It was previously reported that a midregion domain of
parathyroid hormone-related protein (
PTHrP), that is, [67-86]-
amide, is able to restrain growth and promote
matrigel penetration by the 8701-BC cell line, derived from a biopsy fragment of a primary ductal infiltrating
carcinoma of the human breast, and that cell invasion in vitro is drastically impaired by inactivation of
urokinase-
plasminogen activator (uPa). In this study we started a more detailed investigation of the possible effects on gene expression arising from the interaction between
PTHrP [67-86]-
amide and 8701-BC
breast cancer cells by a combination of conventional-, differential display-and semi-quantitative multiplex-polymerase chain reaction (PCR) assays. We present here the first evidence that the upregulation of some stress-related genes, most noticeably heat shock factor binding protein-1 (hsbp1) and
heat shock protein 90 (hsp-90), is involved in the acquisition of an in vitro more invasive phenotype by cells treated with midregion
PTHrP. This is conceivably accomplished by sequestering and inactivating heat shock factor-1 (hsf1) which is able to recognize
Ets transcription-factor-binding sites present in some gene promoters, such as those of uPa and matrix metalloprotease-1 (MMP-1). In fact, our data show that incubation of
PTHrP [67-86]-
amide-treated cells with either antisense hsbp1-oligonucleotide or
geldanamycin, an hsp90-inactivating
antibiotic, results in downregulation of uPa and upregulation of MMP-1, and in a prominent inhibition of cell invasion in
matrigel-containing Transwell chambers. Alternatively, incubation of untreated 8701-BC cells with
quercetin, a
flavonoid known to decrease the amount of free hsf1, is found to induce upregulation of uPa and downregulation of MMP-1, and an increase of
matrigel invasion by cells, thus providing further supporting data of the involvement of hsf unavailability on the modulation of uPa and MMP-1 expression and on cell invasive behaviour. These studies confirm a previous postulate that over-secretion of uPa, rather than of other extracellular
proteases, is a primary condition for the increase of invasive activity triggered by
PTHrP [67-86]-
amide in vitro, and support a role for midregion forms of
PTHrP in potentially affecting pathological mammary growth and differentiation. They also identify two new key protagonists in the complex scenario of
breast tumor cell invasiveness in vitro, that is, hsbp1 and hsp90, which deserve further and more extensive studies as potential and attractive molecular targets for anti-
breast cancer treatments.