Visible-Light-Driven Photocatalytic Initiation of Radical Thiol–Ene Reactions Using Bismuth Oxide

A nontoxic and inexpensive photocatalytic initiation of anti-Markovnikov hydrothiolation of olefins using visible light is reported. This method is characterized by low catalyst loading, thereby enabling a mild and selective method for radical initiation in thiol–ene reactions between a wide scope of olefins and thiols

Determination of Antibody–Drug Conjugate Released Payload Species Using Directed in Vitro Assays and Mass Spectrometric Interrogation

Antibody-drug conjugates (ADC) are currently an active area of research, focused primarily on oncology therapeutics, but also to a limited extent on other areas such as infectious disease. The success of this type of targeted drug delivery is dependent upon many factors, one of which is the performance of the linker in releasing an active drug moiety under the appropriate conditions. As a tool in the development of linker/payload chemistry, we have developed an in vitro method for the identification of payload species released from ADCs in the presence of lysosomal enzymes. This method utilizes commercially available human liver S9 fraction as the source of these enzymes, and this has certain advantages over lysosomal fractions or purified enzymes. This article describes the characterization and performance of this assay with multiple ADCs composed of known and novel linkers and payloads. Additionally, we report the observation of incomplete degradation of mAb protein chains by lysosomal enzymes in vitro, believed to be the first report of this phenomenon involving an ADC therapeutic

Characterization of cGAS enzyme activity.

<p>Measurement of cGAMP production was conducted by LC-MS as described in Methods. (A) Time course of cGAS (15 nM) activity; (B) titration of dsDNA activation of cGAS (1nM) activity; (C) cGAS enzyme titration; (D) inhibition of cGAS (1 nM) activity by CuBr.</p

Binding affinities and <i>in vitro</i> activities of cGAS inhibitors.

<p>Binding affinities and <i>in vitro</i> activities of cGAS inhibitors.</p

Characterization of cGAMP FP assay.

<p>(A) mAb titration with Cy5-cGAMP (2 nM); (B) competition of Cy5-cGAMP (2 nM) binding to mAb 80–2 with: cGAMP, cAMP, cGMP, ATP or GTP; (C) Z’ results of FP assay in subset screen; (D) Distribution of compound activity from subset screen.</p

Characterization of compound 15 binding to cGAS.

<p>(A) 1D ¹H spectra of 2´,3´-cGAMP (top) and ¹H STD of 2´,3´-cGAMP interacting with cGAS (bottom). (B) 1D ¹H spectra of 2´,3´-cGAMP (orange) and compound 15 (green) (top) and ¹H STD of a mixture of 2´,3´-cGAMP and compound <b>15</b> showing compound <b>15</b> has out competed 2´,3´-cGAMP for interacting with cGAS (bottom). (C) SPR sensorgram of compound <b>15</b> with binding fit inset. (D) Compound <b>15</b> in either its hydroxyl (15a) or keto (15b) tautomeric forms. (E) cGAS active site showing residues that interact with compound <b>15</b>; Fo-Fc electron density omit map (green) for compound <b>15</b> (brown) is contoured at 3 Sigma and shows all density within 4 Å of compound <b>15</b>.</p