Infiltration of prostate tumors by eosinophils is ... · Infiltration of prostate tumors by...
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Infiltration of prostate tumors by eosinophils is associated with a lower risk of progression following prostatectomy Oscar Eduardo Molina 1,2 ◦ Fanny Gaignier 1,2 ◦ Molière Nguile Makao 1,2 ◦ Alain Bergeron 1,2 ◦ Vincent Fradet 1,2 ◦ Yves Fradet 1,2 1 Département de chirurgie, Faculté de médecine, Université Laval. 2 Centre de recherche du CHU de Québec-Université Laval, Québec, QC BACKGROUND AND OBJECTIVES: Eosinophils play an important role in the treatment and presentation of antigens from pathogens and allergens, and in the promotion of T cell responses. Their role in cancer is increasingly studied because their infiltration and degranulation in tumors have been associated with clinical outcomes in several types of cancers. The objective of this study was to analyze the infiltration of prostate tumors by eosinophils and to evaluate its prognostic potential. METHODS: Formalin-fixed and paraffin-embedded tumors from 230 intermediate to high-risk prostate cancer patients treated with radical prostatectomy available as tissue microarrays (TMAs) were used for this study. To identify eosinophils, TMA sections were stained with a monoclonal antibody against eosinophil peroxidase (EPX, Mayo Clinic, Clone: MM25-82.2). The density of eosinophil infiltration in the epithelium and tumor stroma was evaluated by semi- automatic analysis of digitalized images using the Calopix software (TRIBVN). Statistical analyses were performed using R to assess a possible correlation between the results and clinical data. RESULTS: Eosinophils showed specific patterns of distribution. The majority of EPX + cells were found in the tumor epithelium but low infiltration in the tumor stroma was also observed. Cox regression analysis showed an association between average percentage of eosinophils in tumor epithelium and a lower risk of biochemical recurrence (HR = 0.603; p = 0.033) for univariate analyses and (HR = 0.353; p = 0.003) for multivariate analyses. CONCLUSION: Eosinophils contribute to an effective antitumor immune response leading to a lower risk of recurrence of prostate cancer. ABSTRACT INTRODUCTION Evaluate the predictive potential value of eosinophil infiltration in prostate tumors Table 1: Characteristics of the cohort of 230 patients Characteristics Numbers Age at diagnosis (yr) <60 73 60 – 70 132 >70 22 PSA at diagnosis (ng/ml) ≤10 162 10 - 20 57 >20 11 Gleason (prostatectomy) ≤6 56 7 141 ≥8 33 T stage (prostatectomy) T2 84 T3-T4 146 Nodal invasion N0 184 N+ 34 Surgical margin Negative 101 Positive 129 Adjuvant hormonotherapy No 165 Yes 65 Biochemical recurrence No 147 Yes 83 Metastasis No 216 Yes 14 Death Prostate cancer 15 Other causes 21 CONCLUSION • Prostate cancer is the most common cancer among Canadian men (excluding non-melanoma skin cancers). • It is the 3rd leading cause of death from cancer in men in Canada. • Gleason score is considered the most accurate predictive marker for disease-specific mortality after primary treatment • The majority of PCa cluster into intermediate group of Gleason (6 and 7) where there is considerable variation in treatment response, disease recurrence and disease-specific death • Intense tumor-associated tissue eosinophilia is associated with a favorable prognostic in oral squamous cell carcinoma 1 • There is a negative role between cancer and eosinophilia when there is evidence of allergies like asthma/lung cancer or atopic dermatitis/skin cancer 2 • Sipuleucel-T treatment is associated with a transient increase in serum eosinophilia that correlated with induced immune response and longer prostate cancer-specific survival 3 Eosinophils may contribute to an effective antitumor immune response in prostate cancer thus, being a determinant factor of long term outcome after prostatectomy they may lead to a lower risk of recurrence. MATERIALS AND METHODS Immunohistochemistry: • 5 μm TMA sections were stained with anti-EPX (Eosinophil Peroxidase) (clone MM25-82.2, dilution 1:1000, Mayo Clinic, Rochester, MN, USA) monoclonal antibody. • Inhibition of endogenous peroxidases was carried out using 3% H2O2 after an antigen retrieval step carried out with Tris/EDTA pH 9 in a PT Link Pre-treatment module from Dako. • Bound antibodies were revealed using the IDetect super strain HRP kit (Empire Genomics). • The slides were counterstained with hematoxylin. Immunohistochemistry evaluation: • Slides were scanned. • The density of eosinophil infiltration in the epithelium and tumor stroma was evaluated by semi-automatic analysis of digitalized images using the Calopix software (TRIBVN). Statistical analyses: • Statistical analyses were performed using R to assess a possible correlation between the results and clinical data (biochemical recurrence). High eosinophils infiltration Low eosinophils infiltration Test Log Rank p = 0.031 (months) Figure1: Analysis of infiltration of prostate cancer TMA by eosinophils using semi-automatic image analysis software Calopix Figure 2: Close-up of the image and semi-automatic cell detection ANALYSES RESULTS Figure 3: Kaplan-Meier curves showing biochemical recurrence-free survival according to the average level of infiltration by eosinophils PROPOSED ROLE REFERENCES How eosinophils may influence immune response… Angiogenesis activation T cell activation MDSC blockade 1. Dorta RG et al. Tumour-associated tissue eosinophilia as a prognostic factor in oral squamous cell carcinomas. Histopathology. 2002 2. Rittmeyer D. and Lorentz A. Relationship between Allergy and Cancer: An Overview. Int Arch Allergy Immunol. 2012 3. McNeel Douglas G. et al. A Transient Increase in Eosinophils is Associated with Prolonged Survival in Men with Metastatic Castration-Resistant Prostate Cancer Who Receive Sipuleucel-T. Cancer Immunol Res. 2014 TIDCs = Dendritic cells antigen presenting cells TAMs = Macrophages regulation of tumor progression TILs = Cytotoxic T lymphocytes adaptative immune response Tregs = T regulators lymphocytes immune response regulation MDSCs = Myeloid-derived suppressor cell immunosuppressive environment High eosinophils infiltration Low eosinophils infiltration Test Log Rank p = 0.054 (months) Table 3: Cox regression analysis of biochemical recurrence according to the average level of eosinophil infiltration in tumor epithelium. Univariate Variable LEVEL HR 95% CI p Average eosinophils Low 1 Infiltration High 0.603 0.379 0.959 0.033 Multivariate* Variable LEVEL HR 95% CI p Average eosinophils Low 1 Infiltration High 0.358 0.181 0.709 0.003 Table 2: Cox regression analysis of biochemical recurrence according to the maximum level of eosinophil infiltration in tumor epithelium. Univariate Variable LEVEL HR 95% CI p Maximum eosinophils Low 1 Infiltration High 0.657 0.427 1.010 0.056 Multivariate* Variable LEVEL HR 95% CI p Maximum eosinophils Low 1 Infiltration High 0.353 0.179 0.689 0.003 Figure 4: Kaplan-Meier curves showing biochemical recurrence-free survival according to the maximum level of infiltration by eosinophils * Adjustment for baseline characteristics * Adjustment for baseline characteristics OBJECTIVE
Transcript of Infiltration of prostate tumors by eosinophils is ... · Infiltration of prostate tumors by...
Infiltration of prostate tumors by eosinophils is associated with a lower risk of progression following prostatectomyOscar Eduardo Molina1,2 ◦ Fanny Gaignier1,2 ◦ Molière Nguile Makao1,2 ◦ Alain Bergeron1,2 ◦
Vincent Fradet1,2 ◦ Yves Fradet1,2
1Département de chirurgie, Faculté de médecine, Université Laval. 2Centre de recherche du CHU de Québec-Université Laval, Québec, QC
BACKGROUND AND OBJECTIVES: Eosinophils play an important role in the treatment and presentation of antigens from pathogens and allergens, and in the promotion of T cell responses. Their role in cancer is increasingly studied becausetheir infiltration and degranulation in tumors have been associated with clinical outcomes in several types of cancers. The objective of this study was to analyze the infiltration of prostate tumors by eosinophils and to evaluate its prognosticpotential. METHODS: Formalin-fixed and paraffin-embedded tumors from 230 intermediate to high-risk prostate cancer patients treated with radical prostatectomy available as tissue microarrays (TMAs) were used for this study. To identifyeosinophils, TMA sections were stained with a monoclonal antibody against eosinophil peroxidase (EPX, Mayo Clinic, Clone: MM25-82.2). The density of eosinophil infiltration in the epithelium and tumor stroma was evaluated by semi-automatic analysis of digitalized images using the Calopix software (TRIBVN). Statistical analyses were performed using R to assess a possible correlation between the results and clinical data. RESULTS: Eosinophils showed specific patterns ofdistribution. The majority of EPX+ cells were found in the tumor epithelium but low infiltration in the tumor stroma was also observed. Cox regression analysis showed an association between average percentage of eosinophils in tumorepithelium and a lower risk of biochemical recurrence (HR = 0.603; p = 0.033) for univariate analyses and (HR = 0.353; p = 0.003) for multivariate analyses. CONCLUSION: Eosinophils contribute to an effective antitumor immune responseleading to a lower risk of recurrence of prostate cancer.
ABSTRACT
INTRODUCTION
Evaluate the predictive potential value of eosinophilinfiltration in prostate tumors
Table 1: Characteristics of the cohort of 230 patients
Characteristics Numbers
Age at diagnosis (yr) <60 73
60 – 70 132
>70 22
PSA at diagnosis (ng/ml) ≤10 162
10 - 20 57
>20 11
Gleason (prostatectomy) ≤6 56
7 141
≥8 33
T stage (prostatectomy) T2 84
T3-T4 146
Nodal invasion N0 184
N+ 34
Surgical margin Negative 101
Positive 129
Adjuvant hormonotherapy No 165
Yes 65
Biochemical recurrence No 147
Yes 83
Metastasis No 216
Yes 14
Death Prostate cancer 15
Other causes 21
CONCLUSION
• Prostate cancer is the most common cancer among Canadian men(excluding non-melanoma skin cancers).
• It is the 3rd leading cause of death from cancer in men in Canada.
• Gleason score is considered the most accurate predictive marker fordisease-specific mortality after primary treatment
• The majority of PCa cluster into intermediate group of Gleason (6and 7) where there is considerable variation in treatment response,disease recurrence and disease-specific death
• Intense tumor-associated tissue eosinophilia is associated with afavorable prognostic in oral squamous cell carcinoma1
• There is a negative role between cancer and eosinophilia when thereis evidence of allergies like asthma/lung cancer or atopicdermatitis/skin cancer2
• Sipuleucel-T treatment is associated with a transient increase inserum eosinophilia that correlated with induced immune responseand longer prostate cancer-specific survival3
Eosinophils may contribute to an effective antitumor immune response in prostate cancer thus, being adeterminant factor of long term outcome after prostatectomy they may lead to a lower risk ofrecurrence.
MATERIALS AND METHODS
Immunohistochemistry:
• 5 μm TMA sections were stained with anti-EPX (Eosinophil Peroxidase) (clone MM25-82.2, dilution 1:1000, Mayo Clinic, Rochester, MN, USA) monoclonal antibody.
• Inhibition of endogenous peroxidases was carried out using 3% H2O2 after an antigen retrieval step carried out with Tris/EDTA pH 9 in a PT Link Pre-treatment module from Dako.
• Bound antibodies were revealed using the IDetect super strain HRP kit (Empire Genomics).
• The slides were counterstained with hematoxylin.
Immunohistochemistry evaluation:• Slides were scanned.• The density of eosinophil infiltration in the epithelium and tumor stroma was
evaluated by semi-automatic analysis of digitalized images using the Calopixsoftware (TRIBVN).
Statistical analyses:
• Statistical analyses were performed using R to assess a possible correlation between the results and clinical data (biochemical recurrence).
High eosinophils infiltration
Low eosinophils infiltration
Test Log Rank p = 0.031
(months)
Figure1: Analysis of infiltration of prostate cancer TMA by eosinophils using semi-automatic image analysis software Calopix
Figure 2: Close-up of the image and semi-automatic cell detection
ANALYSES
RESULTS
Figure 3: Kaplan-Meier curves showing biochemical recurrence-free survival according to the average level
of infiltration by eosinophils
PROPOSED ROLE
REFERENCES
How eosinophils may influence immune response…
Angiogenesis activation
T cell activation
MDSC blockade
1. Dorta RG et al. Tumour-associated tissue eosinophilia as a prognostic factor in oral squamous cell carcinomas. Histopathology. 2002
2. Rittmeyer D. and Lorentz A. Relationship between Allergy and Cancer: An Overview. Int Arch Allergy Immunol. 2012
3. McNeel Douglas G. et al. A Transient Increase in Eosinophils is Associated with Prolonged Survival in Men with Metastatic Castration-Resistant Prostate Cancer Who Receive Sipuleucel-T. Cancer Immunol Res.
2014
TIDCs = Dendritic cells
antigen presenting cells
TAMs = Macrophages
regulation of tumor progression
TILs = Cytotoxic T lymphocytes
adaptative immune response
Tregs = T regulators lymphocytes
immune response regulation
MDSCs = Myeloid-derived suppressor cell
immunosuppressive environment
High eosinophils infiltration
Low eosinophils infiltration
Test Log Rank p = 0.054
(months)
Table 3: Cox regression analysis of biochemical recurrence according to the average level of eosinophil infiltration in tumor epithelium.
Univariate
Variable LEVEL HR 95% CI p
Average eosinophils Low 1
Infiltration High 0.603 0.379 0.959 0.033
Multivariate*
Variable LEVEL HR 95% CI p
Average eosinophils Low 1
Infiltration High 0.358 0.181 0.709 0.003
Table 2: Cox regression analysis of biochemical recurrence according to the maximum level of
eosinophil infiltration in tumor epithelium.
Univariate
Variable LEVEL HR 95% CI p
Maximum eosinophils Low 1
Infiltration High 0.657 0.427 1.010 0.056
Multivariate*
Variable LEVEL HR 95% CI p
Maximum eosinophils Low 1
Infiltration High 0.353 0.179 0.689 0.003
Figure 4: Kaplan-Meier curves showing biochemical recurrence-free survival according to the maximum
level of infiltration by eosinophils
* Adjustment for baseline characteristics
* Adjustment for baseline characteristics
OBJECTIVE
