CMC Regulatory Considerations for Antibody-Drug Conjugates.

Antibody-drug conjugates unite the specificity and long circulation time of an antibody with the toxicity of a chemical cytostatic or otherwise active drug using appropriate chemical linkers to reduce systemic toxicity and increase therapeutic index. This combination of a large biological molecule and a small molecule creates an increase in complexity. Multiple production processes are required to produce the native antibody, the drug and the linker, followed by conjugation of afore mentioned entities to form the final antibody-drug conjugate.

Purification of the extracellular domain of the membrane protein GlialCAM expressed in HEK and CHO cells and comparison of the glycosylation.

Adhesion molecules are essential for a wide range of biological and physiological functions, including cell-cell interactions, cell interactions with the extracellular matrix, cell migration, proliferation and survival. Defects in cell adhesion have been associated with pathological conditions such as neoplasia, and neurodegenerative diseases. We have identified a new adhesion molecule of the immunoglobulin family, GlialCAM.

Tetracycline-regulated expression of VEGF-A in beta cells induces angiogenesis: improvement of engraftment following transplantation.

Early revascularization of pancreatic islet cells after transplantation is crucial for engraftment, and it has been suggested that vascular endothelial growth factor-A (VEGF-A) plays a significant role in this process. Although VEGF gene therapy can improve angiogenesis, uncontrolled VEGF secretion can lead to vascular tumor formation. Here we have explored the role of temporal VEGF expression, controlled by a tetracycline (TC)-regulated promoter, on revascularization and engraftment of genetically modified beta cells following transplantation.

Bovine microvascular endothelial cells immortalized with human telomerase.

Primary cultures of bovine microvascular endothelial cells (BME) isolated from the adrenal cortex, are commonly used to study vascular endothelium, but have a limited life span. To circumvent these limitations, we have immortalized BME cells with either simian virus 40 (SV40) or with a retrovirus containing the coding region of human telomerase reverse transcriptase (hTERT), and have investigated whether the clonal populations obtained, maintain differentiated properties characteristic of microvascular endothelium. Immortalized cells were characterized for maintenance of typical endothelial morphology, marker expression, and functional characteristics including uptake of Acetylated low-density lipoprotein (Ac-LDL), capillary-like tube formation in three-dimensional collagen gels, as well as metalloproteinase (MMP) and plasminogen activator (PA)-mediated extracellular proteolysis.

Generation and characterization of telomerase-transfected human lymphatic endothelial cells with an extended life span.

The study of lymphatic endothelial cells and lymphangiogenesis has, in the past, been hampered by the lack of lymphatic endothelial-specific markers. The recent discovery of several such markers has permitted the isolation of lymphatic endothelial cells (LECs) from human skin. However, cell numbers are limited and purity is variable with the different isolation procedures.

Molecular pathways involved in the antineoplastic effects of calcitriol on insulinoma cells.

We have previously reported that in tumorigenic pancreatic beta-cells, calcitriol exerts a potent antitumorigenic effect by inducing apoptosis, cell growth inhibition, and reduction of solid beta-cell tumors. Here we have studied the molecular pathways involved in the antineoplastic activity of calcitriol on mouse insulinoma beta TC(3) cells, mouse insulinoma beta TC expressing or not expressing the oncogene p53, and beta TC-tet cells overexpressing or not the antiapoptotic gene Bcl2. Our results indicate that calcitriol-induced apoptosis was dependent on the function of p53 and was associated with a biphasic increase in protein levels of transcription factor nuclear factor-kappa B.

Allogeneic beta-islet cells correct diabetes and resist immune rejection.

Allogeneic MHC-incompatible organ or cell grafts are usually promptly rejected by immunocompetent hosts. Here we tested allogeneic beta-islet cell graft acceptance by immune or naive C57BL/6 mice rendered diabetic with streptozotocin (STZ). Fully MHC-mismatched insulin-producing growth-regulated beta-islet cells were transplanted under the kidney capsule or s.

cFLIP protein prevents tumor necrosis factor-alpha-mediated induction of caspase-8-dependent apoptosis in insulin-secreting betaTc-Tet cells.

Type 1 diabetes is characterized by the infiltration of activated leukocytes within the pancreatic islets, leading to beta-cell dysfunction and destruction. The exact role played by interferon-gamma, tumor necrosis factor (TNF)-alpha, and interleukin-1beta in this pathogenic process is still only partially understood. To study cytokine action at the cellular level, we are working with the highly differentiated insulin-secreting cell line, betaTc-Tet.