Xiaobin Zhao

Targeted delivery systems for oligonucleotide therapeutics.

Oligonucleotides including antisense oligonucleotides and siRNA are emerging as promising therapeutic agents against a variety of diseases. Effective delivery of these molecules is critical to their successful clinical application. Targeted systems can greatly improve the efficiency and specificity of oligonucleotides delivery. Meanwhile, an effective delivery system must successfully overcome a multitude of biological barriers to enable the oligonucleotides to reach the site of action and access their biological targets. Several delivery strategies based on different platform technologies and different targeting ligands have been developed to achieve these objectives. This review aims at providing a summary and perspective on recent progress in this very active area of research.

Lenalidomide-mediated enhanced translation of C/EBPα-p30 protein up-regulates expression of the antileukemic microRNA-181a in acute myeloid leukemia

Recently, we showed that increased miR-181a expression was associated with improved outcomes in cytogenetically normal acute myeloid leukemia (CN-AML). Interestingly, miR-181a expression was increased in CN-AML patients harboring CEBPA mutations, which are usually biallelic and associate with better prognosis. CEBPA encodes the C/EBPα transcription factor. We demonstrate here that the presence of N-terminal CEBPA mutations and miR-181a expression are linked. Indeed, the truncated C/EBPα-p30 isoform, which is produced from the N-terminal mutant CEBPA gene or from the differential translation of wild-type CEBPA mRNA and is commonly believed to have no transactivation activity, binds to the miR-181a-1 promoter and up-regulates the microRNA expression. Furthermore, we show that lenalidomide, a drug approved for myelodysplastic syndromes and multiple myeloma, enhances translation of the C/EBPα-p30 isoform, resulting in higher miR-181a levels. In xenograft mouse models, ectopic miR-181a expression inhibits tumor growth. Similarly, lenalidomide exhibits antitumorigenic activity paralleled by increased miR-181a expression. This regulatory pathway may explain an increased sensitivity to apoptosis-inducing chemotherapy in subsets of AML patients. Altogether, our data provide a potential explanation for the improved clinical outcomes observed in CEBPA-mutated CN-AML patients, and suggest that lenalidomide treatment enhancing the C/EBPα-p30 protein levels and in turn miR-181a may sensitize AML blasts to chemotherapy.

A novel liposomal formulation of flavopiridol

Flavopiridol has shown promising activities in hematologic and solid tumor models, as well as in clinical trials in chronic lymphocytic leukemia patients. Flavopiridol has relatively low solubility and high plasma protein-binding. To address these issues and to provide an alternative strategy to achieve clinical efficacy, we encapsulated flavopiridol into a liposomal carrier and characterized its physicochemical and pharmacokinetic properties. The liposomes, comprising hydrogenated soy phosphatidylcholine (HSPC), cholesterol and poly (ethylene glycol) 2000-distearoyl phosphatidylethanolamine (PEG-DSPE), were prepared by polycarbonate membrane extrusion and then loaded with flavopiridol by a pH-gradient driven remote loading procedure. The liposomes had a mean diameter of 120.7 nm and a flavopiridol entrapment efficiency of 70.4%. Pharmacokinetic study in mice after i.v. bolus injection showed that the liposomal flavopiridol had an increased elimination phase half-life (T((1/2)beta), 339.7 min vs. 57.0 min), decreased clearance (CL, 0.012 L/min vs. 0.036 L/min), and increased area under the plasma concentration-time curve (AUC, 10.8 min micromol/L vs. 3.4 min micromol/L) compared to the free drug. This indicates a significant and potentially beneficial change in flavopiridol pharmacokinetics for the liposomal formulation. Further preclinical studies are warranted to define the toxicity and therapeutic efficacy of this novel formulation.

A novel Raji-Burkitt's lymphoma model for preclinical and mechanistic evaluation of CD52-targeted immunotherapeutic agents.

Purpose: To date, efforts to study CD52-targeted therapies, such as alemtuzumab, have been limited due to the lack of stable CD52 expressing transformed B-cell lines and animal models. We describe generation and utilization of cell lines that stably express CD52 both in vitro and in vivo. Experimental Design: By limiting dilution, we have established several clones of Raji-Burkitts lymphoma cell line that express surface CD52. Immunophenotype and cytogenetic characterization of these clones was done. In vivo usefulness of the CD52high cell line to evaluate the therapeutic efficacy of CD52-directed antibody was investigated using a SCID mouse xenograft model. Results: Stable expression of CD52 was confirmed in cells cultured in vitro up to 52 weeks of continuous growth. The functional integrity of the expressed CD52 molecule was shown using alemtuzumab, which induced cytotoxic effects in vitro in the CD52high but not the CD52low clone. Compared with control antibody, alemtuzumab treatment in CD52high inoculated mice resulted in significantly increased median survival. Comparable levels of CD52-targeted direct cytotoxicity, complement-dependent cytotoxicity, and antibody-dependent cytotoxicity and anti-CD52 immunoliposome-mediated delivery of synthetic oligodeoxyribo nucleotides in CD52high clone and primary B-chronic lymphocytic leukemia cells implicated potential in vivo application of this model for evaluation of CD52-targeted antibody and immunoliposomes encapsulating therapeutic agents. Conclusions: These results show the in vitro utility of the cloned Raji cell lines that stably express high levels CD52. The disseminated leukemia-lymphoma mouse model described herein using these stable cell lines can serve as an excellent system for in vivo therapeutic and mechanistic evaluation of existing and novel antibodies directed against CD52 molecule.