Human papillomavirus in ocular malignant tumours: a study from a tertiary eye care centre in North India.

OBJECTIVE: The present study aims to detect the presence of human papillomavirus (HPV) in ocular malignant tumours, including retinoblastoma, eyelid squamous cell carcinoma (SCC), and sebaceous gland carcinoma (SGC), in the North Indian population. DESIGN: Prospective observational non randomized study. PARTICIPANTS: In this study, 142 prospective cases of ocular malignant tumours (retinoblastoma, SGC, and SCC) were included. METHODS: HPV was detected by multiplex PCR using PGMY09/11 primers in 142 patients with ocular malignancies. This was followed by genotyping using linear array (reverse hybridization). RESULTS: Of the 142 tumours studied, 72 were retinoblastoma, 30 SGC, and 40 SCC. The HPV genome was detected in 2.8% (4 of 142) of cases by multiplex PCR; all positive cases (4 of 40) were SCC. Genotyping revealed that all positives belonged to the high-risk HPV16 genotype. HPV-positive SCC patients had better disease-free survival. Retinoblastoma and SGC cases were negative for HPV. CONCLUSIONS: Low prevalence of HPV in ocular malignancies was observed in this study. The HPV genome was detected only in ocular squamous cell carcinoma cases and these patients were associated with better prognosis. HPV may not have a role in retinoblastoma and SGC in the North Indian population.

High-throughput sequencing reveals no viral pathogens in eight cases of ocular adnexal extranodal marginal zone B-cell lymphoma.

The purpose of the present study was to perform a next-generation sequencing (NGS) based analysis of viruses in ocular adnexal extranodal marginal zone B-cell lymphoma (EMZL). Eight patients with extraocular EMZL were identified in the archives of Department of Ophthalmology, Copenhagen University Hospital. All cases were validated according to the World Health Organization classification. We subjected samples to enrichment of virion-associated (encapsidated) nucleic acids which included sample homogenization, filtration, and nuclease treatment. Both DNA and RNA were sequenced, and we analyzed the sequencing data for the presence of viral sequences. We detected no pathogenic viruses likely to be associated to development of EMZL. In one case, we detected human polyomavirus 7 and traces of Epstein-Barr virus (EBV) (human herpesvirus 4 (HHV4)) and a human papillomavirus. In conclusion, no viral pathogens were consistently detected in the extraocular EMZL samples when applying NGS-based methods.

Series of extranodal natural killer/T-cell lymphoma, nasal type, with periorbital involvement.

PURPOSE: To describe the clinical presentation, diagnosis and treatment of periorbital extranodal natural killer/T-cell lymphoma, nasal type. METHODS: Case series of three patients with periorbital involvement of extranodal natural killer/T-cell lymphoma, nasal type, of whom clinical data, orbital imaging and immunohistochemical analysis were collected. For the purpose of this study, all histopathological and immunohistochemical slides were re-examined. RESULTS: All patients presented with painless eyelid swelling and a history of sinonasal disease, of whom one with bilateral panuveitis, not responding to systemic antibiotics. Extraocular muscle involvement was present in 2 cases upon presentation and in 1 case later on. Initial paranasal and orbital biopsies were negative in 2 patients, with only the second orbital biopsy leading to the diagnosis. Natural killer/T-cell and cytotoxic markers were present in all cases, as well as Epstein-Barr virus encoded RNA in situ hybridization. The patients died respectively 5, 9 and 35 months from diagnosis despite treatment with chemotherapy and radiotherapy. CONCLUSION: Extranodal natural killer/T-cell lymphoma, nasal type, should be suspected in a painless periorbital cellulitis with chronic sinusitis, not responding to conventional therapy. A high index of suspicion is necessary in biopsies showing angiodestruction and necrosis. Epstein-Barr virus encoded RNA in situ hybridization and expert hematopathologist consultation is necessary to decrease the delay in diagnosis.

Hypermethylation of the CDKN2A gene promoter is a frequent epigenetic change in periocular sebaceous carcinoma and is associated with younger patient age.

Periocular sebaceous carcinoma is an aggressive neoplasm with significant morbidity and mortality. Its pathogenesis is poorly understood. It is only rarely associated with Muir-Torre syndrome. Previous studies from Asian countries, have suggested that human papillomavirus (HPV) infection plays a role in the pathogenesis and overexpression of p16(INK4a), a surrogate marker of HPV infection, have also been reported. However, data from western countries seem contradictory. In order to clarify and explore the molecular and epigenetic basis of HPV, CDKN2A status and role of microsatellite instability in the development of periocular sebaceous carcinoma, 24 cases of periocular sebaceous carcinoma were analyzed for the expression of p16(INK4a) and mismatch repair proteins (MLH1, MSH2, MSH6 and PMS2) via immunohistochemistry. Nested polymerase chain reaction (PCR) and genechip HPV typing were used to detect HPV infection and decide its genotype when present. PCR amplification using a consensus primer pair was also performed to detect ß-HPV. The methylation status of CDKN2A promoter region was studied by methylation-specific polymerase chain reaction. HPV-positivity was demonstrated in only one of our cases (HPV 16), while another case showed p16(INK4a) overexpression. All cases showed preserved expression of mismatch repair proteins. CDKN2A promoter hypermethylation was noted in nearly half of our cases (11/24) and was associated with younger patient age (P = .013). Our results showed that periocular sebaceous carcinoma is rarely associated with HPV and microsatellite instability. Higher frequency of CDKN2A promoter hypermethylation in younger patients implies a significant epigenetic role in tumor development in this age group.

Dye laser photodynamic therapy for Bowen's disease in a patient with epidermodysplasia verruciformis.

Epidermodysplasia verruciformis (EV) is a rare heritable skin disease that results in unusual susceptibility to infection with specific types of human papillomavirus (HPV). Here we report a 53-year-old man with EV who developed Bowen's disease on his lower eyelid and the chest. Mutation analysis of EVER1 gene revealed homozygous splice acceptor site mutation (IVS8-2, A > T). In this patient, HPV3, HPV14, and HPV38 had been identified from the skin lesions. The Bowen's skin lesion on the left lower eye-lid was treated by photodynamic therapy (PDT) using 5-aminolevulinic acid (ALA) and pulsed dye laser (PDL). After two rounds of the PDT treatment, the skin lesion disappeared and a skin biopsy confirmed the efficacy of the treatment. This method was simple, less invasive than other treatments, and achieved a satisfactory cosmetic result.

Merkel cell carcinoma with heterologous rhabdomyoblastic differentiation: the role of immunohistochemistry for Merkel cell polyomavirus large T-antigen in confirmation.

Merkel cell carcinoma (MCC) represents an uncommon and lethal form of cutaneous malignancy. Historically, the pathogenesis of MCC has been presumed to be linked to ultraviolet light overexposure, but recently, it has been documented that some examples harbor polyomavirus genome, the presence of which is presumed to be of pathogenetic importance. Extremely rare cases of MCC may show heterologous differentiation. We report an example of MCC with heterologous rhabdomyosarcomatous differentiation, the third such case to date, with emphasis on its distinction from fusion-negative alveolar rhabdomyosarcoma. The role of immunohistochemistry for Merkel cell polyomavirus large T-antigen in this differential diagnosis is emphasized.

Hepatitis C virus infection in ocular adnexal lymphomas.

OBJECTIVE: To assess the influence of hepatitis C virus (HCV) infection on disease appearance and outcome of ocular adnexal non-Hodgkin lymphoma (ONHL). DESIGN: Retrospective comparative study (from January 1, 1992, through December 31, 2006). METHODS: The medical records of 129 patients with ONHL were retrospectively reviewed. All the patients were tested serologically for the presence of HCV infection. Patients were divided into 2 groups according to the presence or absence of HCV infection. MAIN OUTCOME MEASURES: Prevalence of HCV infection, staging to evaluate the extent of disease at the onset, and clinical outcome data on overall and disease-free survival. RESULTS: The prevalence of HCV infection among the patients with ONHL was 17.8%. Seropositivity for HCV infection was significantly associated with extraorbital lymphoma at the onset (P = .006). High prevalence of mucosa-associated lymphoid tissue disease (79.8%) was registered. Protocol therapy included radiotherapy and chemotherapy, depending on the stage of the disease. Complete remission was achieved in 99 patients (76.7%). A total of 23.6% of patients with HCV-seronegative status and 21.7% of those with HCV-seropositive status experienced relapse of the lymphomatous disease. No significant differences in the 5-year overall survival and disease-free survival between the 2 groups were observed. CONCLUSIONS: Prevalence of HCV infection in patients with ONHL is a relevant issue, accounting for 17.8% of the examined patients. Infection with HCV may influence the initial appearance of ONHL because it is associated with more widespread disease at the onset. However, the overall and disease-free survival of the infected patients are not statistically different than that of patients who are not infected.

The evaluation of human papillomavirus and p53 gene mutation in benign and malignant conjunctiva and eyelid lesions.

Papillomas and squamous cell carcinomas are the most common conjunctival and eyelid lesions. The etiology is still unclear and recently human papillomavirus infection and p53 gene mutation have been taken into consideration. The aim of our study was the evaluation of HPV DNApresence and p53 gene mutation in 45 benign and 38 malignant squamous lesions of the conjunctiva and eyelid. For HPV detection PCR-RFLP and immunohistochemical reaction were used; for p53 gene mutation PCR-SSCP was used. Only 8.8% papillomas, 9.1% squamous cell cancers and 3.7% basal cell cancers (using PCR-RFLP method) and 26.6% papillomas, 7.4% squamous cell cancers and 9.1% basal cell cancers (using immunohisto-chemical reaction) were HPV positive. p53 gene mutation was evaluated in 24.4% papillomas, 54.5% squamous cell cancers and 22.2% basal cell cancers; most commonly in 6 and 7 exon. Human papillomavirus infection, opposite to p53 gene mutation, is not a significant etiological factor of the benign and malignant conjunctival and eyelid lesions development.

The role of infectious agents in the etiology of ocular adnexal neoplasia.

Given the fact that infectious agents contribute to around 18% of human cancers worldwide, it would seem prudent to explore their role in neoplasms of the ocular adnexa: primary malignancies of the conjunctiva, lacrimal glands, eyelids, and orbit. By elucidating the mechanisms by which infectious agents contribute to oncogenesis, the management, treatment, and prevention of these neoplasms may one day parallel what is already in place for cancers such as cervical cancer, hepatocellular carcinoma, gastric mucosa-associated lymphoid tissue lymphoma and gastric adenocarcinoma. Antibiotic treatment and vaccines against infectious agents may herald a future with a curtailed role for traditional therapies of surgery, radiation, and chemotherapy. Unlike other malignancies for which large epidemiological studies are available, analyzing ocular adnexal neoplasms is challenging as they are relatively rare. Additionally, putative infectious agents seemingly display an immense geographic variation that has led to much debate regarding the relative importance of one organism versus another. This review discusses the pathogenetic role of several microorganisms in different ocular adnexal malignancies, including human papilloma virus in conjunctival papilloma and squamous cell carcinoma, human immunodeficiency virus in conjunctival squamous carcinoma, Kaposi sarcoma-associated herpes virus or human herpes simplex virus-8 (KSHV/HHV-8) in conjunctival Kaposi sarcoma, Helicobacter pylori (H. pylori,), Chlamydia, and hepatitis C virus in ocular adnexal mucosa-associated lymphoid tissue lymphomas. Unlike cervical cancer where a single infectious agent, human papilloma virus, is found in greater than 99% of lesions, multiple organisms may play a role in the etiology of certain ocular adnexal neoplasms by acting through similar mechanisms of oncogenesis, including chronic antigenic stimulation and the action of infectious oncogenes. However, similar to other human malignancies, ultimately the role of infectious agents in ocular adnexal neoplasms is most likely as a cofactor to genetic and environmental risk factors.

Solitary eyelid Kaposi sarcoma in an HIV-negative patient.

PURPOSE: To describe a case of localized Kaposi sarcoma (KS) of the eyelid in an HIV-seronegative patient. METHODS: An 80-year-old man developed an ulcerated nodular tumor-like mass that grew rapidly on his left upper eyelid. There were no similar lesions elsewhere. The eyelid lesion was completely excised and histopathologically examined. Serological analyses and molecular biologic techniques, including polymerase chain reaction, were used. RESULTS: Laboratory examinations were within normal limits, and serology for HIV was negative. Histological sections revealed a vascular proliferation composed predominantly of small slit-like blood vessels and epithelioid spindle cells, supporting the diagnosis of KS. Polymerase chain reaction was positive for human herpesvirus 8. During a 2-year follow-up, no recurrences, development of new lesions, or HIV seroconversions were observed. CONCLUSION: This is a classic KS involving only the eyelid in an HIV-negative patient. Location in the eyelid is a possible, albeit rare, initial solitary manifestation of KS in elderly HIV-negative patients. Surgery is both safe and effective.

Epstein-Barr positive T-cell lymphoma in the ocular region.

PURPOSE: To present two cases of rapidly growing tumors in the ocular adnexa. Both tumors were Epstein-Barr virus (EBV) positive peripheral T-cell lymphoma. METHODS: Case 1 was a 60-year-old man with a non-tender ulcerating tumor involving the lateral third of both upper and lower right eyelid. Case 2 was a 55-year-old man with a swelling of the left eyelid expanding cranially and dislocating the left eye, resulting in proptosis and diplopia. Both patients underwent incisional biopsy that did not disclose the malignant nature of the tumors. Clinical evaluation resulted in suspicion of malignancy and surgical excision was performed. RESULTS: The tumors were found to be consistent with EBV-positive peripheral T-cell lymphoma. CONCLUSIONS: Peripheral T-cell lymphoma is uncommon but a diagnosis to be considered in a patient with a tumorous lesion in the eye region. Furthermore, peripheral T-cell lymphoma may be EBV-positive.

Natural killer/T-cell lymphoma with ocular and adnexal involvement.

PURPOSE: To review the clinical, radiological, and histopathologic features in 8 patients with natural killer/T-cell lymphoma (NKTL) involving the orbit and/or ocular adnexa, and to describe the responses of these patients to various treatment regimens. DESIGN: Retrospective observational case series. PARTICIPANTS: Eight patients (5 male, 3 female) with NKTL involving the orbit and/or ocular adnexa were identified from 1999 through 2005. The mean age at presentation was 45 years (range, 26-65). METHODS: We retrospectively identified patients with NKTL of the ocular adnexa treated in the authors' medical centers from 1999 through 2004 using computerized diagnostic index retrieval. The clinical records and radiologic studies were analyzed to define modes of presentation and progression, response to therapy, and areas of anatomic involvement. Histopathologic findings, including the presence of CD3, CD56, and Epstein-Barr virus-encoded mRNA in each patient, were reviewed. MAIN OUTCOME MEASUREMENTS: Time of survival from presentation to last known follow-up and tumor-related death. RESULTS: Four of the 8 patients (50%) with NKTL involving the orbit or ocular adnexa had systemic involvement at presentation. Five of the 8 patients (62.5%) had concurrent sinonasal involvement, whereas 3 (37.5%) had orbital involvement alone. All lesions demonstrated CD3, CD56, and/or Epstein-Barr virus positivity on immunopathology studies. Therapy consisted of various chemotherapeutic regimens typically employed in the treatment of non-Hodgkins lymphoma, steroids, surgical intervention, and radiation. Seven (87.5%) patients died 5 weeks to 13 months after presentation, and 1 (12.5%) is alive without disease (5-year follow-up). CONCLUSIONS: Natural killer/T-cell orbital lymphoma is a rare Epstein-Barr virus-associated neoplasm that may occur with or without associated sinonasal involvement. Our series, the largest cohort reported to date, demonstrates the high lethality of this condition despite aggressive conventional therapy, suggesting that new treatment options should be considered early in the course of treatment of patients with this disorder.

The expression of P53 protein and infection of human papilloma virus in conjunctival and eyelid neoplasms.

Human papilloma virus (HPV) infection and loss of P53 function have been identified as frequent events in various human tumors. The aim of this study was to evaluate P53 protein expression and to detect HPV in the tissue samples of 45 benign (papillomas) and 38 malignant conjunctival and eyelid lesions (27 basal cell carcinomas and 11 squamous cell carcinomas). We also looked for eventual relationships between P53 expression and clinicopathological features such as age, histological type of tumor, grading and staging. HPV infection was detected using the PCR-RFLP method. Specific primers were engaged and PCR products of HPV 16, 18, and 33, underwent enzymatic digestion at 37 degrees C. We revealed P53 protein expression in 30 out of 45 (66.6%) squamous cell papillomas. In the SCC and BCC groups, P53 was present in 31 out of 38 carcinomas and there was a statistically significant correlation between histological type of tumor and P53 protein expression. Malignant type HPV 16 and 18 were detected in three squamous cell papillomas, two BCCs and one SCC. However, we observed P53 protein expression in only two HPV-positive papillomas and one infiltrative type of BCC. P53 is probably involved in the development of conjunctival and eyelid tumors due to its high rate of presence in both benign and malignant neoplasms of these organs. HPV seems to occur rarely. In some cases its role in the pathogenesis of conjunctival and eyelid tumorigenesis should be considered as auxiliary.

Detection of HPV-20, HPV-23, and HPV-DL332 in a solitary eyelid syringoma.

PURPOSE: To report evidence of many human papillomavirus types occurring in a solitary syringoma clinically appearing as a papilloma. METHODS: A 57-year-old man presented with a 10-year history of an upper eyelid tumor. Histopathology, human papillomavirus-nested polymerase chain reaction, human papillomavirus-DNA cloning into vector pCR2.1, sequencing, and computer-assisted evaluation were performed. RESULTS: Histopathology demonstrated a solitary benign syringoma. HPV-20 and HPV-23 were present in one clone each, and HPV-5-related HPV-DL332 was present in 9 clones. CONCLUSION: Many human papillomavirus types may be detected in an ocular syringoma.

Extranodal lymphomas associated with hepatitis C virus infection.

Hepatitis C virus (HCV) infection may be complicated by non-Hodgkin's lymphoma. We describe eight cases of B-cell extranodal non-Hodgkin's lymphoma occurring during the course of chronic HCV-related hepatic disease (low-grade of mucosa-associated lymphoid tissue [MALT]-type; diffuse large cell; Burkitt; diffuse small cell). Some were localized to the liver (2), liver and spleen (1), spleen (1), peritoneal cavity (1), parotid gland (1); others manifested in the nasopharynx (1) and eyelid (1) but were accompanied by nodal disease. Four lymphomatous specimens available for molecular analysis exhibited clonal immunoglobulin gene rearrangements, lacked bcl-2, bcl-6, c-myc genes and p53 alterations, and did not contain replicative intermediate HCV RNA, as documented by a strand-specific reverse transcriptase-polymerase chain reaction. Low levels of positive-strand HCV RNA were detected in a single hepatic lymphoma, suggesting the presence of the virus in residual hepatocytes. The antigen-driven properties of HCV-associated B-cell malignant neoplasms may be considered for hepatic MALT-type lymphoma, which probably originated from lymphoid tissue acquired during long-standing HCV infection.

Non-Hodgkin lymphoma and Kaposi sarcoma in an eyelid of a patient with acquired immunodeficiency syndrome. Multiple viruses in pathogenesis.

A 36-year-old patient with acquired immunodeficiency syndrome sought care because of an upper eyelid lesion that dramatically increased in size. The histopathologic examination revealed a high-grade diffuse large cell non-Hodgkin lymphoma in continuity with a Kaposi sarcoma. In situ hybridization revealed Epstein-Barr virus in the large cell lymphoma and Kaposi sarcoma-associated herpesvirus in the Kaposi sarcoma lesion. This collision tumor is an unusual presentation of 2 malignant neoplasms in a patient with acquired immunodeficiency syndrome, with in situ hybridization evidence of Epstein-Barr virus and Kaposi sarcoma-associated herpesvirus in the lesion.

Squamous cell carcinoma of the eyelid and the acquired immunodeficiency syndrome.

PURPOSE: We studied a case of squamous cell carcinoma of the eyelid in a 42-year-old homosexual man with the acquired immunodeficiency syndrome (AIDS) to explore the link between squamous cell carcinoma and human Papillomavirus infection. METHOD: A rapidly growing, 3-mm lesion was removed from the right lower eyelid by excisional biopsy. RESULTS: Examination of the lesion confirmed squamous cell carcinoma. Polymerase chain reaction for human Papillomavirus 16 was positive. CONCLUSION: This case suggests a role for human Papillomavirus infection in the cause of squamous cell carcinoma in AIDS patients.

HPV type 16 in conjunctival and junctional papilloma, dysplasia, and squamous cell carcinoma.

AIMS: To clarify the role of human papillomavirus (HPV) infection in the development of papilloma, dysplasia, squamous cell carcinoma, and basal cell epithelioma arising from the eyelids, including the tunica conjunctiva palpebrum (conjunctiva), its junction to epidemis of eyelid skin (junction), and eyelid skin. METHODS: Sixteen cases of papilloma, four of dysplasia, four of squamous cell carcinoma, and 12 of basal cell epithelioma were examined using formalin fixed and paraffin embedded samples. Detection of HPV-DNA was performed by PCR-RFLP and in situ hybridisation (ISH) methods. RESULTS: HPV-16 was detected in 12/16 papillomas (75%), 2/4 dysplasias (50%), and 1/4 squamous cell carcinomas (25%) but in none of the basal cell epitheliomas. No other HPV subtypes were found. ISH assay showed positive signals in only two cases of dysplasia and squamous cell carcinoma. The mean age of HPV-16 positive dysplasia and squamous cell carcinoma cases (81.7 years) was significantly higher than that of HPV-16 positive papilloma cases (p < 0.01). CONCLUSIONS: Based on the presence of HPV-16 in both benign and malignant lesions and the age distribution, it seems likely that HPV-16 alone may be incapable of causing development of conjunctival and junctional dysplasia and squamous cell carcinoma, and that any correlation between the papilloma-squamous cell carcinoma sequence and HPV infection may be due to rare events.