Photodynamic Therapy (PDT) is generally recognized as a safe and effective strategy and is approved to treat several forms of cancer. PDT requires the simultaneous combination of a drug, molecular oxygen, and light of a specific wavelength to generate reactive oxygen species (ROS), which are responsible for the destruction of tumour cells [1
]. The antitumour effect of PDT is related to three distinct effects responsible for tumour cell death: oxidative damage promoted by ROS leading to apoptosis, autophagy or necrosis; shutdown of the tumour blood vessels that interrupts oxygen and nutrient supply; and eventually an antitumor immune response induced by PDT [3
]. A most often presented advantage of PDT over other cancer therapies is the use of drugs—called photosensitizers (PS)—that are inactive in the absence of light. It is expected that the PS used in PDT are not toxic, or immunogenic, before they are activated by light of a wavelength specifically absorbed by the PS. When the PS is administered with an injectable solvent, the concerns on the toxicity extend to the drug formulation. The toxicity of PS and drug formulation must be assessed before entering clinical trials.
Porfimer sodium (Photofrin®
) is currently the most used photosensitizer for systemic administration. It is clinically employed for the treatment of lung, oesophagus, bile duct, bladder, brain and ovarian tumours. Temoporfin (Foscan®
) is a photosensitizer clinically applied in Europe for the treatment of head & neck, lung, brain, skin and bile duct cancer. Verteporfin (Visudyne®
) met with considerable success as a photosensitizer in the treatment of age-related macular degeneration (AMD), and is also applied in PDT of prostate and pancreatic cancers. Every new drug candidate must be subjected to extensive nonclinical toxicology programs to demonstrate that it has acceptable profiles of tolerability and safety, and guarantee the lowest possible level of risk for the participants in the first-in-human trial. The general requirements for these toxicology studies are described in the harmonized guideline ICH M3(R2), from the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use, and are further detailed in the comprehensive set of guidelines related to safety (S guidelines) [5
]. PS for PDT may elicit intrinsic adverse reactions, may employ drug formulations with some toxicity, or may show toxicity as a consequence of the illumination protocol. This last concern is specific of PDT and requires the design of safety evaluation strategies that take into consideration the combination of drug and light [6
We have been involved in the development of a fluorinated sulphonamide bacteriochlorin named redaporfin—5,10,15,20-tetrakis (2,6-difluoro-3-N
-methylsulfamoylphenyl) bacteriochlorin, code name LUZ11—for PDT of cancer. Redaporfin presents near-ideal properties [8
] and proved to be highly effective in the treatment of mice with subcutaneous colon CT26 tumour, with significant antitumor effects on distant metastasis, attributed to its ability to activate the host immune system [9
]. Redaporfin is amphiphilic with an n
-octanol: water partition coefficient of 80 (logPOW
= 1.9) [8
]. Redaporfin is not soluble in water and tends to aggregate when a stock solution of redaporfin in an organic solvent (e.g., ethanol or dimethylsulfoxide) is added to water. In order to avoid aggregation, the redaporfin formulation contains a small amount of Cremophor EL®
(CrEL). CrEL has been extensively used and characterized to deliver drugs with log POW
> 1. CrEL forms micelles in aqueous solutions and its critical micellar concentration is 0.009% (weight/volume) in protein-free aqueous solution [11
]. It was noted that CrEL concentrations >0.03% in human serum lead to lipoprotein degradation [12
]. Hypersensitivity reactions of drugs formulated with CrEL have been associated with concentrations >0.2% in plasma of cancer patients [13
], but the adverse effects can be countered by pre-medication. Considering that the human blood plasma volume is 35 mL/kg, a total safe dose of CrEL should be <0.07 mL/kg. However, the low solubility of very lipophilic drugs may require an increase in the CrEL content in the drug formulation, which then requires pre-medication to control adverse effects. For example, the paclitaxel formulation is associated with the administration of 0.37 mL/kg of CrEL [14
], but hypersensitivity reactions can be avoided with, for example, pre-medication with dexamethasone [15
]. The minimum value of CrEL may alternatively be limited by its critical micellar concentration (CMC > 0.003 mL/kg). As will be shown below, our toxicology study employed a redaporfin formulation in CrEL/EtOH/NaCl 0.9% (1.2:5.7:93.1, v
) with a redaporfin concentration of 0.86 mg/mL to administer a redaporfin dose of 1.5 mg/kg, i.e.
, a CrEL dose of 0.021 mL/kg. This is above the CMC and well within the limits of the safe dose level. On the other hand, the dose escalation study explored redaporfin doses up to 75 mg/kg and the CrEL content of the formulation used in this study had to be increased to the proportion CrEL/EtOH/NaCl 0.9% (5:10:85, v
), leading to the a total CrEL dose of 0.5 mL/kg.
European Medicines Agency (EMA) recently granted the status of orphan drug designation for redaporfin-PDT of biliary track cancer [16
], and redaporfin entered clinical trials with patients with advanced head and neck cancer [17
]. This work reports preliminary safety studies designed to assess the safety and tolerability of redaporfin-PDT in the clinical trial application of redaporfin. The studies cover a dose escalation study in mice to evaluate the Maximum Tolerated Dose (MTD) of redaporfin in its formulation but without illumination, and a systemic toxicity study in rats, with and without illumination. Since the goal is for the redaporfin-PDT in the clinic to be effective with only one treatment, both studies focused on the acute reactions elicited by a single session of PDT. This work also presents an efficacy study designed to test a 1.5 mg/kg dose of redaporfin with a light dose of 74 J/cm2
in vascular-PDT, with a drug-light interval (DLI) of 15 min. These doses are higher than those used in previous studies with redaporfin [9
], and motivated the use of male, rather than female, BALB/c mice in this study, to take advantage of their larger size (25 g vs.
20 g). The use of higher doses in this study explores the relation between the onset of adverse effects and the size of the animal-models.