Coordination of platinum therapeutic agents to met-rich motifs of human copper transport protein1.

Platinum therapeutic agents are widely used in the treatment of several forms of cancer. Various mechanisms for the transport of the drugs have been proposed including passive diffusion across the cellular membrane and active transport via proteins. The copper transport protein Ctr1 is responsible for high affinity copper uptake but has also been implicated in the transport of cisplatin into cells. Human hCtr1 contains two methionine-rich Mets motifs on its extracellular N-terminus that are potential platinum-binding sites: the first one encompasses residues 7-14 with amino acid sequence Met-Gly-Met-Ser-Tyr-Met-Asp-Ser and the second one spans residues 39-46 with sequence Met-Met-Met-Met-Pro-Met-Thr-Phe. In these studies, we use liquid chromatography and mass spectrometry to compare the binding interactions between cisplatin, carboplatin and oxaliplatin with synthetic peptides corresponding to hCtr1 Mets motifs. The interactions of cisplatin and carboplatin with Met-rich motifs that contain three or more methionines result in removal of the carrier ligands of both platinum complexes. In contrast, oxaliplatin retains its cyclohexyldiamine ligand upon platinum coordination to the peptide.

Development of a Novel c-MET-Based CTC Detection Platform.

UNLABELLED: Amplification of the MET oncogene is associated with poor prognosis, metastatic dissemination, and drug resistance in many malignancies. We developed a method to capture and characterize circulating tumor cells (CTC) expressing c-MET using a ferromagnetic antibody. Immunofluorescence was used to characterize cells for c-MET, DAPI, and pan-CK, excluding CD45(+) leukocytes. The assay was validated using appropriate cell line controls spiked into peripheral blood collected from healthy volunteers (HV). In addition, peripheral blood was analyzed from patients with metastatic gastric, pancreatic, colorectal, bladder, renal, or prostate cancers. CTCs captured by c-MET were enumerated, and DNA FISH for MET amplification was performed. The approach was highly sensitive (80%) for MET-amplified cells, sensitive (40%-80%) for c-MET-overexpressed cells, and specific (100%) for both c-MET-negative cells and in 20 HVs. Of 52 patients with metastatic carcinomas tested, c-MET CTCs were captured in replicate samples from 3 patients [gastric, colorectal, and renal cell carcinoma (RCC)] with 6% prevalence. CTC FISH demonstrated that MET amplification in both gastric and colorectal cancer patients and trisomy 7 with gain of MET gene copies in the RCC patient. The c-MET CTC assay is a rapid, noninvasive, sensitive, and specific method for detecting MET-amplified tumor cells. CTCs with MET amplification can be detected in patients with gastric, colorectal, and renal cancers. IMPLICATIONS: This study developed a novel c-MET CTC assay for detecting c-MET CTCs in patients with MET amplification and warrants further investigation to determine its clinical applicability. Mol Cancer Res; 14(6); 539-47. ©2016 AACR.