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

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.

Site-Specific Dolasynthen Antibody-Drug Conjugates Exhibit Consistent Pharmacokinetic Profiles across a Wide Range of Drug-to-Antibody Ratios.

Key defining attributes of an antibody-drug conjugate (ADC) include the choice of the targeting antibody, linker, payload, and the drug-to-antibody ratio (DAR). Historically, most ADC platforms have used the same DAR for all targets, regardless of target characteristics. However, recent studies and modeling suggest that the optimal DAR can depend on target expression level and intratumoral heterogeneity, target internalization and trafficking, and characteristics of the linker and payload. An ADC platform that enables DAR optimization could improve the success rate of clinical candidates. Here we report a systematic exploration of DAR across a wide range, by combining THIOMAB protein engineering technology with Dolasynthen, an auristatin-based platform with monomeric and trimeric variants. This approach enabled the generation of homogeneous, site-specific ADCs spanning a discrete range of DARs 2, 4, 6, 12, and 18 by conjugation of trastuzumab IgG1 THIOMAB constructs with 1, 2, or 3 engineered cysteines to monomeric or trimeric Dolasynthen. All ADCs had physicochemical properties that translated to excellent in vivo pharmacology. Following a single dose of ADCs in a HER2 xenograft model with moderate antigen expression, our data demonstrated comparable pharmacokinetics for the conjugates across all DARs and dose-dependent efficacy of all test articles. These results demonstrate that the Dolasynthen platform enables the generation of ADCs with a broad range of DAR values and with comparable physiochemical, pharmacologic, and pharmacokinetics profiles; thus, the Dolasynthen platform enables the empirical determination of the optimal DAR for a clinical candidate for a given target.

Biomarker Data from the Phase III KATHERINE Study of Adjuvant T-DM1 versus Trastuzumab for Residual Invasive Disease after Neoadjuvant Therapy for HER2-Positive Breast Cancer.

PURPOSE: In KATHERINE, adjuvant T-DM1 reduced risk of disease recurrence or death by 50% compared with trastuzumab in patients with residual invasive breast cancer after neoadjuvant therapy (NAT) comprised of HER2-targeted therapy and chemotherapy. This analysis aimed to identify biomarkers of response and differences in biomarker expression before and after NAT. EXPERIMENTAL DESIGN: Exploratory analyses investigated the relationship between invasive disease-free survival (IDFS) and HER2 protein expression/gene amplification, PIK3CA hotspot mutations, and gene expression of HER2, PD-L1, CD8, predefined immune signatures, and Prediction Analysis of Microarray 50 intrinsic molecular subtypes, classified by Absolute Intrinsic Molecular Subtyping. HER2 expression on paired pre- and post-NAT samples was examined. RESULTS: T-DM1 appeared to improve IDFS versus trastuzumab across most biomarker subgroups, except the HER2 focal expression subgroup. High versus low HER2 gene expression in residual disease was associated with worse outcomes with trastuzumab [HR, 2.02; 95% confidence interval (CI), 1.32-3.11], but IDFS with T-DM1 was independent of HER2 expression level (HR, 1.01; 95% CI, 0.56-1.83). Low PD-L1 gene expression in residual disease was associated with worse outcomes with trastuzumab (HR, 0.66; 95% CI, 0.44-1.00), but not T-DM1 (HR, 1.05; 95% CI, 0.59-1.87). PIK3CA mutations were not prognostic. Increased variability in HER2 expression was observed in post-NAT versus paired pre-NAT samples. CONCLUSIONS: T-DM1 appears to overcome HER2 resistance. T-DM1 benefit does not appear dependent on immune activation, but these results do not rule out an influence of the tumor immune microenvironment on the degree of response.

Tumor biomarkers and efficacy in patients treated with trastuzumab emtansine + pertuzumab versus standard of care in HER2-positive early breast cancer: an open-label, phase III study (KRISTINE).

BACKGROUND: KRISTINE is an open-label, phase III study of trastuzumab emtansine + pertuzumab (T-DM1 + P) versus docetaxel + carboplatin + trastuzumab + pertuzumab (TCH + P) in patients with HER2-positive, stage II-III breast cancer. We investigated the association of biomarkers with clinical outcomes in KRISTINE. METHODS: Patients were randomized to receive neoadjuvant T-DM1 + P or TCH + P and assessed for pathologic complete response (pCR; ypT0/is, ypN0). HER2 status (per central assessment), hormone receptor status, PIK3CA mutation status, HER2/HER3 mRNA levels, tumor-infiltrating lymphocyte levels, PD-L1 status, and NanoString data were analyzed. pCR rates by treatment arm were compared across biomarker subgroups. Analyses were descriptive. RESULTS: Biomarker analyses included data from all 444 patients (T-DM1 + P, n = 223; TCH + P, n = 221) enrolled in KRISTINE. Biomarker distribution was balanced across treatment arms. All subgroups with higher HER2 amplification/expression and immune marker levels showed numerically higher pCR rates in both arms. Mutated versus non-mutated PIK3CA tumors were associated with numerically lower pCR rates in the T-DM1 + P arm but not in the TCH + P arm. In a multivariate analysis, Prediction Analysis of Microarray with the 50-gene classifier (PAM50) HER2-enriched subtype, HER2 gene ratio ≥ 4, and PD-L1-positive status positively influenced the pCR rate. Biomarkers associated with lower pCR rates (e.g., low HER2 levels, positive hormone receptor status, mutated PIK3CA) were more likely to co-occur. Dynamic on-treatment biomarker changes were observed. Differences in the treatment effects for T-DM1 + P versus TCH + P were similar to those observed in the intent-to-treat population for the majority of the biomarker subgroups. CONCLUSIONS: Although our biomarker analysis did not identify a subgroup of patients that benefited from neoadjuvant T-DM1 + P versus TCH + P, the data revealed that patients with higher HER2 amplification/expression and immune marker levels had improved response irrespective of treatment arm. These analyses confirm the role of HER2 tumor biology and the immune microenvironment in influencing pCR in the neoadjuvant setting and reaffirm the molecular diversity of HER2-positive breast cancer. TRIAL REGISTRATION: ClinicalTrials.gov NCT02131064. Registered 06 May 2014.

LRRC15+ myofibroblasts dictate the stromal setpoint to suppress tumour immunity.

Recent single-cell studies of cancer in both mice and humans have identified the emergence of a myofibroblast population specifically marked by the highly restricted leucine-rich-repeat-containing protein 15 (LRRC15)1-3. However, the molecular signals that underlie the development of LRRC15+ cancer-associated fibroblasts (CAFs) and their direct impact on anti-tumour immunity are uncharacterized. Here in mouse models of pancreatic cancer, we provide in vivo genetic evidence that TGFß receptor type 2 signalling in healthy dermatopontin+ universal fibroblasts is essential for the development of cancer-associated LRRC15+ myofibroblasts. This axis also predominantly drives fibroblast lineage diversity in human cancers. Using newly developed Lrrc15-diphtheria toxin receptor knock-in mice to selectively deplete LRRC15+ CAFs, we show that depletion of this population markedly reduces the total tumour fibroblast content. Moreover, the CAF composition is recalibrated towards universal fibroblasts. This relieves direct suppression of tumour-infiltrating CD8+ T cells to enhance their effector function and augments tumour regression in response to anti-PDL1 immune checkpoint blockade. Collectively, these findings demonstrate that TGFß-dependent LRRC15+ CAFs dictate the tumour-fibroblast setpoint to promote tumour growth. These cells also directly suppress CD8+ T cell function and limit responsiveness to checkpoint blockade. Development of treatments that restore the homeostatic fibroblast setpoint by reducing the population of pro-disease LRRC15+ myofibroblasts may improve patient survival and response to immunotherapy.

Targeting ligand-activated ErbB2 signaling inhibits breast and prostate tumor growth.

ErbB2 is a ligand-less member of the ErbB receptor family that functions as a coreceptor with EGFR, ErbB3, and ErbB4. Here, we describe an approach to target ErbB2's role as a coreceptor using a monoclonal antibody, 2C4, which sterically hinders ErbB2's recruitment into ErbB ligand complexes. Inhibition of ligand-dependent ErbB2 signaling by 2C4 occurs in both low- and high-ErbB2-expressing systems. Since the ErbB3 receptor contains an inactive tyrosine kinase domain, 2C4 is very effective in blocking heregulin-mediated ErbB3-ErbB2 signaling. We demonstrate that the in vitro and in vivo growth of several breast and prostate tumor models is inhibited by 2C4 treatment.