Synthesis and in vitro evaluation of an 111In-labeled ST-peptide enterotoxin (ST) analogue for specific targeting of guanylin receptors on human colonic cancers.

BACKGROUND: Human colonic cancer cells are known to express guanylate cyclase C (GC-C) receptors for guanylin and uroguanylin. E. coli ST is a peptide with high metabolic stability that specifically binds to GC-C receptors. An in vitro evaluation of a new synthetic indium-111 labeled ST conjugate for specific targeting of human colonic cancers that express GC-C receptors was performed. MATERIALS AND METHODS: A DOTA conjugated ST analogue DOTA-NCS-6-Ahx-Phe19-ST[1-19] (DOTA-NCS-ST) was synthesized and labeled with indium-111. The non-radioactive indium analogue (In-DOTA-NCS-ST) was also prepared in macroscopic quantities. 111In-DOTA-NCS-ST was produced as a single species (>80% RCP) and purified by HPLC. Human colon cancer CaCO-2 and T-84 cells were used to evaluate the in vitro IC50 values for GC-C receptor binding and determine the cell uptake and retention of radioactivity. RESULTS: The DOTA-NCS-ST and In-DOTA-NCS-ST conjugates exhibit high in vitro binding affinity for GC-C receptors with IC50 values <10 nM. The in vitro cell binding studies with the 111In-DOTA-NCS-ST conjugate demonstrated that 111In-label ST internalizes in human colon cancer cells and exhibits long-term retention. CONCLUSION: The combination of radiolabeling efficacy and specific in vitro cell uptake and retention suggests that the DOTA-NCS-ST construct holds potential for the development of diagnostic or therapeutic radiopharmaceuticals labeled with trivalent radiometals for specific targeting of human colonic cancers.

Fluorescent tissue site-selective lanthanide chelate, Tb-PCTMB for enhanced imaging of cancer.

In-vivo and in-vitro investigations indicate that a newly developed polyazamacrocyclic chelate of Tb(III) has superior properties for use as an abnormal tissue marker. In addition to tissue selectivity, this molecule is unique because of its low toxicity, attractive fluorescent properties, rapid pharmokinetics, and relatively high water solubility. The complex Tb-3,6,9-tris(methylene phosphonic acid n-butyl ester)-3,6,9,15-tetraazabicyclo[9.3.1]-pentadeca-1(15),11,13 -triene (Tb-PCTMB) has also been shown to exhibit strongly shifted emission (delta lambda--280 nm), moving the detection frequency away from autofluorescence backgrounds, and good quantum efficiencies (phi = 0.51), providing high brightness. Fluorescence imaging was used to quantify Tb-PCTMB at the picomolar level in tissues and to show the significant difference in affinity for the chelate by adenocarcinoma cells HT-29 versus normal epithelial cells (IEC-6). Topical application, or lavage introduction, under endoscopy was used to instill a millimolar aqueous solution of Tb-PCTMB into a dimethylhydrizene-treated Sprague Dawley rat large intestine containing a suspect growth. Subsequent in vitro fluorescence detection and standard histological evaluation confirmed enhanced uptake by adenocarcinoma tissue. Semiquantitative signal interrogation was employed to show the potential for using Tb-PCTMB as a contrast enhancement marker for disease detection.

Imaging and quantitation of a tissue-selective lanthanide chelate using an endoscopic fluorometer.

Tissue spectroscopy and endoscopy are combined with a tissue site-selective fluorescent probe molecule to demonstrate in vitro, spatial, remote, quantitative imaging of the rat small intestine. The probe molecule employed, Tb-3,6,9-tris(methylene phosphonic acid n-butyl ester)-3,6,9,15-tetraaza-bicyclo[9.3.1]pentadeca-1(15),11,13-triene (Tb-PCTMB), is shown to bind with the small intestine and provide improved image contrast. High sensitivity is possible due to the absorption-emission Stokes's shift exhibited by the Tb-PTCMB complex. Excitation is centered near 270 nm and multifeatured emission is observed at 490, 550, 590, and 625 nm. Sprague-Dawley rats were dosed with the Tb-PTCMB complex, which shows biodistribution, leading to preferential binding to the inner surface of the small intestine. It is shown that the fluorescent image, taken at 550 nm, can be used to quantify the amount of Tb-PCTMB present in an excised tissue sample. The 3σ detection limits are found to be in the femtomole range. An optical mass balance for Tb-PCTMB-dosed small intestine is performed and along with radiotracer biodistribution, demonstrates that approximately 40% of the marker probe resides in the endothelial tissue of the small intestine inner lumen. This result is of particular interest since most adult colon cancers develop in this region. These results demonstrate the ability to perform spatial, quantitative, in vitro, endoscopic imaging of a complex biological sample using a probe marker. © 1998 Society of Photo-Optical Instrumentation Engineers.

Relaxometry, animal biodistribution, and magnetic resonance imaging studies of some new gadolinium (III) macrocyclic phosphinate and phosphonate monoester complexes.

The Gd3+ complexes of three new phosphorus containing tetraaza macrocycles (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis (methylene ethylphosphonic acid), H4DOTEP; 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis (methylene phosphonic acid monoethylester), H4DOTPME; and the corresponding n-butyl ester, H4DOTPMB) were prepared and examined for possible use as MRI contrast agents. Although thermodynamically and kinetically less stable than Gd(DOTA)- in saline and HSA solution, the stability of these new macrocyclic complexes appears to be sufficiently high for in vivo applications. NMRD relaxivity profiles of the three complexes indicate that the number of inner sphere water molecules for these chelates is < or = 1 and that the more hydrophobic chelate, Gd(DOTPMB), binds to human serum albumin (HSA). Biodistribution studies of the radioactive 153Sm or 159Gd chelates in rats, gamma imaging of the 153Sm chelates in rats, and proton MRI studies of the nonradioactive Gd3+ chelates in rabbits all indicate that the DOTPMB complexes accumulate preferentially in the liver, spleen, and small intestines while the more hydrophilic DOTEP and DOTPME complexes appear to display renal clearances similar to other low molecular weight contrast agents.