1041 - Anti-FcμR Antibody Drug Conjugate as a Potential Therapeutic Agent for the Treatment of Chronic Lymphocytic Leukemia

Historically treatment with naked monoclonal antibodies (mAbs) alone or in combination with standard chemotherapy (e.g. anti-CD52, anti-CD20 plus fludarabine/chlorambucil) have yielded significant clinical response in patients with chronic lymphocytic leukemia (CLL) compared to chemotherapy alone. Later developments include chimeric antigen receptor expressing T cells (CAR-T) targeting cell surface proteins (e.g. CD19, ROR1), small molecule inhibitors such as ibrutinib, idelalisib, venetoclax targeting intracellular proteins (e.g. BTK, PI3K𝜹, BCl-2) and combinations thereof have shown excellent clinical activity in patients. However, many patients do not enjoy long-term progression free survival (PFS > 5 years) and eradication of the disease is uncommon. Various factors contribute to this failure including disease relapse, treatment intolerance, adverse side effects, unfavorable cytogenetics and treatment-emergent mutations. In this report, we investigate antibody drug conjugates (ADCs) to specifically target CLL cells. An advantage of ADCs is their functional independence of host immune effector mechanisms which are often compromised in patients with CLL.

We and others have previously reported over expression of the Fc receptor for IgM (FcμR) in CLL B cells compared to other leukocytes and such over expression of FcμR is an independent indicator of shorter survival in patients with CLL. FcμR binds to human IgM Fc fragment (Fcμ) with high avidity and is rapidly shuttled via clathrin-coated endocytic vesicles into lysosomes. We have also previously demonstrated that IgM derived protein scaffold conjugated to an antimitotic drug could selectively induced cytotoxicity in CLL cells. Commercially available anti-FcμR mAbs failed to internalize. Therefore, we developed recombinant rabbit human chimeric mAbs against FcμR in double variable domain (DVD) formats. The DVD mAbs contain an outer VH domain that targets FcμR on the cell surface and an inner VH domain with an uniquely positioned catalytic (or reactive) lysine residue that allows quick, site-specific conjugation of a cytotoxic drug (e.g. MMAF). We first demonstrated that anti-FcμR-DVDs, both in Fab and IgG formats were able to bind to FcμR positive malignant B cell lines (e.g. Mino, NU-DHL-1) and not to FcμR negative cell lines (Jurkat and HBL-2). Similarly, anti-FcμR-DVDs were able to bind CLL cells, but not other cells present in PBMCs obtained from patients with CLL and in addition did not bind to B cells from healthy donors. Confocal microscopy demonstrated rapid internalization of cell surface bound anti-FcμR-DVDs upon incubation at 370C and their localization to lyzosomes. Anti-FcμR-DVDs conjugated to MMAF induced specific lysis (>80%) of FcμR+ cell lines, while a control ADC (H38C2-DVD-IgG-MMAF) failed to do so. The anit FcμR-DVD-IgG-ADC induced dose dependent cytotoxicity of FcuR+ cell lines (IC50 0.5 – 33nM) but not of their congeneic FcuR-knock out counterparts. In vitro, we observed that FcμR-DVD-IgG-ADC induced selective cytotoxicity of CLL B cells without affecting the viability of T cells and other mononuclear cells. Together these experiments demonstrated the specificity and potency of anti-FcuR-ADC in vitro, and studies are underway to evaluate its efficacy in vivo using a patient-derived xengograft CLL model. We envisage that anti-FcμR-DVD-ADCs may offer a promising modality for the treatment of CLL.