European Journal of Cardiovascular Medicine

UBC (Urinary Bladder Cancer) is the ninth most common cancer of all cancer cases in men, with a male:female ratio of 8.6:1, which is more than the worldwide ratio of about 3.5:1 [1,2].

 It is the fourth most common malignancy in the western world [3]. and worldwide urinary bladder cancer is the 7^th most common cancer [4]. It is the second most common malignancy of the genitourinary tract after prostatic cancer.

When first diagnosed, the majority of urothelial tumours are superficial, and up to 70% of them have a protracted clinical course during which the patient has recurrent recurrences after local excision without tumour advancement [5]. On the other hand, a smaller but statistically significant proportion of patients have tumours that exhibit an aggressive clinical course over a brief duration. As a result, urothelial cell tumours rank among the most expensive illnesses and need continuous monitoring [6,7].

Histopathology plays a crucial role in directing therapy and gauging the prognosis in patients with urothelial cell tumors. Unfortunately, it is subjective to individual variability and has poor specificity.

Many immunohistochemical markers, such as p63, cytokeratin 7, cytokeratin 20, uroplakin, placental S100 (S100P), and thrombomodulin, have been studied in the past in an attempt to aid in proper staging and grading, and to increase specificity at the metastatic site of this disease. However, none of them have proven to be useful [8-10]. Since these markers are also expressed in a significant number of other epithelial malignancies, such as breast, ovary, and lung, their specificity as well as their  role at the metastatic site remain limited.

GATA-3, sometimes referred to as GATA-binding protein 3 and transacting T-cell-specific transcription factor, has been linked to the advancement of many malignancies, including colorectal and breast cancer [11]. Its identification as an immunohistochemical marker linked with urothelial tissue is quite recent. Few studies with inconsistent outcomes have been conducted to examine expression in various histological variations, grades, and stages of urothelial carcinoma [12,13]

With the hypothesis that the immunohistochemical expression of GATA-3 in urothelial cell tumors and its correlation with histopathological parameters can have diagnostic as well as therapeutic implications, along with site specificity, this study was carried out on 40 specimens to identify the utility of GATA-3 expression in genitourinary carcinoma, especially urothelial carcinoma.

Data for the present study was obtained from bladder biopsies r during the period of March 2021 to August 2022.

Following standard processing, 5µm-thick paraffin slices were stained with hematoxylin and eosin for pathological examination. Additionally, using a commercially available GATA3 monoclonal antibody, 4µm slices were cut from a paraffin block of tumour tissue and placed on a glass slide covered with adhesive (silane) for immunohistochemistry to identify GATA3 expression.

The results were interpreted as follows:

The percentage of positive cells labelled by GATA3 is scored as follows:

Score 0 = No tumor cells stained

Score 1 = 1-10% of tumor cells

Score 2 = 11-50% of tumor cells

Score 3 = 51-80% of tumor cells

Score 4 = 81-100% of tumor cells.

The staining intensity of tumor cells labelled by GATA3 is scored as follows:

Staining Score 0 = No tumor cells stained

Staining Score 1 = Weak

Staining Score 2 = Moderate

Staining Score 3 = Strong

Informed consent was obtained from all individual participants included in the study.

Statistical Methods

Version 20.0 of the SPSS (Statistical Package for Social Sciences) software was used for data analysis. For continuous data, descriptive statistics were derived as mean and standard deviation (SD); for categorical variables, frequencies and percentages were computed. For categorical variables, the Chi-square test was used to assess the relationship between the variables. The means of the quantitative variables were compared between the two groups using the unpaired t test. Bar charts and pie charts were used for the visual representation of the analysed data. The level of significance was set at 0.05.

Most urothelial cancer patients, i.e., 37.5%, belonged to the age group of 61–70 years, followed by 30% of patients who were older than 70 years. The mean (SD) age of study participants was 64.98 years (10.06 years).

As for gender distribution, most urothelial cancer patients were of the male gender. Only 8 patients, i.e., 20% of the patients, were female. (Table 1)

Table I: Age Distribution of Study Participants

The predominant symptom, site of the lesion, and diagnosis of the lesion were identified for each urothelial cancer patient, and the results are as shown in Table 2. The most common predominant symptom was hematuria (seen in 31 patients, i.e., 77.5%), followed by dysuria (seen in 6 patients, i.e., 15%), and urgency (seen in 3 patients, i.e., 7.5%).

Table II: Clinical Presentation of Urothelial Cell Tumors

Site of the Lesion

The most common site for urothelial cancers as per the present study was the lateral wall, which grew cancers in 26 patients (65%), followed by the posterior wall (7 patients, i.e., 17.5%). The remaining 7 patients (17.5%) had lesions in both the lateral wall and the posterior wall.

Diagnosis

Four diagnoses which were considered were: PUNLMP (Papillary urothelial neoplasm of low malignant potential), Low grade papillary UC, High grade papillary UC and Invasive UC, in the increasing order of severity. Proportions of patients having these diagnoses were 17.5%, 35%, 17.5% and 30% respectively.

18% of patients had lesions on both the lateral and posterior walls, with the lateral wall being the most common site of lesions in 65% of patients.

High-grade cancers were reported in 18% of patients, and invasive cancers were reported in 30% of patients.

As shown in Table 3, the most common pathological feature was nuclear pleomorphism (noted in 82.5%, or 33 of 40 patients) followed by mitosis (noted in 40%, or 16 of 40 patients), necrosis (noted in 27.5%, or 11 of 40 patients) and tumor infiltrating lymphocytes (noted in 22.5%, or 9 of 40 patients).

Table III: Pathological Findings of Urothelial Cell Tumors

The distribution of study participants as per immunohistochemistry findings are as shown in most patients i.e., 15 out of 40 (37.5%) were of score 3, and this was followed by 14 patients (35%) belonging to a group of score 2.

Almost half the urothelial cancer patients (45%) included in the study had a score of 2 in the GATA3 intensity scoring system. Another quarter of patients (25%) had a score of 3 (highest score) for the GATA3 intensity score.(Table 4)

Table IV: Distribution of Study Participants as per Immunohistochemistry Findings

Combining the above two parameters, a grading system was devised for GATA3 expression. With respect to this, the table shows that almost a third of the patients (35%) were of grade 3 GATA3 positivity, and another third (30%) belonged to grade 2. The highest grade, i.e., grade 4, was seen in 15% of urothelial cancer patients. Grade 1 positivity status (that is, negative for GATA3 expression) was observed in 20% of patients.

The mean ‘total IHC score’ was 8.14, 5.86, 3.71 and 1.75 among PUNLMP (Figure 1a and Figure 1b), low-grade papillary UC (Figure 2a and 2b), high-grade papillary UC (Figure 3a and 3b) and invasive UC patients (Figure 4a and 4b), respectively. A reduction in the mean IHC score with higher grades of UC was observed, implying that the GATA3 expression was lost. (Table 5)

Table V: Average ‘Total IHC Score’ for Different Histopathological Diagnosis

All the tumors that were negative for GATA3 expression were of the invasive type (Figure 4a and 4b). The proportion of tumors with an invasive nature gradually decreased as the IHC grade increased.

All the tumors that were negative for GATA3 expression had strongly positive nuclear pleomorphism. The proportion of tumors with strongly positive nuclear pleomorphism decreased as the IHC grade increased. The proportion of tumors with mitosis also gradually decreased as the IHC grade increased.

All the tumors with no GATA3 expression had necrosis. All tumors with IHC grades 3 and 4 had no necrosis reported.

All IHC grade 3 and 4 tumors were not infiltrated by lymphocytes while 17% of grade 2 IHC tumors and 88% of grade 1 IHC tumors were infiltrated by lymphocytes.

As shown in Table 6, all these differences were statistically significant (p-value < 0.05).

Table VI: Correlation of GATA 3 Expression with Histological Findings of Urothelial Cell Tumors

Figure 4A and 4B: Microscopic image of GATA3 expression showing  total IHC score of 0 [GATA3 score is 0) x (intensity score is 0)] in a case of infiltrating urothelial carcinoma.(4X and 40X).

Figure 1A and 1B: Microscopic image of GATA3 expression showing strong positivity with total IHC score of 9 [GATA3 score is 3) x (intensity score is 3)] in a case of papillary urothelial neoplasm of low malignant potential (PUNLMP) (4X and 40X).

Figure 2A and 2B: Microscopic image of GATA3 expression showing moderate positivity with total IHC score of 6 [GATA3 score is 3) x (intensity score is 2)] in a case of low grade urothelial carcinoma. (4X and 40X )

Figure 3A and 3B: Microscopic image of GATA3 expression showing Weak positivity with total IHC score of 4 [GATA3 score is 2) x (intensity score is 2)] in a case of High grade urothelial carcinoma.(4X and 40X).

Figure 4A: Microscopic image of GATA3 expression showing Total IHC Score of 0 [(GATA3 score is 0) x (Intensity Score is 0)] in a case of Infiltrating urothelial carcinoma,(4X)

Risk stratification of urothelial cancers based on histological classification is essential for the management of these patients.

Immunohistochemistry has been shown to be a good prognostic marker in addition to histopathological grading. In addition to common biological markers like p53 and GATA3, a nuclear transcription factor that is usually positive in more than two-thirds of urothelial cancers may also be used as a prognostic marker. Literature suggests that invasive carcinomas have a tendency to lose GATA3 expression [14].

The present study showed that around two-thirds (67.5%) of the patients were aged more than 60 years, with a mean age of 64.98 years. This is closely similar to the study conducted by El Kholy M et al [15] during 2018-2020. Also, another study conducted by Naik M et al [16] in 2021 reported a maximum number of patients (38.88%) from the age group of 61-70 years. The mean age was relatively higher (68 years) in the study conducted by Liang Y et al [17].

In the present study, urothelial cancer was more common among males (80%). Most other studies, such as those conducted by El Kholy M et al. in 2020 , Naik M et al. in 2020, and Liang Y et al [18].  in 2014, have reported very similar gender distribution.

The present study reported that 80% of urothelial cancer patients have positive GATA3 expression. This finding was similar to the study conducted by El Kholy M et al. in 2020 and by Naik M et al. in 2020.  A slightly higher Grade 1/negative expression was reported in the study conducted by Liang Y et al.  in 2014, which again could be because of the timing of the study.

A contrasting finding was noted in the study conducted by Inoue MDS et al [19] in 2017, where almost half of cancer patients were negative for GATA3 expression.

The present study showed that GATA3 expression differed significantly among different histological or pathological categories of urothelial cancers and that GATA3 expression was significantly associated with diagnosis, mitosis, necrosis and infiltrations. We observed that as the cancer worsens, GATA3 expression is lost. Similar findings were seen in the studies conducted by El Kholy M et al [20]. Naik M et al., and Liang Y et al.

Expression of GATA3 has a strong correlation with the pathological grading of urothelial cancers and hence may be used in diagnosis, assessment of severity and also in prediction of prognosis.

Acknowledgement:

The authors acknowledge Histo-path Technicians of pathology department for their contributions in our research study.

Conflict of Interest:

The authors declare no Conflict Of interest.

Funding source:

There is no funding source to declare.

Authors contributions:

RT, SK contributed in manuscript writing. AK Contributed in statistics work. All authors read ,edited and approved the manuscript.

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