Simon Chivers

ADCT-402, a PBD dimer-containing antibody drug conjugate targeting CD19-expressing malignancies

Human CD19 antigen is a 95-kDa type I membrane glycoprotein in the immunoglobulin superfamily whose expression is limited to the various stages of B-cell development and differentiation and is maintained in the majority of B-cell malignancies, including leukemias and non-Hodgkin lymphomas of B-cell origin. Coupled with its differential and favorable expression profile, CD19 has rapid internalization kinetics and is not shed into the circulation, making it an ideal target for the development of antibody-drug conjugates (ADCs) to treat B-cell malignancies. ADCT-402 (loncastuximab tesirine) is a novel CD19-targeted ADC delivering SG3199, a highly cytotoxic DNA minor groove interstrand crosslinking pyrrolobenzodiazepine (PDB) dimer warhead. It showed potent and highly targeted in vitro cytotoxicity in CD19-expressing human cell lines. ADCT-402 was specifically bound, internalized, and trafficked to lysosomes in CD19-expressing cells and, following release of the PBD warhead, resulted in formation of DNA crosslinks that persisted for 36 hours. Bystander killing of CD19- cells by ADCT-402 was also observed. In vivo, single doses of ADCT-402 resulted in highly potent, dose-dependent antitumor activity in several subcutaneous and disseminated human tumor models with marked superiority to comparator ADCs delivering tubulin inhibitors. Dose-dependent DNA crosslinks and γ-H2AX DNA damage response were measured in tumors by 24 hours after single dose administration, whereas matched peripheral blood mononuclear cells showed no evidence of DNA damage. Pharmacokinetic analysis in rat and cynomolgus monkey showed excellent stability and tolerability of ADCT-402 in vivo. Together, these impressive data were used to support the clinical testing of this novel ADC in patients with CD19-expressing B-cell malignancies.

Abstract 51: Characterization of the mechanism of action, pharmacodynamics and preclinical safety of ADCT-402, a pyrrolobenzodiazepine (PBD) dimer-containing antibody-drug conjugate (ADC) targeting CD19-expressing hematological malignancies

ADCT-402, currently in Phase I clinical trials for B-cell hematological malignancies, is an ADC composed of a recombinant humanized IgG1 against human CD19, stochastically conjugated via a cleavable linker to a PBD dimer cytotoxin (DAR of 2.3). PBD dimers, DNA minor groove interstrand cross-linking agents, are gaining increasing attention and are currently being tested as the ADC warheads in several clinical trials. ADCT-402 has potent and targeted cytotoxicity against a panel of human lymphoma and leukemia cell lines in vitro. In vivo, ADCT-402 demonstrates dose-dependent antitumor activity against Burkitt’s lymphoma xenograft models. Moreover, ADCT-402 is markedly superior to maytansinoid- and auristatin-based CD19-targeting ADCs in the Ramos xenograft model. In a rat toxicology study, a single dose of ADCT-402 at 2 mg/kg is well tolerated with a favorable PK profile and excellent stability in vivo. The current study aimed to further define the mechanism of action (MOA) of ADCT-402 and validate its pharmacology and preclinical safety for clinical development. CD19 is a clinically validated target with restricted normal tissue expression and a widespread expression in the majority of B-cell malignancies. Importantly, we show here the consistent expression of CD19 in matched samples (initial diagnosis and relapsed/refractory) from panels of lymphoma patients, indicating that relapsed/refractory patients are appropriate for treatment with ADCT-402. ADCT-402 was shown to be efficiently internalized by CD19+ cells in vitro. Moreover, in line with the PBD dimer MOA, following a 2 hour exposure to ADCT-402, DNA interstrand cross-links reached a peak between 8 - 12 hours and persisted for up to 36 hours post-treatment. In contrast, the peak of cross-link formation for the PBD dimer warhead alone was observed immediately after 2 hour incubation, while a non-targeted PBD-ADC did not yield any appreciable DNA cross-links. In SCID mice s.c. implanted with Ramos cells, a single dose of ADCT-402 was administered at 0.33 or 1 mg/kg. Twenty-four hours after treatment, excised tumors showed a dose proportional increase in intensity of staining by an anti-PBD payload antibody, as well as in DNA cross-linking and in γ-H2AX formation. In contrast, no DNA cross-linking was observed in matched lymphocyte samples. The toxicity of ADCT-402 was further evaluated in a repeat dose cynomolgus monkey study. ADCT-402 was clinically well tolerated with an acceptable off-target safety profile. The PK of the ADC was consistent with normal antibody clearance with a half-life of about 12 to 17 days. These data confirm the MOA of ADCT-402 and provide relevant pharmacodynamic assays and preclinical safety assessment to guide the clinical development of this promising ADC in B-cell malignancies. Citation Format: Francesca Zammarchi, Simon Corbett, Karin Havenith, Narinder Janghra, Konstantinos Kiakos, Teresa Marafioti, David G. Williams, Simon Chivers, Phil W. Howard, John A. Hartley, Patrick H. van Berkel. Characterization of the mechanism of action, pharmacodynamics and preclinical safety of ADCT-402, a pyrrolobenzodiazepine (PBD) dimer-containing antibody-drug conjugate (ADC) targeting CD19-expressing hematological malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 51. doi:10.1158/1538-7445.AM2017-51

Pre-clinical pharmacology and mechanism of action of SG3199, the pyrrolobenzodiazepine (PBD) dimer warhead component of antibody-drug conjugate (ADC) payload tesirine

Synthetic pyrrolobenzodiazepine (PBD) dimers, where two PBD monomers are linked through their aromatic A-ring phenolic C8-positions via a flexible propyldioxy tether, are highly efficient DNA minor groove cross-linking agents with potent cytotoxicity. PBD dimer SG3199 is the released warhead component of the antibody-drug conjugate (ADC) payload tesirine (SG3249), currently being evaluated in several ADC clinical trials. SG3199 was potently cytotoxic against a panel of human solid tumour and haematological cancer cell lines with a mean GI50 of 151.5 pM. Cells defective in DNA repair protein ERCC1 or homologous recombination repair showed increased sensitivity to SG3199 and the drug was only moderately susceptible to multidrug resistance mechanisms. SG3199 was highly efficient at producing DNA interstrand cross-links in naked linear plasmid DNA and dose-dependent cross-linking was observed in cells. Cross-links formed rapidly in cells and persisted over 36u2009hours. Following intravenous (iv) administration to rats SG3199 showed a very rapid clearance with a half life as short as 8u2009minutes. These combined properties of cytotoxic potency, rapid formation and persistence of DNA interstrand cross-links and very short half-life contribute to the emerging success of SG3199 as a warhead in clinical stage ADCs.

Antitumor Activity of MEDI3726 (ADCT-401), a Pyrrolobenzodiazepine Antibody–Drug Conjugate Targeting PSMA, in Preclinical Models of Prostate Cancer

Prostate-specific membrane antigen (PSMA) is a membrane-bound glutamate carboxypeptidase that is highly expressed in nearly all prostate cancers with the highest expression in metastatic castration-resistant prostate cancer (mCRPC). The prevalence of increased surface expression and constitutive internalization of PSMA make it an attractive target for an antibody–drug conjugate (ADC) approach to treating patients with mCRPC. MEDI3726 (previously known as ADCT-401) is an ADC consisting of an engineered version of the anti-PSMA antibody J591 site specifically conjugated to the pyrrolobenzodiazepine (PBD) dimer tesirine. MEDI3726 specifically binds the extracellular domain of PSMA and, once internalized, releases the PBD dimer to crosslink DNA and trigger cell death. In vitro, MEDI3726 demonstrated potent and specific cytotoxicity in a panel of PSMA-positive prostate cancer cell lines, consistent with internalization and DNA interstrand crosslinking. In vivo, MEDI3726 showed robust antitumor activity against the LNCaP and the castration-resistant CWR22Rv1 prostate cancer cell line xenografts. MEDI3726 also demonstrated durable antitumor activity in the PSMA-positive human prostate cancer patient–derived xenograft (PDX) LuCaP models. This activity correlated with increased phosphorylated Histone H2AX in tumor xenografts treated with MEDI3726. MEDI3726 is being evaluated in a phase I clinical trial as a treatment for patients with metastatic castrate-resistant prostate cancer (NCT02991911). Mol Cancer Ther; 17(10); 2176–86. ©2018 AACR.

Abstract A004: MEDI3726 (ADCT-401), a novel antibody-drug conjugate targeting PSMA, has potent in vivo antitumor activity in prostate cancer patient-derived xenograft models

Prostate specific membrane antigen (PSMA), a type II membrane glycoprotein, is highly expressed in nearly all prostate cancers, with the highest expression in metastatic castration-resistant prostate cancer (mCRPC). The prevalence, increased surface expression in prostate tumors, and constitutive internalization make PSMA an attractive target for an antibody-drug conjugate (ADC) approach in anticancer treatment of patients with mCRPC. MEDI3726 (previously known as ADCT-401) is an ADC consisting of an engineered version of anti-PSMA antibody J591, site-specifically conjugated with DNA cross-linking pyrrolobenzodiazepine (PBD) dimer SG3249 for targeting prostatic cancer cells. Using prostate cancer cell line models, we have previously shown that MEDI3726 specifically binds to the extracellular domain of PSMA and, once internalized, releases the PBD dimer to cross link DNA and achieve potent in vitro and in vivo cytotoxicity. Here we investigated the in vivo activity of MEDI3726 in a series of LuCaP prostate cancer patient-derived xenograft (PDX) models. The selected LuCaP models had varying PSMA expression and heterogeneous genetic and phenotypic backgrounds. In agreement with the earlier cell line xenograft data, dose-dependent antitumor activity was observed in PSMA-positive PDX models with durable tumor regressions in models with high PSMA expression. In the PSMA-negative LuCaP 35CR PDX model, MEDI3726 did not have significant antitumor activity, thus highlighting target-mediated in vivo activity. Increased phosphorylation of histone H2AX was observed in xenografts dosed with MEDI3726, confirming DNA damage induced by interstrand cross-linking PBD dimer as the mechanism of antitumor activity of MEDI3726. In summary, MEDI3726 demonstrated potent and specific in vivo antitumor activity, concurrent with DNA damage, in clinically relevant prostate cancer PDX models. MEDI3726 is being evaluated in phase 1 clinical trial as an anticancer treatment in patients with metastatic castrate-resistant prostate cancer (NCT02991911). Citation Format: Song Cho, Francesca Zammarchi, Noel R. Monks, Kapil Vashisht, Ravinder Tammali, Kevin Schifferli, Patrick Strout, Wanda King, Karma Dacosta, Ryan Fleming, David G. Williams, Karin Havenith, Mary Jane Masson Hinrichs, Simon Chivers, Nazzareno Dimasi, Phil W. Howard, John A. Hartley, Steve Coats, Ronald Herbst, Patrick H. van Berkel, David A. Tice. MEDI3726 (ADCT-401), a novel antibody-drug conjugate targeting PSMA, has potent in vivo antitumor activity in prostate cancer patient-derived xenograft models [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A004.

Abstract 2792A: Preclinical activity of ADCT-601, a novel pyrrolobenzodiazepine (PBD) dimer-based antibody-drug conjugate (ADC) targeting AXL-expressing tumors

AXL, a member of the tyrosine kinase receptor family TAM, is a transmembrane receptor overexpressed in many solid (e.g. lung, breast, pancreas, glioma and esophageal) and hematological malignancies (acute myeloid and chronic lymphocytic leukemia) and its overexpression is maintained in both primary tumors and metastasis. Moreover, expression and activation of AXL is associated with poor clinical prognosis and several studies suggest that overexpression of AXL results in resistance to both conventional chemotherapy and targeted therapies. All these features make AXL an attractive target for the development of an ADC to treat AXL-expressing cancers. ADCT-601 is an ADC composed of a humanized IgG1 antibody against human AXL, site-specifically conjugated using GlycoconnectTM technology to PL1601, which contains a valine-alanine cleavable linker and the PBD dimer cytotoxin SG3199. The purpose of this study was to characterize the in vitro and in vivo anti-tumor activity of ADCT-601 in human cancer cell lines and xenograft models and to determine its safety and tolerability in the rat. In vitro, ADCT-601 demonstrated potent cytotoxicity in a panel of cell lines of different origin and levels of AXL, while its potency was strongly reduced in AXL-negative cell lines. In vivo, ADCT-601 showed potent anti-tumor activity in the human triple-negative breast cancer-derived MDA-MB-231 xenograft, expressing moderate levels of AXL (≈36,000 copies/cell). In this model, a single dose of ADCT-601 at 1 mg/kg resulted in strong and sustained anti-tumor activity and resulted in 5/10 partial responders (PR) and 4/10 tumor-free survivors (TFS) at the end of the study on day 60. In the SN12C model, a human renal cell carcinoma-derived xenograft with high level of AXL expression (≈88,000 copies/cell), single doses of ADCT-601 at 0.3, 0.6 or 1 mg/kg showed dose-dependent anti-tumor activity compared to the vehicle- and isotype control ADC-treated mice. At the highest dose, ADCT-601 resulted in 7/8 and 6/8 complete responder (CR) and TFS, respectively, at the end of the study on day 60. Moreover, ADCT-601 showed potent and durable anti-tumor activity in a pancreatic cancer patient-derived xenograft model expressing heterogeneous levels of AXL where each single dose of ADCT-601 tested (0.3, 0.6 and 1 mg/kg) resulted in complete eradication of the tumors at the end of the study on day 43. Conversely, none of the mice in the vehicle and isotype control ADC groups had any PR, CR or TFS. ADCT-601 was stable, well tolerated and showed a favorable PK profile in the rat with a half-life of 9 days and a MTD of 6 mg/kg. In conclusion, ADCT-601 demonstrated potent and specific in vitro and in vivo anti-tumor activity in various cancer-derived models with different levels of membranous AXL. ADCT-601 was stable and well tolerated in the rat, warranting further development of this ADC into the clinic. Citation Format: Francesca Zammarchi, Karin Havenith, Simon Chivers, Paul W. Hogg, Charlie Britten, Sandamali Dissanayake, Peter Tyrer, Francois Bertelli, Ian Hutchinson, Luke Masterson, Phil Howard, John A. Hartley, Patrick H. van Berkel. Preclinical activity of ADCT-601, a novel pyrrolobenzodiazepine (PBD) dimer-based antibody-drug conjugate (ADC) targeting AXL-expressing tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2792A.

Abstract 3111A: ADCT-401/MEDI3726, a novel pyrrolobenzodiazepine (PBD)-based antibody-drug conjugate (ADC) targeting PSMA-expressing prostate cancers

Prostate-specific membrane antigen (PSMA) is an attractive target for an ADC approach as it is over-expressed by virtually all prostate cancers and its expression is highest in poorly-differentiated, metastatic and castration-resistant cases. PSMA has limited expression in non-prostatic tissues, it is not secreted or cleaved by PSMA-expressing cells, and it is constitutively internalized, a process that may be accelerated by specific antibody binding. ADCT-401/MEDI3726 is an ADC composed of a monoclonal antibody (J591), directed against human PSMA, site-specifically conjugated (drug-to-antibody ratio of 1.8) to a highly cytotoxic DNA cross-linking PBD dimer with a valine-alanine dipeptide linker. In vitro, ADCT-401/MEDI3726 demonstrated potent and specific cytotoxicity in a panel of PSMA-expressing prostate cancer cell lines, whereas its activity was greatly reduced in PSMA-negative cell lines. In vivo, ADCT-401/MEDI3726 showed strong antitumor activity against CWR22Rv1 and LNCaP human-derived prostate cancer xenograft models. In the CWR22Rv1 model, a tumor with low, heterogeneous PSMA expression, ADCT-401/MEDI3726 achieved dose-dependent antitumor activity when administered as single dose at either 0.33 or 1 mg/kg, which resulted in significant increase in survival compared to the vehicle-treated animals. Moreover, a single dose of ADCT-401/MEDI3726 showed remarkable superior antitumor activity compared to multiple doses of another PSMA-targeted ADC stochastically conjugated to the auristatin payload vcMMAE with a DAR of ~4. In the LNCaP model, ADCT-401/MEDI3726 resulted in dose-dependent antitumor activity when dosed once at 0.11, 0.33 or 1 mg/kg. In the PSMA-negative prostate cancer-derived PC3 xenograft model, ADCT-401/MEDI3726 and an isotope-control ADC showed limited tumor growth inhibition highlighting target-mediated antitumor activity. ADCT-401/MEDI3726 demonstrated potent and specific in vitro and in vivo antitumor activity in prostate cancer-derived models of differing levels of PSMA and this warrants further development of this ADC into the clinic. Citation Format: Francesca Zammarchi, Simon Chivers, Karin Havenith, David G. Williams, Lauren Adams, Maria Mellinas-Gomez, Simon Corbett, Peter Tyrer, Francois D9Hooge, Song Cho, Nazzareno Dimasi, Mary Jane Hinrichs, Phil W. Howard, John A. Hartley, Patrick H. van Berkel. ADCT-401/MEDI3726, a novel pyrrolobenzodiazepine (PBD)-based antibody-drug conjugate (ADC) targeting PSMA-expressing prostate cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3111A. doi:10.1158/1538-7445.AM2017-3111A