Essen BioScience (United States)

The emergence of allogeneic, universal chimeric antigen receptor (CAR) T cell therapies requires intensified and scalable manufacturing workflows supported by representative scale-down models (SDMs) to enable efficient process development and future large-scale production of off-the-shelf therapies. Here, we present a 7-day CAR-T cell expansion process intensified via perfusion of serum-free medium in a 2 L Univessel® Single-Use stirred-tank bioreactor (STR), consistently achieving 30 × 106 cells/mL, corresponding to 113 ± 7 anti-CD19 CAR-T doses per batch. Parallel runs in 250 mL Ambr® 250 STRs conducted at equivalent volumetric power input (P/V) of ∼8.78 W/m3 demonstrated comparable process performance and final product quality, with univariate and multivariate analyses of cell growth, phenotype, cytotoxicity, and cytokine secretion validating the Ambr® 250 as a predictive SDM for the 2 L process. Integrating capacitance sensing in the 2 L STR enabled robust monitoring of viable cell concentrations in real-time, with strong correlation to offline measurements (R2 = 0.98). For downstream processing, the Ksep® 400 was used to automate CAR-T cell harvesting, concentration, and washing at the 2 L scale, achieving >90% product recovery and nine-fold volume reduction without impacting product quality attributes compared to manual methods. This study establishes a scalable CAR-T manufacturing workflow supported by a predictive SDM, providing an efficient platform for process development and scale-up to enable future large-scale production of allogeneic CAR-T cell therapies.

Somatostatin analogues (SSA) are efficacious and safe treatments for a variety of neuroendocrine tumors, especially pituitary neuroendocrine tumors (PitNET). Their therapeutic effects are mainly mediated by somatostatin receptors SST2 and SST5. Most SSAs, such as octreotide/lanreotide/pasireotide, are either nonselective or activate mainly SST2. However, nonfunctioning pituitary tumors (NFPTs), the most common PitNET type, mainly express SST3 and finding peptides that activate this particular somatostatin receptor has been very challenging. Therefore, the main objective of this study was to identify SST3-agonists and characterize their effects on experimental NFPT models. Binding to SSTs and cAMP level determinations were used to screen a peptide library and identify SST3-agonists. Key functional parameters (cell viability/caspase activity/chromogranin-A secretion/mRNA expression/intracellular signaling pathways) were assessed on NFPT primary cell cultures in response to SST3-agonists. Tumor growth was assessed in a preclinical PitNET mouse model treated with a SST3-agonist. We successfully identified the first SST3-agonist peptides. SST3-agonists lowered cell viability and chromogranin-A secretion, increased apoptosis in vitro, and reduced tumor growth in a preclinical PitNET model. As expected, inhibition of cell viability in response to SST3-agonists defined two NFPT populations: responsive and unresponsive, wherein responsive NFPTs expressed more SST3 than unresponsive NFPTs and exhibited a profound reduction of MAPK, PI3K-AKT/mTOR, and JAK/STAT signaling pathways upon SST3-agonist treatments. Concurrently, SSTR3 silencing increased cell viability in a subset of NFPTs. This study demonstrates that SST3-agonists activate signaling mechanisms that reduce NFPT cell viability and inhibit pituitary tumor growth in experimental models that expresses SST3, suggesting that targeting this receptor could be an efficacious treatment for NFPTs.