Expression of cancer-testis antigens in the immune microenvironment of non-small cell lung cancer.

The antigenic repertoire of tumors is critical for successful anti-cancer immune response and the efficacy of immunotherapy. Cancer-testis antigens (CTAs) are targets of humoral and cellular immune reactions. We aimed to characterize CTA expression in non-small cell lung cancer (NSCLC) in the context of the immune microenvironment. Of 90 CTAs validated by RNA sequencing, eight CTAs (DPEP3, EZHIP, MAGEA4, MAGEB2, MAGEC2, PAGE1, PRAME, and TKTL1) were selected for immunohistochemical profiling in cancer tissues from 328 NSCLC patients. CTA expression was compared with immune cell densities in the tumor environment and with genomic, transcriptomic, and clinical data. Most NSCLC cases (79%) expressed at least one of the analyzed CTAs, and CTA protein expression correlated generally with RNA expression. CTA profiles were associated with immune profiles: high MAGEA4 expression was related to M2 macrophages (CD163) and regulatory T cells (FOXP3), low MAGEA4 was associated with T cells (CD3), and high EZHIP was associated with plasma cell infiltration (adj. P-value < 0.05). None of the CTAs correlated with clinical outcomes. The current study provides a comprehensive evaluation of CTAs and suggests that their association with immune cells may indicate in situ immunogenic effects. The findings support the rationale to harness CTAs as targets for immunotherapy.

ALK gene rearrangement status in non-squamous non-small cell lung carcinoma in the Middle Eastern population.

BACKGROUND: Worldwide, the frequency of ALK rearrangement ranges between 3-6%, however its prevalence in the Middle Eastern population has not been reported. The aim of this study is to determine the frequency of the ALK rearrangement, as well as the clinico-pathologic characteristics of Lebanese patients with ALK-rearranged lung adenocarcinoma. METHODS: 152 patients diagnosed with non-squamous non-small cell lung carcinomas (NSCLC), at Hôtel-Dieu de France University Hospital between February 2014 and July 2016, were included in the study. ALK gene rearrangement expression was screened by immunohistochemistry (IHC) (D5F3 Clone). Positive cases were then sent for confirmation with Fluorescence in situ hybridization (FISH) technique. RESULTS: On immunohistochemistry, patients were distributed as following: score 0: n=108, score 1+: n=26, score 2+: n= 9, score 3+: n= 9. ALK gene rearrangement was detected in 6 out of 18 (2+ and 3+ score) tested patients by FISH technique. The presence of ALK rearrangement was significantly associated with the female gender (n=6, p=0.003) and with non-smoking status (n=4, p=0.0.18). CONCLUSION: This study confirms that the prevalence of the ALK gene rearrangement in the Middle Eastern region is within the worldwide ranges and is almost exclusive to patients with adenocarcinoma subtype and tends to occur more frequently in women and non-smokers.

Tumor Banks: A Quality Control Scheme Proposal.

Introduction: Tumor banks make a considerable contribution to translational research. Using emerging molecular tests on frozen material facilitates the development of new diagnostic and therapeutic strategies, especially in rare cases. However, standard quality control schemes are lacking in the current literature. Methods: In 2017, we have conducted a robust quality control test on 100 of 15,000 fresh frozen samples collected between 2000 and 2013 at the Jules Bordet Tumor Bank (Brussels). RNA and DNA extraction was done. The quality of RNA, DNA and proteins were evaluated, respectively by measuring RNA Integrity Number (RIN), by checking Electrophoretic Integrity (EI) and by performing Immunohistochemistry staining (IHC). A score, ranging from poor (1) to excellent (4), was attributed based on technical analysis. Results: RNA purity was scored 4 in 97% of the cases, 3 in 2%, and 2 in 1%. RIN scores were similarly 4 in 89%, 3 in 10%, and 2 in 1% of the cases. DNA purity was scored 4 in 94% and 3 in 6%, EI was scored 4 in 100% of the cases. Despite morphology loss after freezing, HER2, ER, and Ki67 IHC stainings yielded a score of 4 in the majority of samples. Furthermore, participating in the ISBER Proficiency Testing helped us validate our techniques and the technician's work. Seven processing schemes were carried out, the scores obtained were very satisfactory (20/27) or satisfactory (7/27). Conclusion: Tumor Banks can be precious for translational research. Nevertheless, firm quality controls should be applied to ensure high quality material delivery. Only then can biobanks contribute to diagnostics, biomarkers discovery and reliable molecular test development.

Chromogenic In Situ Hybridization as a Tool for HPV-Related Head and Neck Cancer Diagnosis.

Human papillomavirus (HPV) infection is a major risk factor for a subtype of oropharyngeal squamous cell carcinoma (OPSCC), which tends to be associated with a better outcome than alcohol- and tobacco-related OPSCC. Chromogenic in situ hybridization (CISH) of HPV viral RNA could allow the semiquantitative evaluation of viral transcripts of the oncogenic proteins E6 and E7 and an in situ visualization with a good spatial resolution. This technique allows the diagnosis of an active infection with the visualization of HPV transcription in the tumoral HPV-infected cells. An advantage of this technique is the avoidance of contamination from nonneoplastic HPV-infected cells adjacent to the tumor. Overall, its good diagnosis performances have it considered to be the gold standard for active HPV infection identification. Since E6 and E7 viral protein interaction with cell proteins pRb and p53 is mandatory for cell transformation, HPV RNA CISH is functionally relevant and acutely reflects active oncogenic HPV infection. This technique is clinically relevant as well since "low" or "high" HPV transcription levels helped the identification of two prognosis groups among HPV-related p16-positive head and neck cancer patients. Here we present the protocol for manual HPV RNA CISH performed on formalin-fixed paraffin-embedded (FFPE) slides with a kit obtained from the manufacturer. Instead of chromogenic revelation, RNA in situ hybridization may also be performed with fluorescent revelation (RNA FISH). It may also be combined with conventional immunostaining.

Immunohistochemical Analysis of Somatostatin Receptors in Endometriosis Tissue Samples: A Retrospective Study.

Three types of endometriosis are described: superficial peritoneal endometriosis (SPE), ovarian endometrioma (OMA), and deep infiltrating endometriosis (DIE). The expression of somatostatin receptors (SSTR1, 2, and 5) in human endometrial tissue and its ectopic form has been previously studied and may be different in each type of endometriosis. The aim of this study was to assess the immunohistochemical expression of SSTR1, 2, and 5 in tissue samples of SPE, OMA, and DIE. We performed a retrospective analysis in the pathology department database. Patients aged <50 yr and diagnosed with endometriosis have been identified and sorted into 3 groups according to their endometriosis type: SPE, OMA, and DIE. For each selected patient, formalin-fixed paraffin-embedded blocks were retrieved in order to make new sections to be incubated with polyclonal rabbit antibodies anti-SSTR1, 2, and 5. Receptor status was considered as positive on the sections when >50% of the cells showed immunostaining. Seventy-six patients were included in the analysis. SSTR1 and 5 were expressed in 95.4% and 77.2% of SPE, respectively, in 95.8% and 83.3% of OMA, respectively, and in 100% and 80% of DIE, respectively. There was no significant difference between SPE, OMA, and DIE with regard to the SSTR1 (P=0.5) and SSTR5 (P=0.9) expression. We observed a significant difference between SPE (9.0%), OMA (16.6%), and DIE (63.3%) with regard to SSTR2 expression (P<0.05). The present study identifies 2 different immunohistochemical patterns of endometriosis lesions with regard to their SSTR expression: SSTR1+/SSTR2-/SSTR5+ for SPE and OMA, and SSTR1+/SSTR2+/SSTR5+ for DIE.

Sporadic colon cancer in Lebanon: A clinicopathological study.

Colon cancer is a heavy public health burden. No data has been previously published on colon cancer epidemiology in Lebanon. The objective of this study was to report the clinical and pathological features of surgically operated colon cancer. From July 2005 to July 2012, 187 sporadic colonic tumors were operated in Hotel-Dieu de France Hospital (Beirut, Lebanon). Demographic, clinical, pathological and staging data was collected. The male: female sex ratio was 1.3 and the mean age at diagnosis was 66.0 years. Most tumors occurred at the right colon (54.0%). Histologically, conventional adenocarcinoma had the biggest proportion (88.2%), followed by mucinous adenocarcinoma (9.1%). Most cases operated belonged to stages II, III or IV, leaving only 8.0% of the cases in stage I. Some characteristics were similar to western countries like the predominance of right tumors while others were similar to developing countries like the sex ratio and the high proportion of stage IV tumors at diagnosis. This study reports for the first time in the middle-eastern populations a clinico-pathological analysis of surgically operated colon cancer.

Inflammatory Stroma of Lymphoepithelioma-like Carcinoma of the Cervix: Immunohistochemical Study of 3 Cases and Review of the Literature.

The aim of this study was to characterize the inflammatory infiltrate of the lymphoepithelioma-like carcinoma, a variant of squamous cell carcinoma clinically associated with a good prognosis. Immunohistochemistry was used to characterize 3 cases of lymphoepithelioma-like carcinoma in the uterine cervix, diagnosed over a period of 3 yr. The patients were between 30 and 50 yr old. Their cervical smears had shown atypical squamous cells of unknown significance or high-grade squamous intraepithelial lesions, and the initial cervical biopsy showed high-grade cervical intraepithelial neoplasia (CIN3) or squamous cell carcinoma. All patients underwent a Wertheim operation, 2 of which were preceded by a cervicectomy. Microscopically, all tumors were characterized by poorly defined sheets of undifferentiated squamous cells with a syncytial pattern and a dense background of infiltrating lymphocytes. Immunohistochemical analysis revealed that the lymphocytes were predominantly CD3 and CD8 T cells. The tumor lobules contained isolated CD8 T cells, whereas CD4 T cells and CD20 B cells surrounded the tumor lobules. CD56 NK cells and CD79 B cells were scattered in the tumor tissue. An in situ hybridization staining for Epstein-Barr encoding region was negative but all cases were immunohistochemically positive for P16. Follow-up varied between 2 mo and 2.7 yr. All 3 patients were disease free. Lymphoepithelioma-like carcinoma of the uterine cervix is a variant of squamous cell carcinoma, known for its better prognosis. The good prognosis of this tumor is potentially explained by the high levels of infiltrating CD8 T cells.

Distribution and Characteristics of Pulmonary Neuroendocrine Tumors: Single Institution Experience in Lebanon.

BACKGROUND: Neuroendocrine tumors represent 20% of primary lung neoplasms in some registries. According to the WHO classification of 2004, reconsidered for 2015, these lung tumors are divided into 4 groups: typical and atypical carcinoid, small cell and large cell neuroendocrine carcinomas. We report in this paper, for the first time in Lebanon, the distribution and the population characteristics of these tumors. MATERIALS AND METHODS: This descriptive retrospective study concerned all the pulmonary neuroendocrine tumors (NET) with their characteristics diagnosed in Hotel Dieu de France in Beirut, Lebanon from 2001 to 2012, with attention to features like age, gender and subgroup. RESULTS: Of 194 patients with pulmonary NET, 12.4% were typical carcinoid tumors, 3.6% atypical carcinoid, 66.5% small cell lung cancer, 7.7% combined small cell carcinomas and 9.8% large cell neuroendocrine tumors. The mean ages of patients were respectively 51.2 years in typical carcinoid, 64 years in atypical carcinoid, 64.2 years in small cell lung cancers, 67.2 in combined small cell lung cancer and 66.9 in large cells neuroendocrine tumors. The M/F sex ratios were respectively 0.3, 1.3, 1.4, 2.7 and 2.2. CONCLUSIONS: The characteristics of lung neuroendocrine tumors in our Lebanese institution are comparable to those reported in the literature.

Digestive Neuroendocrine Tumor Distribution and Characteristics According to the 2010 WHO Classification: a Single Institution Experience in Lebanon.

BACKGROUND: Gastro-entero-pancreatic neuroendocrine neoplasms (GEP-NEN) are relatively rare tumors, not equally distributed in the gastrointestinal system. In 2010, a revised version of the WHO classification of GEP-NENs was published. This study reports for the first time the distribution and characteristics of GEP-NENs in a Lebanese population. MATERIALS AND METHODS: This descriptive retrospective study concerns all the digestive neuroendocrine tumors with their characteristics diagnosed in Hotel Dieu de France in Beirut, Lebanon from 2001 to 2012, all the pathology reports being reanalyzed according to the latest WHO 2010 classification. The characteristics and features of GEP-NEN analyzed in this study were age, gender, grade and site. RESULTS: A total of 89 GEP-NENs were diagnosed, representing 28.2% of all neuroendocrine tumors. The mean age of GEP-NEN patients was 58.7 years and the M/F sex ratio was 1.2. The primary localization was as follows: 21.3%(19) pancreatic, 18% (16) gastric, 15.7% (14) duodenal, 11.2% (10) appendix, 10.1% (9) intestinal, 10.1% (9) colorectal (7.9% colonic and 2.2% rectal), 5.6% (4) hepatic, 2.2% (2) ampulla, 1.1% (1) esophageal and 7.9%(5) NOS digestive (metastatic with unknown primary). Of the 89 patients with GEP-NEN, 56.2% (50) were diagnosed as grade I, 11.2% (10) as grade II, 20.2% (18) as grade III and 12.4% (11) were considered as mixed adeno-neuroendocrine carcinomas (MANEC). CONCLUSIONS: This study, one of the rare examples based on the 2010 WHO classification of neuroendocrine tumors in the literature, indicates that in the Lebanese population, all duodenal and appendicular tumors are G1 and the majority of MANEC tumors are gastric and pancreatic tumors. Moreover, more duodenal tumors and fewer rectal tumors were encountered in our study compared to European reports.

Osteonecrosis of the jaw during biyearly treatment with zoledronic acid for aromatase inhibitor associated bone loss in early breast cancer: A literature review.

Osteonecrosis of the jaw (ONJ) is one of the most relevant and specific complication of biphosphonates. ONJ in patients receiving zoledronic acid every 3 to 4 weeks is frequently described, but the ONJ biyearly regimen used to reduce aromatase inhibitor associated bone loss (AIBL), is rarely reported. A literature review, focusing on the important trials using zoledronic acid to reduce AIBL, found that the mean risk of developing ONJ when zoledronic acid is used biyearly varies between 0.12% and 0.7%.

EGFR mutation status in Middle Eastern patients with non-squamous non-small cell lung carcinoma: A single institution experience.

BACKGROUND: Epidermal growth factor receptor (EGFR) represents an important molecular target in the treatment of lung adenocarcinomas; many anti-EGFR therapies are approved as first line and second-line treatment in patients having metastatic lung adenocarcinomas. The occurrence of this mutation varies in terms of race; it is around 10% in Caucasians and can reach 30% in Asians. Its prevalence in our Middle Eastern region is not well known. METHODS: Patients diagnosed with non-squamous non-small cell lung carcinomas between March 2013 and March 2015 were included. This study was conducted at Hôtel-Dieu de France University Hospital, a tertiary medical center in Lebanon. EFGR mutations were analyzed using real time PCR technique on the Rotor-Gene Q using Scorpions and ARMS technologies. The following data was collected: the patients' characteristics (age, gender, smoking status, stage), the samples' characteristics (histology subtype, TTF-1 and Napsin A immunostainings, the site and the adequacy and the type of the sample), and the mutational EGFR status (presence and type of mutation). These variables were analyzed using SPSS 20. RESULTS: 201 patients were included. The mean age was 65.2 years [31-87]; 40.2% were females. 78.1% of the included patients were smokers or ex-smokers. 12.9% of patients had a localized disease, 17.4% a locally advanced disease and 69.7% a metastatic disease. Adenocarcinoma was the main histologic subtype found in 90.5% of patients, followed by large cell carcinoma (3.5%), adenosquamous carcinoma (3.0%) and non-small cell carcinoma not otherwise specified (3.0%). 11.9% of patients had an EGFR mutation: 48% of them presented a deletion on exon 19, 40.0% a L858R mutation on exon 21, 4.0% a G719X mutation in exon 18, 4.0% an insertion in exon 20, and 4% a T790M mutation in exon 20. The presence of an EGFR mutation was significantly associated with the female gender (two-third) (p<0.05) and the non-smoking status (two-third) (p<0.05). CONCLUSIONS: The prevalence of EGFR mutation (11.9%) detected in our Lebanese population is similar to that observed in the Caucasian population. This mutation is also significantly more frequent in females and non-smokers.

Histologic Distribution of Pulmonary Tumors in Lebanon: A 5-Year Single Institution Experience.

BACKGROUND: To compare the current histologic distribution of lung cancer in Lebanon to the worldwide trends, according to the 2004 WHO Classification. MATERIALS AND METHODS: 1,760 patients with a pulmonary pathology examination at Hotel-Dieu de France University Hospital between July 2009 and July 2014 were included. RESULTS: Some 676 out of the total investigated patients (38.4%) had a lung tumor. In 665 (98.4%) the tumors were malignant, with a mean age at diagnosis of 63.8 years and a male/female (M/F) sex ratio of 1.7:1. Among the malignant tumors, 86.2% were epithelial tumors with a mean age at diagnosis of 64.8 years and an M/F sex ratio of 1.9. Other malignant tumors consisted of metastatic tumors (10.2%), lymphoproliferative tumors (2.1%) and mesenchymal tumors (1.5%). Most common carcinoma subtypes were adenocarcinoma (48.0%), squamous cell carcinoma (23.0%) and small cell carcinoma (13.3%). Carcinoid tumors were the only carcinoma subtype with an M/F sex ratio below 1 (0.7). Salivary gland tumors were the carcinoma with lowest mean age at diagnosis (45.5 years). CONCLUSIONS: The histologic distribution of lung tumors in Lebanon is similar to that in developed countries. We believe this resemblance is due to common smoking habits, known to be responsible for the increase of lung adenocarcinoma at the expense of other subtypes.

Distribution of testicular tumors in Lebanon: a single institution overview.

BACKGROUND: Testicular tumors constitute a rare type of cancer affecting adolescents and young adults with recent reports confirming an increase in incidence worldwide. The purpose of this study was to estimate the epidemiological characteristics and histological subtypes of testicular tumors in the Lebanese population according to the WHO classification of testicular and paratesticular tumors. MATERIALS AND METHODS: In this single institutional retrospective study, all patients diagnosed with a testicular tumor in Hotel-Dieu de France Hospital University in Beirut between 1992 and 2014 were enrolled. The age, subtype based on the 2004 WHO classification and body side of tumor were analyzed. RESULTS: A total of two hundred and forty-four (244) patients diagnosed with a testicular tumor in our institution were included in the study. Two hundred and one patients (82.4% of all testicular tumors) had germ cell tumors (TGCT). Among TGCT, 50% were seminomatous tumors, 48% non-seminomatous tumors (NST) and 2% were spermatocytic seminomas. The NST were further divided into mixed germ cell tumors (63.9%), embryonic carcinomas (18.6%), teratomas (15.4%) and yolk sac tumors (2.1%). The mean age for testicular tumors was 32 years. The mean age for germ cell tumors was 31 years and further subtypes such as seminomatous tumors had a mean age of 34 years, 28 years in non-seminomatous tumors and 56 years in spermatocytic seminoma. Patients with right testicular tumor were the predominant group with 55% of patients. Three patients (1.2%) presented with bilateral tumors. CONCLUSIONS: The distribution of different subgroups and the mean age for testicular tumors proved comparable to most countries of the world except for some Asian countries. Germ cell tumors are the most common subtype of testicular tumors with seminomatous tumors being slightly more prevalent than non-seminomatous tumors in Lebanese patients.

Type distribution of lymphomas in Lebanon: five-year single institution experience.

BACKGROUND: Lymphomas represent the fifth most frequent cancer in Lebanon. However, little is known concerning epidemiologic characteristics and distribution of lymphoid neoplasms according to the 2008 WHO classification. MATERIALS AND METHODS: We conducted a retrospective study of lymphoma cases diagnosed from 2008 till 2012 at Hotel-Dieu de France University Hospital. RESULTS: A total of 502 new cases of lymphoma were diagnosed at our institution during a five year period: 119 cases (24%) were Hodgkin lymphomas (HL) and 383 cases (76%) were non-Hodgkin lymphomas (NHL). HLs were equally distributed in both sexes with a mean age at diagnosis of 30 years. Among NHL, 87% (332 cases) were B cell lymphomas, 9% (34 cases) were T cell lymphomas and 4%(17 cases) were classified as precursor lymphoid neoplasms. Among B cell lymphomas, 44% (147 cases) were diffuse large B cell lymphomas (DLBCL), 20% (65 cases) follicular lymphomas and 8% (27 cases) mantle cell lymphomas. DLBCL were equally distributed in both sexes with a mean age of 58 years. Follicular lymphomas were characterized by a male predominance (57%) and a mean age of 60 years. Mantle cell lymphomas showed a pronounced male predominance (85%) with a mean age of 60 years in men and 70 years in women. Some 72% of patients having T cell lymphomas were men, with a mean age of 57 years in men and 45 years in women, while 65% of patients having precursor lymphoid neoplasms were women with a mean age of 22 years in women and 30 years in men. CONCLUSIONS: The lymphoma subtype distribution in Lebanon is unique when compared to other countries from around the world. In fact, Hodgkin and follicular lymphomas are more frequent than in most Far Eastern, European and American countries, while T-cell lymphomas and DLBCL are less frequent.