Early detection with clinically translatable molecular imaging agents: Novel clinically translatable molecular imaging contrast agents for detecting tumors by T1-weighted magnetic resonance imaging (MRI) or positron emission tomography (PET) will be developed, optimized, and quantitatively characterized. The MRI agents are activatable cell penetrating peptides (ACPPs) conjugated to dendrimers bearing gadolinium chelates. The ACPPs'selectivity for matrix metalloproteinases and the stability of the gadolinium chelates will be optimized. The PET reporters will be reduced-size antibodies bearing novel near-infrared (NIR) fluorophores incorporating 18F-fluoborates. Both MRI and PET agents will be tested in mice on tumors stably expressing genetic reporters for fluorescence (FL), bioluminescence (BL), and positron emission tomography (PET), combining best available versions of far-red fluorescent protein, firefly luciferase, and thymidine kinase, expressed as separate but genetically linked proteins separated by self-cleaving viral 2A sequences. These tests will quantify how reliably can such injectable agents detect solid tumors of different sizes and stages of progression, under what circumstances false positive and negative signals are obtained, when is either combination of probe and imaging modality superior to the other, and whether indolent vs. highly malignant and invasive tumors can be distinguished. Evaluation and improvement of FL-guided surgical resection and adjuvant therapies: Intraoperative FL guidance from injectable agents such as ACPPs conjugated to fluorescent dendrimers, reduced-size antibodies labeled with NIR fluorophores, or 5-aminolevulinate will be compared with FL guidance from the fluorescent protein representing perfect tumor-specific labeling. These comparisons will show which injectable agent performs best under what circumstances, and whether completeness of fluorescence-guided resection is limited more by imperfect biochemical specificity of current contrast agents, or by inability to resolve and manually remove scattered or disseminated tumor cells. Photodynamic therapy (PDT) to kill off residual tumor cells in the surgical field before closure will also be tested. Completeness of initial tumor removal and any recurrence will be assessed postoperatively using the genetic reporters. Comparison with signals from injectable contrast agents and traditional survival measures should quantify how surgical outcomes depend on the thoroughness of tumor cell removal, how much improvement results from current injectable contrast agents vs. genetically perfectly labeling, whether PDT helps, how well can current injectable agents monitor recurrence, and what materials or procedures most need improvement. PUBLIC HEALTH RELEVANCE: This project aims to develop, optimize, and characterize the performance of novel injectable, clinically translatable contrast agents for early detection of tumors, distinction between indolent and aggressive cancers, molecular fluorescence-guided surgery, and postoperative monitoring. The long-term goal is to improve detection of aggressive tumors while they are still small and localized, and to help surgeons remove those tumors accurately completely while sparing as much normal tissue as possible.