Combination anti-CD74 (milatuzumab) and anti-CD20 (rituximab) monoclonal antibody therapy has in vitro and in vivo activity in mantle cell lymphoma

Abstract

Mantle cell lymphoma (MCL) is an aggressive B-cell malignancy with a median survival of 3 years despite chemoimmu-notherapy. Rituximab, a chimeric anti-CD20 monoclonal antibody (mAb), has shown only modest activity as single agent in MCL. The humanized mAb milatu-zumab targets CD74, an integral membrane protein linked with promotion of B-cell growth and survival, and has shown preclinical activity against B-cell malig-nancies. Because rituximab and milatu-zumab target distinct antigens and potentially signal through different pathways, we explored a preclinical combination strategy in MCL. Treatment of MCL cell lines and primary tumor cells with immobilized milatuzumab and rituximab resulted in rapid cell death, radical oxygen species generation, and loss of mitochon-drial membrane potential. Cytoskeletal distrupting agents significantly reduced formation of CD20/CD74 aggregates, cell adhesion, and cell death, highlighting the importance of actin microfilaments in rituximab/milatuzumab-mediated cell death. Cell death was independent of caspase activation, Bcl-2 family proteins or modulation of autophagy. Maximal inhibition of p65 nuclear translocation was observed with combination treatment, indicating disruption of the NF-B pathway. Significant in vivo therapeutic activity of combination rituximab and milatuzumab was demonstrated in a preclinical model of MCL. These data support clinical evaluation of combination milatuzumab and rituximab therapy in MCL. (Blood. 2011; 117(17):4530-4541) Introduction Mantle cell lymphoma (MCL) is a B-cell malignancy with a variable histology and clinical course, distinguished by the characteristic translocation t(11;14)(q13, q32) that results in overexpres-sion of cyclin D 1 and consequent dysregulation of cell-cycle control. 1 In addition, MCL exhibits alterations in cell survival pathways, including constitutive activation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling 2 and nuclear factor-B (NF-B). 3 Despite the hallmark genetic translocation in MCL, the clinical course of MCL is variable with some patients experiencing indolent disease, 4 whereas others exhibit rapid progression. 5 MCL patients have a median overall survival (OS) of approximately 3 years, and no consensus exists for standard first-line therapy. 6-9 Although aggressive therapies including chemoimmunotherapy 10,11 or stem cell transplantation 12,13 have been shown to improve outcomes, no therapy offers the potential for cure. Given the absence of curative therapy and the limited number of options for patients with relapsed/refractory MCL, novel treatment approaches are essential. Rituximab (Genentech), a chimeric anti-human CD20 monoclo-nal antibody (mAb), has been used in multiple strategies to treat patients with MCL. 14 As a single agent, rituximab has been tested in patients with newly diagnosed and relapsed/refractory MCL with response rates (RR) of 27% to 38% and a median response duration of 6 to 12 months. 15,16 Interestingly, the RR obtained in untreated patients was not higher than in relapsed...

Transcription factor ICBP90 regulates the MIF promoter and immune susceptibility locus

“…ICBP90 also may provide a functional link that will increase our understanding of MIF's role in the inflammatory pathogenesis of human tumors (52,53), especially with emerging data suggesting associations between high expression MIF alleles and tumor progression (14,54). Finally, given ongoing efforts to develop MIF-based therapies, which may be most beneficial in high MIF expressors, the present findings open the possibility of the pharmacological targeting of ICBP90 in immunological and oncologic disease (19,40,55 EMSAs. Nuclear extracts from human THP-1 monocytes were analyzed with the LightShift Chemiluminescent EMSA Kit (Thermo Scientific).…”

Dual-targeting immunotherapy of lymphoma: potent cytotoxicity of anti-CD20/CD74 bispecific antibodies in mantle cell and other lymphomas

“…Similar results have been observed with the combination of rituximab and milatuzumab in the presence of a crosslinking Ab in MCL lines. 17 Our studies also indicate that the induction of HA by the anti-CD20/CD74 HexAbs can be blocked without affecting the sustained activation of ERKs and JNK, and neither classic apoptosis nor autophagy is involved in the resulting cell death, which appeared to closely resemble the actin-and lysosome-dependent cell death evoked by tositumomab and hL243 in Raji and SU-DHL4. 49 The present data also indicate that 20-(74)-(74), but not 74- (20)- (20), is internalized into JeKo-1; 74- (20)- (20) However, this advantage is lost in the xenograft model, which may be because of ADCC and CDC activity.…”

“…We have previously noted 20 that one effective approach to converting a type I anti-CD20 mAb to a type II can be achieved by making the type I mAb multivalent, as shown by the HexAb generated from the type I veltuzumab, 20- (20)- (20), which exhibits biologic properties attributable to both type II (eg, weak CDC; negative for calcium mobilization; positive for antiproliferation, apoptosis, and HA) and type I (eg, positive for trafficking to lipid rafts). 20 The strategy to target both CD20 and CD74 with distinct mAbs was reported recently by us in a preclinical study that used a combination of milatuzumab and rituximab plus a crosslinking Ab in MCL lines and primary tumor cells, 17 which showed that the treatment resulted in rapid cell death, generation of ROS, loss of ⌬ m , strong HA, and inhibition of p65 nuclear translocation. The observed cell death was attributed to a nonclassical apoptotic mechanism, because it lacked evidence of autophagy and caspaseactivation but required the participation of actin and lysosomes.…”

“…16 More recently, the potential advantage of targeting both CD20 and CD74 has been reported by us in a preclinical study that involved rituximab and milatuzumab (humanized anti-CD74 IgG 1 , also referred to as hLL1), which in the presence of a crosslinking Ab showed improved antitumor activity in MCL lines and primary patient samples compared with either parental Ab alone. 17 The Dock-and-Lock (DNL) method is a platform technology that combines genetic engineering with site-specific conjugation to enable self-assembly of 2 modular components with each other, resulting in a covalent structure of defined composition with retained bioactivity. 18,19 We have applied DNL to generate various multivalent, multispecific structures that include monospecific and bispecific HexAbs, each comprising a pair of stabilized dimers of Fab linked to a full IgG at the carboxyl termini of the 2 heavy chains, thus conferring 6 Fab-arms and a common Fc entity.…”

MIFallele-dependent regulation of the MIF coreceptor CD44 and role in rheumatoid arthritis

“…of anti-MIF and -CD74 antibodies, with the objective that therapeutic intervention may be guided by an individual's MIF genotype (28)(29)(30).…”