Building and characterizing antibody-targeted lipidic nanotherapeutics.
ABSTRACT Immunoliposomes provide a complementary, and in many instances advantageous, drug delivery strategy to antibody-drug conjugates. Their high carrying capacity of 20,000-150,000 drug molecules/liposome, allows for the use of a significantly broader range of moderate-to-high potency small molecule drugs when compared to the comparably few subnanomolar potency maytansinoid- and auristatin-based immunoconjugates. The multivalent display of 5-100 antibody fragments/liposome results in an avidity effect that can make use of even moderate affinity antibodies, as well as a cross-linking of cell surface receptors to induce the internalization required for intracellular drug release and subsequent activity. The underlying liposomal drug must be effectively engineered for long circulating pharmacokinetics and stable in vivo drug retention in order to allow for the drug to be efficiently delivered to the target tissue and take advantage of the site-specific bioavailability provided for by the targeting arm. In this chapter, we describe the rationale for engineering stable immunoliposome-based therapeutics, methods required for preparation of immunoliposomes, as well as for their physicochemical and in vivo characterization.
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ABSTRACT: The 22nd Annual Antibody Engineering and 9th Annual Antibody Therapeutics international conferences, and the 2011 Annual Meeting of The Antibody Society, organized by IBC Life Sciences with contributions from The Antibody Society and two Scientific Advisory Boards, were held December 5-8, 2011 in San Diego, CA. The meeting drew ~800 participants who attended sessions on a wide variety of topics relevant to antibody research and development. As a preview to the main events, a pre-conference workshop held on December 4, 2011 focused on antibodies as probes of structure. The Antibody Engineering Conference comprised eight sessions: (1) structure and dynamics of antibodies and their membrane receptor targets; (2) model-guided generation of binding sites; (3) novel selection strategies; (4) antibodies in a complex environment: targeting intracellular and misfolded proteins; (5) rational vaccine design; (6) viral retargeting with engineered binding molecules; (7) the biology behind potential blockbuster antibodies and (8) antibodies as signaling modifiers: where did we go right, and can we learn from success? The Antibody Therapeutics session comprised five sessions: (1)Twenty-five years of therapeutic antibodies: lessons learned and future challenges; (2) preclinical and early stage development of antibody therapeutics; (3) next generation anti-angiogenics; (4) updates of clinical stage antibody therapeutics and (5) antibody drug conjugates and bispecific antibodies.mAbs 03/2012; 4(2). DOI:10.4161/mabs.4.2.19495 · 4.73 Impact Factor
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ABSTRACT: The urokinase system is overexpressed in epithelial ovarian cancer (OvCa) cells and is expressed at low levels in normal cells. To develop a platform for intracellular and targeted delivery of therapeutics in OvCa, we conjugated urokinase plasminogen activator (uPA) antibodies to liposomal nanobins. The arsenic trioxide loaded nanobins had favorable physicochemical properties and the ability to bind specifically to uPA. Confocal microscopy showed that the uPA targeted nanobins were internalized by OvCa cells, while both ICP-MS and FACS analyses confirmed >4-fold higher uptake of targeted nanobins when compared to untargeted nanobins. In a co-culture assay, the targeted nanobins showed efficient uptake in OvCa cells but not in the normal primary omental mesothelial cells. Moreover, this uptake could be blocked by either down-regulating uPA receptor expression in the OvCa cells using shRNA or by competition with free uPA or uPA antibody. In proof-of-concept experiments, mice bearing orthotopic ovarian tumors showed a greater reduction in tumor burden when treated with targeted nanobins than with untargeted nanobins (47% versus 27%; p<0.001). The targeted nanobins more effectively inhibited tumor cell growth both in vitro and in vivo compared to untargeted nanobins, inducing caspase-mediated apoptosis and impairing stem cell marker, ALDH1A1, expression. Ex vivo fluorescence imaging of tumors and organs corroborated these results, showing preferential localization of the targeted nanobins to the tumor. These findings suggest that uPA targeted nanobins capable of specifically and efficiently delivering payloads to cancer cells could serve as the foundation for a new targeted cancer therapy utilizing protease receptors.Molecular Cancer Therapeutics 09/2013; 12(12). DOI:10.1158/1535-7163.MCT-13-0204 · 6.11 Impact Factor
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ABSTRACT: Many proteins of interest in basic biology, translational research studies and for clinical targeting in diseases reside inside the cell and function by interacting with other macromolecules. Protein complexes control basic processes such as development, cell division but also abnormal cell growth when mutations occur such as found in cancer. Interfering with protein-protein interactions is an important aspiration in both basic and disease biology but small molecule inhibitors have been difficult and expensive to isolate. Recently, we have adapted molecular biology techniques to develop a simple set of protocols for isolation of high affinity antibody fragments (in the form of single VH domains) that function within the reducing environment of higher organism cells and can bind to their target molecules. The method called Intracellular Antibody Capture (IAC) has been used to develop inhibitory anti-RAS and anti-LMO2 single domains that have been used for target validation of these antigens in pre-clinical cancer models and illustrate the efficacy of the IAC approach to generation of drug surrogates. Future use of inhibitory VH antibody fragments as drugs in their own right (we term these macrodrugs to distinguish them from small molecule drugs) requires their delivery to target cells in vivo but they can also be templates for small molecule drug development that emulate the binding sites of the antibody fragments. This article is part of a Special Issue entitled: Recent advances in molecular engineering of antibody.Biochimica et Biophysica Acta (BBA) - Proteins & Proteomics 01/2014; · 3.19 Impact Factor