P53 Immunostaining Research Articles

Altered p53/p16 expression is linked to urothelial carcinoma progression but largely unrelated to prognosis in muscle-invasive tumors

Background Most inactivating p53 mutations result in a nuclear p53 accumulation – detectable by immunohistochemistry (IHC). p53 alterations leading to a complete lack of p53 protein and absence of immunostaining do also occur – not easily detectable by IHC. p16 is upregulated in p53 inactivated cells. We hypothesized that a positive p16 IHC may help to distinguish p53 inactivation in IHC negative cases. Material and methods We investigated p53 and p16 immunostaining on 2710 urothelial bladder carcinomas in a tissue microarray format to understand their impact in relation to clinicopathological parameters of disease progression and patient outcome. Results p16 immunostaining was absent in normal urothelium but occurred in 63.5% (30.4% strong) of cancers. p16 strongly positive cases increased from pTaG2 low-grade (9.6%) to pTaG3 high-grade tumors (46.5%, p < .0001) but decreased from pTaG3 to pT4 (33.3%; p = .0030). Among pT2-4 carcinomas, p16 positivity was linked to high-grade (p = .0005) but unrelated to overall survival. p53 staining was negative in 8.4%, very weak in 15.4%, weak in 55.3%, strong in 4.7%, and very strong in 16.2% cancers. p53 negative (potentially p53 null phenotype), strong, and very strong p53 positivity increased from pTaG2 low-grade to pTaG3 high-grade tumors (p < .0001) and from pTaG3 to pT2-4 cancers (p = .0007). p53 staining was largely unrelated to histopathological parameters or patient prognosis among pT2-4 carcinomas, except of p53 strong/very strong immunostaining. p16 expression predominated in tumors with very strong, strong, and negative p53 staining and the combination of p53 negative/p16 strongly positive cancers was linked to features of tumor aggressiveness. Conclusion Aberrant p53 and p16 immunostaining increases during grade and stage progression although p53 negative and p16 positive immunostaining lack prognostic significance in pT2–4 carcinomas. Potential diagnostic features are that high level p16 expression is limited to neoplastic urothelium and p53 null phenotype to aggressive cancers (grade 3 and invasive).

Influence of p16 Protein Immunostaining on Histopathological Features of Pleomorphic Adenoma and Carcinoma ex- Pleomorphic Adenoma.

The objective of this study was to evaluate the role of p16 in histologic characteristics and transition of Pleomorphic Adenoma (PA) to Carcinoma ex-PA (CxPA). So, 60 PA and 4 CxPA were histologic reviewed based on microscopic characteristics proposed by Hellquist, Triantafyllou and Dulguerov (PA) and Morais, Antony and Toluie (CxPA). Immunostaining for p16 was associated in different parenchyma and stroma of both tumors and Fisher's/chi-square tests and Mann-Whitney test were performed (SPSS v20.0, p<0.05). In PA the periductal cells were predominantly p16- and that ductal and myoepithelial cells showed a significant increase in p16+ cells (p<0.001). In CxPA, none of the cases showed p16+ in periductal cells, most parotid cases showed p16+ in ductal cells, and one case of parotid and the submandibular case showed mild immunostaining for myoepithelial cells. There was a small reduction in p16+ in CxPA compared to PA (p=0.537), but in both tumors there was less p16+ cells in solid stroma than other (p<0.001). The p16+ cases of PA had a higher capsular thickness (p=0.047). So, the loss of p16 immunostaining does not seem to be associated with the transition from PA to CxPA, but in both tumors the loss of p16+ cells are related to microscopic aggressiveness.

Whole-exome sequencing reveals candidate high-risk susceptibility genes for endometriosis

BackgroundEndometriosis is a common, chronic disease among fertile-aged women. Disease course may be highly invasive, requiring extensive surgery. The etiology of endometriosis remains elusive, though a high level of heritability is well established. Several low-penetrance predisposing loci have been identified, but high-risk susceptibility remains undetermined. Endometriosis is known to increase the risk of epithelial ovarian cancers, especially of endometrioid and clear cell types. Here, we have analyzed a Finnish family where four women have been diagnosed with surgically verified, severely symptomatic endometriosis and two of the patients also with high-grade serous carcinoma.ResultsWhole-exome sequencing revealed three rare candidate predisposing variants segregating with endometriosis. The variants were c.1238C>T, p.(Pro413Leu) in FGFR4, c.5065C>T, p.(Arg1689Trp) in NALCN, and c.2086G>A, p.(Val696Met) in NAV2. The only variant predicted deleterious by in silico tools was the one in FGFR4. Further screening of the variants in 92 Finnish endometriosis and in 19 endometriosis–ovarian cancer patients did not reveal additional carriers. Histopathology, positive p53 immunostaining, and genetic analysis supported the high-grade serous subtype of the two tumors in the family.ConclusionsHere, we provide FGFR4, NALCN, and NAV2 as novel high-risk candidate genes for familial endometriosis. Our results also support the association of endometriosis with high-grade serous carcinoma. Further studies are required to validate the findings and to reveal the exact pathogenesis mechanisms of endometriosis. Elucidating the genetic background of endometriosis defines the etiology of the disease and provides opportunities for expedited diagnostics and personalized treatments.

HPV-Associated Squamous Cell Carcinoma of the Eyelid: Diagnostic Utility of p16 Immunohistochemistry and mRNA In Situ Hybridization.

High-risk (HR) Human papillomavirus (HPV) has been implicated in pathogenesis of squamous cell carcinomas (SCC) at several sites with mucocutaneous junctions, including the head and neck. SCC is the second most common eyelid malignancy. However, its association with transcriptionally active HR-HPV has not been adequately studied. Two index cases of eyelid HPV-associated SCC are described in detail. A retrospective cohort of eyelid SCC was examined for p16 immunoexpression. Cases demonstrating p16 positivity or equivocal staining were subjected to high-risk HPV mRNA in situ hybridization (ISH). Quantitative real-time PCR (qPCR) was performed in mRNA ISH-positive cases for HPV genotyping. The two index patients were older adult females, with upper eyelid tumours. On histology, both tumours were non-keratinizing SCC with trabecular and nested architecture reminiscent of oropharyngeal HPV-associated non-keratinizing SCC, prompting p16 immunohistochemistry, which was positive. HR-HPV mRNA ISH was positive, and qPCR detected HPV16 in both cases. Three of 20 (15%) archival cases showed p16 immunopositivity and two (10%) showed equivocal staining. However, mRNA ISH was negative. All cases showing p16 immunostaining and lacking HR-HPV were keratinizing SCCs. Thus, 9% of all eyelid SCC examined demonstrated HR-HPV. The prevalence of HR-HPV in eyelid SCC is low in Indian patients. HPV-associated SCC may mimic commoner eyelid carcinomas as it lacks overt keratinization. In basaloid-appearing eyelid carcinomas, p16 immunopositivity should be followed by reflex HR-HPV mRNA ISH, as p16 immunohistochemistry alone has low specificity. The prognostic role, if any, of HPV association needs further evaluation.

Correlation of progesterone receptors and P63 to the histological grade of meningiomas: Review and significance in an African population.

Meningiomas, a common neoplasm of the central nervous system, is a widely studied meningeal tumor. According to the World Health Organization (WHO) 2021 classification of meningiomas, there are 15 subtypes that have been grouped into grades 1, 2, and 3. The WHO grade 1 meningiomas are generally grouped as benign while the WHO grades 2 and 3 tumors are grouped as malignant. Progesterone receptors and P63 are common immunohistochemical markers that have proven useful in the diagnosis, grading, and prognostication of many neoplasms such as breast carcinoma, prostate carcinoma, and gastrointestinal tumors in histopathology practice. The application of these immunohistochemical markers to the grading of meningiomas has been reported and their usefulness documented in reports from Africa, Europe, North America, South America, and Asia. This study, therefore, seeks to determine if these findings are applicable to the meningiomas seen in an African population. A 10-year review of results and histologically diagnosed cases of meningiomas received in the Department of Morbid Anatomy, University of Nigeria, Enugu. Immunostaining for progesterone receptors (PgRs) and P63 were done and results compared with histologic grades. The three WHO grades of meningioma were assessed in this study. M: F ratio was 1:1.4 and peak age was 41-50 years age range (SD ± 16.54). The majority of the cases were WHO grade 1 (86.1%) while the WHO grades 2 and 3 tumors were 8% and 5.9%, respectively. The fibrous variant was the most common subtype (27.1%). There was no correlation between progesterone receptor and P63 immunopositivity to the WHO grades of meningioma (P = 0.112 and P = 0.138, respectively). Our study showed that progesterone receptors and P63 immunopositivity did not correlate with the WHO grades of meningiomas. This may be due to the predominant variant of meningioma seen in this study. These findings indicate that PgR antagonist may not be an effective alternative for treatment in patients with inoperable meningiomas. Furthermore, P63 immunopositivity may not be a sufficient grading tool for managing meningiomas in our population.

Effects of bisphenol A on murine salivary glands and human tumor cell lines

Bisphenol A (BPA) is an endocrine-disrupting chemical with a potential role in endocrine cancers. However, the effects of BPA on the salivary glands have been barely explored. We investigated the impact of in vivo sub-chronic exposure to BPA and its in vitro effects on human salivary gland mucoepidermoid carcinoma cell lines. Male and female mice were exposed to BPA (30 mg/kg/day). Sublingual and submandibular salivary glands from an estrogen-deficiency model were also analyzed. BPA concentration in salivary glands was evaluated by gas chromatography coupled to ion trap mass spectrometry. Immunohistochemical analysis using anti-p63 and anti-α-SMA antibodies was performed on mouse salivary gland tissues. Gene expression of estrogen receptors alpha and beta, P63 and α-SMA was quantified in mouse salivary gland and/or mucoepidermoid (UM-HMC-1 and UM-HMC-3A) cell lines. Cell viability, p63 and Ki-67 immunostaining were evaluated in vitro. BPA disrupted the tissue architecture of the submandibular and sublingual glands, particularly in female mice, and increased the expression of estrogen receptors and p63, effects that were accompanied by significant BPA accumulation in these tissues. Conversely, ovariectomy slightly impacted BPA-induced morphological changes. In vitro, BPA did not affect the proliferation of neoplastic cells, but augmented the expression of p63 and estrogen receptors. The present data highlight a potential harmful effect of BPA on salivary gland tissues, particularly in female mice, and salivary gland tumor cells. Our findings suggest that estrogen-dependent pathways may orchestrate the effects of BPA in salivary glands.

Clinico-pathological Factors in Malignant Transformation of RRP.

Various clinico-pathological factors play role in the papilloma proliferation and pathogenesis of Recurrent respiratory papillomatosis (RRP). However, it is not known if they are directly responsible for malignant transformation of these papillomas or not. We did this study to elucidate any such association. The most recent debrided tissue of RRP in 20 patients was evaluated for p16 expression, VEGF estimation (tissue expression and serum levels), and tissue HPV DNA concentration. The final histopathology results were then correlated with these pathological factors and with clinical factors like duration of illness, age of onset of symptoms, extent of disease, etc. Squamous papilloma was seen in 60%, dysplasia in 25%, and squamous cell carcinoma (SCC) in 15% of the patients. Positive immunostaining for p16 (staining in ≥70% of tumor cells) was seen only in one case, which was SCC. There was no statistically significant difference between p16 expression, tissue VEGF expression, serum VEGF levels, and tissue HPV DNA in any of the histological groups. The mean age of disease onset was significantly higher in patients with SCC (p = 0.03). A significantly higher number of patients with dysplasia had tracheobronchial involvement (p = 0.022). We concluded that no single pathological factor is solely responsible for development of malignancy in RRP, whereas clinical factors like tracheobronchial involvement and age of onset may contribute to development of dysplasia or carcinoma.

Apoptosis-related Prognostic Factors in Advanced Colorectal Cancer Determined Using Tissue Microarrays.

Cancer cells evade apoptosis in colorectal cancer (CRC); however, overlap between apoptosis and poor prognosis marker proteins in the invasive front of tumors has not been reported. Here, we aimed to clarify the relationship between apoptosis, apoptosis-related protein expression, and prognosis in the central and invasive front regions of CRC using tissue microarrays. Data of 207 patients with pathological stage 3 CRC, who underwent radical surgery between October 2010 and November 2014, were retrospectively reviewed. We assessed apoptosis using M30 CytoDEATH, CD163, and p53 immunostaining in tumor sections in the center and invasive front using tissue microarrays and correlated the results with the survival outcomes. M30 CytoDEATH staining was negative; 134 cases (64.7%) were apoptosis-negative in the center and 103 (49.8%) were apoptosis-negative at the invasive front. CD163 positivity was observed in 16 cases (7.8%) in the center and in 36 cases (17.6%) at the invasive front; p53 positivity was observed in 33 (15.9%) and 64 (30.9%) cases in the center and invasive front, respectively. CD163 and p53 expression was not associated with survival outcomes; however, the apoptosis-negative group at the invasive front had significantly poorer survival outcomes (overall survival: p=0.044, relapse-free survival: p=0.001). We identified cases with a poor prognosis by combining apoptosis and CD163 expression. A lower apoptosis percentage at the invasive front is associated with a poorer prognosis. CRC cases with a poor prognosis can be identified by evaluating apoptosis and CD163 expression in the invasive front.

Subclonal p53 immunostaining in the diagnosis of endometrial carcinoma molecular subtype.

The significance of subclonal expression of p53 (abrupt transition from wild-type to mutant-pattern staining) is not well understood, and the arbitrary diagnostic cut-off of 10% between NSMP and p53abn molecular subtypes of endometrial carcinoma (EC) has not been critically assessed. Our aim was to characterise subclonal p53 and discrepant p53 expression/TP53 sequencing results in EC and assess their clinical significance. Subclonal p53 immuostaining on whole sections from 957 ECs was recorded. Agreement between TP53 mutational assessment and p53 immunostaining was evaluated. Subclonal p53 IHC staining was seen in 4.0% (38 of 957) of cases, with 23 of 957 (2.4%) showing mutant-pattern p53 staining in ≥10% of tumour cells. It was most commonly seen in POLEmut (nine of 65, 14%) and MMRd (13 of 274, 4.7%) EC ('multiple classifier' ECs), where subclonal p53 staining does not impact the molecular subtype diagnosis. Excluding POLEmut and MMRd EC, 11 of 957 (1.1%) showed ≥10% subclonal p53 from which four patients died of disease, while there were no deaths due to disease in the five patients with <10% mutant-pattern p53 staining. Agreement between p53 immunostaining and TP53 sequencing was 92.6%; most of the discrepant results were in the ultramutated POLEmut or hypermutated MMRd ECs. In NSMP and p53abn EC the agreement between IHC and sequencing was 95.8%. Subclonal p53 staining ≥10% is present in only 1.1% of EC after excluding 'multiple classifier' ECs. The cut-off of ≥10% subclonal p53 staining identified patients at increased risk of dying from EC, supporting its use to diagnose p53abn molecular subtype.

Correlation of p53 Expression with Grade and Stage of Ovarian Surface Epithelial Cancer

Introduction: Advanced ovarian cancer is characterized by a poor prognosis and the development of resistance to chemotherapy. In the absence of definite etiologic factors and effective tools for screening, the only possible means of improving survival would be to develop effective targeted therapy. Expression of p53 in ovarian cancer may serve as a promising therapeutic strategy to treat, prevent visceral metastasis and relapse. Objective: The study was carried out to find out the correlation of p53 expression with the grade and stage of ovarian cancers. Methods: The study had 45 cases. These cases were previously reported as ovarian surface epithelial cancers. Grading was done in all cases. Pathological staging of tumour (pT) was done in 30 cases and pathological staging of lymph node metastasis (pN) was done in 10 cases. Distant metastasis was present in only 1 case. Result: p53 immunostaining was found positively correlated &amp; associated with the grade &amp; stage of ovarian surface epithelial cancers. Conclusion: p53 expression may be important for prognostic outcomes or useful targets of therapeutic intervention. Mediscope 2023;10(2): 42-49

Malignant Proliferating Pilar Tumor: Clinicopathologic, Immunohistochemical, and Molecular Study of 17 Cases.

Proliferating pilar tumors are rare neoplasms that differentiate toward the outer sheath near the isthmus and can rarely undergo malignant transformation. We performed histopathologic evaluation on 26 benign proliferating pilar tumor (BPPT) and 17 malignant proliferating pilar tumor (MPPT). Ki-67 and p53 immunostains were performed on 13 BPPT and 10 MPPT. Six MPPT cases were successfully analyzed by a next-generation sequencing platform which surveyed exonic DNA sequences of 447 cancer genes and 191 regions across 60 genes for rearrangement detection. Patient demographics and clinical characteristics were similar between the BPPT and MPPT groups. Follow-up data of 16 of 17 MPPT (median, 25mo) showed metastasis in 1 MPPT. The histologic features associated with MPPT include size >2.5cm, adjacent desmoplastic stroma, small nests or cords of atypical epithelium in surrounding stroma, irregular infiltration or borders, abnormal keratinization, large hyperchromatic nuclei, prominent nucleoli, severe cytologic atypia, nuclear pleomorphism, necrosis, and increased mitotic figures. MPPT harbors copy number gains of 15q and losses of 6p and 6q, findings previously reported in BPPT. However, MPPT harbors frequent TP53 mutations as molecular markers of progression. Different from cutaneous squamous cell carcinoma, MPPT more frequently demonstrates low tumor mutational burden and typically lacks a UV signature, suggestive of a different etiologic pathway than squamous cell carcinoma. In summary, with a median follow-up of 25 months, this study shows that MPPT is a biologically indolent carcinoma with rare metastasis. Molecular analyses suggest a non-UV-related pathogenesis with frequent TP53 aberration.

Immune Status of Cervical Lymph Nodes in Head and Neck Cancer-A Surgical Oncology Perspective.

Neck dissection for cervical lymph node metastasis is an established procedure for head and neck cancer (HNC). However, with the advent of immunotherapy, head and neck surgical oncologists need to rethink removing all lymph nodes, including those with immune function. We investigated the anti-cancer immune response of the cervical lymph nodes in four patients with human papillomavirus type 16 (HPV16)-positive head and neck squamous cell carcinoma. Using lymphocytes extracted from local, metastatic, and non-metastatic lymph nodes and peripheral blood from these patients, we performed an intracellular flow cytometric cytokine assay using anti-IFNγ and anti-TNF-α monoclonal antibodies to detect HPV16 E6- and E7-specific T cells. HPV status and p16 immunostaining were determined by in situ detection using the HPV RNAscope method and immunohistochemistry. In one case, E6-specific and E7-specific CD8+ T cells were detected in proximal metastatic nodes and distal non-metastatic nodes. This finding suggests that non-metastatic nodes should be preserved for their immune function during neck dissection and that the immune function of non-metastatic lymph nodes is important when administering immunotherapy. In this context, head and neck surgical oncologists treating HNC should consider the place of immunotherapy and neck dissection in the treatment of HNC.

Clinico-Pathological Features and Immunohistochemical Comparison of p16, p53, and Ki-67 Expression in Muscle-Invasive and Non-Muscle-Invasive Conventional Urothelial Bladder Carcinoma.

The identification of bladder detrusor muscle invasion in urothelial cancer is essential for prognosis and management. We studied the clinical, histological, and immunohistochemical expression of p16, p53, and Ki-67 in urothelial detrusor muscle-invasive bladder cancer (MIBC) and urothelial non-detrusor muscle-invasive bladder cancer (NMIBC) in Egyptian patients. Sixty-two bladder urothelial cancer cases obtained through TURBT were included and divided into two groups: (MIBC, stage T2) and NMIBC (T1). Tissue blocks were recut and re-examined microscopically; then, the immunostaining of p16, p53, and Ki-67 was performed to compare both groups and evaluate the 13% cut-off for Ki-67, 20% for p53, and p16 intensity in various conditions aided by telepathology technology. Hematuria was the main clinical first presentation, with no significant difference between either group. The mean age was 61.6 years, with male predominance (52 males and 10 females). The absence of papillary histological pattern was associated with a higher stage, including detrusor muscle invasion (p = 0.000). The overall average percent of p53 immunostaining was 12.9%, revealing no significant difference between MIBC and NMIBC when a cut-off of 20% was implicated. The Ki-67 expression was correlated with higher grade and muscle invasion; however, no association was found with the other two markers' expression. The negative immunostaining of p16 was associated with low grade and NMIBC in the case of the preservation of the papillary pattern. We recommend further studies on the cut-off of widely used markers and more immunohistochemical and genetic studies on the p16(INK4A), taking into consideration the histological pattern of conventional carcinomas.

Hepatic Angiosarcomas With Sinusoidal Growth Patterns.

Hepatic angiosarcomas are aggressive malignant tumors of the liver with variable morphology. One of the rare morphologies is that of the sinusoidal growth pattern, which is challenging to diagnose because of its subtle imaging and morphologic findings. This retrospective study characterizes the clinical, histologic, and immunohistochemical features of sinusoidal hepatic angiosarcomas. Thirteen cases were included in the study, comprising 12 (92.3%) needle core biopsies and 1 wedge biopsy; one of the needle biopsies also had a subsequent resection specimen available for review. Multiple biopsies were needed to make the diagnosis in 4 cases. At least moderate sinusoidal dilatation was seen in 53.8% of cases. Increased cellularity within the sinusoids was seen at both low-power and high-power magnification (69.2% and 84.6%, respectively). Cytologic atypia ranged from mild to marked. Multinucleated tumor cells were present in most cases (10/13 cases) but were often sparse. Mitotic activity was identified in 5/13 cases. ERG immunostains were more reliable than CD31 and CD34 in identifying the tumor cells. Ki-67 proliferative index ranged from 5% to 30%. p53 immunostains were available in 9 cases and c-MYC in 7 cases; they were positive in 62.5% and 33.3% of cases, respectively and had a mutually exclusive staining pattern. In summary, this rare pattern of hepatic angiosarcoma is challenging to diagnose but has distinctive morphologic findings that can be supplemented with immunostains to establish the diagnosis.

Genetic characterisation of Felis catus papillomavirus type 7, a rare infection of cats that may be associated with skin cancer

Six Felis catus papillomavirus (FcaPV) types have been fully sequenceed from domestic cats including some that have been associated with the development of neoplasia. A sequence from a novel FcaPV type was amplified from a basal cell carcinoma that contained unusual histological evidence of PV infection and intense p16CDKN2A protein (p16) immunostaining. The entire 7467 bp genome was amplified using ‘outward facing’ primers. The PV was designated FcaPV7 and contained putative coding regions that were predicted to produce five early proteins and two late ones. The ORF L1 showed 77% similarity to that of FcaPV6. As the novel PV also showed greater than 60% similarity to three other feline Tau-PV types, FcaPV7 is proposed to be classified within this genus. Specific primers were designed but did not amplify FcaPV7 DNA from any of 60 samples from the mouth and skin of cats. FcaPV7 appears to rarely infect cats. However, FcaPV7 may be associated with skin cancer in this species.

The utility of p63, CK7, and CAM5.2 staining in differentiating pagetoid intraepidermal carcinomas.

Extramammary Paget disease (EMPD), pagetoid squamous cell carcinoma in situ (PSCCIS), and Paget disease of the breast (PD) are intraepidermal carcinomas with overlapping histopathologic features. CK7 and CAM5.2 stains are frequently utilized to distinguish PSCCIS from EMPD and PD. However, some cases of PSCCIS can stain positively for CAM5.2 and CK7, indicating a potential pitfall with these stains. p63 has been shown to distinguish PSCCIS from EMPD. We assessed p63 staining in PD and compared it to p63 staining of PSCCIS and EMPD. A retrospective search for 15 examples each of PSCCIS, EMPD, and PD with remaining tissue in the paraffin block was performed. The diagnosis was confirmed by a board-certified dermatopathologist and immunostaining for p63, CK7, and CAM5.2 was performed. Staining >55% was scored as positive. Staining <55% was scored as negative and an approximate percentage of positive cells was recorded. Diffuse nuclear expression for p63 was detected in 100% (15/15) of PSCCIS cases, 0% (0/15) of PD cases, and 0% (0/15) of EMPD cases. CK7 and CAM5.2 stains were positive in 100% of PD. CAM5.2 was positive in 100% of EMPD and CK7 was positive in 93% of EMPD. CAM5.2 was positive in 0% of PSCCIS biopsy specimens, but partial staining was seen in 20%. CK7 was positive in 13%, but partial staining was seen in 47%. p63 immunostaining is a highly sensitive and specific method for differentiating between PSCCIS and PD or EMPD. While CAM5.2 and CK7 are also useful ancillary stains in this differential diagnosis, false-positive and false-negative staining occurs with these two markers.

The effect of P53 expression and smoking/alcohol in P16(+) and P16(–) oropharyngeal carcinoma and risk classification: the Turkish Society of Radiation Oncology Head & Neck Study Group 01-002

The aim of this study is to categorize the risk groups of patients with oropharyngeal carcinoma (OPC) according to p16 and p53 status, smoking/alcohol consumption history, and other prognostic factors. The immunostaining of p16 and p53 of 290 patients was retrospectively evaluated. The history of smoking/alcohol consumption of each patient was noted. p16 and p53 staining patterns were reviewed. The results were compared with demographic findings and prognostic factors. Risk groups have been classified for the p16 status of patients. The median follow-up was 47 months (range 6-240). Five-year disease-free survival (DFS) rates for patients with p16 (+) and (-) were 76% and 36%, and overall survival rates were 83% vs 40%, respectively (HR=0.34 [0.21-0.57], P < .0001), HR=0.22 [0.12-0.40] P < .0001, respectively). p16(-), p53(+), heavy smoking/alcohol consumption, performance status; advanced T and N stages in patients with p16(-), and continuing smoking/alcohol consumption after treatment were found to be unfavorable risk factors. Five-year overall survival rates were 95%, 78%, and 36% for low, intermediate, and high-risk groups, respectively. The results of our study have shown that p16 negativity in patients with oropharyngeal cancer was found to be an important prognostic factor, especially for those with lower p53 expression and not smoking/consuming alcohol.

Abstract 5555: Altered p53/p16 expression is linked to urothelial carcinoma progression but is unrelated to prognosis in muscle-invasive tumors

Abstract Background: Most inactivating p53 mutations result in a nuclear accumulation of the defective p53 protein. However, p53 alterations that result in a complete lack of cellular p53 and complete absence of p53 immunostaining do also occur. As p16 is upregulated in p53 inactivated cells, p16 immunohistochemistry may be a surrogate marker for p53 inactivation. Design: In this study, we investigated p53 and p16 immunostaining on more than 2,500 urothelial bladder carcinomas in a tissue microarray format to better understand their impact in relation to clinicopathological parameters of disease progression and patient outcome. Results: p16 immunostaining was not observed in normal urothelium but occurred in 1,576 (63.5%) of cancers including 755 (30.4%) with a strong staining. The fraction of p16 positive cases increased markedly from pTaG2 low grade (9.6%) to pTaG3 high grade tumors (46.5% strongly positive, p&amp;lt;0.0001 for pTaG2 low vs. pTaG3) but continuously decreased from pTaG3 to pT2 (41.3% strongly positive), pT3 (36.5%) and pT4 (33.3%; p=0.0030). Within pT2-4 carcinomas, p16 positivity was also linked to high grade (p=0.0005) but unrelated to overall survival. p53 staining has been recorded as negative in 203 (8.4%), very weak in 373 (15.4%), weak in 1,341 (55.3%), strong in 115 (4.7%), and very strong in 393 (16.2%) cancers. The fraction of tumors with negative (potentially p53 null phenotype), strong, and very strong p53 positivity increased markedly from pTaG2 low grade to pTaG3 high grade tumors (p&amp;lt;0.0001) and from pTaG3 to muscle-invasive pT2-4 cancers (p=0.0007). p53 staining pattern was unrelated to histopathological parameters of malignancy or patient prognosis within pT2-4 carcinomas, however. There was a significant overall association between p53 and p16 expression but strong p16 expression predominated in tumors with very strong, strong, and negative p53 staining. Subset analyses showed that the combination of p53 negative/p16 strongly positive cancers was particularly linked to features of tumor aggressiveness. Conclusion: Our data show that altered function of p53 and p16 immunostaining increases during grade and stage progression although these alterations lack prognostic significance in pT2-4 carcinomas. That high level p16 expression is limited to neoplastic urothelium and that the p53 null phenotype is largely limited to grade 3 and invasive urothelial carcinomas are features with potential diagnostic utility. Citation Format: Simon Schallenberg, Henning Plage, Sebastian Hofbauer, Kira Kornienko, Sarah Weinberger, Paul G. Bruch, Florian Roßner, Sefer Elezkurtaj, Martina Kluth, Maximilian Lennartz, Tim Mandelkow, Elena Bady, Niclas C. Blessin, Andreas H. Marx, Henrik Samtleben, Margit Fisch, Michael Rink, Marcin Slojewski, Krystian Kaczmarek, Thorsten Ecke, Steffen Hallmann, Stefan Koch, Nico Adamini, Sarah Minner, Ronald Simon, Guido Sauter, David Horst, Tobias Klatte, Thorsten Schlomm, Henrik Zecha. Altered p53/p16 expression is linked to urothelial carcinoma progression but is unrelated to prognosis in muscle-invasive tumors. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5555.

Abstract 6222: Clinical impact of p53 and AXL immunostaining on the efficacy of immune checkpoint inhibitor-based therapy after osimertinib treatment in patients with epidermal growth factor mutant non-small cell lung cancer

Abstract Background: Current evidence indicates that immune checkpoint inhibitors (ICIs) have a limited efficacy in patients with lung cancer harboring epidermal growth factor receptor (EGFR) mutations. However, there is a lack of data on the efficacy of ICIs after osimertinib treatment, and the predictors of ICI efficacy are unclear. Methods: We retrospectively assessed consecutive patients with EGFR-mutant NSCLC who received ICI-based therapy after osimertinib treatment at 10 institutions in Japan, between March 2016 and March 2021. Immunohistochemical staining was used to evaluate the expression of p53 and AXL. The deletions of exon 19 and the exon 21 L858R point mutation in EGFR were defined as common mutations; other mutations were defined as uncommon mutations. Results: A total of 36 patients with advanced or recurrent EGFR-mutant NSCLC were analyzed. In multivariate analysis, p53 expression in tumors was an independent predictor of PFS after ICI-based therapy (p = 0.002). In patients with common EGFR mutations, high AXL expression was a predictor of shorter PFS and overall survival after ICI-based therapy (log-rank test; p = 0.04 and p = 0.02, respectively). Conclusion: The levels of p53 in pretreatment tumors may be a predictor of ICI-based therapy outcomes in patients with EGFR-mutant NSCLC after osimertinib treatment. High levels of AXL in tumors may also be a predictor of ICI-based therapy outcomes, specifically for patients with common EGFR mutations. Further prospective large-scale investigations on the predictors of ICI efficacy following osimertinib treatment are warranted. Citation Format: Kenji Morimoto, Tadaaki Yamada, Shinsaku Tokuda, Koichi Takayama. Clinical impact of p53 and AXL immunostaining on the efficacy of immune checkpoint inhibitor-based therapy after osimertinib treatment in patients with epidermal growth factor mutant non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6222.

Sun-Exposed versus Sun-Protected Cutaneous Basal Cell Carcinoma: Clinico-Pathological Profile and p16 Immunostaining.

Although widespread, BCC is still relatively poorly understood in regards to pathogenesis and prognosis, particularly the lesions formed on anatomical sites away from sun exposure. With the aim of deepening our understanding of the pathogenesis and clinico-pathological correlations of BCCs, we conducted this study. Tissue blocks and data of 52 Egyptian patients diagnosed with BCC were retrieved for clinical information and inclusion criteria, then re-examined histologically; p16 immunostaining was carried out and evaluated for analysis and comparison between the two groups, i.e., sun-exposed and sun-protected. Sex, age, clinical suspicion, tumor size, recurrence status, and histologic variants did not show a significant difference between the sun-protected and sun-exposed groups; however, the mean ages recorded were 67.2 vs. 62.7 for the sun-protected and sun-exposed groups, respectively. A total of 52% of BCCs were positive for p16. The sun-protected lesions showed p16 positivity in 61% of cases, whereas 49% of the sun-exposed lesions were positive with no significant difference. There was a significant difference in p16 expression between the recurrent and non-recurrent lesions. A significant difference was seen in the case of cancer recurrence, where all the recurrent BCCs in this study demonstrated negative p16 immunostaining of the primary lesions; however, the positively stained cases in total were 52% of BCCs. The mean patient age of the sun-protected group was much higher than in previous peer studies. We assume that the biological, prognostic, and clinical aspects of p16 protein expression in BCCs are still far from being clearly understood. Further studies are highly recommended, with more focus on its role in the pathogenesis and the prognostic factors.