ApoSense: a novel technology for functional molecular imaging of cell death in models of acute renal tubular necrosis

Abstract

PurposeAcute renal tubular necrosis (ATN), a common cause of acute renal failure, is a dynamic, rapidly evolving clinical condition associated with apoptotic and necrotic tubular cell death. Its early identification is critical, but current detection methods relying upon clinical assessment, such as kidney biopsy and functional assays, are insufficient. We have developed a family of small molecule compounds, ApoSense, that is capable, upon systemic administration, of selectively targeting and accumulating within apoptotic/necrotic cells and is suitable for attachment of different markers for clinical imaging. The purpose of this study was to test the applicability of these molecules as a diagnostic imaging agent for the detection of renal tubular cell injury following renal ischemia.MethodsUsing both fluorescent and radiolabeled derivatives of one of the ApoSense compounds, didansyl cystine, we evaluated cell death in three experimental, clinically relevant animal models of ATN: renal ischemia/reperfusion, radiocontrast-induced distal tubular necrosis, and cecal ligature and perforation-induced sepsis.ResultsApoSense showed high sensitivity and specificity in targeting injured renal tubular epithelial cells in vivo in all three models used. Uptake of ApoSense in the ischemic kidney was higher than in the non-ischemic one, and the specificity of ApoSense targeting was demonstrated by its localization to regions of apoptotic/necrotic cell death, detected morphologically and by TUNEL staining.ConclusionApoSense technology should have significant clinical utility for real-time, noninvasive detection of renal parenchymal damage of various types and evaluation of its distribution and magnitude; it may facilitate the assessment of efficacy of therapeutic interventions in a broad spectrum of disease states.

From the Gla domain to a novel small-molecule detector of apoptosis

“…Conceptually, low-molecular-weight probes can be advantageous over large proteins such as Annexin-V (MW=37 000 Da) in the aspects of radio-labeling, biodistribution and immunogenicity. Based on these considerations, we have developed and previously reported DDC [9][10][11] and NST-732 [12], which are members of the ApoSense family of recently developed small-molecule apoptosis probes. However, DDC or NST-732 do not comprise a Gla structural motif, which, as revealed in the present study, is unique in being the smallest structural motif for detection of apoptosis known to date.…”

“…Based on this hypothesis and on our previous experience in developing fluorescent small-molecule apoptosis probes of the ApoSense family, such as didansyl-Lcystine (DDC) [9,10,11] and NST-732 [12], we now present ML-10 (2-(5-fluoro-pentyl)-2-methyl-malonic acid), as a rationally designed molecule, with an amphipathic compact structure (MW=206 Da). We also describe the performance of this novel Gla-derived probe as a detector of apoptosis, and its mechanism of action.…”

Activity of aspirin analogues and vanillin in a human colorectal cancer cell line

“…The Apo-Trace compound accumulates in the cytoplasm of apoptotic cells. end-stage apoptosis and death is characterised by cells accumulating the Apo-Trace compound (N,N'-didansyl-L-cystine) (29) and staining positively for PI. PI binds to double-stranded DNA and can only permeate a compromised plasma membrane thus staining only non-viable cells.…”

“…the apoptotic potential of the compounds was tested in vitro using the commercially available Apo-TRACe™ Apoptotic Cell Staining Kit (Sigma-Aldrich), which utilises a small non-toxic compound of the ApoSense family of compounds, according to the manufacturer's instructions (29). Briefly, at 80-90% confluency, SW480 cells were incubated with either 1 mM aspirin, 0.5 mM BCS, carrier amounts of DMSO or 25 µM irinotecan for 24 or 48 h, and the medium containing the drug was removed.…”

Myocardial and Renal Ischemia/Reperfusion Injury and Experimental Models: Review

“…115 Bhalodia ve ark., sıçanlarda her iki böbrek pedikülünün 60 dk süresince sıkıştırılması ve 24 saat reperfüz-yonu ile 24 saat içinde renal yetmezlik geliştir-mişlerdir. 116 117 Foglieni ve ark., sağ renal arter ve venin her ikisini birlikte 60 dk mikrocerrahi klemp ile sıkıştırma sonrası 60 dk reperfüzyon sonucu renal yetmezlik bildirmişlerdir. 118 Baker ve ark., domuzlarda intrarenal aortun standart köşeli arteriyel çapraz klemp (cross-clemp) ile 150 dk süresince oklüzyonunun ardından (Toplam aortik oklüzyonu doğrulamak amacıyla distal aort palpasyonu yapılır) 180 dk reperfüzyon ile renal yetmezlik geliştiğini bildirmiş-lerdir.…”