Immunohistochemical expression of p53 in Type I and II epithelial ovarian cancer among Sudanese women: a cross-sectional study

Introduction

Ovarian cancer is a fatal disease, the mortality rate ranks the highest of all gynecological malignancies¹. It is considered the third most common cancer in the female reproductive system (following uterine cervix and corpus) and the leading cause of death from gynecologic malignancies in the United States as estimated by the American Cancer Society for the year 2019^1,2. Ovarian cancer refers to a group of morphologically and genetically heterogeneous neoplasms^3,4. In Sudan, ovarian cancer represents the fourth most frequent tumor type in women⁵.

Based on clinicopathological and molecular studies, epithelial ovarian cancer (EOC) is classified as type I or type II. Type I tumors are genetically quite stable, typically present at a low stage, and reveal distinct, morphologic differences than type II tumors⁶. These include different histotypes: low-grade serous, endometrioid, clear-cell, and mucinous ovarian carcinoma. Type I tumors are characterized by distinct molecular genetics profiles, such as mutations in KRAS, BRAF, PIK3CA, PTEN and ERBB2, but not TP53⁶. Type II tumors are generally high-grade serous (about 90% of all EOCs). They are highly aggressive, develop rapidly, present in an advanced stage in most cases, genetically unstable and express a mutated TP53^7–12. TP53 mutations have an important role in the prognosis and treatment of ovarian cancer¹³. Mutations in TP53 are found in high grade and rarely in low grade serous ovarian cancers. TP53 encodes the 53 kDa nuclear protein, their mutations leading to gain or loss of function of its protein product. TP53 mutation leads either to overexpression of p53 protein or complete lack of expression, while wild-type p53 is associated with focal expression^14,15.

Immunohistochemical staining for p53 was considered as an essential biomarker for clinical trials targeting mutant p53 and used in the diagnostic workup of carcinomas of multiple sites, including ovarian cancers¹⁶. It is used as a substitute for TP53 mutational analysis, these mutations were global in high-grade serous ovarian cancer (HGSOC) (over 96% were mutated), so were used to discriminates between high- and low-grade serous carcinomas^17–20. Access to TP53 sequencing is not feasible in many low- and middle-income countries; pathologists there used p53 immunohistochemistry, which is quick, easy to perform, inexpensive and can approach 100% specificity for the presence of TP53 mutation. Its high negative predictive value is clinically useful as it can exclude the possibility of a low-grade serous tumor^18–20. To our knowledge, there are no published reports about the frequency of p53 immunostaining in type I and II EOC in Sudan. The study aimed to determine the frequency of p53 overexpression and its relationship with tumor types I and II and tumor grades among Sudanese women with EOC.

Methods

Results

Overexpression of p53

Positive p53 immunostaining was seen in 35.1% (40/114) of ovarian epithelial carcinoma. From the 40 positive cases, staining intensity was as follows: 25 cases exhibited strong staining (+3), and 15 cases were moderate staining (+2). While the remaining 74 cases were considered as negative results (complete absence of p53 expression seen in 45 cases and +1 staining in 29 cases) (Figure 1–Figure 4).

Figure 1. High-grade serous carcinoma showing strong immunostaining (+3 staining) for p53.

Magnification: main image, x40; inset, x10.

Figure 2. High-grade serous carcinoma showed moderate immunostaining (+2 staining) for p53.

Magnification, x40).

Figure 3. High-grade serous carcinoma showed weak immunostaining (+1 staining) for p53.

Magnification, x40.

Figure 4. High-grade serous carcinoma showing absence of immunostaining for p53.

Magnification, 40X.

Overexpression of p53 was associated significantly with the histological subtype (p = 0.004) as seen in Table 1. p53 expression was significantly associated with tumor grade (p = 0.003): it was positive in 49.2% (30/61) of high-grade tumors, 16.7% (1/6) of the non-graded borderline tumors, and 19.15% (9/47) of the low-grade tumors. The association between p53immunohistochemical expression and epithelial ovarian cancer type showed a highly significant association (p = 0.000). p53 was positive in 53.7% (29/54) of Type II and 18.5% (10/54) of Type I tumors (Table 2).

Table 1. Association between p53 expression and histological subtypes of the studied ovarian tumors.

Histological sub-type	Variable	p53 expression		Total
		positive	negative
High grade serous	n	29	25	54
	%	53.7	46.3	100%
Low grade serous	n	0	6	6
	%	0	100	100%
Mucinous carcinoma	n	7	15	22
	%	31.8	68.2	100%
Endometroid	n	2	14	16
	%	12.5	87.5	100%
Clear cell	n	1	6	7
	%	14.3	85.7	100%
Brenner tumor	n	0	3	3
	%	0	100	100%
Borderline	n	1	5	6
	%	16.7	83.3	100%
P value		0.004

Table 2. Association between p53 expression and studied ovarian tumor type and grade.

Clinicopathological characteristic (N)	p53 status		P value
	Positive	Negative
Cancer Type
Type I (54)	10 (18.5%)	44 (81.5%)	0.000
Type II (54)	29 (53.7%)	25 (46.3%)
Borderline (6)	1 (16.7%)	5 (83.3%)
Cancer Grade
High grade (61)	30 (49.2%)	31 (50.8%)	0.003
Low grade (47)	9 (19.1%)	38 (80.9%)
Non-graded (6)	1 (16.7%)	5 (83.3%)

Discussion

Immunohistochemical staining for p53 was used as a surrogate for TP53 mutational analysis to discriminate between high (over 96% was mutated) and low-grade serous carcinomas^17–20.

In the present study, p53 positivity was observed in 35.1% (40/114) of EOC cases, and higher positivity was showed in HGS (53.7%; 29/54); a significant association was found between p53 expression and histopathological diagnosis and with tumor grade and tumor type. This result agreed with those of Tan et al., who found that 64.18% (43/67) of HGS samples analyzed overexpressed p53¹³. Harris et al., found that in 274 cases, 68% of tumors were characterized as p53 mutant (n= 186) and p53-mutant tumors were more likely to be HGS (72%) with significant association between p53 expression and histology and grade of tumor¹⁰. Markowska et al., found that positive expression of p53 protein was observed in 27.8% of EOC²⁴. Yemelyanova et al. found out of the 57 tumors, 36 contained functional mutations and 23 cases (63%) with mutant Tp53 were positive for p53 IHC and 70% of HGS were p53 mutant¹⁶. Kobel et al. showed p53 marker overexpressed in 69% (118/171) of HGS cases¹⁸. Kobel et al. also showed that abnormal Tp53 expression was detected in 904 out of 912 (99.1%) of HGS²⁵. Our results also agreed with those of Matsuo et al., who found that for 121 cases of high-grade serous p53 positivity was (71.4%)²⁶, and Zhang et al., found that strong staining of p53 was observed in (70.8%) of serous carcinomas and showed that the tumor type was closely associated with the expression of p53²⁷. Mozes et al., found p53 overexpression in 83.6% of HGS²⁸. Bell et al. reported that Tp53 mutated in 96% of HGS²⁹. Moreover, Oaknin et al. reported that expression of p53 markers in the various histological types of ovarian carcinoma (HGS 94%, LGS 0%, CCC 12%, EC 15% and MC 61%)¹⁷ were nearly to our results (HGS 53.7%, LGS 0%, CCC 14.3%, EC 12.5% and MC 31.8%). Lim et al., reported that p53 was not expressed in EC in 30 samples, but they agreed with our result in that they found HGS overexpressed p53 in 50% of cases examined³⁰. Alexander et al. reported that p53 was positive for HGS in 68% of cases (52/76) and differed from our result in that they found positivity in LGS (24%; 18/76)¹⁹. Mackenzie et al., reported a higher percentage of p53 in mucinous carcinoma (68 %)³¹.

Furthermore, our result agreed also with Sallum et al., who reported that p53 positivity was found in 68.2% of HGS and significant association was found between p53 expression and tumor type¹⁵. Sundov et al.,³² and Tan et al.,¹³ also found that immunoexpression of p53 was significantly associated with tumor grade. And it disagreed with Brachova et al., who showed that p53 protein expression insignificantly associated with tumor grade³³.

The present study showed that p53 marker was overexpressed in (53.7%) of type II, and (18.5%) of type I. This result agreed with Carter et al., 2018, who reported that p53 was highly expressed in type II EOC (68.8%) than type I (33.3%)⁴. HGSOC is the most frequent type of ovarian cancer and has been associated with a poor clinical outcome³⁴. According to The Cancer Genome Atlas report, mutations in TP53 are the most common events in EOC, especially in HGSCs³⁵.

The results of some of these studies may be conflicting primarily because of the indiscriminate grouping of TP53 mutations, which can result in either loss of function or gain of function³⁶. GOF mutations can convert p53 protein from a tumor suppressor to an oncogene, leading to expression of a mutant p53 protein at a high level, while LOF mutants leading to loss of p53 protein expression³⁷. TP53 mutations are classified according to their function as oncomorphic, loss of function and unclassified. Around 21% of all ovarian cancer patients harbor oncomorphic TP53 mutations, which had the highest p53 protein levels and contribute to chemoresistance and cancer progression, and the tumors with unclassified TP53 mutations express the mutated p53 protein at a fairly high level³³. The differences found between the studies in the frequency of p53 expression may be due to the differences in scoring of p53 expression and interpretation of results: some studies scored this as overexpression (OE), complete absence (CA), cytoplasmic (CY) or normal/wild type (WT)^18,20. Some authors consider complete absence of p53 expression as a mutant also because not all TP53 mutations alter the expression of the protein^15,17,25. Complete absence of p53 expression does not indicate TP53 mutation, as a lack of immunoexpression may be found in normal cells¹⁶. So, we believe that true overexpression (more than 75% of the cells stained positive) was the most important type of mutation to consider due to their importance in clinical practice, as they are chemoresistant mutations³³ and could also be interpreted easily in the immune-slide without any confusion.

Study limitations

The limitations of our study were related to the relatively small sample size. Many cases were found in the lab records but lacking tissue blocks, and some blocks contain less amount of tissue for TMA.

Conclusion

Our study showed that the overexpression of p53 tumor marker is associated with EOC, histological subtype and tumor grade, and found that high-grade serous tumors had a higher percentage of p53 expression in more than 50% of cases, while low grade serous was negative in 100% of the cases. We recommended the use of p53 immunohistochemical staining in the pathologic workup of ovarian carcinomas. Careful attention to laboratory protocols and practical works, including adequate controls, and training in interpretation is needed to make this a reliable test informing diagnosis and subsequent management of ovarian carcinoma.

Data availability