Durable and Specific Inhibition of ROR1 Signaling Associates with Prolonged Progression Free Survival in Patients with Chronic Lymphocytic Leukemia Treated with Cirmtuzumab
Authors: Choi Michael , Widhopf II George F. , Yu Jian , Chen Liguang , Hasan Md. Kamrul , Rassenti Laura Z. , Pittman Emily , Messer Karen , Gutierrez Charlene , Kidwell Reilly Lilianna , Prussak Charles E. , Castro Januario E , Jamieson Catriona , Kipps Thomas J. 
Published: April 2021

Abstract
Small-molecule drugs that inhibit survival signals in leukemia cells have demonstrated high efficacy in the treatment of patients (pts) with chronic lymphocytic leukemia (CLL). Although not directly cytotoxic for leukemia cells, these drugs block key signaling pathways that govern leukemia-cell trafficking, proliferation, and survival. However, non-specific inhibition of enzymes involved in other signaling pathways in non-neoplastic cells may result in toxicity, which can limit the therapeutic index of such drugs, at least in some settings.
Monoclonal antibodies (mAbs) also may be used as antagonists of signaling by interfering with receptor-ligand interactions or reducing the availability of cell-surface receptors. Unlike small-molecule inhibitors, mAbs may function as highly specific inhibitors of their target receptors and have superior pharmacokinetics (PK) due to a longer half-life. Based on this rationale, we developed cirmtuzumab, a humanized mAb specific for ROR1. ROR1 is an oncoembronic cell-surface receptor for Wnt5a, which induces non-canonical Wnt signaling, leading to Rho-GTPase activation and phosphorylation of hematopoietic lineage cell-specific protein 1 (HS1), and enhances leukemia-cell migration, survival, and proliferation. ROR1 is strongly expressed on CLL, but not on virtually all normal postpartum tissues. As such, ROR1 appears to be a selective target for anti-cancer therapy.
We conducted a phase 1, dose-escalation trial in pts with relapsed/refractory CLL. Eligible pts had progressive disease with an indication for therapy, as per iwCLL guidelines. Pts received four biweekly infusions of cirmtuzumab at doses ranging from 0.015 to 20mg/kg. We performed PK studies on plasma samples using an ELISA assay to assess for cirmtuzumab binding to immobilized recombinant ROR1. Pharmacodynamic (PD) studies included analyses of leukemia-cell activation of Rho-GTPases and phosphorylation/activation of HS1.
Twenty-five pts enrolled and received cirmtuzumab. Cirmtuzumab was safe and well-tolerated. There were no drug-related SAEs, DLTs, or infusion-related reactions. At higher doses, PK studies showed the half-life of cirmtuzumab at higher doses was 32.4 days (SD 1.9 days) and cirmtuzumab levels remained detectable in the plasma until approximately 3 months following the final infusion. No patient developed neutralizing autoantibodies, and no circulating ROR1 antigen was detected.
Pharmacodynamic studies confirmed sustained target inhibition. At doses of 2 mg/kg or higher, we detected inhibition of leukemia-cell Rho-GTPase activation within 24 hours of the initial cirmtuzumab dose. Inhibition of leukemia-cell activation of Rho-GTPases was sustained for weeks after the final infusion until the plasma cirmtuzumab levels became negligible (&lt;200 ng/mL). Similar findings were observed with phosphorylated HS-1.
Clinical effects were in line with those of the PD and PK results. Despite limited treatment with just four doses of antibody, there were encouraging signs of clinical activity, including treatment-related reduction in circulating lymphocyte counts, lymph node size, and/or leukemia-cell infiltration of the marrow. Importantly, nearly all patients treated at dose levels ≥2 mg/kg had arrested disease progression, despite having progressive disease upon study entry. Moreover, 16 of 19 evaluable patients had stable disease at the time of response assessment (2 months following the final infusion per iwCLL guidelines). Patients did not require subsequent therapy for prolonged periods. The median time to requiring next treatment due to progressive disease was 259 days (95% CI 168-265 days). This also correlated with the time at which cirmtuzumab levels and PD effects became undetectable.
Specific inhibition of ROR1 signaling by cirmtuzumab inhibits leukemia-cell RhoGTPase activation and HS-1 phosphorylation, and arrests disease progression without significant toxicity. This represents a novel approach to inhibiting growth/survival signaling in CLL, providing an advantageous PK and specific PD relative to that of small-molecule inhibitors, potentially ushering in a new era of immunopharmacology.

Disclosures
Choi: Gilead, Genentech, Abbvie, and PCYC: Speakers Bureau; AbbVie, Genentech, and PCYC: Consultancy; AbbVie: Other: Institutional research funding. Kipps: Celgene: Consultancy, Honoraria, Research Funding; Gilead: Consultancy, Honoraria, Speakers Bureau; Oncternal: Research Funding; Pharmacyclics, LLC, an AbbVie Company: Consultancy, Honoraria; Roche: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria, Research Funding.