Research paper
Characterization of the intra-prostatic immune cell infiltration in androgen-deprived prostate cancer patients

https://doi.org/10.1016/j.jim.2009.06.004Get rights and content

Abstract

Introduction

Our goal was to study the hormonal regulation of immune cell infiltration in prostate cancer patients treated by androgen deprivation therapy (ADT) using an optimized computer-assistance quantification approach.

Methods

The relative density of immune cell subtypes (CD3+, CD8+, CD20+, CD56+, CD68+ and Foxp3+) was analyzed by immunohistochemistry in archived prostate specimens from control patients (radical prostatectomy only, n = 40) and ADT-treated patients (ADT prior to radical prostatectomy, n = 35) using an image analysis software and a whole-slide scanner.

Results

ADT-treated patients had significantly increased relative density of CD3+ (p < 0.001) and CD8+ T lymphocytes (p < 0.001) as well as CD68+ macrophages (p < 0.001). Elevated abundance of CD56+ Natural Killer (NK) cells was associated with a lower risk of prostate cancer progression (p = 0.044), while a high density of CD68+ macrophages was related to an increased risk of biochemical recurrence (p = 0.011).

Conclusions

Our results demonstrate that the infiltration of specific immune cell subtypes is modulated by ADT. Furthermore our data confirm that NK cells have a protective role against tumor progression while macrophages seem to favor the development of advanced prostate cancer.

Introduction

Prostate cancer remains the most frequently diagnosed cancer and third leading cause of cancer related deaths for North American men (Jemal et al., 2008). Androgens participate in the prostate's organogenesis and carcinogenesis (Grossmann et al., 2001). As such, the most common treatment for men with advanced stage or recurrent prostate cancer is androgen deprivation therapy (ADT). ADT promotes the apoptosis of the hormone sensitive prostate epithelial cells, which leads to the involution of the prostate (Montironi and Schulman, 1998, Ohlson et al., 2005). Unfortunately, generally within one to five years following ADT initiation, patients develop hormone refractory prostate cancer, the major contributor to prostate cancer related death, and a disease for which only palliative therapies are currently available (Oefelein et al., 2002, Tannock et al., 2004).

Novel therapeutic protocols are currently emerging with the goal of tackling hormone-refractory prostate cancer, including immune-based therapies. The common rationale between the various immunotherapeutic approaches is the activation of the anti-tumoral immune response within the tumor and/or metastases. To this day, immunotherapeutic protocols in clinical trial have yet to attain the optimistic results demonstrated in animal models. Recent data suggest that the prostate possesses a strong immunoregulatory potential, which may suppress the activation of the anti-tumoral immune response (Miller and Pisa, 2007). Therefore, the interactions between the immune system and prostate cancer cells within the patient's primary tumor needs to be better understood in order to develop clinically effective immunotherapies.

Several publications have demonstrated that different immune cell populations infiltrate the prostate and that, in some cases, the abundance of specific immune cells may correlate with cancer progression (Vesalainen et al., 1994, Irani et al., 1999, Shimura et al., 2000, McArdle et al., 2004, Karja et al., 2005). Nonetheless, in the context of an androgen dependant cancer such as prostate cancer, significant knowledge needs to be obtained on whether androgens can modulate the abundance and activity of the immune cell infiltrate within the primary tumor. It is known that ADT fosters the development of a pro-inflammatory environment within the prostate (Civantos et al., 1996, Guinan et al., 1997, Mercader et al., 2007). Moreover, the immunosuppressive potential of the primary tumor can be dampened following ADT (Drake et al., 2005). With clinical trials combining immune-based therapies and ADT currently being evaluated, it is essential to gain insights on the various immunoregulatory changes present within the prostate following ADT.

Our goal was thus to characterize, using a computer-based approach, the immune cell infiltrate in patients treated by ADT. Using a cohort of 75 patients, we quantified the relative density of various adaptive and innate immune cell populations using a software-assisted protocol coupled to a whole-slide image scanner. The abundance of various immune cell populations was quantified using a freely available image-analysis software. With the idea that immune cell abundance could be used as a prognostic tool for prostate cancer progression, we optimized a system that would allow for the rapid and accurate quantification of various immunohistochemical markers on a large tissue sample thereby eliminating significant biases of analyses on smaller tissue sections. We believe that our method could help standardize the analysis of diverse immune cell populations within primary tumors and thus facilitate the interpretations of independent studies to better understand the biology of the immune system in prostate cancer.

Section snippets

Patients

Paraffin-embedded formalin-fixed primary tumor specimens from 40 control patients (radical prostatectomy only) and 35 ADT patients (ADT prior to radical prostatectomy) who had undergone surgery between 1991 and 2001 were used in this study. The 35 patients in the ADT group had (i) histologic effects attributable to neoadujvant ADT (ii) histologically identifiable areas of tumor remaining within prostate samples. The control group was matched according Gleason score in order to eliminate immune

Image analysis using Image Scope™

Our previous publication demonstrated the accuracy of quantifying immune cell populations by immunohistochemistry in formalin-fixed paraffin-embedded tissues using a digital image analysis software (Gannon et al., 2006). In the present study, whole-slide high-resolution images were analyzed with the freely available Image Scope™ software from Aperio. By changing the various parameters of the pixel-count algorithm (see Materials and methods section), we were able to accurately differentiate

Discussion

Several studies have associated the degree of immune cell infiltration with prostate cancer progression (Vesalainen et al., 1994, Irani et al., 1999, Sari et al., 1999, Shimura et al., 2000, McArdle et al., 2004, Karja et al., 2005). Increased immune cell infiltrate evaluated by H&E coloration correlates with increased rate of tumor recurrence (Irani et al., 1999) and capsular and perineural invasion (Karja et al., 2005), whereas elevated density of CD4+ T lymphocytes (McArdle et al., 2004) and

Acknowledgments

The authors would like to thank Mona Alam Fahmy, Jason Madore and Josh Levin for their technical assistance, Chantale Auger for her work with the prostate tumor bank, as well as Manon de Ladurantaye and Sylvie Dagenais for administrative assistance. F.S. holds the University of Montreal Chair in Prostate Cancer. R.L. is supported by a fellowship from the Fonds de recherche en santé du Québec (FRSQ). P.O.G. is a recipient of a Ph.D. studentship from the FRSQ and received additional support from

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    Research performed at: CRCHUM – Notre-Dame Hospital – Institut du cancer de Montréal, 1560 Sherbrooke East (Y-4609), Montréal, Québec, Canada H2L 4M1.

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