Targeted probes for fluorescence imaging and photodynamic therapy of breast cancer.

Positive surgical margins remain a persistent challenge in breast-conserving surgery (lumpectomy), often necessitating re-excision or additional adjuvant therapies. We investigated a dual molecular targeting strategy that combines fluorescence imaging with photodynamic therapy (PDT), with potential application in improving surgical precision and eradicating residual disease at the margins of surgery. The breast cancer-targeting peptide 18-4 and its cyclic analog c(18-4), which bind the keratin-1 receptor overexpressed in breast cancer cells, were conjugated to Cy5.5 and IRDye78 to generate imaging probes, and to mesopyropheophorbide-a (mPPa) to produce therapeutic PDT probes. Because chlorin-based photosensitizers are highly hydrophobic, a tri-sulfonated water-solubilizing platform was developed to enable intravenous administration of the PDT agents. The resulting probes were evaluated in EMT6 and MDA-MB-231 breast cancer cell lines using confocal microscopy and quantitative fluorescence assays, and in orthotopic murine breast tumor models to assess tumor localization, retention, and therapeutic efficacy. In vitro, targeted probes exhibited enhanced binding relative to untargeted dyes, with cyclic peptide conjugates generally demonstrating stronger uptake than their linear counterparts. In vivo, the dye-peptide conjugate, IRDye78-(18-4) exhibited improved tumor localization and prolonged retention compared with free dye, enabling clear tumor visualization and informing the timing of PDT treatment. The water-soluble PDT probe WS-Lys(mPPa)-c(18-4) produced delayed tumor progression, with complete responses in a subset of treated animals when illumination coincided with peak probe accumulation. Together, these results demonstrate a matched molecular targeting strategy that supports future investigation of fluorescence-guided lumpectomy followed by targeted photodynamic therapy to eradicate residual disease at surgical margins to potentially reduce reoperation rates in the treatment of cancer.