Something new about prognostic factors for lupus nephritis? A systematic review.

BACKGROUND: Lupus nephritis (LN) affects 30-45% of patients with systemic lupus erythematosus (SLE) and causes great morbidity and mortality. About 10-25% of patients will develop chronic kidney disease (CKD), and it has been described a mortality of 10-20% at 10 years. The contribution of clinical and biological markers to the prediction of outcome is unclear. OBJECTIVE: To describe the factors, with measures of association, that predict the main outcomes of LN. MATERIAL AND METHODS: We have conducted a systematic review. Medline, Embase, and Cochrane Library were systematic searched from inception up to Oct 2019, with a strategy that included synonyms of all targeted outcomes of LN: (kidney failure, response to treatment, cardiovascular events, and mortality). Only studies with longitudinal prospective design or with warranties of unbiased recollection of the prognostic factors, where LN was confirmed by biopsy were included. Risk of bias was assessed with the New Castle Ottawa scale. Predictive factors and their effect measures were collected from each study. RESULTS: From 1221 studies identified, 25 studies were included, of which 15 were retrospective, nine prospective, and one was a trial extension study (range from 3 months to 11 years). The main predictive factors of renal response were serum creatinine (SCr) and glomerular filtration rate C3 levels, titer of anti-C1q, and anti-dsDNA antibodies. Renal histological findings such as class type (IV or V), tubulointerstitial or vascular lesions and chronicity index were risk factors for development of chronic kidney disease. The factors associated with persistence of activity were proteinuria, anti-dsDNA, anticardiolipin, anti C1q antibodies, and complement values. The factors associated to cardiovascular events and mortality were age, smoking, amount of proteinuria, and histological findings, such as vascular lesions. Meta-analysis was precluded given the heterogeneity of designs definitions and effect measures. CONCLUSIONS: Nowadays, we do not have new biomarkers that establish the renal prognosis of patients with LN. Classical clinical, renal, and histological markers are used in most studies. It is worth noting the heterogeneity of studies in the definition of renal outcomes, which complicates risk stratification in these patients.

The interaction of anti-DNA antibodies with DNA antigen: Evidence for hysteresis for high avidity binding.

Antibodies to DNA (anti-DNA) are the serological hallmark of systemic lupus erythematosus. Previous studies have indicated that the phosphodiester backbone is the main antigenic target, with electrostatic interactions important for high avidity. To define further these interactions, the effects of ionic strength on anti-DNA binding of SLE plasmas were assessed in association and dissociation assays by ELISA. As these studies demonstrated, increasing ionic strength to a concentration of 1000 mM NaCl reduced antibody binding although the extent of the reduction varied among samples. In dissociation assays, differences among plasmas were also observed. For one of the plasmas, binding to DNA displayed resistance to dissociation by increasing ionic strength even though these concentrations limited binding in association assays. Time course studies showed a gradual change in binding interactions. These studies indicate that anti-DNA binding can involve both electrostatic and non-electrostatic interactions, with binding in some plasmas showing evidence of hysteresis.

Defective Efferocytosis in a Murine Model of Sjögren's Syndrome Is Mediated by Dysfunctional Mer Tyrosine Kinase Receptor.

Sjögren's syndrome (SjS) is a chronic autoimmune disease primarily involving the exocrine glands in which the involvement of the innate immune system is largely uncharacterized. Mer signaling has been found to be protective in several autoimmune diseases but remains unstudied in SjS. Here, we investigated the role of Mer signaling in SjS. Mer knockout (MerKO) mice were examined for SjS disease criteria. SjS-susceptible (SjSS) C57BL/6.NOD-Aec1Aec2 mice were assessed for defective Mer signaling outcomes, soluble Mer (sMer) levels, A disintegrin and metalloprotease 17 (ADAM17) activity, and Rac1 activation. In addition, SjS patient plasma samples were evaluated for sMer levels via ELISA, and sMer levels were correlated to disease manifestations. MerKO mice developed submandibular gland (SMG) lymphocytic infiltrates, SMG apoptotic cells, anti-nuclear autoantibodies (ANA), and reduced saliva flow. Mer signaling outcomes were observed to be diminished in SjSS mice, as evidenced by reduced Rac1 activation in SjSS mice macrophages in response to apoptotic cells and impaired efferocytosis. Increased sMer was also detected in SjSS mouse sera, coinciding with higher ADAM17 activity, the enzyme responsible for cleavage and inactivation of Mer. sMer levels were elevated in patient plasma and positively correlated with focus scores, ocular staining scores, rheumatoid factors, and anti-Ro60 levels. Our data indicate that Mer plays a protective role in SjS, similar to other autoimmune diseases. Furthermore, we suggest a series of events where enhanced ADAM17 activity increases Mer inactivation and depresses Mer signaling, thus removing protection against the loss of self-tolerance and the onset of autoimmune disease in SjSS mice.

Harnessing SLE Autoantibodies for Intracellular Delivery of Biologic Therapeutics.

Intracellular delivery of therapeutic antibodies is highly desirable but remains a challenge for biomedical research and the pharmaceutical industry. Approximately two-thirds of disease-associated targets are found inside the cell. Difficulty blocking these targets with available drugs creates a need for technology to deliver highly specific therapeutic antibodies intracellularly. Historically, antibodies have not been believed to traverse the cell membrane and neutralize intracellular targets. Emerging evidence has revealed that anti-DNA autoantibodies found in systemic lupus erythematosus (SLE) patients can penetrate inside the cell. Harnessing this technology has the potential to accelerate the development of drugs against intracellular targets. Here, we dissect the mechanisms of the intracellular localization of SLE antibodies and discuss how to apply these insights to engineer successful cell-penetrating antibody drugs.

Blockade of anti-dsDNA ameliorates systemic lupus erythematosus in MRL/Faslpr mice through ameliorating inflammation via the PKCδ-NLRC4 axis.

Anti-double-stranded DNA (anti-dsDNA) is closely associated with the inflammatory burden in the brain after ischemic stroke. Here, we studied the inflammatory cascade and investigated the mechanisms behind the pro-inflammatory role of dsDNA in systemic lupus erythematosus (SLE). The serum levels of interleukin-1beta (IL-1ß) and IL-6 in SLE patients and the corresponding controls were evaluated using ELISA, and the expression level of caspase-1 was evaluated using quantitative real-time polymerase chain reaction (qRT-PCR). We found that the serum levels of IL-1ß and IL-6 were increased in the SLE patients. The expression of caspase-1 was upregulated and positively correlated with the levels of pro-inflammatory factors. The level of anti-dsDNA was also elevated and positively correlated with the results for the mean fluorescence intensity (MFI) of caspase-1. Additionally, we evaluated the functions of PRKCD encoding protein kinase c delta (PKCδ) and NLRC4, in vivo, in MRL/Faslpr mice. We found that renal injury was aggravated, and the levels of pro-inflammatory factors were increased in the MRL/Faslpr mice. We also found that increased levels of NLRC4 in the mice exacerbated renal injury and increased the levels of pro-inflammatory factors, whereas inhibition of PKCδ had the opposite results. These findings provide unique perspectives on pathogenesis of SLE and indicate that inhibition of anti-dsDNA could attenuate renal inflammatory burden, representing a promising therapeutic opportunity for SLE.

Bisphenol A modified DNA: A possible immunogenic stimulus for anti-DNA autoantibodies in systemic lupus erythematosus.

Anti-DNA antibodies are now considered as a universal diagnostic feature for the patients with systemic lupus erythematosus (SLE) but the mechanism(s) involved in the generation of these autoantibodies remains to be investigated. Bisphenol A (BPA) is a synthetic phenol extensively used in the manufacturing of polycarbonated plastics. Upon mixing in the diet, it causes several health hazards. This study was undertaken to investigate the contribution of BPA induced DNA damage in SLE patients. Human DNA was modified by BPA in-vitro and the binding characteristics of SLE circulating immunoglobulin Gs (SLE-IgGs) with BPA damaged DNA (BPA-DNA) were screened and compared with the IgGs from normal healthy humans (NH-IgGs). Immunogenicity of BPA-DNA was determined by immunisation in rabbits. DNA from SLE patients (SLE-DNA) or healthy humans (NH-DNA) were isolated and their binding specificity with rabbit anti-BPA-DNA-IgGs was studied. Treatment of human DNA with BPA caused extensive damaged. Circulating SLE-IgGs showed strong recognition of BPA-DNA. BPA-DNA induced high titre antibodies in rabbits. Rabbit anti-BPA-DNA-IgGs showed strong cross reaction with isolated DNA from SLE patients. In short, we concluded that the structural alterations in DNA by BPA, generate neo-epitopes that may be a factor responsible for the induction of anti-DNA autoantibodies in SLE.

CytoLyt fixation significantly inhibits MIB1 immunoreactivity whereas alternative Ki-67 clone 30-9 is not susceptible to the inhibition: Critical diagnostic implications.

BACKGROUND: The Ki-67 proliferation marker has multiple diagnostic and prognostic applications. Although several clones to the Ki-67 antigen are commercially available, the MIB1 clone is widely recommended in the surgical pathology literature for neuroendocrine tumors. In our cytopathology practice, we have encountered unexpectedly low MIB1 immunoreactivity in CytoLyt-fixed cell blocks (CBs). The current study evaluated the impact of fixatives, CB processing, and immunocytochemical (ICC) procedures on Ki-67 immunoreactivity. METHODS: Test CBs were prepared from freshly resected tumors, and multiple variables in the MIB1 ICC procedure were tested, including CytoLyt versus formalin collection media, MIB1 versus other Ki-67 clones including 30-9, and other variables. MIB1 versus Ki-67 30-9 clones were tested in parallel on CytoLyt-fixed CBs from clinical samples of small cell lung carcinoma (SCLC). RESULTS: In the test CBs (n = 10), the mean MIB1 labeling index was 10% in CytoLyt versus 47% in formalin (P = .0116), with a mean loss of reactivity in matched CBs of 37% (up to 70%). None of the procedure modifications tested in 223 individual ICC reactions recovered MIB1 reactivity in CytoLyt except for switching to the Ki-67 30-9 antibody. In CytoLyt-fixed SCLC samples (n = 14), the Ki-67 30-9 antibody demonstrated expected ranges of reactivity (mean, 83%; range, 60%-100%), whereas MIB1 demonstrated markedly inhibited labeling (mean, 60%; range, 10%-95%) (P = .0058). CONCLUSIONS: CytoLyt fixation substantially inhibits MIB1 immunoreactivity, whereas the Ki-67 30-9 clone is not susceptible to inhibition. Markedly discrepant MIB1 reactivity may present a pitfall in the diagnosis of SCLC and may lead to the incorrect prognostic stratification of other tumor types. For laboratories using CytoLyt, we recommend using the Ki-67 30-9 antibody rather than the MIB1 antibody.

Two ways to inactivate the Ki-67 protein-Fragmentation by nanoparticles, crosslinking with fluorescent dyes.

Light can manipulate molecular biological processes with high spatial and temporal precision and optical manipulation has become increasingly popular during the last years. In combination with absorbing dyes or gold nanoparticles light is a valuable tool for cell and protein inactivation with high precision. Here we show distinct differences in the underlying mechanisms whether gold nanoparticles or fluorescent dyes are used for the inactivation of the Ki-67 protein. The proliferation-associated protein Ki-67 was addressed by the antibody MIB-1. In vitro studies showed a fragmentation of the Ki-67 protein after laser irradiation of 15 nm gold nanoparticle antibody conjugates with nanosecond pulsed laser, while continuous wave (cw) irradiation of fluorescein isothiocyanate (FITC)- and Alexa 488-labeled antibodies led to specific crosslinking of Ki-67. The irradiation energy for the gold nanoparticles was above cavitation bubble formation threshold. We observed a fragmentation of the target protein and also of the gold particles. The understanding of the underlying inactivation mechanisms is important for the application and further development of these two techniques, which can harness nanotechnology to introduce molecular selectivity to biological systems.

Anti-Ro52 antibody acts on the S5-pore linker of hERG to chronically reduce channel expression.

AIMS: The human ether-a-go-go-related gene (hERG) encodes the rapidly activating delayed rectifier potassium channel (IKr). Malfunction of hERG/IKr is the primary cause of acquired long QT syndrome (LQTS), an electrical disorder of the heart that can cause arrhythmias and sudden death. Patients with autoimmune diseases display a high incidence of LQTS. While dysfunction of hERG channels induced by autoantibodies such as anti-Ro52 may play a role in this pathology, the underlying mechanisms are not well understood. Here, we investigated the acute and chronic effects of anti-Ro52 antibody on hERG channels stably expressed in human embryonic kidney (hERG-HEK) 293 cells as well as IKr in neonatal rat ventricular myocytes. METHODS AND RESULTS: Using whole-cell patch clamp, western blot analyses, and immunocytochemistry, we found that a 12-h treatment of hERG-HEK cells with patients' sera containing anti-Ro52 autoantibody decreased the hERG current (IhERG) by 32% compared to cells treated with autoantibody-negative patients' sera. Commercial anti-Ro52 antibody at 100 µg/mL did not acutely block IhERG. Instead, a 12-h treatment with anti-Ro52 antibody at a concentration of 4 µg/mL significantly reduced mature hERG protein expression and IhERG. Specifically, anti-Ro52 antibody did not acutely block hERG current but chronically facilitated hERG endocytic degradation. The extracellular S5-pore linker of hERG, which forms the turret of the channel on the outside of the cell, is the target region for anti-Ro52-mediated hERG reduction since its replacement with the analogous region of EAG abolished the anti-Ro52 effect. In neonatal rat ventricular myocytes, 100 µg/mL anti-Ro52 antibody did not acutely block IKr, but a 12-h treatment of cells with 4 µg/mL anti-Ro52 antibody selectively reduced IKr and prolonged the action potential duration. CONCLUSIONS: Our results indicate that anti-Ro52 antibody acts on the hERG S5-pore linker to chronically decrease hERG expression and current. These findings provide novel insights into hERG regulation and anti-Ro52 antibody-associated LQTS.

Why do some ME/CFS patients benefit from treatment with sodium dichloroacetate, but others do not?

Myalgic Encephalopathy/Chronic Fatigue Syndrome (ME/CFS) is an enigmatic disease the pathogenesis of which remains elusive. Pragmatic proof-of-principle of the hypothetical mechanisms causing the clinical symptoms has been delivered, but it is hard to explain why some patients do respond favourably to treatment with sodium dichloroacetate (DCA), which enhances the activity of the mitochondrial enzyme pyruvate dehydrogenase, but other patients experience no benefit from this substance. In a prospective trial including 35 ME/CFS patients, logistic regression analysis with stepwise elimination has identified 6 pre-treatment characteristics allowing for the differentiation between responders (n = 13) and non-reponders (n = 22) with high accuracy (P < 0.0001; area under the ROC-curve = 0.92). A formula was derived generating the probability of belonging to the group of responders. This finding may assist in selecting ME/CFS patients suitable for treatment with DCA, but requires further studies as to the predictive capacity of the derived formula.

Fast RBC loading by fluorescent antibodies and nuclei staining dye and their potential bioanalytical applications.

This study was designed to load different antibodies (Abs) and a fluorescent dye onto the red blood cell (RBC) surface. We have used fluorescein isothiocyanate (FITC)-conjugate anti-human Ab, CD22-PE (B-cell marker-phycoerythrin Ab), and 4',6-diamidino-2-phenylindole (DAPI) for insertion over the RBC surface. In a first step, conjugation experiments were performed: in dimethyl sulfoxide (DMSO), RBCs were conserved and modified by succinic anhydride to create an additional -COOH group, and then activated with 3-(3-dimethylaminopropyl)carbodiimide-N-hydroxysuccinimide (EDC-NHS) in 2-(N-morpholino) ethanesulfonic acid hydrate buffer for insertion of labeled Abs or DAPI. In a second step, fluorescence signals were evaluated by microscopy and the mean fluorescence intensities of cell lysates were measured by spectrofluorometry. The results showed clear evidence for adsorption of FITC- and PE-labeled Abs to activated conserved RBCs. DAPI was adsorbed well also to DMSO-conserved RBCs without the need for an activation step. The DMSO conservation step was enough to create reactive RBCs for insertion of specific Abs and fluorescent dyes. The additional modification by succinic anhydride and activation with EDC-NHS resulted in two- to seven-fold increase in fluorescence signals, indicating a much higher RBC loading capacity. These Ab- and fluorescent dye-functionalized RBCs have potentially high application in developing new biomedical diagnostic and in vitro assay techniques.

Magnetic Beads-Based Sensor with Tailored Sensitivity for Rapid and Single-Step Amperometric Determination of miRNAs.

This work describes a sensitive amperometric magneto-biosensor for single-step and rapid determination of microRNAs (miRNAs). The developed strategy involves the use of direct hybridization of the target miRNA (miRNA-21) with a specific biotinylated DNA probe immobilized on streptavidin-modified magnetic beads (MBs), and labeling of the resulting heteroduplexes with a specific DNA-RNA antibody and the bacterial protein A (ProtA) conjugated with an horseradish peroxidase (HRP) homopolymer (Poly-HRP40) as an enzymatic label for signal amplification. Amperometric detection is performed upon magnetic capture of the modified MBs onto the working electrode surface of disposable screen-printed carbon electrodes (SPCEs) using the H2O2/hydroquinone (HQ) system. The magnitude of the cathodic signal obtained at -0.20 V (vs. the Ag pseudo-reference electrode) demonstrated linear dependence with the concentration of the synthetic target miRNA over the 1.0 to 100 pM range. The method provided a detection limit (LOD) of 10 attomoles (in a 25 µL sample) without any target miRNA amplification in just 30 min (once the DNA capture probe-MBs were prepared). This approach shows improved sensitivity compared with that of biosensors constructed with the same anti-DNA-RNA Ab as capture instead of a detector antibody and further labeling with a Strep-HRP conjugate instead of the Poly-HRP40 homopolymer. The developed strategy involves a single step working protocol, as well as the possibility to tailor the sensitivity by enlarging the length of the DNA/miRNA heteroduplexes using additional probes and/or performing the labelling with ProtA conjugated with homopolymers prepared with different numbers of HRP molecules. The practical usefulness was demonstrated by determination of the endogenous levels of the mature target miRNA in 250 ng raw total RNA (RNAt) extracted from human mammary epithelial normal (MCF-10A) and cancer (MCF-7) cells and tumor tissues.

Clonal evolution and antigen recognition of anti-nuclear antibodies in acute systemic lupus erythematosus.

The evolutional process of disease-associated autoantibodies in systemic lupus erythematosus (SLE) remains to be established. Here we show intraclonal diversification and affinity maturation of anti-nuclear antibody (ANA)-producing B cells in SLE. We identified a panel of monoclonal ANAs recognizing nuclear antigens, such as double-stranded DNA (dsDNA) and ribonucleoproteins (RNPs) from acute SLE subjects. These ANAs had relatively few, but nonetheless critical mutations. High-throughput immunoglobulin sequencing of blood lymphocytes disclosed the existence of sizable ANA lineages shearing critical mutations intraclonally. We further focused on anti-DNA antibodies, which are capable to bind to both single-stranded (ss) and dsDNA at high affinity. Crystal structure and biochemical analysis confirmed a direct role of the mutations in the acquisition of DNA reactivity and also revealed that these anti-DNA antibodies recognized an unpaired region within DNA duplex. Our study unveils the unique properties of high-affinity anti-DNA antibodies that are generated through antigen-driven affinity maturation in acute phase of SLE.

A structural investigation of FISLE-412, a peptidomimetic compound derived from saquinavir that targets lupus autoantibodies.

FISLE-412 is the first reported small molecule peptidomimetic that neutralizes anti-dsDNA autoantibodies associated with systemic lupus erythematosus (SLE) pathogenesis. FISLE-412 is a complex small molecule that involves a challenging synthesis scheme, but has attractive pharmacological activities as a potential small molecule therapeutic in lupus. Therefore, we initiated a Structure-Activity Relationship study to simplify the complexity of FISLE-412. We synthesized a small library of mimetopes around the FISLE-412 structure and identified several analogues which could neutralize anti-DNA lupus antibodies in vitro and ex vivo. Our strategies reduced the structural complexity of FISLE-412 and provide important information that may guide development of potential autoantibody-targeted lupus therapeutics.

Variable Heavy Chain Domain Derived from a Cell-Penetrating Anti-DNA Monoclonal Antibody for the Intracellular Delivery of Biomolecules.

Targeting and modification of important intracellular proteins using efficient vehicles are invaluable in diagnostic and therapeutic fields. Cell-penetrating antibodies and their fragments can be utilized as vehicles for the delivery of modifiers into cells. In this study, we explored the applicability of variable heavy chain (VH) domain as delivery vehicles for mammalian cells. The characteristics of the recombinant VH domain produced from a cell-penetrating monoclonal anti-double stranded DNA antibody 2C10 were analyzed using flow cytometry, confocal microscopy, cell proliferation assay, and cell cycle analysis in various mammalian cell lines. The VH domain penetrated into various cell lines in a time- and dose-dependent manner, although the internalization efficiency varied. The domain was localized in the nuclei as well as the cytoplasm of living cells. It was also internalized into cells mainly through the clathrin-mediated endocytosis pathway. We tested further its efficiency in delivering specific biomolecule(s) using the conjugates of the single domain molecule and small interfering RNA (siRNA) for the testicular nuclear auto-antigenic sperm protein (tNASP). It was found that the siRNA was successfully delivered by the VH domain into cancer cells, and knockdown effects from the delivered tNASP-siRNA were observed. The levels of the RNA transcript and protein of tNASP were decreased and the down-regulated tNASP inhibited cell proliferation and caused G0G1 phase arrest of the cell cycle. These results indicate that the recombinant 2C10 VH domain could be applied as an efficient vehicle capable of delivering valuable biomolecule into the cytoplasm or cell nuclei for clinical uses.

An anti-DNA antibody prefers damaged dsDNA over native.

DNA-protein interactions, including DNA-antibody complexes, have both fundamental and practical significance. In particular, antibodies against double-stranded DNA play an important role in the pathogenesis of autoimmune diseases. Elucidation of structural mechanisms of an antigen recognition and interaction of anti-DNA antibodies provides a basis for understanding the role of DNA-containing immune complexes in human pathologies and for new treatments. Here we used Molecular Dynamic simulations of bimolecular complexes of a segment of dsDNA with a monoclonal anti-DNA antibody's Fab-fragment to obtain detailed structural and physical characteristics of the dynamic intermolecular interactions. Using a computationally modified crystal structure of a Fab-DNA complex (PDB: 3VW3), we studied in silico equilibrium Molecular Dynamics of the Fab-fragment associated with two homologous dsDNA fragments, containing or not containing dimerized thymine, a product of DNA photodamage. The Fab-fragment interactions with the thymine dimer-containing DNA was thermodynamically more stable than with the native DNA. The amino acid residues constituting a paratope and the complementary nucleotide epitopes for both Fab-DNA constructs were identified. Stacking and electrostatic interactions were shown to play the main role in the antibody-dsDNA contacts, while hydrogen bonds were less significant. The aggregate of data show that the chemically modified dsDNA (containing a covalent thymine dimer) has a higher affinity toward the antibody and forms a stronger immune complex. These findings provide a mechanistic insight into formation and properties of the pathogenic anti-DNA antibodies in autoimmune diseases, such as systemic lupus erythematosus, associated with skin photosensibilization and DNA photodamage.

The Use of Poly-L-Lysine as a Capture Agent to Enhance the Detection of Antinuclear Antibodies by ELISA.

Antibodies to nuclear antigens (antinuclear antibodies or ANAs) are the serological hallmark of systemic lupus erythematosus (SLE). These antibodies bind diverse nuclear antigens that include DNA, histones and non-histone proteins as well as complexes of proteins with DNA and RNA. Because of the frequency of ANA expression in SLE, testing is an important component of clinical evaluation as well as determination of eligibility for clinical trials or utilization of certain therapies. Immunofluorescence assays have been commonly used for this purpose although this approach can be limited by issues of throughput, variability and difficulty in determining positivity. ELISA and multiplex assays are also useful approaches although these assays may give an incomplete picture of antibodies present. To develop a sensitive and quantitative ANA assay, we have explored an ELISA platform in which plates are pre-coated with a positively charged nucleic acid binding polymer (NABP) to increase adherence of antigens containing DNA or RNA. As a source of antigens, we have used supernatants of Jurkat cells undergoing apoptosis in vitro. As results presented show, a poly-L-lysine (PLL) pre-coat significantly enhances detection of antibodies to DNA as well as antigens such as histones, SSA, SSB and RNP. Comparison of the ELISA assay with the PLL pre-coat with a multiplex assay using the BioPlex® 2200 system indicated good agreement in results for a panel of lupus sera. Together, these studies indicate that a pre-coat with a positively charged polymer can increase the sensitivity of an ANA ELISA using as antigens molecules released from dead and dying cells. This assay platform may facilitate ANA testing by providing an ensemble of antigens more similar in composition and structure with antigens present in vivo, with a NABP promoting adherence via charge-charge interactions.

A lupus anti-DNA autoantibody mediates autocatalytic, targeted delivery of nanoparticles to tumors.

Strategies to target nanoparticles to tumors that rely on surface modification with ligands that bind molecules overexpressed on cancer cells or the tumor neovasculature suffer from a major limitation: with delivery of toxic agents the amount of molecules available for targeting decreases with time; consequently, the efficiency of nanoparticle delivery is reduced. To overcome this limitation, here we propose an autocatalytic tumor-targeting mechanism based on targeting extracellular DNA (exDNA). exDNA is enriched in the tumor microenviroment and increases with treatment with cytotoxic agents, such as doxorubicin (DOX), due to release of DNA by dying tumor cells. We tested this approach using poly(lactic-co-glycolic acid) (PLGA) nanoparticles surface-conjugated with fragments of 3E10 (3E10EN), a lupus anti-DNA autoantibody. We demonstrated that 3E10EN-conjugated nanoparticles bound to DNA and preferentially localized to tumors in vivo. The efficiency of tumor localization of 3E10EN-conjugated, DOX-loaded nanoparticles increased with time and subsequent treatments, demonstrating an autocatalytic effect. 3E10EN-conjugated DOX-loaded nanoparticles exhibited a significant anti-tumor effect that was superior to all controls. This work demonstrates the promise of autocatalytic drug delivery mechanisms and establishes proof of concept for a new anti-DNA autoantibody-based approach for enhancing delivery of nanoparticles to tumors.