An atlas of bloodstream-accessible bone marrow proteins for site-directed therapy of acute myeloid leukemia

Abstract

The concept of arming antibodies with bioactive payloads for a site-specific therapy of cancer has gained considerable interest in recent years. However, a successful antibody-based targeting approach critically relies on the availability of a tumor-associated target that is not only preferentially expressed in the tumor tissue but is also easily accessible for antibody therapeutics coming from the bloodstream. Here, we perfused the vasculature of healthy and acute myeloid leukemia (AML)-bearing rats with a reactive ester derivative of biotin and subsequently quantified the biotinylated proteins to identify AML-associated bone marrow (BM) antigens accessible from the bloodstream. In total, >1400 proteins were identified. Overall, 181 proteins were >100-fold overexpressed in AML as compared with normal BM. Eleven of the most differentially expressed proteins were further validated by immunohistochemistry and confocal microscopic analyses, including novel antigens highly expressed in AML cells (for example, adaptor-related protein complex 3 β2) and in the leukemia-modified extracellular matrix (ECM) (for example, collagen-VI-α-1). The presented atlas of targetable AML-associated BM proteins provides a valuable basis for the development of monoclonal antibodies that could be used as carriers for a site-specific pharmacodelivery of cytotoxic drugs, cytokines or radionuclides to the BM in AML.

A mass spectrometry guided approach for the identification of novel vaccine candidates in gram-negative pathogens

“…Next, we tested experimental approaches to biochemically capture bacterial cell surface exposed proteins. We evaluated biotinylation of surface proteins and streptavidin purification 14 , which is the most frequently applied technology for surfome identification 15–18 , and identified 565 proteins (Supplementary Fig. 2).…”

“…However, additional studies separating vesicles by filtration or density centrifugation are required to test that hypothesis. In comparison to widely used cell surface biotin labelling 15–18 , our approach results in lower cytosolic background detection for H. pylori and presumably also other gram-negative pathogens. However, DoE based experimental optimization of surface biotinylation with minimal co-labeling of cytosolic proteins may present an interesting future research strategy.…”

Low‐density lipoprotein receptor (LDLR) is an independent adverse prognostic factor in acute myeloid leukaemia

“…Two representative 1‐mm cores from areas of leukaemic infiltration were arrayed per sample. Immunohistochemical staining was performed as described 11,21,22 . Briefly, following de‐paraffinisation and heat‐induced epitope unmasking, 4‐µm tissue sections were incubated with a primary anti‐LDLR antibody (ab30532; Abcam, Cambridge, UK), followed by suitable secondary and tertiary antibodies (Dako, Carpinteria, CA, USA).…”

“…Immunohistochemical staining was performed as described. 11,21,22 Briefly, following de-paraffinisation and heat-induced epitope unmasking, 4µm tissue sections were incubated with a primary anti-LDLR antibody (ab30532; Abcam, Cambridge, UK), followed by suitable secondary and tertiary antibodies (Dako, Carpinteria, CA, USA). Immunoreactions were visualised with a monoclonal alkaline phosphatase-anti-alkaline phosphatase (APAAP) complex and a fuchsin-based substrate-chromogen system (Dako).…”

“…In addition, there were low protein expression values in healthy organs except for the adrenal glands and some fetal tissues. Thus, LDLR might be an attractive target for the specific delivery of bioactive payloads, such as cytotoxic drugs, to the leukaemic blasts using antibody-drug conjugates, 21,22,35 especially in the context of sAML. Bispecific T-cell engagers and chimeric antigen receptor T cells targeting LDLR on AML blasts represent further potential therapeutic applications.…”