Monomeric Fragment Research Articles

The Hamilton Receptor in Supramolecular Polymer Sciences.

Supramolecular polymers are polymeric materials of monomeric fragments, held jointly by reversible and directional non-covalent interactions such as multiple hydrogen-bonding, charge transfer effects, host-guest interactions, metal coordination, and aromatic stacking. This review article on the Hamilton-based supramolecular polymers aims to shed light on the molecular recognition achievements by the Hamilton-based polymeric systems, evaluate Hamilton receptor's future prospects, and capitalize its potential applications in supramolecular chemistry. To the best of our knowledge, this is the first elaborative and sole manuscript in which polymeric Hamilton receptors are being exposed in detail. The first portion of this manuscript is related to the importance and urgency of polymers along with the historic background of Hamilton receptors. The middle section discloses the potential applications of Hamilton-type receptors in various fields, e.g., dendrimers, mechanically polymeric rotaxanes, and self-assemblies. The final section of the manuscript discloses the future aspects and the importance of novel polymer-based Hamilton-type receptors in the modern era. We believe that this first review in this emerging yet immature field will be useful to inspire scientists around the world to find the unseen future prospects, thereby boosting the field related to this valued artificial receptor in the province of supramolecular chemistry and also in other domains of scientific fields and technology, as well.

Targeting GD2-Positive Tumor Cells by Pegylated scFv Fragment-Drug Conjugates Carrying Maytansinoids DM1 and DM4.

Oligomerization of antibody fragments via modification with polyethylene glycol (pegylation) may alter their function and properties, leading to a multivalent interaction of the resulting constructs with the target antigen. In a recent study, we generated pegylated monomers and multimers of scFv fragments of GD2-specific antibodies using maleimide-thiol chemistry. Multimerization enhanced the antigen-binding properties and demonstrated a more efficient tumor uptake in a syngeneic GD2-positive mouse cancer model compared to monomeric antibody fragments, thereby providing a rationale for improving the therapeutic characteristics of GD2-specific antibody fragments. In this work, we obtained pegylated conjugates of scFv fragments of GD2-specific antibodies with maytansinoids DM1 or DM4 using tetravalent PEG-maleimide (PEG4). The protein products from the two-stage thiol-maleimide reaction resolved by gel electrophoresis indicated that pegylated scFv fragments constituted the predominant part of the protein bands, and most of the scFv formed pegylated monomers and dimers. The conjugates retained the ability to bind ganglioside GD2 comparable to that of the parental scFv fragment and to specifically interact with GD2-positive cells. Both induced significant inhibitory effects in the GD2-positive B78-D14 cell line, in contrast to the GD2-negative B16 cell line. The decrease in the B78-D14 cell viability when treated with scFv-PEG4-DM4 was more prominent than that for scFv-PEG4-DM1, and was characterized by a twofold lower half-maximal inhibitory concentration (IC50). Unlike the parental scFv fragment, the product of scFv and PEG4 conjugation (scFv-PEG4), consisting predominantly of pegylated scFv multimers and monomers, induced direct cell death in the GD2-positive B78-D14 cells. However, the potency of scFv-PEG4 was low in the selected concentration range, thus demonstrating that the cytotoxic effect of DM1 and DM4 within the antibody fragment-drug conjugates was primary. The suggested approach may contribute to development of novel configurations of antibody fragment-drug conjugates for cancer treatment.

The Potential of Nanobodies for COVID-19 Diagnostics and Therapeutics.

The infectious severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the causative agent for coronavirus disease 2019 (COVID-19). Globally, there have been millions of infections and fatalities. Unfortunately, the virus has been persistent and a contributing factor is the emergence of several variants. The urgency to combat COVID-19 led to the identification/development of various diagnosis (polymerase chain reaction and antigen tests) and treatment (repurposed drugs, convalescent plasma, antibodies and vaccines) options. These treatments may treat mild symptoms and decrease the risk of life-threatening disease. Although these options have been fairly beneficial, there are some challenges and limitations, such as cost of tests/drugs, specificity, large treatment dosages, intravenous administration, need for trained personal, lengthy production time, high manufacturing costs, and limited availability. Therefore, the development of more efficient COVID-19 diagnostic and therapeutic options are vital. Nanobodies (Nbs) are novel monomeric antigen-binding fragments derived from camelid antibodies. Advantages of Nbs include low immunogenicity, high specificity, stability and affinity. These characteristics allow for rapid Nb generation, inexpensive large-scale production, effective storage, and transportation, which is essential during pandemics. Additionally, the potential aerosolization and inhalation delivery of Nbs allows for targeted treatment delivery as well as patient self-administration. Therefore, Nbs are a viable option to target SARS-CoV-2 and overcome COVID-19. In this review we discuss (1) COVID-19; (2) SARS-CoV-2; (3) the present conventional COVID-19 diagnostics and therapeutics, including their challenges and limitations; (4) advantages of Nbs; and (5) the numerous Nbs generated against SARS-CoV-2 as well as their diagnostic and therapeutic potential.

Computational insight into spatial and electronic structure of bis-diketonate cobalt complexes with triphenodioxazine ligands

A DFT study of monomeric fragments of 1D-polymer chains consisting of bis-acetylacetonate (acac), bis-trifluoromethylacetylacetonate (tfac) and bis-hexafluoroacetylacetonate (hfac) cobalt complexes with heteropentacene triphenodioxazine (TPDO) ligands revealed characteristic structural features observed in the crystal. It was established that elongation of donor–acceptor Co–N bonds stems from the presence of bulky tert-butyl groups in the TPDO, as well as repulsive interactions between the hydrogen atoms of N-donor ligands and oxygen diketonate atoms. Unusual for the high-spin cobalt(II) ion Co–O distances in Co(acac)2TPDO2 can be caused by possible population of the low-spin doublet state at increased temperature.

A label-free T4 polynucleotide kinase fluorescence sensor based on split dimeric G-quadruplex and ligation-induced dimeric G-quadruplex/thioflavin T conformation

The phosphorylation process of DNA by T4 polynucleotide kinase (T4 PNK) plays a crucial role in DNA recombination, DNA replication, and DNA repair. Traditional monomeric G-quadruplex (G4) systems are always activated by single cation such as K+ or Na+. The conformation transformation caused by the coexistence of multiple cations may interfere with the signal readout and limit their applications in physiological system. In view of the stability of dimeric G4 in multiple cation solution, we reported a label-free T4 PNK fluorescence sensor based on split dimeric G4 and ligation-induced dimeric G4/thioflavin T (ThT) conformation. The dimeric G4 was divided into two independent pieces of one normal monomeric G4 and the other monomeric G4 fragment phosphorylated by T4 PNK in order to decrease the background signal. With the introduction of template DNA, DNA ligase, and invasive DNA, the dimeric G4 could be generated and liberated to combine with ThT to show obvious fluorescence signal. Using our strategy, the linear range from 0.005 to 0.5 U mL-1, and the detection limit of 0.0021 U mL-1 could be achieved without the consideration of interference caused by the coexistence of multiple cations. Additionally, research in real sample determination and inhibition effect investigations indicated its further potential application value in biochemical process research and clinic diagnostics.

A symmetry-driven approach toward the total synthesis of dodecahedrane

Herein, we describe our progress toward the total synthesis of dodecahedrane, a complex and highly symmetrical hydrocarbon that bears twelve fused rings arranged in a cage-like architecture. Central to our approach is a late-stage [2+2+2+2+2] polyene cyclization cascade, which is expected to construct five new bonds and ten new rings in a single transformation. Toward this end, we describe efforts to synthesize key monomeric fragments, along with successful dimerization studies using a pinacol coupling approach. Subsequent studies include an attempted olefin metathesis rearrangement cascade in addition to a successful intramolecular photochemical [2+2] reaction. Although attempts to elaborate the photocycloaddition product were unsuccessful, our studies provide access to complex dimeric scaffolds and are expected to help guide our future total synthesis studies.

A hypothalamic pathway for Augmentor α–controlled body weight regulation

Augmentor α and β (Augα and Augβ) are newly discovered ligands of the receptor tyrosine kinases Alk and Ltk. Augα functions as a dimeric ligand that binds with high affinity and specificity to Alk and Ltk. However, a monomeric Augα fragment and monomeric Augβ also bind to Alk and potently stimulate cellular responses. While previous studies demonstrated that oncogenic Alk mutants function as important drivers of a variety of human cancers, the physiological roles of Augα and Augβ are poorly understood. Here, we investigate the physiological roles of Augα and Augβ by exploring mice deficient in each or both Aug ligands. Analysis of mutant mice showed that both Augα single knockout and double knockout of Augα and Augβ exhibit a similar thinness phenotype and resistance to diet-induced obesity. In the Augα-knockout mice, the leanness phenotype is coupled to increased physical activity. By contrast, Augβ-knockout mice showed similar weight curves as the littermate controls. Experiments are presented demonstrating that Augα is robustly expressed and metabolically regulated in agouti-related peptide (AgRP) neurons, cells that control whole-body energy homeostasis in part via their projections to the paraventricular nucleus (PVN). Moreover, both Alk and melanocortin receptor-4 are expressed in discrete neuronal populations in the PVN and are regulated by projections containing Augα and AgRP, respectively, demonstrating that two distinct mechanisms that regulate pigmentation operate in the hypothalamus to control body weight. These experiments show that Alk-driven cancers were co-opted from a neuronal pathway in control of body weight, offering therapeutic opportunities for metabolic diseases and cancer.

Surface-Functionalized Silica-Coated Calcium Phosphate Nanoparticles Efficiently Deliver DNA-Based HIV-1 Trimeric Envelope Vaccines against HIV-1.

Human immunodeficiency virus type 1 (HIV-1) infection remains one of the worst crises in global health. The prevention of HIV-1 infection is a crucial task that needs to be addressed due to the absence of a licensed vaccine against HIV-1. DNA vaccines present a promising alternative approach to combat HIV-1 infection due to their excellent safety profile, lack of severe side effects, and relatively rapid fabrication. Traditional vaccines composed of a monomeric envelope or peptide fragments have been indicated to lack protective efficacy mediated by inducing HIV-1-specific neutralizing antibodies in clinical trials. The immunogenicity and protection against HIV-1 induced by DNA vaccines are limited due to the poor uptake of these vaccines by antigen-presenting cells and their ready degradation by DNases and lysosomes. To address these issues of naked DNA vaccines, we described the feasibility of CpG-functionalized silica-coated calcium phosphate nanoparticles (SCPs) for efficiently delivering DNA-based HIV-1 trimeric envelope vaccines against HIV-1. Vaccines comprising the soluble BG505 SOSIP.664 trimer fused to the GCN4-based isoleucine zipper or bacteriophage T4 fibritin foldon motif with excellent simulation of the native HIV-1 envelope were chosen as trimer-based vaccine platforms. Our results showed that SCP-based DNA immunization could significantly induce both broad humoral immune responses and potent cellular immune responses compared to naked DNA vaccination in vivo. To the best of our knowledge, this study is the first to assess the feasibility of CpG-functionalized SCPs for efficiently delivering DNA vaccines expressing a native-like HIV-1 trimer. These CpG-functionalized SCPs for delivering DNA-based HIV-1 trimeric envelope vaccines may lead to the development of promising vaccine candidates against HIV-1.

The Enantioselective Synthesis of Eburnamonine, Eucophylline, and 16'-epi-Leucophyllidine.

A synthetic approach to the heterodimeric bisindole alkaloid leucophyllidine is disclosed herein. An enantioenriched lactam building block, synthesized through palladium-catalyzed asymmetric allylic alkylation, served as the precursor to both hemispheres. The eburnamonine-derived fragment was synthesized through a Bischler-Napieralski/hydrogenation approach, while the eucophylline-derived fragment was synthesized by Friedländer quinoline synthesis and two sequential C-H functionalization steps. A convergent Stille coupling and phenol-directed hydrogenation united the two monomeric fragments to afford 16'-epi-leucophyllidine in 21 steps from commercial material.

The Enantioselective Synthesis of Eburnamonine, Eucophylline, and 16′‐epi‐Leucophyllidine

AbstractA synthetic approach to the heterodimeric bisindole alkaloid leucophyllidine is disclosed herein. An enantioenriched lactam building block, synthesized through palladium‐catalyzed asymmetric allylic alkylation, served as the precursor to both hemispheres. The eburnamonine‐derived fragment was synthesized through a Bischler–Napieralski/hydrogenation approach, while the eucophylline‐derived fragment was synthesized by Friedländer quinoline synthesis and two sequential C−H functionalization steps. A convergent Stille coupling and phenol‐directed hydrogenation united the two monomeric fragments to afford 16′‐epi‐leucophyllidine in 21 steps from commercial material.

Ruthenium-Benzothiadiazole Building Block Derived Dynamic Heterometallic Ru-Ag Coordination Polymer and Its Enhanced Water-Splitting Feature.

This article deals with the development of the unprecedented redox-mediated heterometallic coordination polymer {[RuIII(acac)2(μ-bis-η1-N,η1-N-BTD)2AgI(ClO4)]ClO4}n (3) via the oxidation of the monomeric building block cis-[RuII(acac)2(η1-N-BTD)2] (1) by AgClO4 (BTD = exodentate 2,1,3-benzothiadiazole, acac = acetylacetonate). Monomeric cis-[RuII(acac)2(η1-N-BTD)2] (1) and [RuII(acac)2(η1-N-BTD)(CH3CN)] (2) were simultaneously obtained from the electron-deficient BTD heterocycle and the electron-rich metal precursor RuII(acac)2(CH3CN)2 in refluxing CH3CN. Molecular identities of 1-3 were authenticated by their single-crystal X-ray structures as well as by solution spectral features. These results also reflected the elusive trigonal-planar geometry of the Ag ion in Ru-Ag-derived polymeric 3. Ru(III) (S = 1/2)-derived 3 displayed metal-based anisotropic EPR with ⟨g⟩/Δg = 2.12/0.56 and paramagnetically shifted 1H NMR. Spectroelectrochemistry in combination with DFT/TD-DFT calculations of 1n and 2n (n = 1+, 0, 1-) determined a metal-based (RuII/RuIII) oxidation and BTD-based reduction (BTD/BTD•-). The drastic decrease in the emission intensity and quantum yield but insignificant change in the lifetime of 3 with respect to 1 could be addressed in terms of static quenching and/or a paramagnetism-induced phenomenon. A homogeneously dispersed dumbbell-shaped morphology and the particle diameter of 3 were established by microscopic (TEM-EDX/SEM) and DLS analysis, respectively. Moreover, the dynamic nature of polymeric 3 was highlighted by its degradation to the η1-N-BTD coordinated monomeric fragment 1, which could also be followed spectrophotometrically in polar protic EtOH. Interestingly, both monomeric 1 and polymeric 3 exhibited efficient electrocatalytic activity toward water oxidation processes (OER, HER) on immobilization on an FTO support, which also divulged the better intrinsic water oxidation activity of 3 in comparison to 1.

The structure-function relationship of a signaling-competent, dimeric Reelin fragment

Reelin operates through canonical and non-canonical pathways that mediate several aspects of brain development and function. Reelin's dimeric central fragment (CF), generated through proteolytic cleavage, is required for the lipoprotein-receptor-dependent canonical pathway activation. Here, we analyze the signaling properties of a variety of Reelin fragments and measure the differential binding affinities of monomeric and dimeric CF fragments to lipoprotein receptors to investigate the mode of canonical signal activation. We also present the cryoelectron tomography-solved dimeric structure of Reelin CF and support it using several other biophysical techniques. Our findings suggest that Reelin CF forms a covalent parallel dimer with some degree of flexibility between the two protein chains. As a result of this conformation, Reelin binds to lipoprotein receptors in a manner inaccessible to its monomeric form and is capable of stimulating canonical pathway signaling.

Lignin First: Confirming the Role of the Metal Catalyst in Reductive Fractionation

Rhodium nanoparticles embedded on the interior of hollow porous carbon nanospheres, able to sieve monomers from polymers, were used to confirm the precise role of metal catalysts in the reductive catalytic fractionation of lignin. The study provides clear evidence that the primary function of the metal catalyst is to hydrogenate monomeric lignin fragments into more stable forms following a solvent-based fractionation and fragmentation of lignin.

New Polyporphyrin Arrays with Controlled Fluorescence Obtained by Diaxial Sn(IV)-Porphyrin Phenolates Chelation with Cu2+ Cation.

New coordination oligomers and polymers of Sn(IV)-tetra(4-sulfonatophenyl)porphyrin have been constructed by the chelation reaction of its diaxialphenolates with Cu2+. The structure and properties of the synthesized polyporphyrin arrays were investigated by 1H Nuclear Magnetic Resonance (1H NMR), Infra Red (IR), Ultra Violet - Visible (UV-Vis) and fluorescence spectroscopy, mass spectrometry, Powder X-Rays Diffraction (PXRD), Electron Paramagnetic Resonance (EPR), thermal gravimetric, elemental analysis, and quantum chemical calculations. The results show that the diaxial coordination of bidentate organic ligands (L-tyrazine and diaminohydroquinone) leads to the quenching of the tetrapyrrole chromophore fluorescence, while the chelation of the porphyrinate diaxial complexes with Cu2+ is accompanied by an increase in the fluorescence in the organo-inorganic hybrid polymers formed. The obtained results are of particular interest to those involved in creating new ‘chemo-responsive’ (i.e., selectively interacting with other chemical species as receptors, sensors, or photocatalysts) materials, the optoelectronic properties of which can be controlled by varying the number and connection type of monomeric fragments in the polyporphyrin arrays.

Laser Desorption/Ionization of Low-Molecular-Weight Lignin Oligomers

The study compares the efficiency of laser desorption/ionization (LDI) of six dimeric compounds, corresponding to the most important types of bonds between monomeric structural fragments of lignin macromolecules, on three types of different stainless steel target plates. It was found that ground steel target plates demonstrate the best results because of the uniform crystallization of analytes and good surface coverage. The most important ionization mechanisms for lignin oligomers with conjugated structures and increased acidity, along with protonation (cationization), involve the formation of molecular ions due to direct photoionization. For all of the studied compounds, partial in-source decay was observed, the contribution of which became significant at high laser fluence. The LDI method was successfully used to obtain mass spectra of low-molecular-weight products of the depolymerization of technical lignins. It was shown that the use of hybrid mass spectrometers with a quadrupole ion trap leads to an increase in the percentage of signals of protonated molecules in the mass spectra of lignin depolymerization products because of the occurrence of side ion-molecular reactions in the plume and ion trap.

Cationic Bola Form Metallosurfactants Based on Isothiouronium, Synthesis and Anti-Microbial Activity

Abstract Two series of well-defined dimeric metallo-bolaamphiphiles (bola surfactants) with the coordinated metal ions (Cu, Co, Zn) were prepared. These oligomeric surfactants consist of simple monomeric cationic surfactant fragments which are coupled via the hydrophilic ammonium chloride head groups by C6 and C12 spacer groups of different lengths. FTIR and 1HNMR identification techniques confirmed the obtained products. Measurements of surface tensions showed that the synthesized Bola amphiphiles have the desired, relatively low critical micelle formation concentrations (CMC). Bola amphiphiles with long spacer groups (C12) have a pronounced surface activity. The properties of these cationic surfactant oligomers in aqueous solution such as micellization and surface activity were discussed in relation to spacer group. In addition, the synthesized compounds were examined for their anti-microbial activity using the agar diffusion technique.

Abstract 2278: CTLA-4 targeted engineered toxin bodies designed to deplete regulatory T cells (Tregs)

Abstract Tumor resident regulatory T cells (Tregs) are important mediators of an immunosuppressive tumor microenvironment (TME) promoting tumor immune evasion. The presence of Tregs, and a higher ratio of Tregs to effector T cells in the TME, are associated with poor prognosis. The depletion of Tregs in the TME is expected to re-expose the tumor to the immune system to allow for tumor control. CLTA-4 is expressed on the cell surface of Tregs and is a clinically validated target. Better responses to CTLA-4 monoclonal antibody (mAb) treatment are correlated with stronger ADCC-mediated Treg depletion in preclinical models, and patient FcγR polymorphism has been reported to correlate with response to CTLA-4 mAb therapy. Towards improving on current therapies, many efforts to increase effective depletion of Tregs (such as by Fc effector modification) are being pursued. Engineered toxin bodies (ETBs) are comprised of a proprietarily engineered form of Shiga-like Toxin A subunit (SLT-A) genetically fused to antibody-like binding domains. ETBs work through novel mechanisms of action and are capable of forcing internalization, self-routing through intracellular compartments to the cytosol, and inducing potent cell-kill via the enzymatic and permanent inactivation of ribosomes. Targeted ETBs are being developed to specifically target and destroy CTLA-4 expressing cells. In vitro, cells expressing CTLA-4 are effectively and specifically killed by targeted ETBs. This direct cell kill activity of the ETB has the potential to deplete Tregs in the TME without a requirement for ADCC mechanisms. ETBs have been designed to bind various CTLA-4 epitopes, and to comprise of different targeting domains formats, including monomeric and diabody single chain variable fragments (scFvs) as well as single domain and biparatopic antibody fragments. The development of a CTLA-4 targeted ETB is ongoing. The entry of a protein with direct cell kill properties into the therapeutic space represents a differentiated mechanism of action to deplete Tregs for ultimate re-invigoration of the anti-tumor immune response, and has the potential to provide benefit to patients, including in the relapsed or refractory setting. Citation Format: Aimee Iberg, Edith Acquaye-Seedah, Lilia A. Rabia, Garrett L. Robinson, Hilario J. Ramos, Joseph D. Dekker, Jay Zhao, Erin K. Willert. CTLA-4 targeted engineered toxin bodies designed to deplete regulatory T cells (Tregs) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2278.

A Mixed-Valence Tetra-Nuclear Nickel Dithiolene Complex: Synthesis, Crystal Structure, and the Lability of Its Nickel Sulfur Bonds

In this study, by employing a common synthetic protocol, an unusual and unexpected tetra-nuclear nickel dithiolene complex was obtained. The synthesis of the [Ni4(ecpdt)6]2− dianion (ecpdt = (Z)-3-ethoxy-3-oxo-1-phenylprop-1-ene-1,2-bis-thiolate) with two K+ as counter ions was then intentionally reproduced. The formation of this specific complex is attributed to the distinct dithiolene precursor used and the combination with the then coordinated counter ion in the molecular solid-state structure, as determined by X-ray diffraction. K2[Ni4(ecpdt)6] was further characterized by ESI-MS, FT-IR, UV-Vis, and cyclic voltammetry. The tetra-nuclear complex was found to have an uncommon geometry arising from the combination of four nickel centers and six dithiolene ligands. In the center of the arrangement, suspiciously long Ni–S distances were found, suggesting that the tetrameric structure can be easily split into two identical dimeric fragments or two distinct groups of monomeric fragments, for instance, upon dissolving. A proposed variable magnetism in the solid-state and in solution due to the postulated dissociation was confirmed. The Ni–S bonds of the “inner” and “outer” nickel centers differed concurrently with their coordination geometries. This observation also correlates with the fact that the complex bears two anionic charges requiring the four nickel centers to be present in two distinct oxidation states (2 × +2 and 2 × +3), i.e., to be hetero-valent. The different coordination geometries observed, together with the magnetic investigation, allowed the square planar “outer” geometry to be assigned to d8 centers, i.e., Ni2+, while the Ni3+ centers (d7) were in a square pyramidal geometry with longer Ni–S distances due to the increased number of donor atoms and interactions.

Biochemical and structural analyses reveal that the tumor suppressor neurofibromin (NF1) forms a high-affinity dimer

Biochemical and structural analyses reveal that the tumor suppressor neurofibromin (NF1) forms a high-affinity dimer

Multimerization through Pegylation Improves Pharmacokinetic Properties of scFv Fragments of GD2-Specific Antibodies.

Antigen-binding fragments of antibodies specific to the tumor-associated ganglioside GD2 are well poised to play a substantial role in modern GD2-targeted cancer therapies, however, rapid elimination from the body and reduced affinity compared to full-length antibodies limit their therapeutic potential. In this study, scFv fragments of GD2-specific antibodies 14.18 were produced in a mammalian expression system that specifically bind to ganglioside GD2, followed by site-directed pegylation to generate mono-, di-, and tetra-scFv fragments. Fractionated pegylated dimers and tetramers of scFv fragments showed significant increase of the binding to GD2 which was not accompanied by cross-reactivity with other gangliosides. Pegylated multimeric di-scFvs and tetra-scFvs exhibited cytotoxic effects in GD2-positive tumor cells, while their circulation time in blood significantly increased compared with monomeric antibody fragments. We also demonstrated a more efficient tumor uptake of the multimers in a syngeneic GD2-positive mouse cancer model. The findings of this study provide the rationale for improving therapeutic characteristics of GD2-specific antibody fragments by multimerization and propose a strategy to generate such molecules. On the basis of multimeric antibody fragments, bispecific antibodies and conjugates with cytotoxic drugs or radioactive isotopes may be developed that will possess improved pharmacokinetic and pharmacodynamic properties.