A CD3 x FRα T-cell engaging bispecific antibody for efficient killing of ovarian cancer cells with minimal cytokine release.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e18050-e18050
Author(s):  
Ben Buelow ◽  
Brian Avanzino ◽  
Aarti Balasubramani ◽  
Andrew Boudreau ◽  
Laura Davison ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Cytokine Production ◽  
Cell Cytotoxicity ◽  
Cytokine Release ◽  
Limited Efficacy ◽  
Human T Cells

e18050 Background: Ovarian Cancer (OvCa) is the leading cause of gynecologic cancer mortality in women. Since the introduction of platinum-based chemotherapy there has been little change in the prognosis of OvCa patients, with < 30% overall survival in advanced disease, creating an urgent medical need for novel therapies. Few ovarian epithelium-specific surface proteins are suited for Ab targeting. However, studies have shown folate receptor α (FRα) to be highly over-expressed in OvCa; expression level and stage correlate, and FRα is absent or minimally expressed in normal tissues. However, naked Ab therapy has shown limited efficacy while CAR-T therapy has been plagued by toxicity and limited efficacy. ADCs have demonstrated some activity but present the risk of toxin-mediated side effects. Using Teneobio’s unique antibody discovery platform, we have developed a CD3 x FRα T-BsAb that retains the potent cytotoxicity of other T-cell redirecting therapies but with significantly reduced cytokine release. Methods: Antibodies targeting CD3 and FRα were generated via immunization of our proprietary transgenic animals. Candidate antibodies were selected by repertoire deep sequencing of B-cells from draining lymph nodes, high-throughput gene assembly, recombinant expression, and functional screening. Bispecific antibodies targeting CD3 and FRα were assembled and evaluated for their ability to selectively activate primary human T-cells and mediate killing of FRα+ tumor cells in vitro and in vivo. T-cell activation surface markers, cytokine production and tumor cell cytotoxicity were measured. Results: Primary human T-cells were activated only in the presence of both the CD3 x FRα T-BsAb and FRα (either recombinant or cell-surface protein). Potent and selective cytotoxicity against FRα+ tumor cells was observed in co-cultures of primary human T-cells and OvCa tumor cell lines. Strikingly, our T-BsAb mediated comparable tumor cell cytotoxicity to CD3 x FRα T-BsAbs containing a high affinity anti-CD3 domain but with significantly reduced cytokine production. Our Ab showed preliminary evidence of tumor growth inhibition in xenograft models of OvCa in vivo. Conclusions: We have created a novel CD3 x FRα T-BsAb that mediates T-cell killing of FRα+ tumor cells with minimal production of cytokines. This molecule may improve safety, efficacy, and offer opportunities for combination therapy to treat OvCa.

Effect of modulation of CD3 binding in a PSMAxCD3 T-cell engaging bispecific antibody on maintenance of efficient tumor cell kill and cytokine release.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e17583-e17583
Author(s):  
Ben Buelow ◽  
Kevin Dang ◽  
Pranjali Dalvi ◽  
Yuping Li ◽  
Alexander Cheung ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Bispecific Antibody ◽  
Cytokine Production ◽  
Bispecific Antibodies ◽  
Cytokine Release ◽  
Human T Cells

e17583 Background: Castration resistant prostate cancer (CRPC) is an incurable disease and represents a significant unmet need. Prostate specific membrane antigen (PSMA) is a protein highly expressed on the surface of prostate cancer cells; expression has been shown to increase with disease progression. Therapies targeting PSMA, such as anti-PSMA radioligand conjugates, have shown promise in clinical trials, validating this target for CRPC. T-cell recruiting bispecific antibodies (T-BsAbs) have demonstrated potent tumor killing activity against multiple tumor types, but immune-mediated toxicities have hampered T-cell redirecting therapies to date. Using Teneobio’s unique antibody discovery platform, we have developed a CD3xPSMA bispecific antibody (TNB-585) that retains the potent cytotoxicity of other T-BsAbs but with significantly reduced cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated via immunization of our proprietary transgenic animals. Candidate antibodies were selected by repertoire deep sequencing of B-cells from draining lymph nodes, followed by high-throughput gene assembly and recombinant expression. Multiple bispecific antibodies targeting CD3 and PSMA were assembled and evaluated for their ability to selectively activate primary human T-cells and mediate killing of PSMA+ tumor cells in vitro, ex vivo, and in vivo. T-cell activation surface markers, cytokine production, and tumor cell cytotoxicity were measured. Results: In co-culture experiments, primary human T-cells were activated only in the presence of both the bispecifics and PSMA+ cells. These bispecifics mediated potent and selective cytotoxicity against PSMA-positive tumor cells, prostate tumor cell lines, or primary human prostate tumor cells isolated from patients. From among these we identified TNB-585, which showed attenuated binding to CD3. TNB-585 mediated comparable tumor cell cytotoxicity to CD3xPSMA T-BsAbs containing a high affinity anti-CD3 domain but with significantly reduced cytokine production. TNB-585 also showed tumor growth inhibition in xenograft models of prostate cancer in vivo. Conclusions: We have developed a novel CD3xPSMA T-BsAb that mediates T-cell killing of PSMA+ tumor cells with minimal production of cytokines. This molecule may improve safety, efficacy, and offer opportunities for combination therapy to treat CRPC. A Phase 1 clinical trial of this compound in CRPC is scheduled to begin in Q1 2021.

Development of human NKG2D-CD3ε chimeric antigen receptor (CAR) for T-cell-mediated cancer immunotherapy.

2017 ◽  
Vol 35 (7_suppl) ◽  
pp. 150-150
Author(s):  
Sergei Kusmartsev ◽  
Johaness Vieweg ◽  
Victor Prima
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Chimeric Antigen Receptor ◽  
Antigen Receptor ◽  
Adaptor Protein ◽  
T Cell Subsets ◽  
Human T Cells

150 Background: NKG2D is a lectin-like type 2 transmembrane receptor that expressed by natural killer cells and some T cell subsets. Stimulation of NKG2D receptor with specific agonistic ligands produces activating signals through signaling adaptor protein DAP10 leading to the enhanced cytokine production, proliferation, and cytotoxicity against tumor cells. There is strong evidence that NKG2D ligands are expressed in many human tumors, including melanoma, leukemia, myeloma, glioma, and carcinomas of the prostate, breast, lung, and colon. Recent studies also demonstrated that T cells bearing chimeric antigen receptor (CAR) NKG2D linked to CD3ζ (zeta) chain produce marked in vitro and in vivo anti-tumor effects. The aim of current study was to determine whether human T cells bearing chimeric antigen receptor (CAR) NKGD2 linked to CD3ε (epsilon) chain could be activated by the NKG2D-specific stimulation and able to kill human cancer cells. Given the important role of CD3ε in activation and survival of T cells, we hypothesized that NKG2D-CDε-bearing T cells could exert strong in vitro and in vivo anti-tumor effects. Methods: NKG2D CAR was produced by linking human NKG2D to DAP10 and the cytoplasmic portion of the CD3ε chain. Original full-length human cDNA clones were obtained from NIH Mammalian Gene Collection (MGC). Functional domain analysis and oligonucleotide design in the in-Fusion system of DNA cloning (Clontech) was used to generate the retroviral expression constructs. Results: Human PBMC-derived T cells were retrovirally transduced with newly generated NKG2D-CD3ε CAR DNA construct. These NKG2D CAR-expressing human T cells responded to NKG2D-specific activation by producing IFN-γ and exhibited significant cellular cytotoxicity against human tumor cells in vitro. In vivo studies demonstrated that NKG2D-CD3ε-bearing cells are capable of inhibiting growth of DU-145 human prostate cancer in the immunodeficient mice. Conclusions: Collectively, our data indicate the feasibility of developing chimeric antigen receptor NKG2D-CD3ε for T cells and suggest that adoptive transfer of T cells bearing NKG2D-CD3ε CAR could be potentially effective for immunotherapy of cancer patients.

ISB 1342: A first-in-class CD38 T cell engager for the treatment of relapsed refractory multiple myeloma.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. 8044-8044
Author(s):  
Marie-Agnès Doucey ◽  
Blandine Pouleau ◽  
Carole Estoppey ◽  
Cian Stutz ◽  
Amelie Croset ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Cell Killing ◽  
Fragment Antigen Binding ◽  
Tumor Cell Killing

8044 Background: ISB 1342 is a bispecific antibody heterodimer based on the Ichnos proprietary Bispecific Engagement by Antibodies based on T cell receptor (BEAT) platform. ISB 1342 is a first-in-class CD38 T cell engager under investigation in subjects with relapsed multiple myeloma refractory to proteasome inhibitors (PIs), immunomodulators (IMiDs) and daratumumab (study ISB 1342-101). Methods: ISB 1342 was engineered with a single chain variable fragment (scFv) arm that specifically recognizes a cluster of differentiation (CD)3-epsilon (CD3ε) and a fragment antigen binding (Fab) arm which specifically recognizes CD38 and does not compete with daratumumab. By co-engaging CD3ε on T cells and CD38 on tumor cells, ISB 1342 redirects T cells to kill CD38-expressing tumor cells. This mechanism of action is differentiated from existing monospecific CD38 targeting therapies and was designed to overcome resistance to daratumumab in multiple myeloma. Results: In vitro, ISB 1342 killed a large range of CD38-expressing tumor cell lines (EC50:12 to 90 pM) with 8 to 239-fold superior efficacy than daratumumab. ISB 1342 was also able to efficiently kill CD38 low-intermediate-expressing tumor cells that were poorly killed by daratumumab. ISB 1342 retained the potency to kill CD38 low-intermediate-expressing tumor cells when used in sequential or concomitant combination with daratumumab. In addition, the presence of soluble CD38 or glucocorticoid did not impact ISB 1342 killing potency. ISB 1342 was constructed with a double LALA mutation that dampens the binding to Fcγ receptors and C1q. Consistently, ISB 1342 showed only residual Fc-mediated effector functions and its mechanism of tumor cell killing critically relies on the engagement and the activation of T lymphocytes. ISB 1342 showed a favorable on target specificity profile in vitro and was unable to activate T cells in the absence of CD38 positive target cells. Further, ISB 1342-induced tumor cell killing was not associated with a detectable T cell fratricide in vitro. Finally, the potency of ISB 1342 was assessed in vivo in a therapeutic model of a subcutaneously established Daudi tumor co-xenografted with human PBMCs. In marked contrast to daratumumab, which induced only a partial tumor control, ISB 1342 induced complete tumor eradication when injected intravenously weekly at 0.5 mg/kg. As anticipated, the ISB 1342 control molecule (ISB 1342_13DU) made of an irrelevant CD38 binder failed to control tumor growth. The release of the Granzyme A and B, TNF-alpha and CXCL-10 in the tumor micro-environment one week post-treatment was strongly and significantly increased by ISB 1342 but not by daratumumab and ISB 1342_13DU; this represents a correlate of anti-tumor immunity associated with ISB 1342 efficacy in vivo. Conclusions: Hence the higher potency of ISB 1342 relative to daratumumab supports the ongoing clinical development in multiple myeloma patients.

Evaluation of monovalent versus biparatopic CD3xPSMA bispecific antibodies for t-cell mediated killing of prostate tumor cells with minimal cytokine release.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. e16519-e16519
Author(s):  
Ben Buelow ◽  
Starlynn Clarke ◽  
Kevin Dang ◽  
Jacky Li ◽  
Chiara Rancan ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Bispecific Antibodies ◽  
Cytokine Release ◽  
Binding Domains ◽  
Prostate Tumor Cells

e16519 Background: Castration resistant prostate cancer (CRPC) remains an incurable disease and new treatments are needed. Therapies directed against Prostate specific membrane antigen (PSMA) -such as radiolabeled antibodies, chimeric antigen receptor T cells (CAR-Ts) and T-cell engaging bispecific antibodies (T-BsAbs)- have shown promising efficacy but also induce significant toxicity. In particular T-cell redirection leads to efficient killing of tumor cells but induces cytokine release-related toxicities. We have developed a panel of monovalent and biparatopic CD3xPSMA bispecific antibodies that eliminate prostate tumor cells while minimizing cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated in transgenic rats (UniRat™, OmniFlic™) followed by deep sequencing of the antibody repertoire from draining lymph nodes in immunized animals, and high-throughput gene assembly/expression. PSMA x CD3 T-BsAbs were assembled and evaluated for stability, pharmacokinetics, and T cell activation and ability to eliminate PSMA+ tumor cells in vitro and in vivo. Results: Bispecific CD3xPSMA Abs. incorporating either monovalent or biparatopic anti-PSMA binding domains activated T-cells in the presence of PSMA (plate-bound or cell surface), while no T cell activation occurred in the absence of either PSMA antigen or bispecific antibody. Potent/selective cytotoxicity against PSMA+ cells was observed in co-cultures of primary human T cells and tumor cells treated with CD3xPSMA T-BsAbs. Similar results were observed in in vivo Xenograft models of prostate cancer. Strikingly, CD3xPSMA bispecifics containing a novel low affinity anti-CD3 domain produced similar levels of tumor cytotoxicity compared to those with a traditional high affinity anti-CD3 domain, but with reduced cytokine production. Conclusions: We have created novel CD3xPSMA bispecific antibodies incorporating both monovalent and biparatopic anti-PSMA binding domains that mediate T-cell killing of PSMA+ tumor cells with minimal production of cytokines. Such T-BsAbs may improve safety, efficacy, and opportunities for combination therapy to treat CRPC.

Bispecific SCORPION™ Molecules Effectively Redirect T-Cell Cytotoxicity Toward CD19-Expressing Tumor Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3722-3722
Author(s):  
Ruth A. Chenault ◽  
Rebecca Gottschalk ◽  
Gabriela Hernandez-Hoyos ◽  
Jennifer Wiens ◽  
Brian Gordon ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Mouse Serum ◽  
Cell Cytotoxicity ◽  
Employment Equity ◽  
Equity Ownership ◽  
T Cell Cytotoxicity

Abstract Abstract 3722 Background: Despite advances in treatments for B-cell leukemias and lymphomas, many patients ultimately relapse and succumb to disease following multiple courses of therapy. Bispecific antibody fragments that can simultaneously engage T cells and tumor cells have been shown, in the literature, to destroy tumor cells by effectively redirecting the cytotoxic function of T cells. T-cell engaging bispecific molecules linking anti-CD19 and anti-CD3 binding domains in the context of novel SCORPION™ (multi-specific protein therapeutic) proteins were evaluated both in vitro and in vivo for function and stability. Methods: Redirected T-cell cytotoxicity (RTCC) was measured by combining CD19 positive or negative cell lines with SCORPION proteins in the presence of human T cells. In a similar assay context, CFSE-labeled T cells were monitored for activation and proliferation. Functional RTCC assays were also used to analyze serum stability of SCORPION molecules in vitro and to complete an in vivo pharmacokinetic analysis. In vivo efficacy was assessed by monitoring the rate of tumor outgrowth of Ramos xenografts co-implanted with human peripheral blood mononuclear cells (PBMC) in NOD/SCID mice after treatment with SCORPION molecules. Results: SCORPION molecules potently mediate target-specific T-cell cytotoxicity toward tumor cell lines presenting cell surface CD19, with EC50 values for cytotoxicity at low pM concentrations. These molecules also demonstrate induction of T-cell activation and proliferation in the presence of target-bearing tumor cells but not in the absence of target expression. SCORPION molecules retain stable function following incubation at 37°C in mouse serum for up to a week in vitro, and pharmacokinetic analysis of SCORPION protein function in BALB/c mouse serum following intravenous administration resulted in half-life estimates of 69–84 hours. In efficacy studies conducted in NOD/SCID mice, SCORPION proteins significantly inhibited the outgrowth of Ramos tumor xenografts in the presence of human effector cells. Conclusion: SCORPION molecules targeting CD19 and CD3 effectively harness the cytotoxic activity of T cells to kill CD19 positive tumor cells both in vitro and in vivo and show potential for further investigation as possible therapeutic agents for B-cell malignancies. Disclosures: Chenault: Emergent BioSolutions: Employment. Gottschalk:Emergent BioSolutions: Employment. Hernandez-Hoyos:Emergent BioSolutions: Employment. Wiens:Emergent BioSolutions: Employment. Gordon:Emergent BioSolutions: Employment. Klee:Emergent BioSolutions: Employment, Equity Ownership. Bienvenue:Emergent BioSolutions: Employment. Dasovich:Emergent BioSolutions: Employment. Kumer:Emergent BioSolutions: Employment. Aguilar:Emergent BioSolutions: Employment. Bannink:Emergent BioSolutions: Employment, Equity Ownership. McMahan:Emergent BioSolutions: Employment, Equity Ownership. Natarajan:Emergent BioSolutions: Employment, Equity Ownership. Algate:Emergent BioSolutions: Employment, Equity Ownership. Blankenship:Emergent BioSolutions: Employment, Equity Ownership, Patents & Royalties.

A novel CD3xPSMA bispecific antibody for efficient T cell mediated killing of prostate tumor cells with minimal cytokine release.

2019 ◽  
Vol 37 (7_suppl) ◽  
pp. 324-324 ◽  
Author(s):  
Starlynn Clarke ◽  
Kevin Dang ◽  
Yuping Li ◽  
Preethi Sankaran ◽  
Duy Pham ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Prostate Tumor ◽  
Bispecific Antibodies ◽  
Therapeutic Window ◽  
Cytokine Release ◽  
Human T Cells ◽  
Prostate Tumor Cells

324 Background: Castration resistant prostate cancer (CRPC) remains an incurable disease and new therapeutics are urgently needed. Prostate specific membrane antigen (PSMA) is expressed on the surface of prostate cancer cells and expression increases with disease progression. Therapies directed against PSMA such as radiolabeled antibodies and T cell redirecting therapies including chimeric antigen receptor T cells (CAR-Ts) and T-cell engaging bispecific antibodies (T-BsAbs) have shown promising efficacy in clinical trials but also induce significant toxicity. In particular CAR-Ts and T-BsAbs potently kill tumor cells but induce cytokine release-related toxicities. Novel anti-CD3 engaging domains may be required to create T-BsAbs with a broader therapeutic window. We have developed fully human CD3xPSMA bispecific antibodies that efficiently eliminate prostate tumor cells while minimizing cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated in transgenic rats that produce human antibodies (UniRat, OmniFlic) followed by repertoire deep sequencing of lymph nodes isolated from immunized animals and high-throughput gene assembly/expression. CD3xPSMA T-BsAbs were assembled and evaluated for T cell activation and ability to eliminate PSMA+ tumor cells in vitro. Results: Primary human T cells were activated only in the presence of both bispecific CD3xPSMA antibodies and PSMA (either plate-bound or on the surface of tumor cells). Potent and selective cytotoxicity against PSMA+ prostate tumor cells was observed in co-cultures of primary human T cells and tumor cells treated with CD3xPSMA bispecific antibodies. Strikingly, CD3xPSMA bispecifics containing a novel low affinity anti-CD3 domain produced similar levels of tumor cell cytotoxicity compared to CD3xPSMA bispecifics containing a traditional high affinity anti-CD3 domain, but with reduced cytokine production. Conclusions: We have created novel CD3xPSMA bispecific antibodies that mediate T-cell killing of PSMA+ tumor cells with minimal production of cytokines. Such T-BsAbs may improve safety, efficacy, and opportunities for combination therapy to treat CRPC.

scholarly journals Preclinical and Nonclinical Characterization of HPN217: A Tri-Specific T Cell Activating Construct (TriTAC) Targeting B Cell Maturation Antigen (BCMA) for the Treatment of Multiple Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3225-3225 ◽  
Author(s):  
Che-Leung Law ◽  
Wade Aaron ◽  
Rick Austin ◽  
Manasi Barath ◽  
Evan Callihan ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Half Life ◽  
Cell Activation ◽  
The Us ◽  
Dose Dependent

Abstract About 31,000 new cases of multiple myeloma (MM) will be diagnosed in the US in 2018. In addition to chemotherapeutic agents, several targeted therapies utilizing distinct mechanisms of action, e.g., proteasome inhibitors (bortezomib, carfilzomib, ixazomib), HDAC inhibitors (panobinostat), Cullin-RING E3 ubiquitin ligase activators (thalidomide, lenalidomide, pomalidomide), and antibodies (daratumumab, elotuzumab) have become available for treating MM. However, MM remains an incurable disease. Patients who relapse after or are refractory to standard of care treatments generally have poor prognosis. In 2018, close to 13,000 patients will die of the disease in the US. Targeting the B cell maturation antigen (BCMA), a BAFF/BLyS and APRIL receptor, for treating MM patients can provide a new treatment approach complementary to existing therapies. CAR-T therapies and an antibody-drug conjugate targeting BCMA have demonstrated early clinical success in the treatment of relapsed refractory MM (RRMM). HPN217 is a tri-specific T cell activating construct (TriTAC) consisting of three binding domains: an N-terminal single domain antibody (sdAb) that binds to human BCMA, a middle sdAb that binds to human serum albumin (HSA), and a C-terminal single chain Fv (scFv) that binds to CD3ε of the T cell receptor (TCR) complex. HPN217 is a highly stable single polypeptide of ~ 53 kDa expressed by CHO cells. Simultaneous engagement of BCMA on a target MM cell and CD3 on a T cell results in T cell activation, functional differentiation and the eventual lysis of the target MM cell. Engineering of an HSA binding domain into HPN217 represents a unique strategy in extending serum half-life, giving the TriTAC molecule a small molecular size and flexibility. This approach is different from Fc-engineering applied in other CD3-based bispecific T cell engaging molecules. The KD of HPN217 binding to recombinant human BCMA, HSA, and recombinant human CD3ε was determined to be 5.5 nM, 6 nM, and 17 nM, respectively, as measured by biolayer interferometry. Flow cytometric analysis on a panel of T cells from normal donors and BCMA positive and BCMA negative tumor cell lines confirmed binding of HPN217 to its native targets expressed on cell surface. The in vitro pharmacological activity of HPN217 was evaluated by T cell-dependent cellular cytotoxicity (TDCC) assays. In co-cultures of T cells from normal human or cynomolgus monkey donors, target tumor cells, and HSA, HPN217 mediated dose-dependent and BCMA-dependent cytotoxicity with EC50 values ranging from 0.05 to 0.7 nM. Killing was dependent on expression of BCMA on target tumor cells. Concomitant with target tumor cell killing, HPN217 also mediated dose-dependent upregulation of CD25 and CD69 on T cells in the TDCC co-cultures when BCMA positive tumor cells were presence. Consistent with the mechanism of action of CD3-based T cell engaging molecules, T cell derived cytokines, e.g., TNFα and IFNγ, were detected. Similar T cell activation could be observed using human or cynomolgus monkey whole blood as a source of T cells. Nonclinical in vivo properties of HPN217 were evaluated in xenograft models and a single dose pharmacokinetic (PK) study in cynomolgus monkeys. HPN217 mediated dose-dependent growth suppression against the RPMI-8226 MM model and Jeko-1 mantle cell lymphoma model expressing relatively low levels of 5,600 and 2,200 copies of BCMA per cell, respectively. In the PK study, a single dose of HPN217 at 0.01, 0.1, or 1 mg/kg was given to cynomolgus monkeys. HPN217 exhibited linear PK behavior over this dose range. Serum half-life was in the range of 64 to 85 hours. Serum half-life, volume of distribution, and clearance appeared to be independent of dose. HPN217 was demonstrated to be stable and remained intact up to 3 weeks in vivo as demonstrated by a functional ligand binding assay using recombinant CD3ε and BCMA, respectively, to capture and detect HPN217. Importantly, serum samples collected one week after dosing were as potent as stock HPN217 in MM tumor cell killing in TDCC assays. Collectively, preclinical and nonclinical characterization suggests that HPN217 is an efficacious novel therapeutic candidate that can provide a convenient dosing schedule for patients. A first-in-human phase 1 clinical trial is planned to evaluate HPN217 in RRMM. Disclosures Law: Harpoon Therapeutics: Employment. Aaron:Harpoon Therapeutics: Employment. Austin:Harpoon Therapeutics: Employment. Barath:Harpoon Therapeutics: Employment. Callihan:Harpoon Therapeutics: Employment. Evans:Harpoon Therapeutics: Employment. Gamez Guerrero:Harpoon Therapeutics: Employment. Hemmati:Harpoon Therapeutics: Employment. Jones:Harpoon Therapeutics: Employment. Kwant:Harpoon Therapeutics: Employment. Lao:Harpoon Therapeutics: Employment. Lemon:Harpoon Therapeutics: Employment. Patnaik:Harpoon Therapeutics: Employment. Sexton:Harpoon Therapeutics: Employment. Wesche:Harpoon Therapeutics: Employment. Xiao:Harpoon Therapeutics: Employment. Yu:Harpoon Therapeutics: Employment. Yu:Harpoon Therapeutics: Employment.

scholarly journals Preclinical Characterization of an ANTI-CD38/CD3 T CELL-Redirecting Bispecific Antibody

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4463-4463
Author(s):  
Xiao He ◽  
Yanliang Zhang ◽  
Yun Wei Lai ◽  
Stephanie Baguley ◽  
Yan Li ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
T Cell Activation ◽  
Cytokine Release ◽  
Employment Equity ◽  
Equity Ownership ◽  
Anti Tumor Activity

Introduction: Multiple Myeloma (MM) and Non-Hodgkin Lymphoma (NHL) are hematologic malignancies that remain difficult to treat. While autologous CAR-T cell therapies have shown promise in treating these diseases, these therapies are not without issues, including lack of response in many patients, lengthy time to produce CAR-T cells, occasional production failures, as well as high manufacturing costs. As an alternative approach, protein-based T cell engaging and redirecting bispecific antibodies (BsAbs) have been developed. We have generated anti-CD38/CD3 BsAbs to redirect T cells against CD38, a clinically validated antigen in MM and studied their ability to elicit target-dependent tumor cell lysis. The lead molecule is a humanized, stability-engineered CD3-engaging and CD38 antigen affinity-optimized BsAb with reduced effector function to mitigate antigen-independent T cell toxicity. Preclinical data demonstrate potent anti-tumor activity in vitro assays and in vivo studies against CD38-expressing lymphoma and MM cell lines. Methods: Anti-CD38/CD3 BsAbs were generated by CH3 Fc domain interface engineering for heterodimerization of a CD38-targeting Fab arm and anti-CD3-scFv-Fc fusion chain with hinge mutations for reduced FcR affinities. Novel bispecific molecules that bind to CD38 with various affinities/binding kinetics were evaluated in a series of in vitro and in vivo studies, including target-specific redirected T cell cytotoxicity (RTCC) against cancer cell lines. T cell response profiles, and cytokine release. The lead CD38/CD3 BsAb was selected and further evaluated for its ability to inhibit tumor growth and prolong survival in a disseminated luciferase-expressing Raji xenograft mouse model co-implanted with primary human peripheral blood mononuclear cells (hPBMC). Results: Our lead CD38/CD3 BsAb possesses the desired CD38 and CD3, affinities resulting in effective tumor antigen and T cell engagement for RTCC. The CD38/CD3 BsAb induced potent T cell-dependent lysis of CD38-positive cancer cells in vitro, with the CD38 antigen density positively correlating with the cytotoxicity potency. Antigen dependent and dose-dependent T cell activation and cytokine release were studied in vitro, with the level of T cell activation and cytokine release being indicative of the anti-tumor potency but not necessarily anti-CD3 affinity. In an in vivo study, we evaluated the impact of CD38 affinity of the BsAb on anti-tumor activity of the BsAbs. The data showed that a balanced CD38 vs CD3 affinity was shown to be preferred for T cell stimulation and prolonged anti-tumor activity. In preclinical cytotoxicity assays against a cancer cell line panel using hPBMC from healthy donors, our lead CD38/CD3 BsAb was benchmarked against daratumumab, a marketed anti-CD38 antibody for MM, and demonstrated more potent tumor cell killing. These data suggest a more robust anti-tumor activity exerted by the CD38/CD3 BsAb through RTCC than daratumumab through antibody-dependent cellular cytotoxicity (ADCC). In Raji tumor cell-bearing NSG mice implanted with previously unstimulated hPBMCs, our CD38/CD3 BsAb induced tumor growth inhibition and prolonged survival compared to control BsAb or hPBMCs-only treated animals. Conclusions: Our preclinical data demonstrate that our lead CD38/CD3 BsAb recruits T cells against CD38-positive tumor MM and lymphoma cells in a potent target and dose-dependent manner in preclinical studies. These preclinical characterizations support the rationale for clinical investigation of the lead BsAb in selected CD38-positive malignancies. Disclosures He: Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties. Zhang:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties. Lai:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties. Baguley:Sorrento Therapeutics, Inc.: Employment, Equity Ownership. Li:Sorrento Therapeutics, Inc.: Employment, Equity Ownership. Cao:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties. Yan:Sorrento Therapeutics, Inc.: Employment, Equity Ownership. Takeshita:Sorrento Therapeutics, Inc.: Employment, Equity Ownership. Zeldis:Sorrento Therapeutics Inc: Employment, Equity Ownership. Ji:Sorrento Therapeutics Inc: Employment, Equity Ownership, Patents & Royalties; Celularity, Inc.: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Kaufmann:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties.

scholarly journals Inducible MyD88/CD40 Enhances Proliferation and Survival of PRAME-Specific TCR-Engineered T Cells and Increases Anti-Tumor Effects in Myeloma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1886-1886 ◽  
Author(s):  
Tsvetalina Hoang ◽  
Aaron Foster ◽  
Jeannette Crisostomo ◽  
An Lu ◽  
Annemarie Moseley ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Mhc Class I ◽  
Class I ◽  
Tumor Control ◽  
Employment Equity ◽  
Human T Cells ◽  
Equity Ownership

Abstract Introduction: Use of T cells engineered to express antigen-specific T cell receptors (TCRs) has shown promise as a cancer immunotherapy treatment; however, durable responses have been limited by poor T cell persistence and expansion in vivo. Additionally, MHC class I downregulation on tumor cells weakens T cell recognition, further reducing therapeutic efficacy. To address these deficiencies, we co-expressed in human T cells a novel, small molecule (rimiducid)-dependent T cell activation switch, inducible MyD88/CD40 (iMC), along with PRAME-specific TCR to allow control of T cell expansion and activation, while upregulating MHC class I expression on tumor cells. Methods: Human T cells were activated with anti-CD3/CD28 and transduced with retrovirus encoding TCR α and β chains recognizing PRAME-derived, HLA-A*201-restricted peptide SLLQHLIGL (SFG-PRAME) or a polycistronic vector encoding the PRAME-specific TCR along with tandem rimiducid (AP1903)-binding domains (FKBP12v36) cloned in-frame with MyD88 and CD40 signaling domains (SFG-iMC-PRAME). Proliferation, cytokine production and cytotoxicity of modified T cells was assessed using peptide-pulsed T2 cells or against PRAME-expressing, HLA-A2+ U266 myeloma tumor cells with or without rimiducid (10 nM) stimulation. MHC class I expression on tumor cells was measured by flow cytometry using a transwell assay. In vitro tumor killing was analyzed using T cell and tumor coculture assays with various effector to target ratios over a 7-day period. In vivo efficacy was determined using immune-deficient NSG mice engrafted s.c. with U266 cells and treated i.v. with 1x107 transduced T cells. iMC was activated in vivo by weekly i.p. injections of 5 mg/kg rimiducid. Tumor size and T cell expansion was measured using in vivo luciferase bioluminescence imaging and flow cytometric phenotyping. Results: Both PRAME and iMC-PRAME retroviral vectors efficiently transduced activated human T cells (81±2.1% and 89±2.8%, respectively) and showed antigen-specific IFN-g production and cytolytic function against peptide-pulsed T2 cells and PRAME+ U266 myeloma cells. However, both TCR ligation and rimiducid-dependent costimulation were required for IL-2 production (PRAME, 217±256 pg/ml; iMC-PRAME, 23±56 pg/ml; iMC-PRAME + rimiducid, 5417±2599 pg/ml) against peptide-pulsed T2 cells. Coculture assays against PRAME-expressing U266 myeloma cells showed that tumor elimination was optimized with concurrent rimiducid-driven iMC activation, and this effect was accompanied by increased IL-2 secretion and robust T cell proliferation (PRAME, 0.18-fold; iMC-PRAME, 0.28-fold; iMC-PRAME + rimiducid, 7.7-fold). Further, iMC activation produced IFN-g independently of TCR ligation, which significantly increased MHC class I expression on tumor cells (no T cells, 61±3 MFI; PRAME, 1256±493 MFI; iMC-PRAME, 6747±656 MFI; iMC-PRAME + rimiducid, 9096±1583 MFI). In NSG mice engrafted with PRAME+ U266 myeloma tumors, PRAME TCR-modified T cells showed significant tumor control compared to non-transduced control T cells (p-value = 0.01, 0.01 and 0.0001 for PRAME, iMC-PRAME and iMC-PRAME + rimiducid, respectively) and rimiducid activation of iMC-PRAME-modified T cells showed significant tumor control compared to T cells transduced with only the PRAME TCR (p = 0.005). Importantly, weekly injections of rimiducid dramatically expanded PRAME TCR-expressing T cell numbers by 473-fold 4 weeks post-injection compared to T cells expressing the PRAME TCR only (p = 0.02). Summary: iMC is a novel "Go" switch that utilizes rimiducid, a small molecule dimerizer, to drive activation and expansion of PRAME-specific TCR-engineered T cells while sensitizing tumor to TCR-mediated recognition by upregulating MHC class I via IFN-g, thereby increasing antitumor efficacy and durability. Thus, iMC-PRAME is the prototype of a class of novel "Go-TCR" engineered T cell therapies that may increase efficacy, safety and durability of adoptive T cell therapies. Disclosures Hoang: Bellicum Pharmaceuticals: Employment. Foster:Bellicum Pharmaceuticals: Employment. Crisostomo:Bellicum Pharmaceuticals: Employment. Lu:Bellicum Pharmaceuticals: Employment. Moseley:Bellicum Pharmaceuticals: Employment, Equity Ownership. Slawin:Bellicum Pharmaceuticals: Employment, Equity Ownership. Spencer:Bellicum Pharmaceuticals: Employment, Equity Ownership.

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