The insulinoma-associated 1: a novel promoter for targeted cancer gene therapy for small-cell lung cancer

Abstract

The insulinoma-associated 1 (INSM1) gene is expressed exclusively during early embryonal development, but has been found reexpressed at high levels in neuroendocrine tumors. The regulatory region of the INSM1 gene is therefore a potential candidate for regulating expression of a therapeutic gene in transcriptionally targeted cancer gene therapy against neuroendocrine tumors. We analyzed expression of a reporter gene from a 1.7 kb region of the INSM1 promoter in a large number of small-cell lung cancer (SCLC) cell lines. This INSM1 promoter region showed very high levels of expression in most of the SCLC cell lines and expression was absent in cell lines of non-neuroendocrine origin. Inclusion of the general transcriptional enhancer from SV40 compromised the specificity of the promoter and did not enhance transcription in most of the SCLC cell lines. For comparison, the region of the gastrin releasing peptide (GRP) previously suggested for SCLC gene therapy was analyzed in a similar manner. High expression was observed for a number of cell lines, but unlike for the INSM1 promoter, reporter gene expression from the GRP promoter did not correlate to the relative GRP mRNA levels, demonstrating that this region may not contain all necessary regulatory elements. Expression of the suicide gene herpes simplex virus thymidine kinase (HSV-TK) from the INSM1 promoter in combination with treatment with the prodrug ganciclovir (GCV) caused a significant increase in GCV sensitivity specifically in INSM1-expressing cell lines. The INSM1 promoter is therefore a potential novel tool for transcriptionally targeted gene therapy for neuroendocrine tumors.

Estrogen Induces Repression of the Breast Cancer and Salivary Gland Expression Gene in an Estrogen Receptor α–Dependent Manner

“…An altered expression in some types of cancer has also been reported for INSM1. For example, it is reexpressed in neuroendocrine tumors (23) and its expression is increased in small cell lung cancer (24). However, no link between INSM1 expression and breast cancer tumors has been reported thus far.…”

Modifications to the INSM1 promoter to preserve specificity and activity for use in adenoviral gene therapy of neuroendocrine carcinomas

“…In the same study, transient transfection of the INSM1 promoter-driven HSV-TK construct into the GLC-16 and H69 SCLC cell lines treated with increasing doses of ganciclovir showed an 80% kill rate in vitro. 21 We exploited the spatial and temporal regulatory features of the INSM1 promoter to develop an adenoviral delivered HSV-TK suicide gene therapy for the treatment of pediatric neuroendocrine tumors retinoblastoma, neuroblastoma and medulloblastoma. 22 In our studies, we demonstrated specific activation of the HSV-TK gene in Y79, IMR-32 and D283Med cells.…”

“…19 Use of a nonviral INSM1 promoter-driven suicide gene therapy for SCLC has been demonstrated. 21 We developed an adenoviral INSM1 promoter-driven suicide gene therapy approach for the potential treatment of a variety of INSM1-positive pediatric neuroendocrine tumors. 22 In this study, in vivo imaging of the Ad-INSM1 promoter luciferase2-IRES-HSV-TK suicide gene therapy revealed off-target activity of the INSM1 promoter in mouse pancreas, kidney and lung.…”

A chimeric fusion of the hASH1 and EZH2 promoters mediates high and specific reporter and suicide gene expression and cytotoxicity in small cell lung cancer cells

“…12 Transient luciferase reporter gene transfection assay Cells (adherent cells plated 1 day prior to transfection) were transfected in duplicate with 0.5 mg plasmid DNA using Lipofectamine 2000 in OPTIMEM (Invitrogen) in 24-well dishes. For DMS53 and DMS456, 0.2 Â 10 6 cells per well were plated and for all other cell lines 0.1 Â 10 6 cells were plated per well.…”

“…Furthermore, the INSM1 promoter was capable of inducing high and SCLC-specific cytotoxicity in a suicide gene assay in vitro. 12 Another gene highly upregulated in SCLC encodes the basic helix-loop-helix transcription factor human achaetescute homolog 1 (alias hASH1, ASH1 and ASCL1 in humans and MASH1 in rodents). 13 Global gene expression profiling indicates expression levels of hASH1 to be 5-to 8-fold higher than INSM1 in SCLC suggesting the hASH1 promoter to be a strong regulatory candidate.…”