The HER2-directed antibody-drug conjugate DHES0815A in advanced and/or metastatic breast cancer: preclinical characterization and phase 1 trial results

Abstract

Approved antibody-drug conjugates (ADCs) for HER2-positive breast cancer include trastuzumab emtansine and trastuzumab deruxtecan. To develop a differentiated HER2 ADC, we chose an antibody that does not compete with trastuzumab or pertuzumab for binding, conjugated to a reduced potency PBD (pyrrolobenzodiazepine) dimer payload. PBDs are potent cytotoxic agents that alkylate and cross-link DNA. In our study, the PBD dimer is modified to alkylate, but not cross-link DNA. This HER2 ADC, DHES0815A, demonstrates in vivo efficacy in models of HER2-positive and HER2-low cancers and is well-tolerated in cynomolgus monkey safety studies. Mechanisms of action include induction of DNA damage and apoptosis, activity in non-dividing cells, and bystander activity. A dose-escalation study (ClinicalTrials.gov: NCT03451162) in patients with HER2-positive metastatic breast cancer, with the primary objective of evaluating the safety and tolerability of DHES0815A and secondary objectives of characterizing the pharmacokinetics, objective response rate, duration of response, and formation of anti-DHES0815A antibodies, is reported herein. Despite early signs of anti-tumor activity, patients at higher doses develop persistent, non-resolvable dermal, ocular, and pulmonary toxicities, which led to early termination of the phase 1 trial.

Fig. 1. Design and characterization of PBD derivatives.

a Structures of PBD, PBD-monoamine and PBD-monoamide, and summary table for DNA binding and drug activity (free drugs and ADCs in vitro and in vivo). In the left panel, circles denote reactive imine moiety and changes thereof to generate monoamine and monoamide derivatives. For the summary table: large cell line screens were performed to assess potency of the PBD dimer, PBD-monoamine and -monoamide. The PBD dimer was tested in 7 different cell line screens; the PBD-monoamide in 3 screens; and the PBD-monoamine in 2 screens (n = 3 wells per treatment group for all). The number of cell lines tested per screen ranged from 50-643 for the PBD; 73-146 for the PBD-monoamide and 72-147 for the PBD-monoamine. Data shown are the pooled mean IC₅₀ values ± standard error from the respective screens. ADC IC₅₀ was determined in SK-BR-3 cells (data are mean IC₅₀ for 3 independent experiments, n = 4 wells per treatment group for each; see Fig. 1c for graph). DNA alkylation percent is derived from data shown in part 1b. For determining MED, multiple xenograft models were used, with n = 5 to n = 10 mice per group. MED = minimum efficacious dose, defined as a single injection dose that results in tumor stasis at day 21 in mouse xenograft models. b DNA binding of PBD dimer, PBD-monoamine and PBD-monoamide as assessed by alkylation of double-stranded DNA oligonucleotides (each point represents oligonucleotide reactions, from 2 separate experiments). c Activity of DHES0815A compared to conjugated PBD dimer or PBD-monoamine (same antibody and linker) in SK-BR-3 cells in vitro and in MMTV-HER2 Fo5 model in vivo (PBD and PBD-monoamide conjugates only, n = 8 mice per group). d Structure of DHES0815A, comprised of 7C2 THIOMAB (MHES0448A), methyl-disulfide linker and PBD-monoamide. Source data are provided as a Source Data file.

Fig. 2. In vitro characterization of DHES0815A.

a DHES0815A activity is HER2-dependent and is differentiated from T-DM1 (n = 4 individual wells per treatment group, data points are mean ± standard error of the mean pooled from 3 independent experiments). b DHES0815A induces ADCC in HER2+ SK-BR-3 and KPL-4 cells in the presence of PBMCs (n = 3 individual wells per treatment group; data points are mean of individual replicates $\pm$ standard error of the mean from 2 independent experiments). c DHES0815A mediates bystander activity, as indicated by killing of HER2- MCF7/NucLight Red in the presence of HER2+ SK-BR-3/H2B-GFP and ADCs with reducible linkers (T-SPP-DM1, DHES0815A). Data points are from 3 separate experiments. Source data are provided as a Source Data file.

Fig. 3. DHES0815A induces apoptosis and markers of DNA damage in SK-BR-3 cells.

a Kinetics of caspase 3 activation for DHES0815A compared to T-DM1, MHES0448A and trastuzumab (n = 6 individual wells for each treatment group, the study was repeated 4 times). b Time-dependent PARP cleavage after treatment with antibodies (trastuzumab, MHES0488 A) or ADCs (T-DM1 or DHES0815A). c Induction of DNA damage markers (phospho-H2AX, phospho-/total p53 and phospho-CHK2) with free PBD-monoamide and DHES0815A. d Time-dependent induction of DNA damage and mitotic (phospho-histone H3) markers after treatment with unconjugated antibodies and ADCs. Western blot studies in (b)–(d), were performed 2 times with the same results. Source data are provided in Source Data file.

Fig. 4. In vivo efficacy of DHES0815A in HER2+ breast and gastric cancer models, HER2-low breast PDX models, and combinability with standard-of-care therapies.

a Dose-dependent tumor growth inhibition after single dose DHES0815A treatment in HER2+ MMTV-HER2 Fo5 (n = 8 per group) and HCC1569 X2 (n = 5 mice per group), and comparison to T-DM1 in the WHIM 8 PDX model (n = 10 per group). b Efficacy of DHES0815A compared to T-DM1 in HER2+ STO410 and STO41 gastric cancer PDX models (n = 10 mice per group). c Efficacy of DHES0815A vs. T-DM1 in HER2-low (IHC 1+ or 2+ /ISH-) breast cancer PDX models BC207, BC197, BC128 and BC085 (n = 10 mice per group for all studies). d Enhanced efficacy of DHES0815A combined with T-DM1 or docetaxel compared to single agent activity in MMTV-Fo5 (n = 5 mice per group for T-DM1 combination; n = 7 mice per group for docetaxel combination) and HCC1569 X2 (n = 8 mice per group for T-DM1 combination; n = 8 mice per group for docetaxel combination) models. Source data are provided as a Source Data file.

Fig. 5. Comparison of free drug and ADC activity for DHES0815A and trastuzumab deruxtecan (T-DXd).

a In vitro potency of different ADC payloads (DM1, PBD dimer, PBD-monoamide, DXd and SN38) in KPL-4 and SK-BR-3 cells (n = 4 individual wells per data point; data are represented as mean ± standard error from 3 independent experiments). b In vivo efficacy of DHES0815A compared to T-DXd in the WHIM 8 HER2+ PDX model (n = 6 mice per group). Source data are provided as a Source Data file.

Fig. 6. Clinical activity and pharmacokinetics of DHES0815A.

a Waterfall plot for phase 1 dose-escalation, including patient histories. Doses shown are the highest administered (initially assigned dose for some patients, highest dose following intrapatient escalation for other patients). b Pharmacokinetic analysis from dose-escalation for total antibody, antibody-conjugated PBD-monoamide (acPBD-ma) and unconjugated PBD-monoamide. *PD due to new lesions. Source data are provided as a Source Data file.