Objective

To credential Stathmin 1 (STMN1) and p16(INK4A) (p16) as adjunct markers for the diagnosis of serous tubal intraepithelial carcinoma (STIC), and to compare STMN1 and p16 expression in p53-positive and p53-negative STIC and invasive high-grade serous carcinoma (HGSC).

Methods

Immunohistochemistry (IHC) was used to examine STMN1 and p16 expression in fallopian tube specimens (n=31) containing p53-positive and p53-negative STICs, invasive HGSCs, and morphologically normal FTE (fallopian tube epithelium). STMN1 and p16 expression was scored semiquantitatively by four individuals. The semiquantitative scores were dichotomized, and reported as positive or negative. Pooled siRNA was used to knockdown p53 in a panel of cell lines derived from immortalized FTE and HGSC.

Results

STMN1 and p16 were expressed in the majority of p53-positive and p53-negative STICs and concomitant invasive HGSCs, but only scattered positive cells were present in morphologically normal FTE. Both proteins were expressed consistently across multiple STICs from the same patient and in concomitant invasive HGSC. Knockdown of p53 in immortalized FTE cells and in four HGSC-derived cell lines expressing different missense p53 mutations did not affect STMN1 protein levels.

Conclusions

This study demonstrates that STMN1 and p16 are sensitive and specific adjunct biomarkers that, when used with p53 and Ki-67, improve the diagnostic accuracy of STIC. The addition of STMN1 and p16 helps to compensate for practical limitations of p53 and Ki-67 that complicate the diagnosis in up to one third of STICs.

Recent insights supporting the fallopian tube epithelium (FTE) and serous tubal intraepithelial carcinomas (STIC) as the tissue of origin and the precursor lesion, respectively, for the majority of high-grade serous ovarian carcinomas (HGSOC) provide the necessary context to study the mechanisms that drive the development and progression of HGSOC. Here, we investigate the role of the E3 ubiquitin ligase RNF20 and histone H2B monoubiquitylation (H2Bub1) in serous tumorigenesis and report that heterozygous loss of RNF20 defines the majority of HGSOC tumors. At the protein level, H2Bub1 was lost or downregulated in a large proportion of STIC and invasive HGSOC tumors, implicating RNF20/H2Bub1 loss as an early event in the development of serous ovarian carcinoma. Knockdown of RNF20, with concomitant loss of H2Bub1, was sufficient to enhance cell migration and clonogenic growth of FTE cells. To investigate the mechanisms underlying these effects, we performed ATAC-seq and RNA-seq in RNF20 knockdown FTE cell lines. Loss of RNF20 and H2Bub1 was associated with a more open chromatin conformation, leading to upregulation of immune signaling pathways, including IL6. IL6 was one of the key cytokines significantly upregulated in RNF20- and H2Bub1-depleted FTE cells and imparted upon these cells an enhanced migratory phenotype. These studies provide mechanistic insight into the observed oncogenic phenotypes triggered by the early loss of H2Bub1. SIGNIFICANCE: Loss of RNF20 and H2Bub1 contributes to transformation of the fallopian tube epithelium and plays a role in the initiation and progression of high-grade serous ovarian cancer.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/4/760/F1.large.jpg.