Human anti-nucleolin recombinant immunoagent for cancer therapy

Abstract

Nucleolin (NCL) is a nucleocytoplasmic protein involved in many biological processes, such as ribosomal assembly, rRNA processing, and mRNA stabilization. NCL also regulates the biogenesis of specific microRNAs (miRNAs) involved in tumor development and aggressiveness. Interestingly, NCL is expressed on the surface of actively proliferating cancer cells, but not on their normal counterparts. Therefore, NCL is an attractive target for antineoplastic treatments. Taking advantage of phage-display technology, we engineered a fully human single-chain fragment variable, named 4LB5. This immunoagent binds NCL on the cell surface, it is translocated into the cytoplasm of target cells, and it abrogates the biogenesis of NCLdependent miRNAs. Binding of 4LB5 to NCL on the cell surface of a variety of breast cancer and hepatocellular carcinoma cell lines, but not to normal-like MCF-10a breast cells, dramatically reduces cancer cell viability and proliferation. Finally, in orthotopic breast cancer mouse models, 4LB5 administration results in a significant reduction of the tumor volume without evident side effects. In summary, here we describe, to our knowledge, the first anti-NCL single-chain fragment variable displaying antineoplastic activity against established solid tumors, which could represent the prototype of novel immunebased NCL-targeting drugs with clinical potential as diagnostic and therapeutic tools in a wide variety of human cancers.is one of the most abundant nonribosomal proteins in the nucleolus (1), first identified in ribosomal RNA processing (2). Additional studies have demonstrated that NCL is a multifunctional nucleocytoplasmic protein, involved in ribosomal assembly, chromatin decondensation, transcription, nucleo-cytoplasmic import/export, and chromatin remodeling (3, 4). NCL is frequently up-regulated in cancer and in cancerassociated endothelial cells compared with normal tissues (5, 6), where it is also present on the cell surface (7,8). Altered NCL expression and localization results in oncogenic effects, such as stabilization of AKT, Bcl-2, Bcl-XL, and IL-2 mRNAs (9-11). Moreover, surface-NCL acts as a receptor for several oncogenic ligands (12-15) and viruses (16). Recently, we reported that NCL has a critical protumorigenic function regulating the biogenesis of selected microRNAs (miRNAs), a class of noncoding single-stranded RNAs 19-22 nt in length (17) that regulate gene expression at the posttranscriptional level by targeting mRNAs in a sequence-specific manner (18). In fact, NCL enhances the maturation of specific miRNAs (including miR-21, miR-221, and miR-222) causally involved in cancer pathogenesis and resistance to several antineoplastic treatments (19-23). Our findings demonstrated that NCL modulates the biogenesis of these miRNAs at the posttranscriptional level, enhancing their maturation from pri-to premiRNAs, identifying a novel NCL-dependent oncogenic mechanism (19).Because of its oncogenic role and specific expression on cancer cells surface, NCL represents an attractive target f...

Druggable Nucleolin Identifies Breast Tumours Associated with Poor Prognosis That Exhibit Different Biological Processes

“…Overall, our data open up interesting opportunities in clinics. First, several promising anti-NCL drugs have been developed to fight liquid and solid cancers that are currently evaluated in clinical trials phase II [ 2 , 5 , 6 , 7 , 8 , 9 , 10 ]. Compared to intermediate and low NCL -expressing tumours, tumours expressing high NCL are the most prone to respond to anti-NCL drugs.…”

“…Interestingly, most of the anti-NCL therapies have been evaluated in vitro and in vivo using triple-negative breast cancer models and showed strong cytotoxic effects, making NCL a powerful candidate for future evaluation of anti-NCL in breast cancer [ 6 ]. Indeed, the antibody 4LB5 inhibiting NCL activity decreased MDA-MB-231 cell viability, clonogenicity, and tumour growth in xenografts while inducing apoptosis [ 8 ]. In addition, the peptides F3 and HB19 have been shown to significantly reduce tumour growth using MDA-MB-435 and MDA-MB-231 xenografts models, respectively [ 9 , 10 ].…”

Skeletal loading regulates breast cancer-associated osteolysis in a loading intensity-dependent fashion

“…Fibronectin is a high molecular weight glycoprotein that binds to integrins. 31 Nucleolin is known to be involved in cell differentiation, adhesion, inflammation and tumor development, 32 while Vimentin is an intermediate filament protein that is used as a marker of mesenchymal-derived cells. 33 In this study, we focused on the role of OPN in the responses to mechanical stimulation, since OPN is known to be involved in tumor progression.…”

Satellite-like Gold Nanocomposites for Targeted Mass Spectrometry Imaging of Tumor Tissues

“…Most human breast cancer cells overexpress nucleolin; however, MCF-7 cells show lower overexpression of nucleolin than that of MDA-MB-231 cells 45 , 64 . Reports show that, nucleolin located on the breast cell membrane acts as a receptor for several oncogenic ligands, and transformation in the expression and localization of nucleolin can induce many oncogenic effects, such as stabilization of AKT, Bcl-2, Bcl-XL, and IL-2 mRNAs 64 , 65 . Analysis of tumor cells by other mass spectrometry techniques require sample pretreatment, labeling and purification, and are time consuming, and difficult for specific targeting of tumor cells for quantitation.…”