Targeting Mitochondrial Guanine Quadruplexes for Photoactivatable Chemotherapy in Normoxic and Hypoxic Environments.

A first example of a mitochondrial G-quadruplex (mitoG4s) targeted Ru(II) photooxidant complex is reported. The complex, Ru-TAP-PDC3 induces photodamage toward guanine quadruplexes (G4s) located in the mitochondrial genome under hypoxic and normoxic conditions. Ru-TAP-PDC3 shows high affinity for mitoG4s and localises within mitochondria of live HeLa cells. Immunolabelling with anti-G4 antibody, BG4, confirms Ru-TAP-PDC3 associates with G4s within the mitochondria of fixed cells. The complex induces depletion of mtDNA in live cells under irradiation at 405 nm, confirmed by loss of PicoGreen signal from mitochondria. Biochemical studies confirm this process induces apoptosis. The complex shows low dark toxicity and an impressive phototoxicity index (PI) of >89 was determined in Hela under very low intensity irradiation, 5 J/cm2. The phototoxicity is thought to operate through both Type II singlet oxygen and Type III pathways depending on normoxic or hypoxic conditions from live cell imaging and plasmid DNA cleavage. Overall, we demonstrate targeting mitoG4s and mtDNA with a photooxidant is a potent route to achieving apoptosis under hypoxic conditions that can be extended to phototherapy.

Phototoxicity of Tridentate Ru(II) Polypyridyl Complex with Expanded Bite Angles toward Mammalian Cells and Multicellular Tumor Spheroids.

Tridentate ligand-coordinated ruthenium (II) polypyridyl complexes with large N-Ru-N bite angles have been shown to promote ligand field splitting and reduce singlet-triplet state mixing leading to dramatically extended emission quantum yields and lifetimes under ambient conditions. These effects are anticipated to enhance their photoinduced singlet oxygen production, promoting prospects for such complexes as type II phototherapeutics. In this contribution, we examined this putative effect for [Ru(bqp)(bqpCOOEt)]2+, Ru-bqp-ester, a heteroleptic complex containing bqp = [2,6-bi(quinolin-8-yl)pyridine], a well-established large bite angle tridentate ligand, as well as its peptide conjugates [Ru(bqp)(bqpCONH-ahx-FrFKFrFK(Ac)-CONH2)]5+ (Ru-bqp-MPP) and [Ru(bqp) (bqp)(CONH-ahx-RRRRRRRR-CONH2)]10+ (Ru-bqp-R8) that were prepared in an effort to promote live cell/tissue permeability and targeting of the parent. Membrane permeability of both parent and peptide conjugates were compared across 2D cell monolayers; A549, Chinese hamster ovary, human pancreatic cancer (HPAC), and 3D HPAC multicellular tumor spheroids (MCTS) using confocal microscopy. Both the parent complex and peptide conjugates showed exceptional permeability with rapid uptake in both 2D and 3D cell models but with little distinction in permeability or distribution in cells between the parent or peptide conjugates. Unexpectedly, the uptake was temperature independent and so attributed to passive permeation. Both dark and photo-toxicity of the Ru(II) complexes were assessed across cell types, and the parent showed notably low dark toxicity. In contrast, the parent and conjugates were found to be highly phototoxic, with impressive phototoxic indices (PIs) toward HPAC cell monolayers in particular, with PI values ranging from ∼580 to 760. Overall, our data indicate that the Ru(II) parent complex and its peptide conjugates show promise at both cell monolayers and 3D MCTS as photosensitizers for photodynamic therapy.

In Cellulo Light-Induced Dynamics in a BODIPY-Perylene Dyad.

BODIPY-based donor-acceptor dyads are widely used as sensors and probes in life science. Thus, their biophysical properties are well established in solution, while their photophysical properties in cellulo, i. e., in the environment, in which the dyes are designed to function, are generally understood less. To address this issue, we present a sub-ns time-resolved transient absorption study of the excited-state dynamics of a BODIPY-perylene dyad designed as a twisted intramolecular charge transfer (TICT) probe of the local viscosity in live cells.