The Non-Coding Oncofetal H19 Gene in Brain Tumors

Abstract

H19 is an imprinted non-coding RNA gene that encodes for perhaps one of the most abundant mRNA-like transcript in fetal life expressed predominantly in tissues of mesodermal and endodermal origin and down regulated at birth. It is located within the imprinted cluster on chromosome 11p15.5 in human and shares common regulatory sequences with other genes within the cluster, including insulin-like growth factor-2 (IGF2). H19 gene is either highly expressed and/or shows aberrant allelic pattern of expression in a large array of human cancers, while not expressed in the corresponding normal tissues. It possesses diagnostic, prognostic and therapeutic values in many cancers and its essential role in tumorigenesis has been demonstrated by using RNA interference and knockdown technologies. Modulation of the imprinting status of H19 and IGF2 may play an important role in the development of brain tumors including meningiomas, medulloblastomas, and gliomas. High levels of H19 expression have been shown by utilizing gene array in glioblastoma-derived cell lines, which are related to CD133 positive cells. This might be related to the modulation of H19 by hypoxia through upregulation of its non-coding RNA. There appears to be a crucial link between the well-established tumor suppressor p53 gene and the oncogenic H19 RNA that could determine vital processes in the tumorigenic pathway including response to hypoxic stress. The expression of H19 in a large array of human tumors and at different stages of tumor development, its hypoxic induction and its tight association with mutant form of p53, make it an ideal new target for cancer therapy, including low grade astrocytomas, which show frequent p53 mutations, and glioblastomas, which show unique hypoxia-driven angiogenesis (microvascular proliferation) and necrosis (pseudopalisading necrosis). Clinical studies and human compassionate use treatments using a DNA plasmid containing H19 gene regulatory sequences that drive the expression of an intracellular toxin [diphtheria toxin A (DTA) chain] and preclinical studies with siRNAs specifically targeting the H19 RNA have demonstrated promising results in several types of carcinomas. In preliminary studies, we have found expression of H19 and IGF2 by RT-PCR in several glioma cell lines and also significant expression of H19 by in situ hybridization performed on paraffin sections from several brain tumors including high-grade astrocytic neoplasms. In some of the glioblastomas, expression of H19 was observed in neoplastic cells as well as in proliferating microvasculture. The A172 human astrocytoma cell line has shown high expression of H19, IGF2-P3 and IGF2-P4 by using semiquantitative RT-PCR. Significant reduction in the number of cells was observed after transfection with a double promoter expressing vector, carrying two separate genes expressing DTA on a single construct, from two different regulatory sequences, namely H19-DTA-P4-DTA. In conclusion, H19-DTA-P4-DTA plasmid, which is based upon the toxic efficacy of diphtheria-toxin and H19-siRNA, which appears to be a promising therapeutic agent, are putative therapeutic candidates for treating brain tumors either with intra-tumoral convection-enhanced delivery or with better tolerated and more efficient modes of delivery yet to be developed.