Background: Topical 5-Fluorouracil (5-FU) exhibits suboptimal efficacy for non-melanoma skin cancer, attributed to insufficient intracutaneous penetration. This study investigates the impact of ablative fractional laser (AFXL) at different laser-channel depths on cutaneous 5-FU pharmacokinetics and biodistribution. Methods: In vitro porcine skin underwent AFXL-exposure using a fractional 10,600 nm CO₂-laser, generating microscopic ablation zones (MAZ) reaching the dermoepidermal junction (MAZ-ED), superficial-(MAZ-DS), or mid-dermis(MAZ-DM). 5-FU in AFXL-exposed and control skin was measured in Franz diffusion cells at 4 and 24 hours (n = 55). HPLC quantified 5-FU in full-thickness skin, specific skin depths of 100μm-1500μm, and transcutaneous receiver-compartments. Qualitative matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) visualized 5-FU in selected samples. Results: Overall, AFXL enhanced and accelerated 5-FU uptake versus unexposed controls, with increased accumulation in deep skin layers (p < 0.01). While total, 24-hour 5-FU uptake in control skin was 0.096 mg/cm³ (0.19% of applied concentration), AFXL delivered up to 4.707 mg/cm³ (MAZ-DM; 9.41% uptake, 49-fold enhancement) (p = 0.002; 24 hours). Indicating accelerated delivery, 5-FU in laser-exposed samples at 4 hours was at least 10-fold that of 24-hour controls (p = 0.002). Deeper laser-channels increased delivery throughout the skin (MAZ-ED vs. MAZ-DM; p<0.01). MALDI-MSI confirmed enhanced, accelerated, deeper and more uniform 5-FU distribution after AFXL versus controls. Conclusions: AFXL offers laser-channel depth-dependent, enhanced and accelerated 5-FU uptake, with increased deposition in deep skin layers.

Inhibitor of nicotinamide phosphoribosyltransferase APO866 is a promising cancer drug currently in phase II clinical trials in oncology. Here, we present a strategy for increasing the therapeutic potential of APO866 through the rescue of normal tissues by coadministration of nicotinic acid (Vitamin B₃). We examined the toxicity profile of APO866 in B6D2F1 mice and the effect of oral administration of nicotinic acid on tissue toxicity. Nicotinic acid (50 mg/kg) protects mice from death and severe toxicity from an APO866 dose (60 mg/kg) four times the monotherapy maximum tolerated dose (15 mg/kg). In a panel of six cancer cell lines, we find that three (including ML-2 cells) are protected by nicotinic acid in vitro, whereas the cytotoxicity of APO866 remains unaffected in the remaining three (including A2780 cells). A selective biomarker for the protection by nicotinic acid was subsequently identified by quantitative RT-PCR. The expression of nicotinic acid phosphoribosyltransferase is low in the cell lines not rescued from APO866 by nicotinic acid compared with protected cell lines. The findings in cell lines translated into xenograft models in which the combination of 50 mg/kg nicotinic acid and 50 mg/kg APO866 in mouse xenografts of A2780 cells increased life span by >3-fold compared with standard treatment of 15 mg/kg, and the effect of APO866 was clearly decreased when using the same treatment paradigm in ML-2 xenografts. In conclusion, the combination of high doses of APO866 with rescue by nicotinic acid may significantly increase the therapeutic potential in a subset of cancers with low expression of nicotinic acid phosphoribosyltransferase. Mol Cancer Ther; 9(6); 1609–17. ©2010 AACR.

Inhibitors of nicotinamide phosphoribosyltransferase (NAMPT) are promising cancer drugs currently in clinical trials in oncology, including APO866, CHS-828 and the CHS-828 prodrug EB1627/GMX1777, but cancer cell resistance to these drugs has not been studied in detail. Here, we introduce an analogue of CHS-828 called TP201565 with increased potency in cellular assays. Further, we describe and characterize a panel of cell lines with acquired stable resistance towards several NAMPT inhibitors of 18 to 20,000 fold compared to their parental cell lines. We find that 4 out of 5 of the resistant sublines display mutations of NAMPT located in the vicinity of the active site or in the dimer interface of NAMPT. Furthermore, we show that these mutations are responsible for the resistance observed. All the resistant cell lines formed xenograft tumours in vivo. Also, we confirm CHS-828 and TP201565 as competitive inhibitors of NAMPT through docking studies and by NAMPT precipitation from cellular lysate by an analogue of TP201565 linked to sepharose. The NAMPT precipitation could be inhibited by addition of APO866. We found that CHS-828 and TP201565 are competitive inhibitors of NAMPT and that acquired resistance towards NAMPT inhibitors can be expected primarily to be caused by mutations in NAMPT.