Redox-inducible Radiomimetic Photosensitizers Selectively Suppress Cancer Cell Proliferation by Damaging DNA through Radical Cation Chemistry.

Radiotherapy leverages ionizing radiation to kill cancer cells through direct and indirect effects, and direct effects are considered to play an equal or greater role. Several photosensitizers have been developed to mimic the direct effects of radiotherapy, generating radical cations in DNA models, but none has been applied in cellular studies. Here, we design a radiomimetic photosensitizer, producing DNA radical cations in cells for the first time. To reduce adverse effects, several redox-inducible precursors are prepared as cancer cells have elevated levels of GSH and H2O2. These precursors respond to GSH or H2O2, releasing the active photosensitizer that captures DNA abasic (AP) sites and generates DNA radical cations upon photolysis, without disrupting the redox state of cells. DNA radical cations migrate freely and are eventually trapped by H2O and O2 to yield DNA lesions, thus triggering DNA damage response. Our study suggests that direct effects of radiotherapy suppress cancer cell proliferation mainly by inducing G2/M phase cell cycle arrest, rather than promoting apoptosis. Synergistic effects of the precursor and chemotherapeutic agents are also observed in combination phototherapy. Beyond highlighting an alternative strategy for phototherapy, this proof-of-concept study affords a facile cellular platform to study the direct effects of radiotherapy.

Hydrogen Peroxide-Inducible PROTACs for Targeted Protein Degradation in Cancer Cells.

Proteolysis-targeting chimeras (PROTACs) provide a powerful technique to degrade targeted proteins utilizing the cellular ubiquitin-proteasome system. The major concern is the host toxicity resulting from their poor selectivity. Inducible PROTACs responding to exogenous stimulus, such as light, improve their specificity, but it is difficult for photo-activation in deep tissues. Herein, we develop H2 O2 -inducible PROTAC precursors 2/5, which can be activated by endogenous H2 O2 in cancer cells to release the active PROTACs 1/4 to effectively degrade targeted proteins. This results in the intended cytotoxicity towards cancer cells while targeted protein in normal cells remains almost unaffected. The higher Bromodomain-containing protein 4 (BRD4) degradation activity and cytotoxicity of 2 towards cancer cells is mainly due to the higher endogenous concentration of H2 O2 in cancer cells (A549 and H1299), characterized by H2 O2 -responsive fluorescence probe 3. Western blot assays and cytotoxicity experiments demonstrate that 2 degrades BRD4 more effectively and is more cytotoxic in H2 O2 -rich cancer cells than in H2 O2 -deficient normal cells. This method is also extended to estrogen receptor (ER)-PROTAC precursor 5, showing H2 O2 -dependent ER degradation ability. Thus, we establish a novel strategy to induce targeted protein degradation in a H2 O2 -dependent way, which has the potential to improve the selectivity of PROTACs.

Light and hydrogen peroxide dual-responsive DNA interstrand crosslink precursors with potent cytotoxicity.

Arylboronic acid/esters and phenyl selenides-based quinone methide (QM) precursors were reported to induce DNA interstrand crosslink (ICL) formation upon reaction with the inherently high concentrations of H2O2 in cancer cells. However, some normal cells (such as macrophages) also contain high-levels of H2O2, which may interfere with precursors' selectivity. In order to enhance the spatiotemporal specificity by the photolysis, we developed photo- and H2O2- dual-responsive DNA ICL precursors 1-3, bearing a photo-responsive coumarin moiety and a H2O2 inducible phenyl selenide group. Precursors 1-3 are efficiently activated by photoirradiation and H2O2 to generate reactive QMs crosslinking DNA. Moreover, the reactivity of precursors can be modulated by the introduction of aromatic substituents (OMe, F), and the electron donating group (OMe) displays a more pronounced promoting effect on DNA ICL formation. A subsequent piperidine heat stability study confirmed that the formed QMs primarily alkylate dAs, dGs and dCs in DNA. Furthermore, 1-3 inhibit lung cancer cell (H1299) growth by inducing DNA damage and producing toxic reactive oxygen species (ROS) upon photolysis of released coumarin. This study illustrates the potent cytotoxicity achieved by novel photo/H2O2 dual-responsive QM precursors 1-3, affording a novel strategy for the development of inducible DNA interstrand cross-linkers.

Dual-responsive probe and DNA interstrand crosslink precursor target the unique redox status of cancer cells.

Elevated GSH and H2O2 in cancer cells is sometimes doubted due to their contrary reactivities. Here, we construct a dual-responsive fluorescent probe to confirm the conclusion, and employ this to exploit a redox-inducible DNA interstrand crosslink (ICL) precursor. It crosslinks DNA upon activation by GSH and H2O2, affording an alternative dual-responsive strategy.

Assessment of Phenylboronic Acid Nitrogen Mustards as Potent and Selective Drug Candidates for Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) has limited treatment options and the worst prognosis among all types of breast cancer. We describe two prodrugs, namely, CWB-20145 (1) and its methyl analogue FAN-NM-CH3 (2) that reduced the size of TNBC-derived tumors. The DNA cross-linking of nitrogen mustard prodrugs 1 and 2 was superior to that of chlorambucil and melphalan once activated in the presence of H2O2. The cellular toxicity of 1 and 2 was demonstrated in seven human cancer cell lines. The TNBC cell line MDA-MB-468 was particularly sensitive toward 1 and 2. Compound 2 was 10 times more cytotoxic than chlorambucil and 16 times more active than melphalan. An evaluation of the gene expression demonstrated an upregulation of the tumor suppressor genes p53 and p21 supporting a transcriptional mechanism of a reduced tumor growth. Pharmacokinetic studies with 1 showed a rapid conversion of the prodrug. The introduction of a methyl group generated 2 with an increased half-life. An in vivo toxicity study in mice demonstrated that both prodrugs were less toxic than chlorambucil. Compounds 1 and 2 reduced tumor growth with an inhibition rate of more than 90% in athymic nude mice xenografted with MDA-MB-468 cells. Together, the in vivo investigations demonstrated that treatment with 1 and 2 suppressed tumor growth without affecting normal tissues in mice. These phenylboronic acid nitrogen mustard prodrugs represent promising drug candidates for the treatment of TNBC. However, the mechanisms underlying their superior in vivo activity and selectivity as well as the correlation between H2O2 level and in vivo efficacy are not yet fully understood.