Two-photon excitation has attracted the attention of biologists, especially after the development of two-photon excited microscopy in the nineties. Since then, new applications have rapidly emerged such as the release of biologically active molecules and photodynamic therapy (PDT) using two-photon excitation. PDT, which requires a light-activated drug (photosensitiser), is a clinically approved and minimally invasive treatment for cancer and for non-malignant diseases. This feature article focuses on the engineering of molecular two-photon photosensitisers for PDT, which should bring important benefits to the treatment, increase the treatment penetration depth with near-infrared light excitation, improve the spatial selectivity and reduce the photodamage to healthy tissues. After an overview of the two-photon absorption phenomenon and the methods to evaluate two-photon induced phototoxicity on cell cultures, the different classes of photosensitisers described in the literature are discussed. The two-photon PDT performed with historical one-photon sensitisers are briefly presented, followed by specifically engineered cyclic tetrapyrrole photosensitisers, purely organic photosensitisers and transition metal complexes. Finally, targeted two-photon photosensitisers and theranostic agents that should enhance the selectivity and efficiency of the treatment are discussed.

A molecular theranostic agent for magnetic resonance imaging (MRI) and photodynamic therapy (PDT) consisting of four [GdDTTA](-) complexes (DTTA(4-) = diethylenetriamine-N,N,N″,N″-tetraacetate) linked to a meso-tetraphenylporphyrin core, as well as its yttrium(III) analogue, was synthesized. A variety of physicochemical methods were used to characterize the gadolinium(III) conjugate 1 both as an MRI contrast agent and as a photosensitizer. The proton relaxivity measured in H2O at 20 MHz and 25 °C, r1 = 43.7 mmol(-1) s(-1) per gadolinium center, is the highest reported for a bishydrated gadolinium(III)-based contrast agent of medium size and can be related to the rigidity of the molecule. The complex displays also a remarkable singlet oxygen quantum yield of ϕΔ = 0.45 in H2O, similar to that of a meso-tetrasulfonated porphyrin. We also evidenced the ability of the gadolinium(III) conjugate to penetrate in cancer cells with low cytotoxicity. Its phototoxicity on Hela cells was evaluated following incubation at low micromolar concentration and moderate light irradiation (21 J cm(-2)) induced 50% of cell death. Altogether, these results demonstrate the high potential of this conjugate as a theranostic agent for MRI and PDT.

A molecular theranostic agent designed for photodynamic therapy (PDT) treatment in the near-infrared and for imaging tissue tumors with magnetic resonance imaging (MRI) is reported. It consists of a linear p-conjugated Zn(II)-porphyrin dimer linked at each extremity to a GdDOTA-type complex. This agent has shown very promising potential for PDT applications with good singlet oxygen generation in DMSO and high linear absorption in the near-infrared (lmax = 746 nm, e ≈ 10 5 M -1 cm -1 ). Moreover, this molecule has a propensity for two-photon excited PDT with high two-photon cross-sections (≈ 8000 GM in 880-930 nm range), which should allow for deeper tumor treatments and higher spatial precision as compared to conventional one-photon PDT. Regarding the MRI contrast agent properties, the molecule has shown superior relaxivity (14.4 mM -1 s -1 at 40 MHz, 298 K) in comparison to clinical contrast agents and ability to be internalized in cells, thanks to its amphiphilic character.Irradiation of HeLa cells using either one-photon (740 nm) or two-photon excitation (910 nm) has led in both cases to important cell death.