Integrated Hedgehog signaling is induced following castration in human and murine prostate cancers

Abstract

Background: The interplay between androgen and Hedgehog (Hh) signaling pathways may be associated with prostate cancer progression and resistance to therapy.

Methods: Tissue microarrays from prostatectomy specimens were derived from 53 patients treated preoperatively with androgen ablation (AA) with or without chemotherapy, and from 26 stage- and grade-matched controls. A previously characterized androgen-regulated human prostate cancer xenograft was used to conduct parallel murine studies. Expression of markers of interest was determined on both untreated and castrated tumors.

Results: Four-month exposure to AA or AA with chemotherapy led to a uniform increase in Hh signaling as compared to controls, paired with an inverse trend of androgen receptor (AR) and CYP17 expression in clinically derived specimens. Changes in the expression profiles of Hh signaling were observed in the epithelium and stroma, in response to genotoxic stress of androgen ablation and chemotherapy. A reduced expression of KI67 and increased bcl2 expression was observed in the malignant epithelial compartment.

Conclusion: To our knowledge, this is the first clinical evidence that Hh signaling is induced by AA or the combination of AA and chemotherapy and, by inference, contributes to castrate-resistant progression of prostate cancer as supported by parallel human and murine studies. These data are in agreement with previous reports that implicate Hh signaling in castrate-resistant progression of prostate cancer. Based on these findings, we are pursuing parallel clinical and murine investigations to determine if Hh signaling inhibition combined with AA will be more effective than AA alone.

Figure 1a. Hedgehog signaling in untreated control tumors

Representative images of 3 different untreated tumors. Active Hh signaling is heterogeneous and limited compared to that of treated tumors (Figure 1b-c) as illustrated by the expression of Gli2 and smoothened.

Figure 1b. Increased Hh signaling in residual tumors following AA

Representative images of 3 different radical prostatectomy specimens with varied extent of residual tumor. Expression of all components of Hh signaling assessed (gli2, smoothened, and sonic hedgehog) is higher than in untreated controls (Figure 1a). Hedgehog signaling is active both in the residual tumor epithelium and stroma as indicated by the nuclear expression of the transcription factor gli2. Three cases are included to represent heterogeneity in histology.

Figure 1c

Increased Hh signaling in residual tumor following AA and KAVE chemotherapy.

Figure 2. Androgen receptor expression

There was a trend for lower AR expression in the tumor epithelium following AA (middle panel) and chemotherapy (right panel) for 16 weeks versus untreated control specimens (left panel). Interestingly when this occurred, adjacent stroma exhibited an increase in AR expression (arrows).

Figure 3. Hedgehog signaling expression in human xenograft model

An increase is shown in stromal mRNA expression of Hh signaling components following castration (blue bars) in a castrate-responsive tumor xenograft as compared to non castrate (red bars). These include the transcription factors gli1, gli2 and the ligand Shh.