Antitumor activity of AZD0754, a dnTGFβRII-armored, STEAP2-targeted CAR-T cell therapy, in prostate cancer
Peter Zanvit, Dewald van Dyk, Christine Fazenbaker, Kelly McGlinchey, Weichuan Luo, Jessica M. Pezold, John Meekin, Chien-ying Chang, Rosa A. Carrasco, Shannon Breen, Crystal Sao-Fong Cheung, Ariel Endlich-Frazier, Benjamin Clark, Nina J. Chu, Alessio Vantellini, Philip L. Martin, Clare E. Hoover, Kenesha Riley, Steve M. Sweet, David Chain, Yeoun Jin Kim, Eric Tu, Nathalie Harder, Sandrina Phipps, Melissa Damschroder, Ryan N. Gilbreth, Mark Cobbold, Gordon Moody, Emily E. Bosco
Peter Zanvit, Dewald van Dyk, Christine Fazenbaker, Kelly McGlinchey, Weichuan Luo, Jessica M. Pezold, John Meekin, Chien-ying Chang, Rosa A. Carrasco, Shannon Breen, Crystal Sao-Fong Cheung, Ariel Endlich-Frazier, Benjamin Clark, Nina J. Chu, Alessio Vantellini, Philip L. Martin, Clare E. Hoover, Kenesha Riley, Steve M. Sweet, David Chain, Yeoun Jin Kim, Eric Tu, Nathalie Harder, Sandrina Phipps, Melissa Damschroder, Ryan N. Gilbreth, Mark Cobbold, Gordon Moody, Emily E. Bosco
View: Text | PDF
Research Article Oncology

Antitumor activity of AZD0754, a dnTGFβRII-armored, STEAP2-targeted CAR-T cell therapy, in prostate cancer

Abstract

Prostate cancer is generally considered an immunologically “cold” tumor type that is insensitive to immunotherapy. Targeting surface antigens on tumors through cellular therapy can induce a potent antitumor immune response to “heat up” the tumor microenvironment. However, many antigens expressed on prostate tumor cells are also found on normal tissues, potentially causing on-target, off-tumor toxicities and a suboptimal therapeutic index. Our studies revealed that six-transmembrane epithelial antigen of prostate-2 (STEAP2) was a prevalent prostate cancer antigen that displayed high, homogeneous cell surface expression across all stages of disease with limited distal normal tissue expression, making it ideal for therapeutic targeting. A multifaceted lead generation approach enabled development of an armored STEAP2 chimeric antigen receptor T cell (CAR-T) therapeutic candidate, AZD0754. This CAR-T product was armored with a dominant-negative TGF-β type II receptor, bolstering its activity in the TGF-β–rich immunosuppressive environment of prostate cancer. AZD0754 demonstrated potent and specific cytotoxicity against antigen-expressing cells in vitro despite TGF-β–rich conditions. Further, AZD0754 enforced robust, dose-dependent in vivo efficacy in STEAP2-expressing cancer cell line–derived and patient-derived xenograft mouse models, and exhibited encouraging preclinical safety. Together, these data underscore the therapeutic tractability of STEAP2 in prostate cancer as well as build confidence in the specificity, potency, and tolerability of this potentially first-in-class CAR-T therapy.

Authors

Peter Zanvit, Dewald van Dyk, Christine Fazenbaker, Kelly McGlinchey, Weichuan Luo, Jessica M. Pezold, John Meekin, Chien-ying Chang, Rosa A. Carrasco, Shannon Breen, Crystal Sao-Fong Cheung, Ariel Endlich-Frazier, Benjamin Clark, Nina J. Chu, Alessio Vantellini, Philip L. Martin, Clare E. Hoover, Kenesha Riley, Steve M. Sweet, David Chain, Yeoun Jin Kim, Eric Tu, Nathalie Harder, Sandrina Phipps, Melissa Damschroder, Ryan N. Gilbreth, Mark Cobbold, Gordon Moody, Emily E. Bosco

×

Figure 4

In vitro cytolytic activity of 40A3Bz dnTGFβRII CAR-Ts.

Options: View larger image (or click on image) Download as PowerPoint
In vitro cytolytic activity of 40A3Bz dnTGFβRII CAR-Ts. (A) Components o...
(A) Components of the STEAP2 unarmored and armored lentivirus constructs used in generation of the CAR-Ts. (B) Viable cell expansion of CAR-Ts was assessed for 10 days after lentivirus transduction in n = 3 donors. Data represent mean ± SEM. (C) The 40A3Bz cells and 40A3Bz dnTGFβRII STEAP2 CAR-Ts were evaluated by flow cytometry at day 9 after transduction to assess CAR positivity and cell surface expression of dnTGFβRII, compared with untransduced T cells from the same donor. (D) CAR-Ts from C were stained for phenotypic surface markers including CD45RO and CD62L and analyzed by flow cytometry. Naive (CD45RO–CD62L+), central memory (CD45RO+CD62L+), effector memory (CD45RO+CD62L–), and effector (CD45RO–CD62L–) T cells were used. (E) CAR-Ts from C were cocultured with antigen-positive (Ad293 STEAP3-2 and LNCaP) and antigen-negative (Ad293 and LNCaP STEAP2 CRISPR) cell lines. Killing of target cells was measured over 100 hours with the xCELLigence impedance assay. Data are an average of duplicate. (F) Supernatants from the same coculture experiments were collected 24 hours after addition of CAR-Ts, and cytokines (IFN-γ, TNF-α, and IL-2) were measured by Meso Scale Discovery (MSD) electrochemiluminescence assay. Data are an average of duplicate. (G) STEAP2 CAR-Ts were subjected to FACS for CAR positivity and starved overnight before stimulation with recombinant human TGF-β treatment. Cell lysates were generated during the indicated time course, and Western blotting was performed to evaluate levels of p-SMAD2/3, total SMAD2/3, and β-actin in each sample. (H) STEAP2 CAR-Ts were cocultured with antigen-positive cells (C4-2 cells stably expressing mKate2 red fluorescent protein) at a 0.3:1 ratio in the presence of 30 ng/mL recombinant TGF-β. C4-2 cell viability was monitored over 120 hours with the Incucyte live cell analysis system (Sartorius) in triplicate. Data represent mean ± SEM. Data in B–F and H are representative of the results obtained in 3 or more independent experiments with CAR-Ts prepared from 3 healthy donors, and G was performed twice with 2 donors.