Identification of a Pan-Gamma-Secretase Inhibitor Response Signature for Notch-Driven Cholangiocarcinoma.

Cholangiocarcinoma (CCA) mortality rates are increasing as a result of rising incidence and limited curative treatment(s) for patients with advanced disease. NOTCH pathway reactivation has been reported in biliary malignancies to conflicting degrees, hindering prioritization of key therapeutic targets within the network and identification of candidate responder patients for NOTCH-directed therapies. We analyzed genomic data from 341 patients with CCA and identified NOTCH1 significantly increased in a subgroup characterized by distinct stromal infiltration. Network-wide imbalance of the NOTCH pathway was seen in CCA, including correlation of NOTCH1 with NOTCH3 and NOTCH ligands. Given the diversity of observed NOTCH receptor engagement, γ-secretase modulation was rationalized as a therapeutic option. Indeed, subcutaneous transplantation of sensitive and resistant CCA cell lines pretreated with a γ-secretase inhibitor (GSi) cocktail demonstrated the antineoplastic effects of GSi in a subset of CCA and led to the development of a 225-gene responder signature. This signature was validated in an independent cohort of 119 patients. Further, this signature was enriched in liver tumors initiated by hydrodynamic injections of activated-NOTCH as compared with the AKT-RAS-driven tumors. Candidate GSi-responder patients were characterized by distinct transcriptomes overlapping with previous hepatobiliary metastasis and stemness, unique stromal properties, and dysfunctional intratumoral immune infiltration. Pan-cancer analysis identified 41.9% of cancer types to harbor prospective GSi-responder patients, which was adapted into a 20-gene GSi-sensitivity score metric capable of discriminating nanomolar versus micromolar sensitivity to a cell-permeable GSi (Z-LLNle-CHO) across 60 diverse tumor lines (area under the curve = 1). Conclusion: We have established a GSi-responder signature with evidence across several patient cohorts, as well as in vitro and in vivo models, to enable precision medicine application of NOTCH-directed therapy in CCA as well as prospectively across diverse malignancies.

Bone marrow cell homing to sites of acute tibial fracture: 89Zr-oxine cell labeling with positron emission tomographic imaging in a mouse model.

BACKGROUND: Bone fracture healing is dependent upon the rapid migration and engraftment of bone marrow (BM) progenitor and stem cells to the site of injury. Stromal cell-derived factor-1 plays a crucial role in recruiting BM cells expressing its receptor CXCR4. Recently, a CXCR4 antagonist, plerixafor, has been used to mobilize BM cells into the blood in efforts to enhance cell migration to sites of injury presumably improving healing. In this study, we employed zirconium-89 (89Zr)-oxine-labeled BM cells imaged with positron emission tomography (PET)/computed tomography (CT) to visualize and quantitate BM cell trafficking following acute bone injury and to investigate the effect of plerixafor on BM cell homing. Unilateral 1-mm incisions were created in the distal tibia of mice either on the same day (d0) or 24 h (d1) after 89Zr-oxine-labeled BM cell transfer (n = 4-6, 2-2.3 × 107 cells at 9.65-15.7 kBq/106 cells). Serial microPET/CT imaging was performed and migration of 89Zr-labeled cells to the bone injury was quantified. The effects of three daily doses of plerixafor on cell trafficking were evaluated beginning on the day of fracture generation (n = 4-6). The labeled cells localizing to the fracture were analyzed by flow cytometry and immunohistochemistry. RESULTS: In d0- and d1-fracture groups, 0.7% and 1.7% of administered BM cells accumulated within the fracture, respectively. Plerixafor treatment reduced BM cell migration to the fracture by approximately one-third (p < 0.05 for both fracture groups). Flow cytometry analysis of donor cells collected from the injured site revealed a predominance of CD45+ stem/progenitor cell populations and subsequent histological analysis demonstrated the presence of donor cells engrafted within sites of fracture repair. CONCLUSION: 89Zr-oxine labeling enabled visualization and quantitation of BM cell recruitment to acute fractures and further demonstrated that plerixafor plays an inhibitory role in this recruitment.