Knockdown of HMGA1 inhibits human breast cancer cell growth and metastasis in immunodeficient mice

Highlights

• Overexpression of HMGA1 has been previously implicated in breast carcinogenesis.

• An RNAi-based HMGA1-silencing construct with co-expression of EmGFP was generated.

• HMGA1-silenced MDA-MB-231 human breast cancer cells were generated.

• Knockdown of HMGA1 impairs anchorage-independent growth and tumorsphere formation.

• Knockdown of HMGA1 impairs xenograft growth and metastasis in immunodeficient mice.

Abstract

The high mobility group A1 gene (HMGA1) has been previously implicated in breast carcinogenesis, and is considered an attractive target for therapeutic intervention because its expression is virtually absent in normal adult tissue. Other studies have shown that knockdown of HMGA1 reduces the tumorigenic potential of breast cancer cells in vitro. Therefore, we sought to determine if silencing HMGA1 can affect breast cancer development and metastatic progression in vivo. We silenced HMGA1 expression in the human breast cancer cell line MDA-MB-231 using an RNA interference vector, and observed a significant reduction in anchorage-independent growth and tumorsphere formation, which respectively indicate loss of tumorigenesis and self-renewal ability. Moreover, silencing HMGA1 significantly impaired xenograft growth in immunodeficient mice, and while control cells metastasized extensively to the lungs and lymph nodes, HMGA1-silenced cells generated only a few small metastases. Thus, our results show that interfering with HMGA1 expression reduces the tumorigenic and metastatic potential of breast cancer cells in vivo, and lend further support to investigations into targeting HMGA1 as a potential treatment for breast cancer.

Introduction

Breast cancer is the most frequent type of cancer and a leading cause of cancer death among women worldwide [1], [2], [3]. The proteins of the high mobility group A (HMGA) family have been previously implicated in breast carcinogenesis [4]. These are non-histone, DNA-binding proteins often referred to as architectural transcription factors. They contain basic A-T hook domains that mediate binding to the minor groove of AT-rich regions of chromosomal DNA. Upon binding to DNA, HMGA proteins regulate gene expression by organizing the transcriptional complex through protein–protein and protein–DNA interactions (reviewed in [5], [6], [7]). The HMGA family includes the products of the HMGA1 and HMGA2 genes. Of these, HMGA1 can generate three different protein isoforms through alternative splicing (HMGA1a, b and c). HMGA1a and HMGA1b are the most abundant isoforms, and differ by only 11 amino acids that are present in HMGA1a but not in HMGA1b [4]. HMGA1 is expressed almost exclusively during embryonic development [8], but has been found to be abnormally expressed in several types of cancer, including leukemia [9], pancreatic [10], [11], thyroid [12], colon [13], breast [14], [15], [16], lung [17], ovarian [18], endometrial [19], prostate [20], and head and neck cancer [21]. Several studies have also shown that overexpression of HMGA1 induces transformation both in vitro and in animal models (reviewed in [6], [7]).

The causal role of HMGA1 in breast cancer development and metastasis is supported by studies in cell lines [14], [22], [23] as well as by the analysis of clinical specimens [15], [16]. For example, elevated HMGA1 protein expression has been reported in breast carcinomas and hyperplastic lesions with cellular atypia, in contrast with normal breast tissue where HMGA1 was not detected [15], [16]. Similarly, HMGA1 overexpression has been observed in human breast cancer cell lines, with the highest levels in known metastatic lines, such as Hs578T and MDA-MB-231 [14], [22], [23]. Moreover, exogenous overexpression of HMGA1a was shown to induce transformation of the human non-tumorigenic mammary myoepithelial cell line Hs578Bst in vitro [14] and to increase the metastatic ability of MCF7 breast cancer cells in vivo [22]. Conversely, decreasing HMGA1 expression in Hs578T breast cancer cells was shown to cause a reduction in anchorage-independent growth, which is a typical feature of cancer cells [14].

HMGA1 is considered an attractive target for therapeutic intervention because its expression is virtually absent in normal adult tissue and knockdown of HMGA1 has been shown to interfere with the tumorigenic growth of multiple cancer cell lines [6], [7]. We therefore sought to determine if silencing HMGA1 can affect breast cancer development and metastatic progression in vivo using a human xenograft mouse model.