eluvixtamab (AMG 330)

CRS was mitigated with stepwise dosing of AMG 330, prophylactic dexamethasone, and early treatment with tocilizumab. Among 60 evaluable patients, eight achieved complete remission or morphologic leukemia-free state; of the 52 non-responders, 37% had ≥50% reduction in AML bone marrow blasts. AMG 330 is a promising CD33-targeted therapeutic strategy for R/R AML.

P1, N=1, Terminated, Amgen | Phase classification: P1b --> P1

We propose a novel mechanism by which AMG 330-activated T cells prime and sensitize AML target cells in a forward feedback loop towards STING activation, leading to increased type-I-IFN production and induction of ISGs. The beneficial effect of physiological cGAMP in enhancing AMG 330-mediated cytotoxicity was accompanied by the pronounced expression of effector cytokines and an overall cytotoxic T-cell phenotype. We established a key role for interferon gamma in AMG 330-mediated cytotoxicity of AML cells and in rendering AML cells responsive to STING agonism.

T-cell fitness was assessed by T-cell function assays in co-cultures and immune synapse formation by applying a CD33 BiTE molecule (AMG 330). The addition of the immunomodulatory drug (IMiD) lenalidomide to co-cultures led to stabilization of immune synapses and improved subsequent T-cell responses. We conclude that target cells modulate CD33 BiTE molecule-dependent T-cell activation and hence, combinatorial strategies might contribute to enhanced efficacy.

We hypothesized that combining the CD33 BiTE construct AMG 330 with a cGAS-STING agonist has the potential to reverse immunosuppressive mechanisms and augment anti-leukemic activity...This leads to pronounced expression of effector cytokines and an overall cytotoxic T-cell phenotype, contributing to the beneficial effect of cGAMP in enhancing BiTE construct-mediated lysis. Ethics Approval Peripheral blood or bone marrow samples were collected from healthy donors and patients with acute myeloid leukemia at initial diagnosis, relapse, or complete remission after written informed consent was received in accordance with the Declaration of Helsinki and approval was granted by the Institutional Review Board of the Ludwig-Maximilian-Universität (Munich, Germany, reference number: 216-08).

Using antibody single-chain variable region (scFv) sequences derived from amatuximab-recognizing MSLN, and from either blinatumomab or AMG330 targeting CD3, we engineered and expressed two MSLN/CD3-targeting BsAbs: MSLN-CD3 and MSLN-CD3, respectively. These data validate the in vivo efficacy and specificity of MSLN-targeting BsAbs. Because prior MSLN-directed therapies appeared safe in humans, MSLN-targeting BsAbs could be ideal immunotherapies for MSLN-positive pediatric AML patients.

Currently, two CD33xCD3 BiTE ® antibody constructs (AMG 330 & AMG 673) are being investigated in phase I dose escalation trials in patients with relapsed/refractory Acute Myeloid Leukemia (AML) with early evidence of acceptable safety and anti-leukemic activity (Ravandi et al., ASH 2020; Subklewe et al., EHA 2020). They support the notion that T cell co-signaling receptors like CD86 and PD-L1 modulate T-cell response in an early event manner. Prospective analyses in clinical trials are needed to validate the relevance of checkpoint molecule expression on target cells as a potential predictive biomarker for response.

Additionally, T-cell function was assessed after stimulation with 1) CD3/CD28 beads; 2) AMG 330, a CD33/CD3 specific BiTE ® construct, after incubation with OCI-AML3 target cells; or 3) AMG 330 in an autologous ex vivo long-term culture system after incubation with primary AML cells (pAML)...These observations may highlight the significant role of the AML target cells in shaping a T-cell response. To this end, we will further analyze the longitudinal communication between T cells and their corresponding AML blasts.

Antibody single-chain variable region (scFv) sequences derived from amatuximab recognizing MSLN and from either blinatumomab or AMG330 targeting CD3 were used to engineer and express two MSLN/CD3-targeting BsAbs: MSLN AMA -CD3 L2K and MSLN AMA -CD3 AMG respectively...Chemotherapy (DA) consisted of 3 doses of 1.5 mg/kg daunorubicin iv and 5 doses of 50 mg/kg cytarabine ip...Conclusion These data validate the efficacy of MSLN-targeting BsAbs in PDX models with endogenous MSLN expression. Because prior MSLN-directed therapies appeared safe in humans, MSLN-targeting BsAbs could be ideal immunotherapies for MSLN-positive pediatric AML patients.

Gemtuzumab ozogamicin was the first and only CD33-directed antibody-drug conjugate to be US Food and Drug Administration approved for AML...Promising new strategies include cellular therapy mechanisms and linker molecules. This is an exciting target that requires a considerable amount of precision to yield clinical benefit.

Preliminary response assessment showed a correlation with lower tumor burden at baseline and with a trend towards higher CD8+ lymphocyte count and E:T ratio at baseline. Conclusion AMG 330 dosed up to 720 μg/day provided early evidence of acceptable safety profile, drug tolerability and anti-leukemic activity, and supports further dose escalation.

AMG 330 dosed up to 720 μg/day provided early evidence of acceptable safety profile, drug tolerability and anti-leukemic activity, and supports further dose escalation. Research Funding: Amgen Inc.

Although the use of gemtuzumab ozogamicin (GO) has validated the concept of antibody directed therapy for AML targeting CD33, complications from myelosuppression and opportunistic infections due to on-target toxicity to normal CD33+ hematopoietic cells have limited this approach. B) Quantification of CD14+/CD33+ and CD14+/CD33− cells from peripheral blood of AMG330-treated animals in Cas9 only (n=3) and CRISPR/Cas9 (n = 3) groups. In A) and B), ‘pre’ is before AMG330 treatment and ‘post’ is 5 days after administration.

Using scFv sequences derived from Amatuximab, which recognizes the N-terminal domain of the GPI-linked ectodomain of MSLN, targeting region 1 of MSLN, and from Blinatumomab/AMG-330 targeting CD3, we engineered and expressed two kinds of BiTE molecules – a canonical BiTE and an IgG BiTE, a larger molecule with improved serum half life in vivo. 1H), suggesting that IgG BiTE was far more efficacious than canonical BiTEs (P<0.01). Taken together, these data indicate that MSLN-targeting BiTEs could be used as novel immunotherapy for pediatric AML with MSLN expression.

Proof of concept was shown by Blinatumomab, a CD19xCD3 BiTE® antibody construct, for treatment of r/r B-cell precursor ALL...However, phenotypic and genetic heterogeneity of AML cell lines could mask the influence of p53 mutation on the pharmacodynamic activity of AMG 330...Bulk RNA sequencing of p53 kd AML cell lines compared to p53 wt revealed downregulation of a co-stimulatory ligand 4-1BBL with log2 fold change of - 0.33, -0.30, -0.39 in Molm-13, MV4-11 and OCI-AML3, respectively...In a next step, we will try to identify possible secreted mediators of T-cell modulation by mass. We will also characterize primary patient samples for their T-cell-inhibiting capacities, e.g. p53-mutated disease.