Therapeutic potential of CD22-specific antibody-targeted chemotherapy using inotuzumab ozogamicin (CMC-544) for the treatment of acute lymphoblastic leukemia.

CMC-544 (inotuzumab ozogamicin) is a CD22-specific cytotoxic immunoconjugate of calicheamicin intended for the treatment of B-lymphoid malignancies. This preclinical study investigated antitumor activity of CMC-544 against CD22+ acute lymphoblastic leukemia (ALL). CMC-544 inhibited in vitro growth of ALL cell lines more potently than that of Ramos B-lymphoma cells. When administered to nude mice with established sc xenografts of REH ALL, CMC-544 caused dose-dependent inhibition of xenograft growth producing complete tumor regression and cures in tumor-bearing mice at the highest dose of 160 microg/kg of conjugated calicheamicin. In contrast, a nonbinding control conjugate was 16-fold less effective than CMC-544 in inhibiting growth of REH ALL xenografts. When REH cells were injected intravenously in scid mice and allowed to disseminate systemically, mice developed hind-limb paralysis that was effectively prevented by treatment with CMC-544. Flow cytometric analysis of cells recovered from the bone marrow from mice with disseminated disease verified the presence of engrafted ALL cells. Significantly reduced numbers of ALL cells were recovered from the bone marrow of CMC-544-treated mice than from vehicle-treated mice with disseminated disease. The anti-leukemia activity of CMC-544 demonstrated here further supports clinical evaluation of CMC-544 for the treatment of CD22+ leukemia.

Angiotensin II receptors in human preadipocytes: role in cell cycle regulation.

The role of angiotensin II (AII) in human preadipocyte physiology has been investigated in primary cultures from human adipose tissue. Receptor binding studies indicated that human preadipocytes express a high affinity AII binding site of the AT1 subtype, as binding of 125I-labeled [Sar1,Ile8]AII was rapid, saturable, and specific. As AII has previously been demonstrated to affect the cell cycle in adrenal and cardiac cells, the effect of AII on regulation of cycle progression was examined in human preadipocytes. Stimulation of preadipocytes with AII resulted in G1 phase progression of the cell cycle, as determined by flow cytometric analysis. AII treatment was associated with induction of expression of the messenger RNA for the cell cycle regulatory protein cyclin D1 in a dose-dependent manner. Pretreatment of cells with subtype-selective AT receptor ligands before AII stimulation indicated that the cyclin response was mediated via the AT1 receptor. The identity of the cells as preadipocyte was verified by culture in a defined differentiation medium, observing both leptin message expression and triglyceride accumulation by flow cytometry. These findings indicate that AII has early, receptor-mediated effects on cell cycle progression in human preadipocytes that may contribute to differentiation to the adipocyte phenotype.

Distribution of angiotensin II receptors in rat and human adipocytes.

Angiotensin II (AII) receptor binding assays were performed in rat adipocytes from three separate anatomic depots. Fat cells were isolated by collagenase digestion, and plasma membranes were prepared from the epididymal, mesenteric, and retroperitoneal fat depots of male Sprague-Dawley rats at 100 days of age. Binding of 125I-labeled [Sar1,Ile8]AII was rapid, saturable, and specific in membranes from all depots, identifying a receptor with a similar affinity of approximately 1 nM. Site-associated differences in receptor number were observed, with epididymal and mesenteric fat cell membranes exhibiting significantly more receptors than retroperitoneal fat cells when binding was expressed per unit of membrane protein. When corrected for cell volume, the number of receptors per cell ranked epididymal > retroperitoneal > mesenteric. Inhibitory constants for the peptide agonists AII and AIII and the peptide antagonist [Sar1,Ala8]AII indicated similar affinities in all three depots. Because the receptor has been classified pharmacologically into two subtypes, the AT1 selective antagonist losartan, and the AT2 selective antagonist PD 123,319 were used to classify the adipocyte receptor, indicating an AT1 subtype with an affinity for losartan in the mesenteric and retroperitoneal adipocytes that was significantly greater than the epididymal. Similar studies were performed in adipocyte membranes obtained from human omental and subcutaneous adipose tissue, revealing the presence of an AII receptor in both depots with an affinity of approximately 10 nM for losartan. These data indicate site-specific differences in AII receptor number in fat cell membranes from rats and the existence of human adipocyte AII receptors, suggesting that the adipocyte is significant for the peripheral metabolism of components of the renin-angiotensin system.

Identification of the KDR tyrosine kinase as a receptor for vascular endothelial cell growth factor.

Vascular endothelial cell growth factor (VEGF), also known as vascular permeability factor, is an endothelial cell mitogen which stimulates angiogenesis. Here we report that a previously identified receptor tyrosine kinase gene, KDR, encodes a receptor for VEGF. Expression of KDR in CMT-3 (cells which do not contain receptors for VEGF) allows for saturable 125I-VEGF binding with high affinity (KD = 75 pM). Affinity cross-linking of 125I-VEGF to KDR-transfected CMT-3 cells results in specific labeling of two proteins of M(r) = 195 and 235 kDa. The KDR receptor tyrosine kinase shares structural similarities with a recently reported receptor for VEGF, flt, in a manner reminiscent of the similarities between the alpha and beta forms of the PDGF receptors.