The Role p53 Protein Expression in Urothelial Neoplasm

The urinary bladder had a line with transitional epithelium. Urothelial neoplasms are the majority among the bladder neoplasms. Urothelial neoplasms were more common in males than in females. Urothelial neoplasms usually occur in the elderly age group. The majority of the tumor size was more significant than 3 cm, and papillary type is majorly found among the tumors. The present study aimed to identify the grade and staging of the p53 in urothelial neoplasms. The bladder carcinoma (54) cases from Department of Pathology, Sree Balaji Medical College and hospital (during September 2015 to September 2017) were analysed. The median age for bladder carcinoma in the present study was 66 years. The clinical parameter studied was the size of the lesion based on the cytoscopic or radiological findings (3133). Based on tumor size, lesions were classified into two groups, less than equal to 3cm and greater than 3cm, which was found to be 42% and 58%, respectively This study concludes that p53 is useful in differentiating benign and malignant neoplasms in morphologically difficult cases. Immunohistochemistry for p53 is useful adjunct to histomorphology. Original Research Article Sahoo et al.; JPRI, 33(21B): 85-99, 2021; Article no.JPRI.66769 86


INTRODUCTION
Bladder cancer is the seventh common cancer worldwide and the sixth most public in developed countries and comprises 3.2% of all cancers [1,2]. It is more common in westernized countries like North America, United Kingdom and Australia. The incidence of bladder cancer in India is 3 .2% in males and 0.7% in females among all malignancies. Ninety-eight percent of urinary bladder cancers are of epithelial origin, among which 90% are urothelial carcinomas [3][4]. Bladder cancer forms when the DNA in cells in the bladder change, deactivating the functions that control cell growth. The immune system criticizes these mutated cells. But some mutated cells may escape the immune system and grow out of control, forming a tumor in the bladder. Bladder cancer is mainly caused by smoking and industrial exposure [5][6][7]. By epidemiological studies, tobacco smoking has been occupied in 50% in men and 30% in women of bladder cancer cases [4][5][6]. Ninety eight percent of urinary bladder cancers are of epithelial origin, among which 90% are urothelial carcinomas. It usually presents as a non-invasive papillary tumor swelling from the surface of the mucosa. About one -third of bladder cancers presents as non-papillary, solid tumors which arise from in situ dysplasia and carcinoma in situ. These tumors attack the bladder wall and are prone for metastasis [8][9][10][11]. The 2016 WHO classification of tumors of the urothelial tract provides a review of morphology of urothelial neoplasms, emphasizing on their divergent differentiation, multiple morphologic variants and a diverse genomic landscape.
Rating is important in noninvasive disease, specifically papillary neoplasms. Noninvasive tumors divided into two categories, papillary and flat. As in 2004, the 2016 WHO classification continues to recommend the application of the grading classification first put forth by ISUP in 1997. According to the 2016 WHO classification, papillary urothelial neoplasms have been categorized as: low grade papillary urothelial carcinoma (LGPUC), high grade papillary urothelial carcinoma (HGPUC), papillary urothelial neoplasm of low malignant potential urothelial papilloma (UP), inverted urothelial papilloma (IUP), urothelial proliferation of uncertain malignant potential urothelial dysplasia (UD) [10,[11][12][13][14].
Urothelial carcinoma is well known for its different variation. The most common are glandular and squamous differentiation. These replacements are major to find as it has diagnostic, prognostic and therapeutic considerations [11,[15][16][17][18]. There are several prognostic factors in bladder carcinoma; histological stage and grade of the tumor have been reflected as vital factors. In some cases, tumors with greater rating and stage behave in an indolent way, whereas tumors with lower rating and stage show a high incidence of recurrence [19][20][21][22][23]. Hence additional prognostic information is required to guide clinicians in the management of these patients. Urothelial playing is the most directing projecting pointer in urothelial carcinoma and is a major defining parameter in the management of this disease. (TNM Scheme for bladder carcinoma), [24][25][26]. The important factors while assigning a T -stage include the size of the lesion and extent of invasive carcinoma. Urothelial neoplasms are generally classified into four categories according to the WHO/ ISUP 2004 consensus classification system [25,[26][27][28][29][30].
 Hyperplasia  Flat lesion with atypia  Papillary neoplasms  Invasive neoplasms Classifying urothelial neoplasm based on histomorphology alone has certain diagnostic dilemmas because of overlap in morphological features between: -Carcinoma in situ and flat urothelial hyperplasia, -Non invasive low grade papillary urothelial carcinoma and papillary neoplasms of low malignant potential, -Non invasive low grade/ invasive high grade urothelial carcinoma and non invasive high grade urothelial carcinoma. Immunohistochemical markers along with histomorphology can be used to differentiate various urothelial neoplasms. p53 is a tumor suppressor gene that maps to the human chromosome 17p13. The mutation of which is associated with tumor progression [13]. Popov et al. studied p53 expression in 114 cases of urothelial carcinomas and found a significant correlation between histological grade and stage (p<0. 001) [13]. Apoptosis can be induced by the binding of Caspase 9 to cytochrome c and Apaf1. P53 may activate the expression of Apaf1 and Bax. Bax can then stimulate the release of cytochrome c from mitochondria. The identification of the factor for facilitating the prognosis is a challengeable one to analyze the risks of the patients. The present study aimed to identify the grade and staging of the p53 in urothelial neoplasms.

MATERIALS AND METHODS
54 cases were bladder cases. Among them, 2 specimens were radical cystectomies, 51 specimens were TURBT and 1 specimens were other type biopsies. The total number of nonneoplastic and malignant cases of the urinary bladder was 1 and 53 respectively.

Source of Data
The bladder carcinoma cases reported in the Department of Pathology, Sree Balaji Medical College and hospital from September 2015 to September 2017 and had been sent by the Department of Urology for analysis.

Inclusion Criteria
All the cases of urothelial neoplasms reported in bladder specimens irrespective of patient's age and sex and the procedure done were included for the study.

Exclusion Criteria
• Carcinomas other than urothelial carcinomas.
• Cases with inadequate material. • Cases without deep muscle biopsy.

Method of Data Collection
Detailed history of the cases regarding age, sex, and history, type of procedure, site, size, stage, previous surgery details and urinary cytology were obtained for all the urothelial carcinomas reported during the study period. 4 μ thick sections of paraffin tissue specimen are stained with Hematoxylin and Eosin. Age, gender, tumour size and tumour location (base, lateral, posterior walls and trigone) were the clinical and pathological parameters evaluated. For the malignant lesion staging was done according to TNM staging for bladder carcinomas.

Statistical Analysis
The various histological parameters were correlated with immunohistochemical profile. The immunohistological profiles were correlated with grade and stage. Association between alteration of marker expression and grade and stage were examined using Pearson Chi-square test. Results were considered significant if the p-value <0.05.

RESULTS
Fifty four specimen of newly diagnosed cases of bladder carcinomas were received from the department of urology during the study period. The urothelial neoplasms constituted 92. 59 % of newly diagnosed bladder carcinomas, whereas the non urothelial neoplasms were 7.40%.
The distribution of bladder carcinomas is shown in Among the non-urothelial neoplasms squamous cell carcinoma were 75% and adenocarcinoma were 25%.

Sex Distribution
The incidence of urothelial neoplasm in males accounted for 83. 01% whereas in females it was about 16. 98 % and the ratio of male: female ratio 3.1:1 (Fig. 1).

Age Distribution in Urothelial Neoplasm
The median age for urothelial neoplasm in our study group was 66 years. The age of patients ranged from 24 to 82 years. Majority of the cases were between the 5 th to 8th decade (Fig. 2).

Size of the Tumor
The size of the lesions was noted from the cystoscopic/ radiological findings, the tumor size was the largest of that noted in the three dimensions, and the largest of that is taken as tumor size. The largest tumor size was considered in case of multiple lesions. The tumors were grouped into two based on the size (greater than 3 and less than or equal to 3). In our study majority of the cases, 58 % were greater than 3cm (Fig. 3).

Tumor Location
Based on the cystoscopic and findings majority (48%) of the located in the lateral walls posterolateral, right lateral and left was in the anterior and 18% in posterior although less in frequency few were bladder neck and trigone (Fig. 4).

Papillary Neoplasm
All the urothelial neoplasms were papillary study which includes only one benign rest were malignant neoplasms.

Analysis
features of the and correlated this study. studied were prominent nucleoli and 5-10, >10). of mitosis were between the benign grade neoplasms varying degrees of nuclear pleomorphisms and prominent In classical cases, differentiating and high grade carcinoma is not certain limitations. Preserved umbrella is considered to be a feature carcinomas, but in this study, around grade carcinomas had preserved layer. A number of mitosis were differentiating low grade and carcinomas. Low grade papillary carcinomas mitosis less than 5/10 hpf, whereas carcinomas mitosis were 0 -30 grade carcinomas had 1 -2 prominent (Tables 2 and 3).

Immunohistochemistry
In the present study, immunohistochemistry performed in 27 cases diagnosed neoplasm and p53 marker expression studied.

p53 in Urothelial Neoplasms
In this study, the benign case i. e., negative (<10%) for p53. Twenty percent of PUNLMP was positive (>10%) and the percentage of positivity was which is considered as grade 1. low grade carcinoma and (11/ 12) grade carcinomas showed expression. p 53 expression was grades (grade 0, 1, 2 ,3,4).  A-Absent, p-value is significant between low grade and high grade carcinomas for number of mitosis.

Expression of p 53 in Low
High -Grade Carcinomas 91.6 % of high grade carcinomas showed p53 expression. Percentage of cells p53 was not different between the The number of cases was distributed between the grades of p53. p-value significant (p=0.2). The intensity expression and level of p53 positivity differentiating the two grades. Diffuse intense positivity was noted in carcinomas. The difference in p53 was not significant between low grade grade carcinomas (Table 4).

Staging in Urothelial Neoplasm
Staging was performed in all cases specimens which included low grade   Kalantari et al. [28], studied p53 expression with tumor grades and stage of urothelial carcinomas. The author found that all PUNLMP cases were negative for p53 and the difference between in p53 expression was statistically significant between different types (PUNLMP, low grade, high grade carcinomas). In this present study,3 out of 4 cases showed negativity for p53, 1 case showed grade 1 positivity for p 53 and none of the cases showed >25% positivity for p 53 in the PUNLMP category.

CONCLUSION
Morphology plays a major role in the differential diagnosis of urothelial neoplasms. Negative p53 staining in PUNLMPs and high p53 index in high grade papillary urothelial carcinomas and invasive carcinomas supported the involvement of p53 mutation in the development of urothelial neoplasms but plays a crucial role in progression of the malignancy.

CONSENT
As per international standard or university standard, patients' written consent has been collected and preserved by the author(s).

ETHICAL APPROVAL
The study was approved by the Institutional Ethics Committee.