Design and selection of optimal ErbB-targeting bispecific antibodies in pancreatic cancer

The ErbB family of receptor tyrosine kinases is a primary target for small molecules and antibodies for pancreatic cancer treatment. Nonetheless, the current treatments for this tumor are not optimal due to lack of efficacy, resistance, or toxicity. Here, using the novel BiXAb™ tetravalent format platform, we generated bispecific antibodies against EGFR, HER2, or HER3 by considering rational epitope combinations. We then screened these bispecific antibodies and compared them with the parental single antibodies and antibody pair combinations. The screen readouts included measuring binding to the cognate receptors (mono and bispecificity), intracellular phosphorylation signaling, cell proliferation, apoptosis and receptor expression, and also immune system engagement assays (antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity). Among the 30 BiXAbs™ tested, we selected 3Patri-1Cetu-Fc, 3Patri-1Matu-Fc and 3Patri-2Trastu-Fc as lead candidates. The in vivo testing of these three highly efficient bispecific antibodies against EGFR and HER2 or HER3 in pre-clinical mouse models of pancreatic cancer showed deep antibody penetration in these dense tumors and robust tumor growth reduction. Application of such semi-rational/semi-empirical approach, which includes various immunological assays to compare pre-selected antibodies and their combinations with bispecific antibodies, represents the first attempt to identify potent bispecific antibodies against ErbB family members in pancreatic cancer

Highly Specific and Effective Targeting of EGFRvIII-Positive Tumors with TandAb Antibodies

To harness the cytotoxic capacity of immune cells for the treatment of solid tumors, we developed tetravalent, bispecific tandem diabody (TandAb) antibodies that recognize EGFRvIII, the deletion variant III of the epidermal growth factor receptor (EGFR), and CD3 on T-cells, thereby directing immune cells to eliminate EGFRvIII-positive tumor cells. Using phage display, we identified scFv antibodies selectively binding to EGFRvIII. These highly EGFRvIII-specific, fully human scFv were substantially improved by affinity maturation, achieving KDs in the picomolar range, and were used to construct a set of bispecific EGFRvIII-targeting TandAbs with a broad range of binding and cytotoxic properties. These antibodies exhibited an exquisite specificity for a distinguished epitope in the N-terminal portion of EGFRvIII, as shown on recombinant antigen in Western Blot, SPR, and ELISA, as well as on antigen-expressing cells in FACS assays, and did not bind to the wild-type EGFR. High-affinity EGFRvIII/CD3 TandAbs were most potent in killing assays, displaying cytotoxicity toward EGFRvIII-expressing CHO, F98 glioma, or human DK-MG cells with EC50 values in the range of 1–10 pM in vitro. They also demonstrated dose-dependent growth control in vivo in an EGFRvIII-positive subcutaneous xenograft tumor model. Together with the tumor-exclusive expression of EGFRvIII, the EGFRvIII/CD3 TandAbs’ high specificity and strictly target-dependent activation with no off-target activity provide an opportunity to target tumor cells and spare normal tissues, thereby reducing the side effects associated with other anti-EGFR therapies. In summary, EGFRvIII/CD3 TandAbs are highly attractive therapeutic antibody candidates for selective immunotherapy of EGFRvIII-positive tumors

Initial Dose Escalation of ISB 1442, a Novel CD38 Biparatopic x CD47 Bispecific Antibody, in Patients with Relapsed / Refractory Multiple Myeloma (RRMM)

Introduction: ISB 1442 is a fully human bispecific, biparatopic antibody that targets CD38 and CD47, generated using Ichnos' Bispecific Engagement by Antibodies based on the T cell receptor (BEAT ®) platform. ISB 1442 is designed to kill CD38-expressing tumor cells through multiple mechanisms of action including blocking CD47-signal regulatory protein alpha (SIRPα) axis to increase several antibody effector functions: antibody-dependent cellular phagocytosis (ADCP), antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) through optimized architecture, affinity to targets, and Fc engineering. ISB 1442 has 2 Fab domains binding to distinct CD38 epitopes that do not compete functionally with daratumumab. ISB 1442 is expected to have optimized tolerability with low potential for adverse effects on red blood cells (RBC) such as hemagglutination, platelet aggregation and RBC depletion (Sammicheli at al., ASH 2021, Blood (2021) 138 (Supplement 1): 73). We report here initial findings from the early dose-escalation portion of an ongoing, multi-center, open-label, single-agent phase 1/2 study (NCT05427812) of ISB 1442 in patients with RRMM. Methods: Adult patients with relapsed refractory multiple myeloma (RRMM) to prior therapies, including a proteasome inhibitors (PIs), an immunomodulatory drugs (IMiDs), and an anti-CD38 antibodies received subcutaneous (SC) doses of ISB 1442 weekly (QW) in 28-day cycles. Patients had measurable disease per the International Myeloma Working Group (IMWG) criteria (2016). Dose escalation began at 6 mg dose with single patient accelerated titration phase for the first 3 cohorts, followed by a standard titration phase with a “3 + 3” design. The primary study objective for phase 1 is to assess the safety and tolerability to determine the maximum tolerated dose (MTD) and select the recommended phase 2 dose (RP2D) of ISB 1442. For phase 2 the primary study objective is to evaluate efficacy of ISB 1442. Secondary objectives include evaluation of pharmacokinetics (PK) and immunogenicity of ISB 1442. Exploratory objectives include assessment of minimal residual disease (MRD), assessment of cellular biomarkers in blood and bone marrow, and soluble factors in blood, and their correlation with efficacy, safety and other clinical endpoints of interest. Results: As of July 18, 2023, based on preliminary data from ongoing clinical database, 10 subjects had received once weekly SC injections of ISB 1442 in 4 dose-escalation groups from 6 mg to 150 mg. The majority were male (60%) and white (90%). The median age was 67 years (range 57-79). The median number of prior anti-myeloma lines of therapy was 6 (range 3-7); 70% were exposed to 5 drugs (2PIs, 2IMiDs, and CD38). The median number of ISB 1442 cycles was 1(range 1-2). Eight subjects (80%) experienced treatment-related adverse events (TRAEs), all were grade 1 or 2: cytokine release syndrome (CRS) (50%), injection site reactions (injection site erythema 20%, injection site bruising 10%), anemia (10%, 1 subject, grade 2) (Table 1). No grade 5 TRAE was observed. Following QW SC injection, ISB 1442 was slowly absorbed into the systemic circulation with T max generally occurring on day 2 of dosing. The ISB 1442 serum concentrations generally remained quantifiable over the entire dosing duration from 20 mg and above. The available PK data suggest an approximately dose-linear increase in serum concentration up to DL 3 (60 mg), followed by a supra-proportional increase in serum levels in subjects treated at DL 4 (150 mg). To date, 5 subjects treated at DL4 (150 mg) have experienced clinical symptoms of CRS (Grade 1-2) following the first dose of ISB 1442. Assessment of a panel of 63 soluble factors (including multiple cytokines, chemokines and growth factors) in the peripheral blood revealed that several subjects at DL3-4 exhibited transient increases (>10-fold) in macrophage inflammatory protein-1b (MIP-1b/CCL4) within 24h after treatment with ISB 1442, consistent with a macrophage-associated mechanism of action. Conclusions: Treatment with ISB 1442 was well tolerated at the dose levels evaluated. The observed clinical CRS events were moderate and potentially related to macrophage activation following ISB 1442 administration. Updated clinical, biomarker and PK data will be presented for this ongoing study