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1.
Biochemistry ; 63(5): 644-650, 2024 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-38350078

RESUMO

The concept of tag-free protein modification has attracted considerable interest in chemical biology because of its flexible and straightforward reaction process. In 2021, a groundbreaking approach using lipoate ligase A (LplA) for tag-free enzymatic modification of antibodies was unveiled, demonstrating its potential for the generation of precise antibody conjugates. In this study, to further explore LplA-mediated antibody-drug conjugate (ADC) synthesis, we performed initial biological evaluations of ADCs synthesized using LplA. Using the anti-HER2 antibody trastuzumab, we introduced octanoic acid azide using LplA and subsequently obtained an ADC using click chemistry with the drug DBCO-VC-PAB-MMAE. The bioactivity of the synthesized anti-HER2-ADC was evaluated using HER2-positive SKBR-3 and HER2-negative MCF7 cells. Its toxicity and selectivity were found to be comparable to those of the FDA-approved Kadcyla. In addition, a stability study involving rat and human plasma demonstrated the stability of the LplA-mediated ADC. Additionally, the affinity for the neonatal Fc receptor (FcRn) was retained after conjugation. These preliminary in vitro evaluations suggested that LplA-derived ADCs can have considerable pharmaceutical potential. Our results can set the stage for further in vivo evaluations and safety assessments. We suggest that the integration of tag-free LplA methods into the production of ADCs can offer a novel and promising approach for biopharmaceutical manufacturing.


Assuntos
Antineoplásicos , Imunoconjugados , Ratos , Animais , Humanos , Ligases , Imunoconjugados/farmacologia , Antineoplásicos/farmacologia , Células MCF-7 , Trastuzumab/farmacologia , Linhagem Celular Tumoral
2.
J Am Chem Soc ; 2023 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-36757843

RESUMO

Complexity in supramolecular polymer systems arises from interactions between different components, including solvent molecules. By varying their concentration or temperature in such multicomponent systems, complex phenomena can occur such as thermally bisignate and dilution-induced assembly of supramolecular polymers. Herein, we demonstrate that both these phenomena emerge from the same underlying interaction mechanism between the components. As a model system, amide-decorated supramolecular polymers of porphyrins were investigated in combination with aliphatic alcohols as hydrogen-bond scavengers, and thermodynamic mass-balance models were applied to map the three-dimensional assembly landscapes. These studies unveiled that the interaction between hydrogen-bond scavengers and monomers is temperature-dependent and becomes dominant at high monomer concentrations. With these insights, we could exploit competitive monomer-alcohol interactions to prompt the dilution-induced assembly of various common monomers as well as bisignate assembly events. Moreover, kinetic insights were obtained by navigating through the assembly landscape. Similar to phase diagrams of covalent polymers, these assembly landscapes provide a comprehensive picture of supramolecular polymerizations, which helps to precisely regulate the system properties. The generality of this approach using assembly landscapes makes it relevant for any supramolecular system, and this enhanced control will open the door to build complex and functional supramolecular polymer systems.

3.
J Am Chem Soc ; 144(16): 7080-7084, 2022 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-35385273

RESUMO

Using a photochemically fluttering thiophene-fused cyclooctatetraene derivative (COT) as a nonplanar chiral monomer, we have succeeded in remotely suspending the supramolecular polymerization in a temporal manner by a completely new strategy. The COT monomer with an 8π electron core adopts a saddle shape in the ground state and flutters 5.8 × 103 times faster upon photoirradiation than in the dark as a result of the stabilized planar conformation by the excited-state aromaticity (Baird aromaticity). Detailed investigation revealed that without photoirradiation the rate constant of the fluttering motion is 1/560 times smaller than that of the chain elongation, indicating that the fluttering of COT does not affect the chain elongation in the dark. In contrast, under photoirradiation (365 nm), the fluttering of COT is at least 11 times more rapid than the chain elongation, thereby suppressing the elongation event. The rapid fluttering of COT to suspend the chain elongation is not accompanied by a decrease in active monomer concentration, leading to depolymerization.


Assuntos
Tiofenos , Polimerização , Tiofenos/química
4.
Phys Chem Chem Phys ; 24(27): 16586-16595, 2022 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-35789351

RESUMO

The initial states of the secondary processes of X-ray emission spectroscopy (XES) and resonant inelastic X-ray scattering (RIXS) are highly excited eigenstates with a deep core hole after a X-ray photoelectron spectroscopy (XPS) process and a X-ray photoabsorption spectroscopy (XAS) process, respectively, so that the XES and RIXS calculation offers a good example of extended quasiparticle theory (EQPT) (K. Ohno, S. Ono and T. Isobe, J. Chem. Phys., 2017, 146, 084108) which is applicable to any initial exited eigenstate. We apply the standard one-shot GW + Bethe-Salpeter equation (BSE) approach in MBPT to this problem on the basis of EQPT and analyze XES and RIXS spectra for CH4, NH3, H2O, and CH3OH molecules. We also suggest a simpler approach only using the GW calculation without solving the BSE to compute the XES and RIXS energies, although it cannot give the spectral intensity. Moreover, according to extended Kohn-Sham theory (T. Nakashima, H. Raebiger and K. Ohno, Phys. Rev. B, 2021, 104, L201116), we give a justification and comment of applying the method relying on time-dependent density functional theory as well as the one-shot GW + BSE approach to this problem.

5.
J Am Chem Soc ; 143(13): 5121-5126, 2021 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-33755459

RESUMO

In stacking-based supramolecular polymerization, chiral hydrogen bonding (H-bonding) monomers often prefer to adapt a homochiral monomer sequence. Herein, we investigated the polymerization of a chiral thiophene-fused cyclooctatetraene (COT) as a novel nonplanar-core monomer and found the first example of the formation of an alternating heterochiral supramolecular copolymer. Although single enantiomer (-) or (+)-COT alone did not polymerize, when (-) and (+)-COT were mixed together, supramolecular polymerization took place to give a stereochemically alternating copolymer. By means of the microcrystal electron crystallography of a shorter side-chained COT analogue, we found that the resulting heterochiral supramolecular copolymer adapted an alternating arrangement of H-bonded and polar π-stacked parts. A computational study using density-functional theory (DFT) suggested that such an alternating heterochiral preference occurs because it allows two thiophene amide moieties facing each other to effectively cancel their in-plane dipole moments.

6.
Dev Biol ; 434(1): 96-107, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29208373

RESUMO

In early zebrafish development, the program for dorsal axis formation begins soon after fertilization. Previous studies suggested that dorsal determinants (DDs) localize to the vegetal pole, and are transported to the dorsal blastomeres in a microtubule-dependent manner. The DDs activate the canonical Wnt pathway and induce dorsal-specific genes that are required for dorsal axis formation. Among wnt-family genes, only the wnt8a mRNA is reported to localize to the vegetal pole in oocytes and to induce the dorsal axis, suggesting that Wnt8a is a candidate DD. Here, to reveal the roles of maternal wnt8a, we generated wnt8a mutants by transcription activator-like effector nucleases (TALENs), and established zygotic, maternal, and maternal zygotic wnt8a mutants by germ-line replacement. Zebrafish wnt8a has two open reading frames (ORF1 and ORF2) that are tandemly located in the genome. Although the zygotic ORF1 or ORF2 wnt8a mutants showed little or no axis-formation defects, the ORF1/2 compound mutants showed antero-dorsalized phenotypes, indicating that ORF1 and ORF2 have redundant roles in ventrolateral and posterior tissue formation. Unexpectedly, the maternal wnt8a ORF1/2 mutants showed no axis-formation defects. The maternal-zygotic wnt8a ORF1/2 mutants showed more severe antero-dorsalized phenotypes than the zygotic mutants. These results indicated that maternal wnt8a is dispensable for the initial dorsal determination, but cooperates with zygotic wnt8a for ventrolateral and posterior tissue formation. Finally, we re-examined the maternal wnt genes and found that Wnt6a is an alternative candidate DD.


Assuntos
Proteínas do Citoesqueleto/metabolismo , Embrião não Mamífero/embriologia , Fases de Leitura Aberta/fisiologia , RNA Mensageiro/metabolismo , Proteínas Wnt/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Animais , Animais Geneticamente Modificados/embriologia , Animais Geneticamente Modificados/genética , Proteínas do Citoesqueleto/genética , RNA Mensageiro/genética , Proteínas Wnt/genética , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
7.
ACS Med Chem Lett ; 14(12): 1767-1773, 2023 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-38116449

RESUMO

Bispecific antibodies (BisAbs) are biotherapeutics that amalgamate the specificities of two distinct antibodies into one molecule, however, their engineering requires genetic modification and remains time-consuming. Therefore, we used AJICAP second-generation technology, which drives the production of site-specific conjugation without genetic modification requirements, to generate BisAbs. Using haloketone chemistry as an alternative to maleimide chemistry, we successfully produced site-specific antibody conjugates. Pharmacokinetic studies revealed that the haloketone-based antibody conjugate was stable in the rat plasma. The resultant BisAbs were rigorously evaluated, and surface plasmon resonance measurements and flow cytometry analyses confirmed that the antigen binding remained intact. Additionally, the affinity for the neonatal Fc receptor (FcRn) was retained after conjugation. Further cytotoxicity evaluation emphasized the pronounced activity of the generated BisAbs. This novel approach introduces a fully chemical, site-specific strategy capable of producing BisAbs, heralding a new era in the field of biotherapeutics.

8.
Chem Asian J ; 17(10): e202200223, 2022 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-35338598

RESUMO

Here, we report a medium-to-polymer anomalous chiral transfer in the supramolecular polymerization of a tetraphenylporphyrin-based achiral hydrogen-bonding monomer (TPP) in a chiral medium of 5-cyanobiphenyl CB*. A mixture of TPP in (R)-CB* ([TPP]=7.7 mol %) at 40 °C gave a columnar oblique LC mesophase, where the individual columns were composed of an optically active helical supramolecular polymer of TPP as a consequence of a successful medium-to-polymer chiral transfer. Meanwhile, upon dilution of CB* with achiral 5-cyanobiphenyl CB, the optical activity of the system showed an anomalous bell-shaped dependency on the composition of CB*/CB, where the gabs value of 0.049 at CB*/CB=50/50 was 6.0 times larger than the gabs value of CB* alone. Such anomalous chiroptical amplification in CD is most likely due to a change in the stacking geometry of TPP in the oblique columnar LC upon lateral compression.


Assuntos
Polímeros , Ligação de Hidrogênio , Polimerização
9.
Science ; 376(6594): 738-743, 2022 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-35549437

RESUMO

Ultrafast water permeation in aquaporins is promoted by their hydrophobic interior surface. Polytetrafluoroethylene has a dense fluorine surface, leading to its strong water repellence. We report a series of fluorous oligoamide nanorings with interior diameters ranging from 0.9 to 1.9 nanometers. These nanorings undergo supramolecular polymerization in phospholipid bilayer membranes to form fluorous nanochannels, the interior walls of which are densely covered with fluorine atoms. The nanochannel with the smallest diameter exhibits a water permeation flux that is two orders of magnitude greater than those of aquaporins and carbon nanotubes. The proposed nanochannel exhibits negligible chloride ion (Cl-) permeability caused by a powerful electrostatic barrier provided by the electrostatically negative fluorous interior surface. Thus, this nanochannel is expected to show nearly perfect salt reflectance for desalination.


Assuntos
Aquaporinas , Nanotubos de Carbono , Politetrafluoretileno , Purificação da Água , Água , Aquaporinas/química , Interações Hidrofóbicas e Hidrofílicas , Bicamadas Lipídicas/química , Nanotubos de Carbono/química , Permeabilidade , Sais/isolamento & purificação , Propriedades de Superfície
10.
J Phys Condens Matter ; 30(21): 21LT01, 2018 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-29651994

RESUMO

It has been highly desired to provide an accurate and reliable method to calculate core electron binding energies (CEBEs) of crystals and to understand the final state screening effect on a core hole in high resolution x-ray photoelectron spectroscopy (XPS), because the ΔSCF method cannot be simply used for bulk systems. We propose to use the quasiparticle calculation based on many-body perturbation theory for this problem. In this study, CEBEs of band-gapped crystals, silicon, diamond, ß-SiC, BN, and AlP, are investigated by means of the GW approximation (GWA) using the full ω integration and compared with the preexisting XPS data. The screening effect on a deep core hole is also investigated in detail by evaluating the relaxation energy (RE) from the core and valence contributions separately. Calculated results show that not only the valence electrons but also the core electrons have an important contribution to the RE, and the GWA have a tendency to underestimate CEBEs due to the excess RE. This underestimation can be improved by introducing the self-screening correction to the GWA. The resulting C1s, B1s, N1s, Si2p, and Al2p CEBEs are in excellent agreement with the experiments within 1 eV absolute error range. The present self-screening corrected GW approach has the capability to achieve the highly accurate prediction of CEBEs without any empirical parameter for band-gapped crystals, and provide a more reliable theoretical approach than the conventional ΔSCF-DFT method.

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