Establishment of patient-derived xenografts of retinoblastoma and choroidal melanoma on the avian chorioallantoic membrane.

Purpose: To develop a viable in vivo chorioallantoic membrane (CAM) model to study the growth and invasion of patient-derived retinoblastoma (RB) and choroidal melanoma (CM) xenografts (PDXs). The study utilizes primary tumor samples instead of cancer cell lines, which provides a more authentic representation of tumors due to conserved morphology and heterogeneity. Methods: Fertilized chicken eggs were procured, windowed, and their CAM layers were dropped. On embryonic development day (EDD) 10, freshly cut patient-derived CM and RB tumors were implanted on the CAM layer and the setup was incubated for 7 days. The tumor-embedded CAM layer was harvested on EDD 17, and the extracted tumor samples were subjected to hematoxylin and eosin staining and immunohistochemical analysis to evaluate the extent of tumor invasion. Results: Significant changes in the vascularity around the RB and CM PDXs were observed, indicating an angiogenic environment. The cross-sectional histological view of the tumor implant site revealed the invasion of both the tumors into the CAM mesoderm. Invasion of CM into CAM mesoderm was visualized in the form of pigmented nodules, and that of RB was indicated by synaptophysin and Ki-67 positivity in Immunohistochemistry (IHC). Conclusion: The CAM xenograft model was successfully able to support the growth of CM and RB PDXs and their invasion in CAM, thus presenting as a feasible alternative to mammalian models for studying tumorigenicity and invasiveness of ocular tumors. Moreover, this model can further be utilized to develop personalized medicine by inoculating patient-specific tumors for preclinical drug screening.

Eyelid sebaceous gland carcinoma, validation of AJCC 8th edition T staging- a retrospective study from North India.

PURPOSE: This study compares the 8th edition staging of AJCC for prognosis of eyelid Sebaceous Gland Carcinoma (SGC) patients with respect to the 7th edition. METHODS: A retrospective study was undertaken over a period of 100 months. Ninety-nine histopathologically proven cases of eyelid SGC available for follow-up were recruited. Patients were staged by both the 7th and 8th editions of AJCC and followed up at six monthly intervals after surgery. RESULTS: Of the 99 eyelid SGC patients recruited, recurrence and orbital invasion developed in 22%, lymph node metastasis in 21% and 4% had distant metastasis and also died. High-risk features include tumour size>20 mm, orbital invasion, exenteration and staging by both the 7th and 8th editions of AJCC. Cox regression analysis revealed that staging by AJCC 8th edition was associated with greater risk for local recurrence (HR 3.01,95% CI-1.65-5.51%, p < 0.01) lymph node metastasis (HR 8.26, 95% CI 3.96-17.19%, p < 0.01) and disease-free survival (HR 4.61, 95% CI 2.81-7.54). The Kaplan-Meir survival curves of eyelid SGC patients by the 8th edition AJCC staging were also significantly associated with lymph node metastasis (p < 0.01), tumour-related deaths (p < 0.01) and reduced disease-free survival (p = 0.07). The higher Harrell's values by the 8th edition signify better predictive value for lymph node metastasis and DFS (disease-free survival). The lower AIC values indicate a better monotonicity of gradients for lymph node metastasis, recurrence and DFS. CONCLUSION: Staging by the 8th AJCC edition is, therefore, recommended for eyelid SGC as it gives a better perspective about disease outcome. The orbital extension was the single most important predictor of lymph node metastasis, recurrence and death.

Microbial Journey: Mount Everest to Mars.

A rigorous exploration of microbial diversity has revealed its presence on Earth, deep oceans, and vast space. The presence of microbial life in diverse environmental conditions, ranging from moderate to extreme temperature, pH, salinity, oxygen, radiations, and altitudes, has provided the necessary impetus to search for them by extending the limits of their habitats. Microbiology started as a distinct science in the mid-nineteenth century and has provided inputs for the betterment of mankind during the last 150 years. As beneficial microbes are assets and pathogens are detrimental, studying both have its own merits. Scientists are nowadays working on illustrating the microbial dynamics in Earth's subsurface, deep sea, and polar regions. In addition to studying the role of microbes in the environment, the microbe-host interactions in humans, animals and plants are also unearthing newer insights that can help us to improve the health of the host by modulating the microbiota. Microbes have the potential to remediate persistent organic pollutants. Antimicrobial resistance which is a serious concern can also be tackled only after monitoring the spread of resistant microbes using disciplines of genomics and metagenomics The cognizance of microbiology has reached the top of the world. Space Missions are now looking for signs of life on the planets (specifically Mars), the Moon and beyond them. Among the most potent pieces of evidence to support the existence of life is to look for microbial, plant, and animal fossils. There is also an urgent need to deliberate and communicate these findings to layman and policymakers that would help them to take an adequate decision for better health and the environment around us. Here, we present a glimpse of recent advancements by scientists from around the world, exploring and exploiting microbial diversity.

Immunohistochemical and mutational status of telomerase reverse transcriptase in conjunctival squamous cell carcinoma.

PURPOSE: Mutations in human telomerase reverse transcriptase (TERT) are associated with increased telomerase activity in cutaneous melanomas. Conjunctival squamous cell carcinoma, also referred to as ocular surface squamous cell carcinoma, is cancer on the surface of the eye. Recent studies have identified UV signat`ure mutations in TERT promoters in ocular melanoma and ocular surface squamous neoplasia. However, its immunohistochemical status has not been reported in ocular surface squamous cell carcinoma. This study aimed to explore the immunohistochemical and mutational status of TERT in ocular surface SCC. METHODS: The immunohistochemical expression of TERT and mutational status of TERT promoter was evaluated in 19 ocular surface squamous cell carcinoma cases. Conjunctival melanoma tissue was used as a positive control. RESULTS: The cytoplasmic overexpression of TERT was detected in 11/19 (57%), and TERT promoter mutations were identified in 6/19 (31%) of ocular surface squamous cell carcinoma. Out of these, 66% had a C228T mutation, and 33% had a C250T mutation. The TERT expression was found to be associated with a high (≥T3) AJCC category (P = 0.023), and TERT immunoexpression was significantly correlated with reduced disease-free survival (P = 0.024, log-rank analysis) in ocular surface squamous cell carcinoma patients. CONCLUSION: The present study demonstrates that TERT promoter mutations with UV signatures are frequent in ocular surface squamous cell carcinoma. The increased expression of TERT could be of biological significance in aggressive ocular surface squamous cell carcinoma.

Evaluation of PD-L1 and PD-1 expression in aggressive eyelid sebaceous gland carcinoma and its clinical significance.

Purpose: Eyelid sebaceous gland carcinoma (SGC) is an aggressive but rare malignancy of ocular region. Over-expression of PD-L1 and PD-1 has been demonstrated in a variety of solid tumors including conjunctival melanoma. PD-L1 is an immunoinhibitory molecule that suppresses the effective T cells response against tumor antigen leading to the progression of tumors. Inhibitors of the interaction of PD-L1 and PD-1 are associated with good clinical response various carcinomas. The prognostic value of the PD-1/PD-L1 axis in SGC remains unexplored. The purpose of this study was to evaluate expressions of PD-1 and its ligand PD-L1 in SGC and correlate its expression with clinicopathological features and patients survival. Methods: The immunohistochemical expression of PD-L1 and PD-1 was evaluated in 30 SGC cases. Results: PD-L1 immunopositivity was detected in 41.9% of the SGC cases. PD-1 expression in tumor infiltrative lymphocytes (TILs) was observed in 53.3% samples. Tumor PD-L1 positivity, PD-1 expression in TILs and tumor size (>10 mm) was associated with reduced disease-free survival. On multivariate analysis only tumor size (>10 mm) and a combined positivity of PD-L1 in tumor cells and PD-1 in TILs with an odds ratio of 5.212 (95% confidence interval 1.449-18.737) continued to be significantly associated with SGC recurrence. Conclusion: PD-L1 is overexpressed in 50% of SGC cases. The combined tumor PD-L1 positivity and TILs showing PD-1 expression within the same SGC patient's samples predict high-risk SGC, suggesting that the up-regulation of PD-L1 in tumor cells and PD-1 positivity within the same SGC patient may aggravate tumor recurrence.

Immunohistochemical evaluation of stress-responsive protein sestrin2 and its correlation with p53 mutational status in eyelid sebaceous gland carcinoma.

BACKGROUND: p53 is a stress-activated tumour suppressor gene, and its mutation has been associated with solid tumours including non-melanoma skin cancers. Sestrin2 expression is associated with DNA damage and oxidative stress and has been described as a downstream target of p53 network. However, its role in sebaceous gland carcinoma (SGC) remains unexplored. OBJECTIVES: To determine the role of p53 and its downstream target gene sestrin2 expression and p53 gene mutation status in SGC. METHODS: Twenty cases of eyelid SGC tumour and circulating cell-free DNA (ccfDNA) were subjected to mutational analysis of p53 gene. p53 and sesrin2 expression was evaluated by immunohistochemistry. Results were correlated with the clinicopathological features of eyelid SGC. RESULTS: p53 gene mutations was detected in 25% of the SGC cases. A C>T transition was identified in exon 6 in a single patient in both tumour and ccfDNA. A G>T transversion leading to amino acid change D259Y was seen in four patients. A splice site mutation affected a single case in exon 6. p53 expression was observed in 55% SGC. Loss of sestrin2 in 55% SGC cases correlated with poor tumour differentiation (P=0.0001), upper eyelid involvement (P=0.004), p53 mutation (P=0.039) and with mutant p53 expression (P=0.0001). CONCLUSION: Sestrin2 expression was found to be significantly reduced in p53 mutated SGC cases and in cases with strong p53 nuclear immunopositivity, suggesting that loss of sestrin2 may be of biological significance in the development of SGC and as a key downstream component of p53 tumour suppression network in eyelid SGC.

Immunohistochemical expression of X-linked inhibitor of apoptosis in eyelid sebaceous gland carcinoma predicts a worse prognosis.

PURPOSE: Overexpression of the inhibitors of apoptosis proteins have been demonstrated in a variety and of solid tumors including melanomas and nonmelanomas skin cancers. X-linked inhibitor of apoptosis protein (XIAP) is an inhibitor of apoptosis which prevents apoptosis by inhibiting caspases 9, 7, and 3. The prognostic value of XIAP in sebaceous gland carcinoma (SGC) remains unexplored. METHODS: The immunohistochemical expression of XIAP was evaluated in 29 SGC cases. RESULTS: The cytoplasmic overexpression of XIAP was detected in 62% SGC cases. XIAP expression was found to be significantly associated with advanced age, large tumor size, and with reduced disease-free survival (P = 0.0174). XIAP expression and advance tumor Grade III emerged as significant risk factors on univariate analysis. On stepwise multivariate analysis, both increased cytoplasmic XIAP expression and high tumor grade were found to be significantly associated with recurrence. Patients with low XIAP immunoexpression had a longer disease-specific survival than those with high expression in the 5-year follow-up. CONCLUSION: The present study demonstrates at the immunohistochemical level that XIAP is overexpressed in SGC and that high expression could be of biological significance in the development of eyelid SGC. Our finding suggests that up-regulation of XIAP may aggravate tumor metastasis in SGC.

Clinical relevance of cyclooxygenase 2 and peroxisome proliferator-activated receptor γ in eyelid sebaceous gland carcinoma.

AIMS: Sebaceous gland carcinoma (SGC) is a malignancy associated with the pilosebaceous unit, and occurs at ocular or non-ocular sites. Cyclooxygenases (COXs) are enzymes that are crucial for lipid metabolism. COX-2 is overexpressed in various cancers, and its inhibition by non-steroidal anti-inflammatory drugs is known to reduce the risk of many cancers. Peroxisome proliferator-activated receptor (PPAR)-γ is a transcription factor involved in adipogenesis. PPAR-γ is a potential therapeutic target for the treatment of malignant tumours, including colon carcinoma. The aim of this study was to explore the status of COX-2 and PPAR-γ as prognostic markers in human eyelid SGC. METHODS AND RESULTS: The immunohistochemical expression of COX-2 and PPAR-γ was evaluated in 31 SGC cases. Cytoplasmic expression of COX-2 was detected in 80% of the SGC cases, and nuclear expression of PPAR-γ in 87%. There were significant correlations of PPAR-γ expression with well-differentiated SGC [19/21 (90%)] and of COX-2 overexpression with reduced disease-free survival (P = 0.0441, log rank analysis). COX-2 expression [odds ratio (OR) 3.82, 95% confidence interval (CI) 1.02-14.33, P = 0.046] and lymph node metastasis (OR 0.17, 95% CI 0.04-0.65, P = 0.009) emerged as significant risk factors in the univariate analysis. However, COX-2 expression did not emerge as a significant independent prognostic factor in multivariate analysis. CONCLUSIONS: COX-2 is a potential marker for identifying high-risk SGC patients. Expression of PPAR-γ in eyelid SGC cases reflects terminal sebaceous differentiation. Inhibitors of COX-2 signalling and PPAR-γ agonists are both prospective novel therapeutic targets in the management of eyelid SGC patients.

Does ß-catenin have a role in pathogenesis of sebaceous cell carcinoma of the eyelid?

AIM: Expression of ß-catenin in sebaceous cell carcinoma (SbCC) of the eyelid and its correlation with histopathological features. METHOD: 48 cases of SbCC were analysed immunohistochemically using monoclonal ß-catenin antibody and the results correlated with tumour size, histopathological differentiation, orbital invasion and pagetoid spread. RESULTS: Cytoplasmic overexpression of ß-catenin was seen in 66% cases of SbCC which correlated positively with tumour size, orbital invasion and pagetoid spread. This correlation was found to be significant in tumour size > 2 cm (p = 0.242). Nuclear staining was not observed in any of the cases. CONCLUSION: Cytoplasmic overexpression of ß-catenin was observed in the majority of the cases of SbCC of eyelid, and this correlated significantly with tumour size. The authors therefore hypothesise that ß-catenin overexpression in SbCC of eyelid may be a result of Wnt/ß-catenin pathway dysregulation. However, its role both in the development of sebaceous cell carcinoma of the eyelid as well as its prognosis needs to be explored further.