DFMT is a new paradigm in nanomedicine for the treatment of hematological B cell malignancies [1,2]. Apoptosis is induced by receptors crosslinking without need for low molecular weight drug. DFMT consists of two nanoconjugates: a) the bispecific engager: an antibody or antibody Fab’ fragment conjugated with a 25-base morpholino oligonucleotide MORF1; and b) the crosslinking effector: human serum albumin (HSA) or N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer grafted with multiple copies of the complementary oligonucleotide MORF2. MORF1-MORF2 hybridization results in crosslinking of surface receptors (e.g., CD20, CD38), calcium influx, mitochondrial depolarization and caspase activation, leading to apoptosis [3]. Fascinatingly, hyper-crosslinking of CD20-bound type II antibodies creates a new therapeutic that combines Type I & II mechanisms of action producing greater antitumor activity [4].Moreover, CD20 and CD38 hetero-receptor crosslinking induces synergistic levels of apoptosis in CD20+/CD38+ Raji cells and significantly increases animal survival compared to single-target DFMT treated mice (Fig. 1) [5]. Enhanced efficacy of dual-target DFMT was also attained in the treatment of samples isolated from patients diagnosed with chronic lymphocytic leukemia [6]. Interestingly, DFMT is efficient also in the treatment of autoimmune disease in animal model [7].
Figure 1. Dual-target (CD20 + CD38) DFMT efficacy. (A) Mechanism of action. (B) In vitro apoptosis induction in Raji cells by single-target and dual-target DFMT [9]. (C) Efficacy of triple-dose single-target and dual-target DFMT on xenograft Non-Hodgkin Lymphoma in mice [5].
MATCH: Inspired by the demonstrated ability of complementary MORF1 andMORF2 hybridization to bring two biomolecules into close proximity both in vitro and in vivo, we designed MATCH. We hypothesized that MORF hybridization could be harnessed to recruit and activate T cells against malignant B cells. In MATCH, a mini-library of cancer cell-targeting motifs, comprising antibody fragment Fab’-MORF1 conjugates (e.g., Fab’BCMA-MORF1, Fab’CD38-MORF1, Fab’GPRC5D-MORF1, Fab’CD20-MORF1, Fab’CD19-MORF1), selectively pairs with a T cell-engaging Fab’ conjugated to the complementary MORF2 (Fab’CD3-MORF2). The rapid, high-fidelity heterodimerization of these complementary nucleotide strands enables self-assembly of conjugates that function like bispecific antibodies. However, MATCH’s two-component “split-antibody” design offers a modular, customizable platform for designing bi- and multi-specific T cell recruitment therapies for lymphoma, leukemia, and multiple myeloma [8-10]. Its key advantages include: (a) cell-specific targeting through interchangeable cancer cell-targeting motifs to address MM heterogeneity; (b) reduced cytokine release and T cell exhaustion; and (c) personalized therapeutic strategies (Fig. 2).
Figure 2. Multi-Antigen T Cell Hybridizers provide cancer cell specific T cell activation by utilizing a two-component system of complementary Fab’ motifs: i) Fab’CD3-MORF2 for T cell engagement, and ii) various Fab’B cell-MORF1 for cancer cell engagement. The two Fab’s then recognize each other to dimerize and complete the bispecific construct [8].
An example of in vivo efficacy of MATCH in human non-Hodgkin lymphoma (NHL) xenografts (at E:T of 5:1) in mice is shown in Fig. 3. Raji cells (4×105) and healthy human T cells were i.v. inoculated into C.B-17 SCID mice. The mice were treated with Fab’CD20-MORF1 dose of 60 µg followed by a T cell engager (Fab’CD3-MORF2) with varying doses (60, 20, 6, or 2 µg). One group of mice received 60 µg of premixed Fab’CD20-MORF1/ Fab’CD3-MORF2 (1:1). Blinatumomab and saline were used as control. Notably, consecutive administration of nanoconjugates was more effective than the premixed one, suggesting the importance of the two-step administration of MATCH. Interestingly, the best therapeutic outcome was achieved when the T-cell engager dose of ~10-fold lower than the B-cell engaging dose was used [9]. Blinatumomab was ineffective in this pilot in vivo study, which is likely due to its short half-life and accelerated clearance.
Figure 3. In vivo efficacy of CD20-MATCH against a human NHL xenograft. A co-culture of luciferase-expressing Raji cells (4×105) and healthy donor, naïve T-cells (2×106) were inoculated via tail vein into female C.B-17 SCID mice. Cells were allowed to disseminate for 1 h before dosing began [9].
References
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