.