Antibodies against Anthrax Toxins: A Long Way from Benchlab to the Bedside.

Anthrax is an acute disease caused by the bacterium Bacillus anthracis, and is a potential biowarfare/bioterrorist agent. Its pulmonary form, caused by inhalation of the spores, is highly lethal and is mainly related to injury caused by the toxins secretion. Antibodies neutralizing the toxins of B. anthracis are regarded as promising therapeutic drugs, and two are already approved by the Federal Drug Administration. We developed a recombinant human-like humanized antibody, 35PA83 6.20, that binds the protective antigen and that neutralized anthrax toxins in-vivo in White New Zealand rabbits infected with the lethal 9602 strain by intranasal route. Considering these promising results, the preclinical and clinical phase one development was funded and a program was started. Unfortunately, after 5 years, the preclinical development was cancelled due to industrial and scientific issues. This shutdown underlined the difficulty particularly, but not only, for an academic laboratory to proceed to clinical development, despite the drug candidate being promising. Here, we review our strategy and some preliminary results, and we discuss the issues that led to the no-go decision of the pre-clinical development of 35PA83 6.20 mAb. Our review provides general information to the laboratories planning a (pre-)clinical development.

Preclinical study of Ublituximab, a Glycoengineered anti-human CD20 antibody, in murine models of primary cerebral and intraocular B-cell lymphomas.

PURPOSE: Primary cerebral lymphoma (PCL) and primary intraocular lymphoma (PIOL) belong to the systemic diffuse large B-cell lymphoma family and are characterized by the presence of CD20(+) lymphoma B cells in the brain or the eye. These highly aggressive malignancies have a poor prognosis and no specific therapy. The presence of effector immune cells in the damaged brain and vitreous suggests that treatment with anti-human CD20 (hCD20) monoclonal antibodies might be effective. We developed murine models of PCL and PIOL to assess the intracerebral and intraocular antitumor effect of ublituximab, a promising glycoengineered anti-hCD20 mAb with a high affinity for FcγRIIIa (CD16) receptors. METHODS: The murine lymphoma B-cell line A20.IIA-GFP-hCD20 (H-2(d)) was injected into the right cerebral striatum or the vitreous of immunocompetent adult BALB/c mice (H-2(d)). Four to 7 days later, ublituximab was injected intracerebrally or intravitreously into the tumor site. Rituximab was the reference compound. Survival was monitored for injected mice; histopathological and flow cytometric analyses were performed to study tumor growth and T-cell infiltration. RESULTS: Single doses of ublituximab, injected intracerebrally or intravitreously, had a marked antitumor effect, more pronounced than that obtained with the same dose of rituximab in these conditions. The reduction in tumor cells was correlated with an increased proportion of CD8(+) T cells. This efficacy was observed only against lymphoma B cells expressing hCD20. CONCLUSIONS: These in vivo results confirm the potential of the glycoengineered anti-hCD20 mAb ublituximab as an innovative therapeutic approach to treat primary central nervous system lymphoma and other B-cell lymphomas.

The SHP-1 protein tyrosine phosphatase negatively modulates glucose homeostasis.

The protein tyrosine phosphatase SHP-1 is a well-known inhibitor of activation-promoting signaling cascades in hematopoietic cells but its potential role in insulin target tissues is unknown. Here we show that Ptpn6(me-v/me-v) (also known as viable motheaten) mice bearing a functionally deficient SHP-1 protein are markedly glucose tolerant and insulin sensitive as compared to wild-type littermates, as a result of enhanced insulin receptor signaling to IRS-PI3K-Akt in liver and muscle. Downregulation of SHP-1 activity in liver of normal mice by adenoviral expression of a catalytically inert mutant of SHP-1, or after small hairpin RNA-mediated SHP-1 silencing, further confirmed this phenotype. Tyrosine phosphorylation of CEACAM1, a modulator of hepatic insulin clearance, and clearance of serum [125I]-insulin were markedly increased in SHP-1-deficient mice or SHP-1-deficient hepatic cells in vitro. These findings show a novel role for SHP-1 in the regulation of glucose homeostasis through modulation of insulin signaling in liver and muscle as well as hepatic insulin clearance.

Distinct Rho GTPase activities regulate epithelial cell localization of the adhesion molecule CEACAM1: involvement of the CEACAM1 transmembrane domain.

CEACAM1 is an intercellular adhesion glycoprotein. As CEACAM1 plays an important role in epithelial cell signaling and functions, we have examined its localization in epithelial cells. We have observed that distribution at cell contacts is not always seen in these cells, suggesting that CEACAM1 localization might be regulated. In Swiss 3T3 cells, the targeting of CEACAM1 at cell-cell boundaries is regulated by the Rho GTPases. In the present study, we have used the MDCK epithelial cells to characterize the effects of the Rho GTPases and their effectors on CEACAM1 intercellular targeting. Activated Cdc42 and Rac1 or their downstream effector PAK1 targeted CEACAM1 to sites of cell-cell contacts. On the other hand, neither activated RhoA nor activated Rho kinase directed CEACAM1 to cell boundaries, resulting in a condensed distribution of CEACAM1 at the cell surface. Interestingly, inhibition of this pathway resulted in CEACAM1 intercellular localization suggesting that a tightly regulated balance of Rho GTPase activities is necessary to target CEACAM1 at cell-cell boundaries. In addition, using CEACAM1 mutants and chimeric fusion constructs containing domains of the colony-stimulating factor receptor, we have shown that the transmembrane domain of CEACAM1 is responsible for the Cdc42-induced targeting at cell-cell contacts.