Single‐Molecule Force Measurement Guides the Design of Multivalent Ligands with Picomolar Affinity

Yang, Zhen; Jiang, Sheng; Li, Feng; Qiu, Yunli; Gu, Jianhua; Pettigrew, Roderic I.; Ferrari, Mauro; Hamilton, Dale J.; Li, Zheng

doi:10.1002/ange.201814347

Cited by 4 publications

(4 citation statements)

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“…As linker length dictates the mechanism of multivalent inhibition, multivalent benzamidines of lower valency that have short and flexible PEG linkers were specifically synthesized to promote statistical rebinding and minimize clustering and other modes of multivalent inhibition. [28,1] Benzamidine has also been shown to exhibit weak subsite binding to the light chain and kringle 5 of plasmin. [29] However, prior comparison of benzamidine inhibition across both plasmin and delta-plasmin, a recombinant plasmin variant possessing only the kringle 1 domain and active site, demonstrates that the active site is the primary benzamidine binding site and therefore, subsite binding impact has minimal effect.…”

Section: Resultsmentioning

confidence: 99%

Multivalent Benzamidine Molecules for Plasmin Inhibition: Effect of Valency and Linker Length

2022

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There is an emerging interest in utilizing synthetic multivalent inhibitors that comprise of multiple inhibitor moieties linked on a common scaffold to achieve strong and selective enzyme inhibition. As multivalent inhibition is impacted by valency and linker length, in this study, we explore the effect of multivalent benzamidine inhibitors of varying valency and linker length on plasmin inhibition. Plasmin is an endogenous enzyme responsible for digesting fibrin present in blood clots. Monovalent plasmin(ogen) inhibitors are utilized clinically to treat hyperfibrinolysis‐associated bleeding events. Benzamidine is a reversible inhibitor that binds to plasmin's active site. Herein, multivalent benzamidine inhibitors of varying valencies (mono‐, bi‐ and tri‐valent) and linker lengths (∼1–12 nm) were synthesized to systematically study their effect on plasmin inhibition. Inhibition assays were performed using a plasmin substrate (S‐2251) to determine inhibition constants (Ki). Pentamidine (shortest bivalent) and Tri‐AMB (shortest trivalent) were the strongest inhibitors with Ki values of 2.1±0.8 and 3.9±1.7 μM, respectively. Overall, increasing valency and decreasing linker length, increases effective local concentration of the inhibitor and therefore, resulted in stronger inhibition of plasmin via statistical rebinding. This study aids in the design of multivalent inhibitors that can achieve desired enzyme inhibition by means of modulating valency and linker length.

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Section: Resultsmentioning

confidence: 99%

Multivalent Benzamidine Molecules for Plasmin Inhibition: Effect of Valency and Linker Length

2022

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“…Figure 1A shows the IC 50 of the synthetic diZD to VEGFR2 protein as 0.31 ± 0.09 nM, which was consistent with the previously reported 100fold improvement of binding affinity as compared to the parental drug ZD6474 (vandetanib) as measured IC 50 of 40 nM with the same assay. 17,18 A similar cell-binding assay was also employed to validate the K d of 177 Lu-DOTA-diZD. Figure 1B shows the saturation curves of the 177 Lu-DOTA-diZD binding to VEGFR-positive HUVECs cells, and the one-site binding mathematical model calculated the K d of 177 Lu-DOTA-diZD as 0.54 ± 0.05 nM, which was comparable to the 64 Cu-DOTA-diZD.…”

Section: ■ Resultsmentioning

confidence: 99%

“…Herein, we radiolabeled diZD, a vascular endothelial growth factor receptor (VEGFR)-targeting agent previously reported by us, , with 177 Lu for an angiogenesis-based TRT and 64 Cu for PET imaging in a preclinical mouse model of TNBC. A remarkable antitumor efficacy was observed on orthotopic murine 4T1 tumor model, resulting in both improved local tumor control and prolonged survival.…”

Section: Introductionmentioning

confidence: 99%

⁶⁴Cu/¹⁷⁷Lu-DOTA-diZD, a Small-Molecule-Based Theranostic Pair for Triple-Negative Breast Cancer

Huang

Yang

et al. 2021

J. Med. Chem.

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Despite advances in targeted therapies, the prognosis for patients with triple-negative breast cancer (TNBC) is poor because there are few actionable molecular targets. The dependence of solid tumor growth on angiogenesis prompted our development of angiogenic-receptor-targeted radionuclide therapy (TRT) to treat TNBC by targeted delivery of therapeutic doses of ionizing radiation to tumors. A high-affinity vascular endothelial growth factor receptor (VEGFR)-targeted agent, diZD, was synthesized and labeled with 177 Lu and 64 Cu by 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator giving the TRT agent, 177 Lu-DOTA-diZD, and PET imaging agent, 64 Cu-DOTA-diZD. We showed that " 64 Cu/ 177 Lu"-DOTA-diZD radiotracers are a promising theranostic pair for TNBC. 4T1-bearing mice treated with 177 Lu-DOTA-diZD-based TRT survived with a median of 28 days, which was significantly longer than that of control mice as 18 days. Anti-PD1 immunotherapy resulted in a shorter median survival of 16 days. This work presents for the first time that small-molecule VEGFR-oriented TRT is a promising therapeutic option to treat "immunogenic cold" TNBC.

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“…PAMAM ‑Cy5.5 was synthesized as reported. , Briefly, Cy5.5-NHS (16 mg) was dissolved in 2 mL of DMSO. The Cy5.5-NHS solution was added dropwise into the PAMAM (100 mg) solution in 5 mL of DMSO and stirred at room temperature for 4 h. The free Cy5.5-NHS was removed by an ultrafiltration centrifuge tube (MWCO = 3000 Da).…”

Section: Materials and Methodsmentioning

confidence: 99%

Small Morph Nanoparticles for Deep Tumor Penetration via Caveolae-Mediated Transcytosis

Zhang

Wang

Yang

et al. 2020

ACS Appl. Mater. Interfaces

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The tumor penetration of nanomedicines constitutes a great challenge in the treatment of solid tumors, leading to the highly compromised therapeutic efficacy of nanomedicines. Here, we developed small morph nanoparticles (PDMA) by modifying polyamidoamine (PAMAM) dendrimers with dimethylmaleic anhydride (DMA). PDMA achieved deep tumor penetration via an active, energy-dependent, caveolae-mediated transcytosis, which circumvented the obstacles in the process of deep penetration. PDMA remained negatively charged under normal physiological conditions and underwent rapid charge reversal from negative to positive under acidic conditions in the tumor microenvironment (pH < 6.5), which enhanced their uptake by tumor cells and their deep penetration into tumor tissues in vitro and in vivo. The deep tumor penetration of PDMA was achieved mainly by caveolae-mediated transcytosis, which could be attributed to the small sizes (5–10 nm) and positive charge of the morphed PDMA. In vivo studies demonstrated that PDMA exhibited increased tumor accumulation and doxorubicin-loaded PDMA (PDMA/DOX) showed better antitumor efficacy. Overall, the small morph PDMA for enhanced deep tumor penetration via caveolae-mediated transcytosis could provide new inspiration for the design of anticancer drug delivery systems.

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Single‐Molecule Force Measurement Guides the Design of Multivalent Ligands with Picomolar Affinity

Cited by 4 publications

References 23 publications

Multivalent Benzamidine Molecules for Plasmin Inhibition: Effect of Valency and Linker Length

Multivalent Benzamidine Molecules for Plasmin Inhibition: Effect of Valency and Linker Length

⁶⁴Cu/¹⁷⁷Lu-DOTA-diZD, a Small-Molecule-Based Theranostic Pair for Triple-Negative Breast Cancer

Small Morph Nanoparticles for Deep Tumor Penetration via Caveolae-Mediated Transcytosis

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Single‐Molecule Force Measurement Guides the Design of Multivalent Ligands with Picomolar Affinity

Cited by 4 publications

References 23 publications

Multivalent Benzamidine Molecules for Plasmin Inhibition: Effect of Valency and Linker Length

Multivalent Benzamidine Molecules for Plasmin Inhibition: Effect of Valency and Linker Length

64Cu/177Lu-DOTA-diZD, a Small-Molecule-Based Theranostic Pair for Triple-Negative Breast Cancer

Small Morph Nanoparticles for Deep Tumor Penetration via Caveolae-Mediated Transcytosis

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⁶⁴Cu/¹⁷⁷Lu-DOTA-diZD, a Small-Molecule-Based Theranostic Pair for Triple-Negative Breast Cancer