Bioorg Med Chem Lett. 2018 May 1;28(8):1363-1370. doi: 10.1016/j.bmcl.2018.03.005. Epub 2018 Mar 3. Site-specific and hydrophilic ADCs through disulfide-bridged linker and branched PEG Abstract Kadcyla® (T-DM1), an antibody-drug conjugates (ADCs) for HER2+ breast cancer treatment, has been approved by the Food and Drug Administration (FDA) in 2013. An ADC of random lysine conjugation, it has difficulties in DAR control and unsatisfactory PK due to uneven DAR distribution. It also gives rise to aggregation during conjugation because of the hydrophobicity nature of the cytotoxin, DM1. The linker-drug in T-DM1, SMCC-DM1 is hydrophobic and requires certain percentage of organic solvent such as DMA in the conjugation solution, limiting the manufacturing process in an organic-solvent-compatible device and adding extra costs. To address these problems, a site-specific conjugation method was developed involving full reduction of antibody and full conjugation with the bridge-like conjugator-drug, based on the work of Caddick and co-workers, to obtain a site-directed antibody-drug conjugate with DAR 4. The bridge-like conjugator was assembled with SMCC-DM1 and different lengths of hydrophilic polyethylene glycol (PEG) moiety. By applying PEG moiety in the side chain of the linker-drug, the organic solvent used in the conjugation can be reduced. When the PEG length is about 26 units, organic solvent is no longer needed in the conjugation. Reducing the amount of organic solvent in conjugation could also diminish the aggregation occurrence during the conjugation. Moreover, the conjugation configuration with the designed conjugator was also discussed in the article. The binding affinity of the resulting ADCs did not show significant decrease and the cell based assay and animal study have shown the comparable results with T-DM1. Keywords: Antibody-drug conjugates; Maytansine DM1; PEGlyation; Site-specific. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

ACS Omega. 2023 Aug 23;8(35):32043-32052. doi: 10.1021/acsomega.3c03952. eCollection 2023 Sep 5. Cytocompatibility Evaluation of PEG-Methylsulfone Hydrogels Abstract Methylsulfone derivatized poly(ethylene) glycol (PEG) macromers can be biofunctionalized with thiolated ligands and cross-linked with thiol-based cross-linkers to obtain bioactive PEG hydrogels for in situ cell encapsulation. Methylsulfonyl-thiol (MS-SH) reactions present several advantages for this purpose when compared to other thiol-based cross-linking systems. They proceed with adequate and tunable kinetics for encapsulation, they reach a high conversion degree with good selectivity, and they generate stable reaction products. Our previous work demonstrated the cytocompatibility of cross-linked PEG-MS/thiol hydrogels in contact with fibroblasts. However, the cytocompatibility of the in situ MS-SH cross-linking reaction itself, which generates methylsulfinic acid as byproduct at the cross-linked site, remains to be evaluated. These studies are necessary to evaluate the potential of these systems for in vivo applications. Here we perform an extensive cytocompatibility study of PEG hydrogels during in situ cross-linking by the methylsulfonyl-thiol reaction. We compare these results with maleimide-thiol cross-linked PEGs which are well established for cell culture and in vivo experiments and do not involve the release of a byproduct. We show that fibroblasts and endothelial cells remain viable after in situ polymerization of methylsulfonyl-thiol gels on the top of the cell layers. Cell viability seems better than after in situ cross-linking hydrogels with maleimide-thiol chemistry. The endothelial cell proinflammatory phenotype is low and similar to the one obtained by the maleimide-thiol reaction. Finally, no activation of monocytes is observed. All in all, these results demonstrate that the methylsulfonyl-thiol chemistry is cytocompatible and does not trigger high pro-inflammatory responses in endothelial cells and monocytes. These results make methylsulfonyl-thiol chemistries eligible for in vivo testing and eventually clinical application in the future. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

J Control Release. 2024 Nov:375:74-89. doi: 10.1016/j.jconrel.2024.08.049. Epub 2024 Sep 5. Optimization of a pendant-shaped PEGylated linker for antibody-drug conjugates Abstract In this work, we conceived and developed antibody-drug conjugates (ADCs) that could efficiently release the drug after enzymatic cleavage of the linker moiety by tumoral proteases. The antibody-drug linkers we used are the result of a rational optimization of a previously reported PEGylated linker, PUREBRIGHT® MA-P12-PS, which showed excellent drug loading capacities but lacked an inbuilt drug discharge mechanism, thus limiting the potency of the resulting ADCs. To address this limitation, we chose to incorporate a protease-sensitive trigger into the linker to favor the release of a "PEGless" drug inside the tumor cells and, therefore, obtain potent ADCs. Currently, most marketed ADCs are based on the Val-Cit dipeptide followed by a self-immolative spacer for releasing the drug in its unmodified form. Here, we selected two untraditional peptide sequences, a Phe-Gly dipeptide and a Val-Ala-Gly tripeptide and placed one or the other in between the drug on one side (N-terminus) and the rest of the linker, including the PEG moiety, on the other side (C-terminus), without a self-immolative group. We found that both linkers responded to cathepsin B, a reference lysosomal enzyme, and liberated a PEG-free drug catabolite, as desired. We then used the two linkers to generate ADCs based on trastuzumab (a HER2-targeting antibody) and DM1 (a microtubule-targeted cytotoxic agent) with an average drug-to-antibody ratio (DAR) of 4 or 8. The ADCs showed restored cytotoxicity in vitro, which was proportional to the DM1 loading and generally higher for the ADCs bearing Val-Ala-Gly in their structure. In an ovarian cancer mouse model, the DAR 8 ADC based on Val-Ala-Gly behaved better than Kadcyla® (an approved ADC of DAR 3.5 used as control throughout this study), leading to a higher tumor volume reduction and more prolonged median survival. Taken together, our results depict a successful linker optimization process and encourage the application of the Val-Ala-Gly tripeptide as an alternative to other existing protease-sensitive triggers for ADCs. Keywords: Antibody-drug conjugate; Anticancer therapy; Cleavable peptide linker; Drug delivery; Optimization; PEG. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

Molecules. 2022 Dec 3;27(23):8501. doi: 10.3390/molecules27238501. Development of Phosphoramidite Reagents for the Synthesis of Base-Labile Oligonucleotides Modified with a Linear Aminoalkyl and Amino-PEG Linker at the 3'-End Abstract Oligonucleotides with an amino linker at the 3'-end are useful for the preparation of conjugated oligonucleotides. However, chemically modified nucleosides, which are unstable under basic conditions, cannot be incorporated into oligonucleotides using the conventional method entailing the preparation of oligonucleotides bearing a 3'-amino linker. Therefore, we designed Fmoc-protected phosphoramidites for the synthesis of base-labile oligonucleotides modified with a 3'-amino linker. The resultant phosphoramidites were then successfully incorporated into oligonucleotides bearing a 3'-amino linker. Various basic solutions were investigated for protecting group removal. All the protecting groups were removed by treating the oligonucleotides with 40% aqueous methylamine at room temperature for 2 h. Thus, the deprotection time and temperature were significantly reduced compared to the conventional conditions (28% NH3 aq., 55 °C, 17 h). In addition, the oligonucleotide protecting groups could be removed using a mild base (e.g., 50 mM potassium carbonate methanol solution). Furthermore, base-labile oligonucleotides bearing an amino linker at the 3'-end were successfully synthesized using the developed phosphoramidite reagents, highlighting the utility of our strategy. Keywords: 3′-modification; amino linker; base-labile oligonucleotide; conjugate. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

Br J Haematol. 2020 May;189(3):442-451. doi: 10.1111/bjh.16254. Epub 2019 Dec 27. Acute lymphoblastic leukaemia patients treated with PEGasparaginase develop antibodies to PEG and the succinate linker Abstract Polyethylene glycol (PEG) conjugated asparaginase (PEGasparaginase) is essential for treatment of paediatric acute lymphoblastic leukaemia. We developed an assay identifying antibodies against the PEG-moiety, the linker and the drug itself in patients experiencing hypersensitivity reactions to PEGasparaginase. Eighteen patients treated according to the DCOG ALL-11 protocol, with a neutralizing hypersensitivity reaction to PEGasparaginase to the first PEGasparaginase doses in induction (12 patients) or during intensification after interruption of several months (6 patients) were included. ELISA was used to measure antibodies, coating with the succinimidyl succinate linker conjugated to BSA, PEGfilgrastim and Escherichia coli asparaginase, and using hydrolysed PEGasparaginase and mPEG5,000 for competition. Anti-PEG antibodies were detected in all patients (IgG 100%; IgM 67%) of whom 39% had anti-PEG antibodies exclusively. Pre-existing anti-PEG antibodies were also detected in patients who not previously received a PEGylated therapeutic (58% IgG; 21% IgM). Antibodies against the SS-linker were predominantly detected during induction (50% IgG; 42% IgM). Anti-asparaginase antibodies were detected in only 11% during induction but 94% during intensification. In conclusion, anti-PEG and anti-SS-linker antibodies predominantly play a role in the immunogenic response to PEGasparaginase during induction. Thus, switching to native E. coli asparaginase would be an option for adequate asparaginase treatment. Keywords: PEGasparaginase; acute lymphoblastic leukemia; antibodies. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

Review Nanomedicine (Lond). 2016 Apr;11(7):851-65. doi: 10.2217/nnm.16.28. Gold nanoparticle surface functionalization: mixed monolayer versus hetero bifunctional peg linker Abstract To create a clinically relevant gold nanoparticle (AuNP) treatment, the surface must be functionalized with multiple ligands such as drugs, antifouling agents and targeting moieties. However, attaching several ligands of differing chemistries and lengths, while ensuring they all retain their biological functionality remains a challenge. This review compares the two most widely employed methods of surface cofunctionalization, namely mixed monolayers and hetero-bifunctional linkers. While there are numerous in vitro studies successfully utilizing both surface arrangements, there is little consensus regarding their relative merits. Animal and preclinical studies have demonstrated the effectiveness of mixed monolayer functionalization and while some promising in vitro results have been reported for PEG linker capped AuNPs, any potential benefits of the approach are not yet fully understood. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

J Mater Chem B. 2021 Apr 7;9(13):2993-2997. doi: 10.1039/d0tb02911d. Epub 2021 Mar 16. A short PEG linker alters the in vivo pharmacokinetics of trastuzumab to yield high-contrast immuno-PET images Abstract The prolonged blood circulation of the radiolabeled antibody conjugates is problematic when using immuno-PET imaging due to the increased radiation exposure and longer hospitalization required until sufficient contrast develops. In contrast to the prevailing belief that PEGylation prolongs blood retention time, we observed that a PEGylated antibody with a short PEG8 linker cleared much faster from the blood while maintaining tumor uptake compared to its non-PEGylated counterpart. Breast tumors were clearly visualized with a very high tumor-to-background ratio as early as 24 h after injection in immuno-positron emission tomography (PET) imaging. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com

Biosens Bioelectron. 2019 Sep 15:141:111477. doi: 10.1016/j.bios.2019.111477. Epub 2019 Jun 25. Novel thiolated-PEG linker molecule for biosensor development on gold surfaces Abstract The surface modifying linker molecules can directly influence the performance and longevity of biosensors. They must allow the attachment of biological recognition layer on the sensor surface, as well as the protection of the surface from fouling effects. Recent advances in this field identified several key factors that can increase the efficiency, stability and the anti-fouling effect of a layer formed by surface modifying linker molecules. Herein, this work presents a simple synthetic procedure, characterization, and application of a novel thiolated-PEG surface modifying molecule (DSPEG2) that could act as a multi-purpose linker for gold surfaces. The analyses of the molecular spatial distribution of DSPEG2 on gold surfaces were performed using time-of-flight secondary ion mass spectrometry (TOF-SIMS) imaging and X-ray photoelectric spectroscopy (XPS). The immobilization of DSPEG2 on gold surfaces was examined using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and surface plasmon resonance (SPR). Our preliminary results demonstrated that DSPEG2 is a promising novel linker molecule that can be applied in a wide range of biosensors based on gold surfaces. Keywords: Anti-fouling; Biosensor; Cyclic voltammetry; Electrochemical impedance spectroscopy; Non-specific adsorption; PEG; Surface plasmon resonance; Synthetic linker. For more product information, please contact us at: US Tel: 1-844-782-5734 US Tel: 1-844-QUAL-PEG CHN Tel: 400-918-9898 Email: sales@sinopeg.com