Synthesis and biological evaluation of a monocyclic Fc-binding antibody-recruiting molecule for cancer immunotherapy.
Ontology highlight
ABSTRACT:
SUBMITTER: Sasaki K
PROVIDER: S-EPMC8130626 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Redirecting endogenous antibodies in the bloodstream to tumor cells using synthetic molecules is a promising approach to trigger anti-tumor immune responses. However, current molecular designs only enable the use of a small fraction of endogenous antibodies, limiting the therapeutic potential. Here, we report Fc-binding antibody-recruiting molecules (Fc-ARMs) as the first example addressing this issue. Fc-ARMs are composed of an Fc-binding peptide and a targeting ligand, enabling the exploitation of endogenous antibodies through constant affinity to the Fc region of antibodies, whose sequence is conserved in contrast to the Fab region. We show that Fc-ARM targeting folate receptor-α (FR-α) redirects a clinically used antibody mixture to FR-α+ cancer cells, resulting in cancer cell lysis by natural killer cells in vitro. Fc-ARMs successfully interacted with antibodies in vivo and accumulated in tumors. Furthermore, Fc-ARMs recruited antibodies to suppress tumor growth in a mouse model. Thus, Fc-ARMs have the potential to be a novel class of cancer immunotherapeutic agents.
| S-EPMC8157400 | biostudies-literature
Project description:Antibody therapeutics are a promising drug class due to their high specificity and favorable pharmacokinetics. While there are many methods for the development of antibodies specific to disease associated antigens, selecting antibodies against functional epitopes with high specificity and affinity can be difficult for certain epitopes. We describe a generalizable method for synthesizing antibody mimetics by site specifically conjugating small molecules (with high affinity and specificity to disease associated antigens) to an Fc fragment to develop drugs with the benefits of an antibody. As a proof of concept, an E269pAcPhe Fc antibody Fc fragment was produced and subsequently site-specifically labeled with a linker-modified folic acid compound to generate an Fc-folic acid antibody-mimetic. This was chosen as the model system because the high-affinity folate receptor FR-α is highly expressed in a number of cancer types including breast and ovarian cancer. The specificity of the Fc-folic acid conjugate was assessed via flowcytometry with the folate-receptor positive breast cancer cell line MDA-MB-231 by measuring Fc-folic acid binding in both the absence and presence of an excess of folic acid. Fc-small molecule conjugates could be developed into a unique class of antibody-like therapeutics.
| S-EPMC6667356 | biostudies-literature
Project description:The redirection of T cell activity using bispecific antibodies is one of the most promising cancer immunotherapy approaches currently in development, but it is limited by cytokine storm-related toxicities, as well as the pharmacokinetics and tumor-penetrating capabilities of current bispecific antibody formats. Here, we have engineered the ATTACK (Asymmetric Tandem Trimerbody for T cell Activation and Cancer Killing), a novel T cell-recruiting bispecific antibody which combines three EGFR-binding single-domain antibodies (VHH; clone EgA1) with a single CD3-binding single-chain variable fragment (scFv; clone OKT3) in an intermediate molecular weight package. The two specificities are oriented in opposite directions in order to simultaneously engage cancer cells and T cell effectors, and thereby promote immunological synapse formation. EgA1 ATTACK was expressed as a homogenous, non-aggregating, soluble protein by mammalian cells and demonstrated an enhanced binding to EGFR, but not CD3, when compared to the previously characterized tandem bispecific antibody which has one EgA1 VHH and one OKT3 scFv per molecule. EgA1 ATTACK induced synapse formation and early signaling pathways downstream of TCR engagement at lower concentrations than the tandem VHH-scFv bispecific antibody. Furthermore, it demonstrated extremely potent, dose-dependent cytotoxicity when retargeting human T cells towards EGFR-expressing cells, with an efficacy over 15-fold higher than that of the tandem VHH-scFv bispecific antibody. These results suggest that the ATTACK is an ideal format for the development of the next-generation of T cell-redirecting bispecific antibodies.
| S-EPMC5739562 | biostudies-literature
Project description:HIV/AIDS is a global pandemic for which new treatment strategies are desperately needed. We have designed a novel small molecule, designated as ARM-H, that has the potential to interfere with HIV survival through two mechanisms: (1) by recruiting antibodies to gp120-expressing virus particles and infected human cells, thus enhancing their uptake and destruction by the human immune system, and (2) by binding the viral glycoprotein gp120, inhibiting its interaction with the human protein CD4 and preventing virus entry. Here we demonstrate that ARM-H is capable of simultaneously binding gp120, a component of the Env surface viral glycoprotein (found on the surface of both HIV and virus-infected cells) and anti-2,4-dinitrophenyl antibodies (already present in the human bloodstream). The ternary complex formed between the antibody, ARM-H, and gp120 is immunologically active and leads to the complement-mediated destruction of Env-expressing cells. Furthermore, ARM-H prevents virus entry into human T-cells and should therefore be capable of inhibiting virus replication through two mutually reinforcing mechanisms (inhibition of virus entry and antibody-mediated killing). These studies demonstrate the viable anti-HIV activity of antibody-recruiting small molecules and have the potential to initiate novel paradigms in HIV treatment.
| S-EPMC2783809 | biostudies-literature
Project description:The low-molecular-weight compound JRC-II-191 inhibits infection of HIV-1 by blocking the binding of the HIV-1 envelope glycoprotein gp120 to the CD4 receptor and is therefore an important lead in the development of a potent viral entry inhibitor. Reported here is the use of two orthogonal screening methods, gold docking and ROCS shape-based similarity searching, to identify amine-building blocks that, when conjugated to the core scaffold, yield novel analogs that maintain similar affinity for gp120. Use of this computational approach to expand SAR produced analogs of equal inhibitory activity but with diverse capacity to enhance viral infection. The novel analogs provide additional lead scaffolds for the development of HIV-1 entry inhibitors that employ protein-ligand interactions in the vestibule of gp120 Phe 43 cavity.
| S-EPMC3049263 | biostudies-literature
Project description:Antibody production for ADCs (or in general) is commonly performed by CHO-based platforms and limited by volumetric productivity, expensive downstream purification, and extended optimization timelines. The Conamax platform is a novel microbial-based protein production and secretion system. A suite of synthetic biology tools have enabled high volumetric productivity (>1 g/L/d) and glycoengineering to produce simple and consistent human-like post-translational modifications. Conamax can be engineered to secrete genuine, functional monoclonal antibodies that have been successfully used to make antibody drug conjugates (ADCs) via cysteine-linked conjugation. Specifically, we evaluated ADCs derived from both a Conamax-produced anti-HER2 antibody and comparable commercially sourced Chinese hamster ovary (CHO)-produced material in an NCI-N87 gastric cancer xenograft model. Conjugation efficiency and resulting analytical data indicated comparable ADC quality and attributes. No statistical difference was observed between Conamax- and CHO-derived test articles thereby indicating similar efficacy and function. These results further demonstrate the potential of Conamax as a useful platform for the discovery and production of therapeutic antibodies and ADCs.
| S-EPMC7143411 | biostudies-literature
Project description:Histone acetylation is an extensively investigated post-translational modification that plays an important role as an epigenetic regulator. It is controlled by histone acetyl transferases (HATs) and histone deacetylases (HDACs). The overexpression of HDACs and consequent hypoacetylation of histones have been observed in a variety of different diseases, leading to a recent focus of HDACs as attractive drug targets. The natural product largazole is one of the most potent natural HDAC inhibitors discovered so far and a number of largazole analogs have been prepared to define structural requirements for its HDAC inhibitory activity. However, previous structure-activity relationship studies have heavily investigated the macrocycle region of largazole, while there have been only limited efforts to probe the effect of various zinc-binding groups (ZBGs) on HDAC inhibition. Herein, we prepared a series of largazole analogs with various ZBGs and evaluated their HDAC inhibition and cytotoxicity. While none of the analogs tested were as potent or selective as largazole, the Zn2+-binding affinity of each ZBG correlated with HDAC inhibition and cytotoxicity. We expect that our findings will aid in building a deeper understanding of the role of ZBGs in HDAC inhibition as well as provide an important basis for the future development of new largazole analogs with non-thiol ZBGs as novel therapeutics for cancer.
| S-EPMC5494147 | biostudies-literature
Project description:The neonatal Fc receptor (FcRn) promotes antibody recycling through rescue from normal lysosomal degradation. The binding interaction is pH-dependent with high affinity at low pH, but not under physiological pH conditions. Here, we combined rational design and saturation mutagenesis to generate novel antibody variants with prolonged half-life and acceptable development profiles. First, a panel of saturation point mutations was created at 11 key FcRn-interacting sites on the Fc region of an antibody. Multiple variants with slower FcRn dissociation kinetics than the wildtype (WT) antibody at pH 6.0 were successfully identified. The mutations were further combined and characterized for pH-dependent FcRn binding properties, thermal stability and the Fc?RIIIa and rheumatoid factor binding. The most promising variants, YD (M252Y/T256D), DQ (T256D/T307Q) and DW (T256D/T307W), exhibited significantly improved binding to FcRn at pH 6.0 and retained similar binding properties as WT at pH 7.4. The pharmacokinetics in human FcRn transgenic mice and cynomolgus monkeys demonstrated that these properties translated to significantly prolonged plasma elimination half-life compared to the WT control. The novel variants exhibited thermal stability and binding to Fc?RIIIa in the range comparable to clinically validated YTE and LS variants, and showed no enhanced binding to rheumatoid factor compared to the WT control. These engineered Fc mutants are promising new variants that are widely applicable to therapeutic antibodies, to extend their circulation half-life with obvious benefits of increased efficacy, and reduced dose and administration frequency.
| S-EPMC6748615 | biostudies-literature
Project description:Developing monoclonal antibodies (mAbs) for cancer immunotherapy is expensive and complicated. Nanobodies are small antibodies possessing favorable pharmacological properties compared with mAbs, but have limited anticancer efficacy due to the lack of an Fc region and poor pharmacokinetics. In this context, engineered universal endogenous antibody-recruiting nanobodies (UEAR Nbs), as a general and cost-effective approach, were developed to generate functional antibody-like nanobodies that could recapitulate the Fc biological functions for cancer immunotherapy. The UEAR Nbs, composed of the IgG binding domain and nanobody, were recombinantly expressed in E. coli and could recruit endogenous IgGs onto the cancer cell surface and trigger potent immune responses to kill cancer cells in vitro. Moreover, it was proved that UEAR Nbs displayed significantly improved half-lives in vivo. The in vivo antitumor efficacy of UEAR Nbs was demonstrated in a murine model using EGFR positive triple-negative breast cancer (TNBC).
| S-EPMC8179521 | biostudies-literature
Project description:Therapeutic immunoglobulin G (IgG) antibodies have comparatively long half-lives because the neonatal Fc receptor (FcRn) binds to the IgG Fc at acidic pH in the endosome and protects IgG from degradation. To further prolong the half-lives, amino acid-substituted antibodies having high affinity to FcRn are being developed, and one such therapeutic antibody (ravulizumab) has been approved. In this study, we investigated the binding property to FcγR and the conformation of seven FcRn affinity-modulated adalimumab variants to clarify the impact of the amino acid substitutions on the function and conformation of IgG Fc. The amino acid substitutions in T254-P261 caused a change in deuterium uptake into some regions of Fc in HDX-MS analysis, but those at T311, M432 and N438 did not cause such a change. The conformations around F245-L255 (FLFPPKPKDTL) were particularly influenced by the amino acid substitution in M256-P261, and the conformational changes of this region were correlated with the decrease of the affinity to FcγRIIIa. Additionally, we investigated the conformational difference of Fc between a Fc fusion protein (etanercept) and a native IgG (adalimumab). Although the Fc fusion proteins were expected to have similar FcRn affinity to IgGs, the affinity of etanercept to FcRn was lower than that of adalimumab, and its half-life was shorter than those of the IgG antibodies. Differences in deuterium uptakes were observed in the two regions where they were also detected in the adalimumab variants, and the conformational differences appeared to be an important factor for the low FcRn affinity of etanercept.
| S-EPMC8158039 | biostudies-literature