TAp63 and ΔNp63 (p40) in prostate adenocarcinomas: ΔNp63 associates with a basal-like cancer stem cell population but not with metastasis

Abstract

Like other malignancies, prostate tumors are thought to contain cancer stem-like cells (CSCs) that are responsible for growth, metastasis, and therapy resistance. ΔNp63 (also called p40) is a regulator of normal prostate stem/progenitor cell activities and a marker of normal basal epithelial cells. The levels of ΔNp63 are reduced in prostate adenocarcinomas, although there is also evidence that ΔNp63 is involved in CSC regulation and drives metastasis to the bone. We studied metastatic deposits of prostate cancers with isoform-specific ΔNp63 and TAp63 antibodies. We identified p63-positive cells in only 3 of 42 metastatic prostate tumors (7%), including 2/38 (5.3%) “usual-type” adenocarcinomas. ΔNp63 and TAp63 isoforms were present in the nuclei of a small subpopulation (< 1%) of tumor cells in these metastases. ΔNp63-positive cells showed a basal-like cell phenotype (cytokeratin 8- and androgen receptor-negative, high molecular weight cytokeratin- and cytokeratin 19-positive), distinct from the tumor bulk. TAp63-positive cells were similar but were sometimes cytokeratin 8-positive. A subset of ΔNp63-positive tumor cells were CD44-positive, a marker of “basal” CSCs but were not positive for the “epithelial” CSC marker ALDH1. TAp63 was not associated with either of these CSC markers. None of the tumors containing p63-positive cells showed evidence of bone metastasis, compared with 28% of the p63-negative tumors. These data show that both ΔNp63 and TAp63 are present in only a small proportion of prostate adenocarcinomas and do not associate with metastasis. The data suggest heterogeneity of CSCs in prostate cancer, similar to other cancer types.

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Data availability

Scanned digital slide images from which the photomicrographs in this article are derived are available on request.

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Funding

This work was supported by the Czech Science Foundation (19-06530S), the European Regional Development Fund - Project ENOCH (No. CZ.02.1.01/0.0/0.0/16_019/0000868) and the Ministry of Health, Czech Republic—conceptual development of research organization (MMCI, 00209805). The MMCI biobank is supported by grant LM2018125 from the Ministry of Education, Youth and Sports and co-funded by ADOPT BBMRI-ERIC, supported by EU Horizon 2020 (grant agreement No. 676550). MG was a Brno Ph.D. Talent Scholarship Holder, funded by the Brno City Municipality.

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MG performed immunohistochemistry and was a major contributor to preparing the manuscript. RN was involved in study concept and design, selected patient material, and interpreted immunohistochemical staining and clinicopathological data. ZP performed immunohistochemistry and assisted with analysis. BV was involved in study design and supervision. PC designed the study, interpreted data, and was a major contributor to writing the manuscript. All authors were involved in revising the manuscript and all authors read and approved the final version.

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Correspondence to Philip J. Coates.

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Conflict of interest

BV is a consultant for and RN is a co-owner of Moravian Biotechnology, who originally produced the p63 monoclonal antibodies used in this study. The company did not provide financial support or have any influence over the design, execution, or interpretation of the data. MG, ZP, and PJC declare that they have no competing interests.

Ethics approval

The study was performed retrospectively on redundant excess tissues. All patient data were anonymized. In accordance with the Declaration of Helsinki and the regulations of the European Union, the use of excess and redundant tissues was approved following local ethical committee review by the Biobanking and Biomolecular resources Research Infrastructure (BBMRI) at the Masaryk Memorial Cancer Institute, Brno. Patient consent is not required for the retrospective use of anonymized, redundant excess tissues.

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Fig. S1
figure5

ΔNp63 immunostaining in three lymph node metastases. Sections were stained with immunoperoxidase (brown) and nuclei were counterstained with hematoxylin (blue). Panels on the right show a different field of the same section at higher magnification. (a) Prostate adenocarcinoma sample where only a few neoplastic cells were ΔNp63+. (b) In the other adenocarcinoma sample, ΔNp63+ cells were concentrated in one region of the lymph node tumor deposit, where they represented ~10% of the tumor cells. (c) Carcinosarcoma with ~5% ΔNp63+ tumor cells occurring in one region of the lymph node metastasis. Arrows indicate examples of ∆Np63+ cells. (PNG 1803 kb)

Fig. S2
figure6

ΔNp63 immunostaining in primary tumor samples in which the corresponding metastasis samples did not contain ΔNp63+ cells. Sections were stained for ΔNp63 with immunoperoxidase (brown) and nuclei were counterstained with hematoxylin. The photomicrographs show four examples of ΔNp63+ cells in radical prostatectomy tissues. In each photomicrograph, whilst the majority of cells are basally located in benign appearing glands, there are occasional cells that are difficult to assign. In all cases, regions such as those shown were found towards the edge of the invasive tumor and were always found together with normal gland structures. (PNG 4493 kb)

Fig. S3
figure7

ΔNp63 co-localization with markers of epithelial differentiation in benign prostate hyperplasia. (a) Dual-peroxidase labelling for ΔNp63 (brown) and the indicated cytokeratins (gray). Nuclei are counterstained with nuclear fast red. The majority of ΔNp63+ cells co-localize with HMWCK and variably with CK19. ΔNp63+ cells are negative for CK8. (b) Immunofluorescence. p63+ cells (red) are negative or weakly positive for AR (green). The merged image shows counterstaining with DAPI (blue). (PNG 1319 kb)

Fig. S4
figure8

ΔNp63 co-localization with cancer stem cell markers in benign prostate hyperplasia. ΔNp63+ cells (brown) are negative for ALDH1 (gray) and are mostly positive for CD44 (gray). (PNG 770 kb)

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Galoczova, M., Nenutil, R., Pokorna, Z. et al. TAp63 and ΔNp63 (p40) in prostate adenocarcinomas: ΔNp63 associates with a basal-like cancer stem cell population but not with metastasis. Virchows Arch (2020). https://doi.org/10.1007/s00428-020-02944-z

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Keywords

  • p63
  • p40
  • Prostate cancer
  • Cancer stem cells
  • CD44
  • Metastasis