Evaluation of Matrix Metalloproteases by Artificial Intelligence Techniques in Negative Biopsies as New Diagnostic Strategy in Prostate Cancer

Abstract

Usually, after an abnormal level of serum prostate-specific antigen (PSA) or digital rectal exam, men undergo a prostate needle biopsy. However, the traditional sextant technique misses 15-46% of cancers. At present, there are problems regarding disease diagnosis/prognosis, especially in patients' classification, because the information to be handled is complex and challenging to process. Matrix metalloproteases (MMPs) have high expression by prostate cancer (PCa) compared with benign prostate tissues. To assess the possible contribution to the diagnosis of PCa, we evaluated the expression of several MMPs in prostate tissues before and after PCa diagnosis using machine learning, classifiers, and supervised algorithms. A retrospective study was conducted on 29 patients diagnosed with PCa with previous benign needle biopsies, 45 patients with benign prostatic hyperplasia (BHP), and 18 patients with high-grade prostatic intraepithelial neoplasia (HGPIN). An immunohistochemical study was performed on tissue samples from tumor and non-tumor areas using specific antibodies against MMP -2, 9, 11, and 13, and the tissue inhibitor of MMPs -3 (TIMP-3), and the protein expression by different cell types was analyzed to which several automatic learning techniques have been applied. Compared with BHP or HGPIN specimens, epithelial cells (ECs) and fibroblasts from benign prostate biopsies before the diagnosis of PCa showed a significantly higher expression of MMPs and TIMP-3. Machine learning techniques provide a differentiable classification between these patients, with greater than 95% accuracy, considering ECs, being slightly lower when considering fibroblasts. In addition, evolutionary changes were found in paired tissues from benign biopsy to prostatectomy specimens in the same patient. Thus, ECs from the tumor zone from prostatectomy showed higher expressions of MMPs and TIMP-3 compared to ECs of the corresponding zone from the benign biopsy. Similar differences were found for expressions of MMP-9 and TIMP-3, between fibroblasts from these zones. The classifiers have determined that patients with benign prostate biopsies before the diagnosis of PCa showed a high MMPs/TIMP-3 expression by ECs, so in the zone without future cancer development as in the zone with future tumor, compared with biopsy samples from patients with BPH or HGPIN. Expression of MMP -2, 9, 11, and 13, and TIMP-3 phenotypically define ECs associated with future tumor development. Also, the results suggest that MMPs/TIMPs expression in biopsy tissues may reflect evolutionary changes from prostate benign tissues to PCa. Thus, these findings in combination with other parameters might contribute to improving the suspicion of PCa diagnosis.

Keywords: automatic learning techniques; cancer diagnosis; cancer-associated fibroblast; matrix metalloproteinases; tumor stroma.

Figure 1

Immunohistochemical staining of MMP-2, -9, -11, -13, and TIMP-3 in benign (benign prostatic hyperplasia (BHP), high-grade prostatic intraepithelial neoplasia (HGPIN)) and malignant prostate tissues, before (Pre-CaP) and after prostate cancer (PCa) diagnosis. Magnification: ×200.

Figure 2

Expression of MMPs and TIMP-3 by epithelial cells from benign prostatic hyperplasia (BHP), high-grade prostatic intraepithelial neoplasia (HGPIN), benign prostate biopsies before the diagnosis of prostate cancer (non-tumor zones and zones with future tumor), and from paired zones from prostatectomies by prostate cancer (non-tumor zone and zone with tumor). The values represent the percentages of cases. Columns represent the number of cases of each series.

Figure 3

Expression of MMPs and TIMP-3 by fibroblasts from benign prostatic hyperplasia (BHP), high-grade prostatic intraepithelial neoplasia (HGPIN), benign prostate biopsies before the diagnosis of prostate cancer (non-tumor zones and zones with future tumor), and from paired zones from prostatectomies by prostate cancer (non-tumor zone and zone with tumor). The values represent the percentages of cases. Columns represent the number of cases of each series.

Figure 4

Schematic comparative study of the expression of MMPs and TIMP-3 by epithelial cells and by fibroblasts from benign prostatic hyperplasia (BHP), benign prostate biopsies before the diagnosis of prostate cancer (non-tumor zones and zones with future tumor), and from paired zones from prostatectomies by prostate cancer (non-tumor zone and zone with tumor). +: lower expression; +++: higher expression.

Figure 5

Schematic comparative study of the expression of MMPs and TIMP-3 by epithelial cells and by fibroblasts from high-grade prostatic intraepithelial neoplasia (HGPIN), benign prostate biopsies before the diagnosis of prostate cancer (non-tumor zones and zones with future tumor), and from paired zones from prostatectomies by prostate cancer (non-tumor zone and zone with tumor). +: lower expression; +++: higher expression.

Figure 6

Schematic comparative study of the expression of MMPs and TIMP-3 by epithelial cells and by fibroblasts from benign prostate biopsies before the diagnosis of prostate cancer (non-tumor zones and zones with future tumor), and from paired zones from prostatectomies by prostate cancer (non-tumor zone and zone with tumor). +: lower expression; ++: middle expression; +++: higher expression.

Figure 7

Schematic representation of the studied areas of the samples of patients with PCa.