EBV-LMP1-targeted DNAzyme restrains nasopharyngeal carcinoma growth in a mouse C666-1 xenograft model

Abstract

Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) has been demonstrated to be linked to the pathogenesis of nasopharyngeal carcinoma (NPC), one of the most common cancers in southeast Asia as well as in the areas of southern China. RNA-cleaving DNAzymes are catalytic nucleic acids that bind and cleave a target RNA and have been increasingly used to inhibit gene expression. In this study, we aimed to explore the effects of down-regulation of LMP1 by DNAzymes on the NPC growth using a mouse xenograft model derived from human NPC C666-1 cells that constitutively express the LMP1. A specific LMP1-targeted DNAzyme, named DZ509, was synthesized, showing high activities in the inhibition of LMP1 expression, while the control DNAzyme (mutDZ509) had little effect on LMP1 expression. Knockdown of the LMP1 expression by DZ509 reduced cell proliferation and increased apoptosis compared to the cells transfected with mutDZ509. Intratumoral injections of DZ509 into C666-1 xenografts caused a significant suppression of tumor growth compared to mutDZ509-treated xenografts. Examination of the LMP1 expression in the xenografts demonstrated a marked inhibition of LMP1 by DZ509. An antisense oligonucleotide targeting the same site of the LMP1 transcripts was employed in parallel and produced a comparable inhibition on cell proliferation in vitro and tumor growth in vivo. These data demonstrate that LMP1-targeted DNAzyme is efficient in controlling tumor growth in vivo, and thus may have therapeutic implications in the treatment of NPC.

An Orthotopic Model of Metastatic Nasopharyngeal Carcinoma and Its Application in Elucidating a Therapeutic Target That Inhibits Metastasis

“…[8][9][10][11][12] To date, most NPC xenograft studies have involved the growth of a mass of tumor cells, often subcutaneously, that do not recapitulate human disease because they remain discrete and encapsulated. [8][9][10] In comparison, clinically advanced NPC is characterized by aggressive invasion with erosion and remodeling of facial and skull base cranial bones 13 as well as distant metastasis to solid organs, most commonly bone, lung, and liver. 14,15 To more accurately replicate advanced human disease, we have developed an orthotopic mouse xenograft model of NPC by inserting luciferasetagged tumor cells into the nasopharyngeal region and then following patterns of growth and metastasis over time.…”

“…Much of the information we have about NPC has been derived from studies of well-characterized cell lines in vitro 7 and in vivo using immunocompromised mice. 8-12 To date, most NPC xenograft studies have involved the growth of a mass of tumor cells, often subcutaneously, that do not recapitulate human disease because they remain discrete and encapsulated. 8-10 In comparison, clinically advanced NPC is characterized by aggressive invasion with erosion and remodeling of facial and skull base cranial bones 13 as well as distant metastasis to solid organs, most commonly bone, lung, and liver.…”

DNA enzymes as potential therapeutics: towards clinical application of 10-23 DNAzymes

“…ENV [55][56][57] TAT, TAT-REV [58][59][60][61] TAR [62] GAG, NEF [63] HIV-1 Integrase [64] Vpr [65] Hepatitis B X [66,67] S, C [68,69] DR1 and PA regions of pre-genomic RNA [70] Hepatitis C Core [19,71,72] NS3 [73,74] NS5B [19] IRES [17] Influenza A PB2 [75,76] M1 [77] Influenza B BM2 [21] Respiratory syncytial virus (RSV) N [15,78] Severe acute respiratory syndrome coronavirus (SARS-CoV) 5'-Untranslated region of a highly conserved fragment of SARS genome [79] Epstein-Barr virus LMP1 [80][81][82][83][84][85] Human papilloma virus type 16 E6/E7 [48] Japanese encephalitis virus 3'-Non-coding regions of RNA genome [16] Bacteria Ampicillin-resistant bacteria TEM1, TEM3 b-Lactamase [14] Methicillin-resistant Staphylococcus aureus blaR1 [86] MecR1 [87] Mycobacterium tuberculosis…”

“…Another Dz Dz509 targeting the same LMP1 RNA was identified for potential therapeutic application for treatment of NPC [85]. Dz509 showed high activity for inhibition of LMP1 expression as compared to a control mutant Dz mutDz509.…”

DCE-MRI assessment of the effect of Epstein-Barr virus-encoded latent membrane protein-1 targeted DNAzyme on tumor vasculature in patients with nasopharyngeal carcinomas

“…Previously, a 33-mer oligonucleotide LMP1-targeted DNAzyme containing three phosphorothioate linkages at its 5’ and 3’ ends was developed to specifically target LMP1 mRNA [ 11 ]. Down-regulation of LMP1 expression using this LMP1-targeted DNAzyme was found to inhibit the growth of NPC cells both in vivo and in vitro by suppressing cell proliferation and inducing apoptosis [ 2 , 12 ]. Moreover, in a recent study, LMP1-targeted DNAzyme was found to enhance the radiosensitivity of LMP1-positive NPC cells by inhibiting telomerase activity [ 13 ].…”