Melanin binding of small molecule drugs can lead to targeted disposition to the pigmented tissues and prolonged pharmacological responses in the eye Melanin binding of drugs in vitro correlates with in vivo binding but current workflows for binding affinity require multiple slow steps and analytical method development and or they ...More |Related Solutions: μPulse^®

Melanin binding of small molecule drugs can lead to targeted disposition to the pigmented tissues and prolonged pharmacological responses in the eye. Melanin binding of drugs in vitro correlates with in vivo binding, but current workflows for binding affinity require multiple slow steps, and analytical method development, and/or they may result in high data variability. We developed tangential flow filtration-based methodology to produce size-specific fractions of water-soluble melanin nanoparticles (MNPs), reducing production time from 2-3 days to just a few hours and yielding MNPs with enhanced fluorescence signal. Improved MNPs enabled modifications to a previously published microscale thermophoresis-based melanin binding protocol, shifting analytical focus toward thermophoretic behavior, reducing data variability, and improving reproducibility. The process was tested with nine compounds with varying melanin binding affinities, and the results were consistent with literature, confirming the ability of the method to differentiate compounds based on melanin binding. Fast and reliable workflow will be useful in screening binding affinity for therapeutics and new drug candidates to melanin thereby facilitating ocular drug discovery and construction of predictive pharmacokinetic simulation models. Less |Related Solutions: μPulse^®

Abstract Background Recombinant human NELL- rhNELL- is a potent osteogenic protein with therapeutic potential in regenerative medicine A stable formulation is essential to prevent aggregation during production filling storage and clinical use Methodology A four-stage rational formulation strategy was used identify intrinsic aggregation risks of rhNELL- screen polysorbate- and cyclodextrin-based ...More |Related Solutions: μPulse^®

Abstract
Background
Recombinant human NELL-1 (rhNELL-1) is a potent osteogenic protein with therapeutic potential in regenerative medicine. A stable formulation is essential to prevent aggregation during production, filling, storage, and clinical use.

Methodology
A four-stage rational formulation strategy was used: (1) identify intrinsic aggregation risks of rhNELL-1; (2) screen polysorbate- and cyclodextrin-based formulations to enhance colloidal and conformational stability; (3–4) test lead candidates under agitation, freeze/thaw, pH shifts, and elevated temperature. Analytical techniques included PEG challenge, differential scanning fluorimetry (DSF), isothermal chemical denaturation (ICD), and dynamic light scattering (DLS). Aggregation was assessed via visible particles (VP), opalescence, subvisible particles (SVP, Micro Flow Imaging), SDS-PAGE, and ultra-high performance size exclusion chromatography (UP-SEC).

Results
rhNELL-1 was prone to self-association via hydrophobic and electrostatic interactions. Polysorbate 20 (PS20) and hydroxypropyl beta cyclodextrin (HPB-LB-BCD) improved protein stability. PS20 markedly reduced VP and SVP formation. While HPB-LB-BCD alone did not further reduce SVP beyond PS20, it enhanced thermal stress resistance. PS20 was more effective under agitation.

Conclusions
Two lead formulations containing potassium phosphate/Tris buffer, sorbitol, PS20, and HPB-LB-BCD demonstrated strong resistance to aggregation under multiple stresses. PS20 mitigated interfacial stress, while HPB-LB-BCD suppressed solution-phase aggregation, especially at high temperatures. This systematic approach offers a framework for stabilizing other aggregation-prone proteins. Less |Related Solutions: μPulse^®

Antibody-drug conjugates ADCs are a promising therapeutic modality that enables the delivery of cytotoxic drugs to the target cells that express the corresponding antigen However the purification of ADCs while ensuring product safety homogeneity and stability is a challenging task due to their complex and fragile structure Size exclusion chromatography ...More |Related Solutions: μPulse^®

Antibody-drug conjugates (ADCs) are a promising therapeutic modality that enables the delivery of cytotoxic drugs to the target cells that express the corresponding antigen. However, the purification of ADCs while ensuring product safety, homogeneity, and stability is a challenging task due to their complex and fragile structure. Size exclusion chromatography (SEC), the conventional method for ADC purification, is time-consuming as it requires multiple column washes and equilibration steps. Moreover, subsequent formulation of ADCs, typically using dead-end filtration (DEF), further complicates the production workflow. We compared SEC+DEF with the µPulse®, a miniaturized and automated tangential flow filtration system, for purification and formulation of ADCs. Quality analysis revealed that both approaches were equally gentle as comparable drug-to-antibody ratios (DARs) and monomer purities were observed in the purified samples. Most importantly, both methods exhibited equivalent cleanup efficiency with a 99.8% reduction in free linker-drug concentration. The endotoxin loads comprised 0.11 EU mg-1 for the µPulse and 0.07 EU mg-1 for SEC+DEF, ensuring validation of the safe application of purified ADCs in living systems. However, the µPulse performed purification and formulation of ADCs simultaneously as compared to SEC+DEF, which required multiple manual interventions. Our results indicate that the µPulse is a gentle, single-step, and walk-away approach for the purification of ADCs. Less |Related Solutions: μPulse^®

Reinvigoration of tumor-reactive T-cells using co-stimulatory bispecific antibodies bsAbs targeting CD or CD is emerging as a promising therapeutic strategy Conditional tumor-specific recruitment can offer a necessary layer of control and specificity We developed pH-selective CD xVISTA bsAbs to act specifically within the acidic tumor microenvironment TME aiming for enhanced ...More |Related Solutions: μPulse^®

Reinvigoration of tumor-reactive T-cells using co-stimulatory bispecific antibodies (bsAbs) targeting CD28 or CD137 is emerging as a promising therapeutic strategy. Conditional, tumor-specific recruitment can offer a necessary layer of control and specificity. We developed pH-selective CD28xVISTA bsAbs to act specifically within the acidic tumor microenvironment (TME), aiming for enhanced T-cell-mediated cancer cell killing while minimizing systemic T-cell activation and Cytokine Release Syndrome (CRS) risk. CD28 agonism by CD28xVISTA bsAbs relies on pH-selective engagement of VISTA, a protein robustly expressed on myeloid cells highly prevalent in most solid tumors. This modality avoids engagement of tumor-associated antigens (TAAs) with the potential to provide highly tumor specific activity with minimal on-target/off-tumor side effects.

We report the identification of a lead candidate with pH-dependent simultaneous engagement of both targets, and VISTA-dependent CD28 signaling in a reporter cell line. CD28xVISTA avidly bound VISTA-positive cells, and co-stimulation was shown in vitro by its ability to activate and expand T-cells and enhance T-cell mediated cancer cell killing in co-cultures of human PBMCs and cancer cells in the presence of a TAA-targeted anti-CD3 T-cell engager. Interestingly, our findings support both signaling in cis (between T-cell and cell displaying peptide-MHC complex) and in trans with stimulation occurring through CD28 clustering outside of the immune synapse. Our lead candidate displayed efficient tumor growth inhibition of human VISTA-expressing MC38 cells in a humanized CD28 syngeneic mouse model in combination with PD-1 blockade. Importantly, our CD28xVISTA bsAb showed no signs of superagonistic properties in several in vitro assays geared towards revealing induction of CRS. Our data supports clinical development in combination with anti-PD-1 or any TAA-targeted anti-CD3 T-cell engagers developed for solid tumors. Less |Related Solutions: μPulse^®

Abstract Background Chronic inflammation and oxidative stress are central to the pathophysiology of Type Diabetes Mellitus T DM contributing to the progression of metabolic dysfunction and related complications Objective The aim of this study was to explore the therapeutic potential of combining hypoxia-preconditioned mesenchymal stem cell SH-MSC with alkaline water ...More |Related Solutions: μPulse^®

Abstract
Background:
Chronic inflammation and oxidative stress are central to the pathophysiology of Type 2 Diabetes Mellitus (T2DM), contributing to the progression of metabolic dysfunction and related complications.

Objective:
The aim of this study was to explore the therapeutic potential of combining hypoxia-preconditioned mesenchymal stem cell SH-MSC with alkaline water in a T2DM rat model.

Methods:
T2DM was induced in Wistar rats through a high-fat diet (HFD) followed by streptozotocin (STZ) administration. A total of 30 healthy male Wistar rats were randomly assigned to five groups: healthy control, T2DM, T2DM + Metformin, T2DM + SH-MSC, and T2DM + SH-MSC + alkaline water.

Results:
The combination of SH-MSC and alkaline water significantly reduced malondialdehyde (MDA) levels, a key indicator of lipid peroxidation, and suppressed the expression of p65 mRNA, a crucial component of the NF-κB signaling pathway. Notably, the most pronounced reduction in p65 mRNA expression was observed in the group receiving both SH-MSC and alkaline water, suggesting a synergistic effect in mitigating oxidative stress and inflammation.

Conclusion:
These findings highlight the potential of SH-MSC and alkaline water as a novel therapeutic strategy for alleviating T2DM. Less |Related Solutions: μPulse^®