Molecular Imaging of bcl-2 Expression in Small Lymphocytic Lymphoma Using ¹¹¹In-Labeled PNA–Peptide Conjugates

Abstract

The bcl-2 gene is overexpressed in non-Hodgkin's lymphoma (NHL), such as small lymphocytic lymphoma (SLL), and many other cancers. Noninvasive imaging of bcl-2 expression has the potential to identify patients at risk for relapse or treatment failure. The purpose of this study was to synthesize and evaluate radiolabeled peptide nucleic acid (PNA)-peptide conjugates targeting bcl-2 gene expression. An 111 In-labeled PNA complementary to the translational start site of bcl-2 messenger RNA was attached to Tyr 3 -octreotate for somatostatin receptor-mediated intracellular delivery. Methods: DOTA-anti-bcl-2-PNA-Tyr 3 -octreotate (1) and 3 control conjugates (DOTA-nonsense-PNATyr 3 -octreotate (2), DOTA-anti-bcl-2-PNA-Ala[3,4,5,6]-substituted congener (3), and DOTA-Tyr 3 -octreotate (4) [DOTA is 1,4,7,10-tetraazacyclododecane-N,N9,N$,N%-tetraacetic acid]) were synthesized by standard solid-phase 9-fluorenylmethoxycarbonyl (Fmoc) chemistry. In vitro studies were performed in Mec-1 SLL cells, which express both bcl-2 messenger RNA and somatostatin receptors. Biodistributions and microSPECT/CT studies were performed in Mec-1-bearing SCID (severe combined immunodeficiency) mice, a new animal model of human SLL. Results: 111 In-Labeled conjugate 1 was taken up by Mec-1 cells through a somatostatin receptor-mediated mechanism. Biodistribution studies showed specific tumor uptake of conjugate 1, the somatostatin analog 4, and the PNA nonsense conjugate 2, but not of the mutant peptide conjugate 3. Mec-1 tumors could be detected by microSPECT/CT using 111 In-labeled DOTA-Tyr 3 -octreotate (4) and the targeted anti-bcl-2 conjugate (1), but not using the 2 negative control conjugates 2 and 3. Conclusion: A new 111 In-labeled antisense PNA-peptide conjugate demonstrated proof of principle for molecular imaging of bcl-2 expression in a new mouse model of human SLL. This imaging agent may be useful for identifying NHL patients at risk for relapse and conventional treatment failure.

Preparation, 99mTc-labeling and biodistribution studies of a PNA oligomer containing a new ligand derivative of 2,2′-dipicolylamine

“…This behavior is comparable to plain radiolabeled PNAs [23,24]. Conjugation of targeting peptide moieties to PNA significantly increases the kidney accumulation of PNA constructs [14,[17][18][19]29,30,44,87,88]. A preliminary SPECT study in a NMRI, nu/nu mouse was also performed with [ 99m Tc(CO) 3 (Dpa-PNA)].…”

“…In such constructs, the role of the PNA is to direct the bioconjugate to a specific mRNA -a unique or a uniquely over-expressed mRNA in a diseased cell -while the function of the radioactive probe is to allow imaging to possibly provide information on cellular gene expression pattern and detect molecular changes at relatively early stages [13,42]. Hence, a number of in vivo biodistribution/hybridization studies of radiolabeled PNAs were undertaken [14,[17][18][19][20]22,[24][25][26][27][28][29][30][31][32][33]36,39,41,[43][44][45][46]. The majority of these reports involved the use of 99m Tc, probably due its ideal nuclear properties and the convenient availability from a commercial generator [47][48][49][50].…”

Chemical Modification of Oligonucleotides for Therapeutic, Bioanalytical and other Applications

“…In vivo imaging by microSPECT/ CT was also investigated and showed that the tumours could be imaged, and that imaging agreed with the biodistribution data. [99] Nanostructures using chemically modified oligonucleotide have been explored for use, for example, in nanotubes, arrays and machinery. Chaput et al demonstrated the use of glycol nucleic acids in the formation of two mirror image 4-helix junctions.…”

DNMT Inhibitors and HDAC Inhibitors Regulate E-Cadherin and Bcl-2 Expression in Endometrial Carcinoma in vitro and in vivo

“…Bcl-2 has been described as an oncogene whose overexpression results in tumorigenesis, suppression of apo-ptosis [16,17,18], and resistance to a number of cytotoxic drugs [19]. Downregulation of Bcl-2 mRNA in leukemic patients using antisense technology resulted in increased apoptosis and potentiation of the effect of antineoplastic drugs in these patients [20].…”