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BRIEF REPORT
Year : 2023 | Volume
: 41
| Issue : 2 | Page : 116-120
The reduced expression of trimethylation of histone H3 at lysine 27 in primary cutaneous mucinous carcinoma
Shu-Hao Li1, Chien-Chun Chiou2, Chien-Chin Chen3
1 Department of Dermatology, Chang Gung Memorial Hospital Linkou Branch, Taoyuan, Taiwan
2 Department of Dermatology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chiayi, Taiwan
3 Department of Pathology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chiayi; Department of Cosmetic Science, Chia Nan University of Pharmacy and Science, Tainan; Ph.D. Program in Translational Medicine, Rong Hsing Research Center For Translational Medicine, National Chung Hsing University, Taichung; Department of Biotechnology and Bioindustry Sciences, College of Bioscience and Biotechnology, National Cheng Kung University, Tainan, Taiwan
Correspondence Address:
Dr. Chien-Chin Chen
Department of Pathology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, No. 539, Zhongxiao Rd., East Dist., Chiayi 600
Taiwan
Dr. Chien-Chun Chiou
Department of Dermatology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, No. 539, Zhongxiao Rd., East Dist., Chiayi 600
Taiwan
Source of Support: None, Conflict of Interest: None
DOI: 10.4103/ds.DS-D-22-00103
Primary cutaneous mucinous carcinoma (PCMC) is rare, and its carcinogenesis is unclear. Trimethylation of histone H3 at lysine 27 (H3K27me3) is a key regulator in chromatin remodeling-controlled transcription. Focusing on the epigenetic mechanism, we aimed to investigate the expression of H3K27me3 in PCMC by immunohistochemistry. A retrospective cohort of PCMC patients from a tertiary hospital in Taiwan was enrolled to evaluate the clinicopathologic features, treatment outcome, and protein expression. Immunohistochemistry for H3K27me3 was performed on all PCMCs and a comparison group of colonic mucinous adenocarcinoma and pure mucinous carcinoma of the breast. The percentage of H3K27me3-negative tumor cells was calculated and analyzed. Three patients with PCMC were recruited. All PCMCs were solitary and slow growing, arising from the head-and-neck region. All PCMCs had tumor excision without local recurrence or metastasis. The loss of H3K27me3 expression was significant in PCMCs (mean ± standard deviation [SD]: 21.0% ± 6.6%) compared to other mucinous carcinomas (mean ± SD: 3.8% ± 1.7%) (P = 0.019). In conclusion, we report a reduction in H3K27me3 expression in PCMC. In contrast, H3K27me3 expression is retained or mildly reduced in other mucinous carcinomas. This is the first study to indicate a possible role of epigenetic events in the pathobiology of PCMC.
Keywords: Dermatology, epigenetic, histone, mucinous carcinoma, skin, trimethylation of histone h3 at lysine 27
Primary cutaneous mucinous carcinoma (PCMC) is a rare malignant adnexal neoplasm.[1] Although the clinical appearance of PCMC is nonspecific, these tumors are mostly present in the sixth or seventh decade, with a predilection for the face, particularly the eyelids.[2] A predominance of Caucasian, male, and elderly patients has been found for PCMC.[2] Histological features are believed to be of eccrine origin, even though some cases have apocrine differentiation.[3] Although PCMC generally has a good prognosis, local recurrence, and occasional regional node metastasis are not uncommon.[2]
Differentiating the diagnosis of primary mucinous carcinoma of the skin from metastatic mucinous carcinoma can be difficult since the histological findings overlap significantly. Immunochemistry staining has reportedly been helpful in diagnosis and enhancing margin control in Mohs surgery.[4] However, a systemic oncologic workup is warranted due to the high proportion of metastatic lesions. Furthermore, additional diagnostic markers would be useful in this differential diagnosis.
In recent explorations, epigenetic changes have been shown to play a crucial role in developing skin cancers. Trimethylation of histone H3 at lysine 27 (H3K27me3) is considered an inactive methylation marker. H3K27me3 regulates chromatin compaction and promotes the transcriptional inactivation of genes.[5] Previous studies have already accentuated the significant role of H3K27me3 in the pathomechanism of cancer.[6],[7],[8] However, there are no studies regarding the relationship between PCMC and the expression of H3K27me3 in the current literature. Therefore, our study aimed to investigate the expression of H3K27me3 in PCMC.
Materials and MethodsPatients and specimens
A retrospective cohort of patients with PCMC in the archive of the Department of Pathology of Ditmanson Medical Foundation Chia-Yi Christian Hospital from 2000 to 2017 was established. A PCMC case should meet all of the following criteria: (1) no metachronous or synchronous tumor; (2) mucinous carcinoma and mucinous components comprised more than 90% of the neoplasm; (3) a cutaneous lesion with or without subcutaneous involvement; (4) tumor excision for total evaluation. Survival data, clinical characteristics, and treatment modalities were obtained from medical charts. Moreover, all PCMC cases were histopathologically reviewed, including the pattern and expression of immunophenotypes. In addition, three colonic mucinous adenocarcinomas and three pure mucinous carcinomas of the breast were randomly retrieved without clinical data and utilized as a comparison group for H3K27me3 expression.
Representative formalin-fixed paraffin-embedded (FFPE) tumor tissues were prepared for further immunohistochemistry. The study was approved by the Institutional Review Board of Ditmanson Medical Foundation Chia-Yi Christian Hospital (No. 106065) and all patient informed consent was obtained. This study was complied with the principles of the Declaration of Helsinki.
Immunohistochemistry and evaluation
Immunohistochemistry was performed on 4 μm thick sections of FFPE specimens after deparaffinization, the removal of xylene, hydration, and microwave-based antigen retrieval using an autostainer (Bond-Max autostainer; Leica Biosystems Newcastle Ltd., Melbourne, Australia) with hematoxylin as a counterstain. The primary antibodies, anti-H3K27me3 (C36B11, 1:200) (Cell Signaling Technology, Switzerland) and anti-Ki-67 (RM-9106-S, 1:100) (LabVision, Sweden), were rabbit monoclonal antibodies. Appropriate positive and negative controls were used. Expression levels were calculated as the proportion of nuclear-positive cells in each case and were graded by a pathologist blinded to the patients' information. One hundred tumor cells per patient were counted, and tumor cells that exhibited nuclear positivity and at least moderate intensity were considered immunohistochemically positive.
Statistical analysis
All data were analyzed by SPSS 25.0 (SPSS Inc., Chicago, IL, USA). Appropriate statistical tests were used to examine the relationships and correlations between variables, including the χ2 test or Fisher's exact test (n < 5) for categorical variables and the Mann‒Whitney U test for continuous variables. All tests were two-sided, and P < 0.05 indicated a statistically significant difference. Finally, we compared the percentage of loss of nuclear expression for H3K27me3 between PCMC and the comparison group using the Mann‒Whitney U test.
ResultsThree PCMC patients were retrospectively analyzed with histologic confirmation, clinical information, and follow-up data. There were two men and one woman with a median age of 58 years (range: 57–64 years old). All tumors were solitary and slow growing. They arose from the head-and-neck region, and the mean size was 1.73 cm (range: 0.8–3.2 cm). All tumors were excised with safe margins and showed no recurrence or metastasis with a mean follow-up of 40.7 months (range: 10–80 months). All patients were alive at the last follow-up. [Table 1] summarizes the patients' clinicopathologic characteristics.
Table 1: Clinical and histologic characteristics of patients with primary cutaneous mucinous carcinomaHistopathologically, all PCMCs had solid nests or micropapillary clusters of neoplastic cells with abundant extracellular mucin production [Figure 1]a, [Figure 1]c, and [Figure 1]e. The Ki-67 proliferation index ranged from 6% to 10% in PCMC. All PCMCs had a heterogeneous and variable proportion of nuclear positivity of H3K27me3 expression [Figure 1]b, [Figure 1]d, and [Figure 1]f. Since H3K27me3 was retained or only mildly reduced in colonic mucinous adenocarcinomas and pure mucinous carcinomas of the breast (the percentage of loss of H3K27me3, mean ± standard deviation [SD]: 3.8% ± 1.7%), the loss of H3K27me3 nuclear positivity in PCMC was significantly higher (mean ± SD: 21.0% ± 6.6%) (P = 0.019) [Table 2]. [Figure 2]a, [Figure 2]b, [Figure 2]c, [Figure 2]d demonstrates representative results of the comparison group.
Figure 1: The reduced nuclear expression of H3K27me3 in PCMC. (a) The histopathology of PCMC 1 revealed trabecular nests of atypical epithelial cells with abundant extracellular mucin production, (b) H3K27me3 staining of PCMC 1 revealed reduced nuclear positivity, (c) PCMC 2 showed micropapillary and papillary clusters of neoplastic epithelial cells floating in the mucin pools with occasionally vacuolated cytoplasm and rare mitoses, (d) Reduced H3K27me3 staining is shown in PCMC 2, (e) PCMC 3 displayed cribriform and solid sheets of tumor cells with abundant extracellular mucin, and (f) H3K27me3 staining of PCMC 3 revealed reduced nuclear positivity (a, c, and e: H and E, ×40; b, d, and f: H3K27me3, ×400). H3K27me3: Trimethylation of histone H3 at lysine 27, PCMC: Primary cutaneous mucinous carcinoma.
Figure 2: The intact nuclear expression of H3K27me3 in mucinous carcinoma of the breast and colonic mucinous adenocarcinoma. (a) A mucinous carcinoma of the breast revealed floating clusters and nests of intermediately differentiated tumor cells in extracellular mucin pools separated by fibrous septa, (b) H3K27me3 staining of the mucinous carcinoma of the breast demonstrated intact nuclear positivity, (c) A colonic mucinous adenocarcinoma showed strips and clusters of neoplastic columnar cells floating in the extracellular mucin pools, and (d) The tumor cells mostly expressed H3K27me3 (a and c: H and E, ×40; b and d: H3K27me3, ×400). H3K27me3: Trimethylation of histone H3 at lysine 27.
Table 2: The percentage of loss of trimethylation of histone H3 at lysine 27 positivity in primary cutaneous mucinous carcinoma and mucinous carcinoma of the colon or breast Discussion PCMC is a rare, slow-growing malignant neoplasm that originates from sweat glands.[1] These tumors have an indolent clinical course and commonly arise in elderly male patients.[2] Historically, there is a higher prevalence of PCMC in the Caucasian population than in Asian or African populations. Clinically, this neoplasm presents as a solitary, slow growing, flesh-colored, erythematous or blue, and usually painless nodule, cyst, or ulceration in the head-and-neck region, particularly the eyelids, face, and scalp.[1] Histologically, a typically characteristic of PCMCs are nests of epithelial cells floating in lakes of extracellular mucin.[3] Unfortunately, the histological distinction between PCMC and metastatic mucinous carcinoma, especially from the breast and gastrointestinal tract, is modest. Immunohistochemical staining may assist in ruling out cutaneous metastatic mucinous tumors. CDX-2 can be used to indicate colorectal origin, and TTF-1 can indicate pulmonary origin. However, with a characteristic CK7+/CK20− phenotype, most PCMC neoplastic cells also express epithelial membrane antigen, carcinoembryonic antigen, E-cadherin, GCDFP15, low-molecular-weight cytokeratins, estrogen, and progesterone receptor.[3] Subsequently, immunohistochemical panels may help differentiate PCMC from metastatic mucinous carcinomas, but a more specific marker for PCMC is needed.
Accumulating evidence supports that epigenetic regulation plays a crucial role in the progression of various diseases by modulating the activation and inactivation of gene transcription. Epigenetic modifications are demonstrating increasing importance in various cancers, including cutaneous cancers.[5] The aberrant pattern in these epigenetic processes alters the expression of genes involved in the cell cycle, cell motility, and apoptosis.[5] Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase and core catalytic subunit of polycomb repressive complex. EZH2 methylates H3K27 and is responsible for the epigenetic silencing of many genes.[9] The EZH2-mediated pathway is involved in the aggressive phenotype of cancer cells by repressing a group of tumor suppressor genes.[10] Notably, H3K27me3 is a well-established histone marker for chromatin compaction and promotes the formation of heterochromatic regions, which decreases transcriptional activity.[7] The levels of H3K27me3 expression can be assessed in FFPE tissues by immunohistochemical techniques to evaluate its role in different neoplasms.[7],[8] Interestingly, increased and decreased activity of enzymes controlling H3K27 methylation may be devoted to carcinogenesis and poor prognosis.[7]
It has been reported that repressive H3K27me3 may repress differentiation genes in proliferative and undifferentiated keratinocytes in epidermal development.[11] Therefore, the aberrant expression of H3K27me3 in cutaneous neoplasms could be distinctive from that in normal skin tissues. Since PCMC with neuroendocrine differentiation is not uncommon, it is interesting that reduced H3K27me3 expression could induce neuroendocrine differentiation in prostate cancer cells.[12] Moreover, pure Merkel cell carcinoma, a primary neuroendocrine skin cancer, has been reported to strongly reduce H3K27me3 staining.[7] In comparison, our PCMC cases had a variable degree of neuroendocrine differentiation, which was immunohistochemically shown by synaptophysin, and probably correlated with reduced H3K27me3 expression [Table 1].
In this study, we hypothesized that there are epigenetic modifications during the carcinogenesis of PCMC. To explore the potential role of epigenetic events in PCMC, we examined H3K27me3 expression levels in PCMC and its histological counterpart, mucinous carcinomas of the breast and colon. Our results indicate that the decreased labeling of H3K27me3 is strongly associated with PCMC but not with mucinous carcinomas of the breast and colon. Combining the evidence discussed above, epigenetic modulations, such as histone modifications, that affect H3K27me3 expression may play a potential role in the carcinogenesis of PCMC. These epigenetic modifications may be able to differentiate tumor origin and direct novel and potential applications for targeted therapy in the future.
However, our report has multiple limitations. First, we had only three PCMC cases retrospectively recruited from a single institute. The small case number cannot yield conclusive evidence. Second, all FFPE blocks from the PCMC cases were stored for at least 10 years, and antigen decay in archival FFPE tissues might occur. Third, there was only one approach and one target protein level change demonstrated by immunohistochemistry in our brief report. Molecular tests for extensive epigenomic targets in a large-scale case study can further provide comprehensive results.
ConclusionTo the best of our knowledge, this is the first study to demonstrate that epigenetic regulation through histone posttranslational modifications may play a role in the tumorigenesis of PCMC. The low aberrant expression of H3K27me3 might help distinguish PCMC from pathological mimics. A more extensive set of cases is necessary to assess the biological significance of H3K27me3 expression and its potential diagnostic value in PCMC. Notably, further investigation of H3K27me3 for prognosis prediction and therapeutic targeting might be promising for PCMC.
Data availability statement
The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.
Financial support and sponsorship
Nil.
Conflicts of interest
There are no conflicts of interest.
References
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