Biospecimen Repositories in Low- and Middle-Income Countries: Insights From an American University of Beirut and Memorial Sloan Kettering Collaboration.

PURPOSE: Biobanking helps source tissue and blood for studying cancer genomics. Access to biorepository resources in low- and middle-income countries is lacking. Memorial Sloan Kettering Cancer Center (MSK) and the American University of Beirut (AUB) established a joint tissue biorepository at AUB in Beirut, Lebanon. The undertaking encountered key challenges that were unanticipated. MATERIALS AND METHODS: Patients age 18 years or older were eligible for enrollment at AUB. After consent, biospecimens were obtained at the time of routine diagnostic and/or therapeutic interventions. Both normal and abnormal tissue and solid and/or liquid specimens were collected from varied body sites. Early on, declining consent was frequently observed, and this was highlighted for investigation to understand potential participants reasoning. RESULTS: Of 850 patients approached, 704 (70.8%) elected to consent and 293 (29.5%) declined participation. The number of declined consents led to an amendment permitting the documentation of reasons for same. Of 100 potential participants who declined to consent and to whom outreach was undertaken, 63% indicated lack of research awareness and 27% deferral to their primary physician or family member. A financial gain for AUB was cited as concern by 5%, cultural boundaries in 4%, and 1% expressed concern about confidentiality. Of the patients who elected to consent, 682 biospecimens were procured. CONCLUSION: The AUB-MSK biospecimen repository has provided a unique resource for interrogation. Patient participation rate was high, and analyses of those who elected not to consent (29%) provide important insights into educational need and the local and cultural awareness and norms.

Novel therapeutic diiminoquinone exhibits anticancer effects on human colorectal cancer cells in two-dimensional and three-dimensional in vitro models.

BACKGROUND: Colorectal cancer (CRC) is the second leading cause of cancer-related mortality. Cancer stem cells (CSCs) in CRC, which are spared by many chemotherapeutics, have tumorigenic capacity and are believed to be the reason behind cancer relapse. So far, there have been no effective drugs to target colon CSCs. Diiminoquinone (DIQ) has shown promising effects on targeting colon cancer. However, there is limited research on the effects of DIQ on eradicating CSCs in CRC. AIM: To investigate the anticancer potential of DIQ on colon CSCs in two-dimensional (2D) and three-dimensional (3D) models using colonospheres and patient-derived organoids. METHODS: Various 2D methods have been used to assess the effect and the mechanism of DIQ on HCT116 and HT29 cell lines including cell proliferation and viability assays, migration and invasion assays, immunofluorescence staining, and flow cytometry. The potency of DIQ was also assessed in 3D culture using the sphere formation assay and colon cancer patient-derived organoid model. RESULTS: Our results showed that DIQ significantly inhibited cell proliferation, migration, and invasion in HCT116 and HT29 cell lines. DIQ treatment induced apoptosis along with an accumulation of HCT116 and HT29 cancer cells in the sub-G1 region and an increase in reactive oxygen species in both CRC cell lines. DIQ reduced sphere-forming and self-renewal ability of colon cancer HCT116 and HT29 stem/progenitor cells at sub-toxic doses of 1 µmol/L. Mechanistically, DIQ targets CSCs by downregulating the main components of stem cell-related -catenin, AKT, and ERK oncogenic signaling pathways. Potently, DIQ displayed a highly significant decrease in both the count and the size of the organoids derived from colon cancer patients as compared to control and 5-fluorouracil conditions. CONCLUSION: This study is the first documentation of the molecular mechanism of the novel anticancer therapeutic DIQ via targeting CSC, a promising compound that needs further investigation.

CD147 Promotes Tumorigenesis via Exosome-Mediated Signaling in Rhabdomyosarcoma.

Rhabdomyosarcoma (RMS) is an aggressive childhood soft-tissue tumor, with propensity for local invasion and distant metastasis. Exosomes are secreted vesicles that mediate paracrine signaling by delivering functional proteins and miRNA to recipient cells. The transmembrane protein CD147, also known as Basigin or EMMPRIN, is enriched in various tumor cells, as well as in tumor-derived exosomes, and has been correlated with poor prognosis in several types of cancer, but has not been previously investigated in RMS. We investigated the effects of CD147 on RMS cell biology and paracrine signaling, specifically its contribution to invasion and metastatic phenotype. CD147 downregulation diminishes RMS cell invasion and inhibits anchorage-independent growth in vitro. While treatment of normal fibroblasts with RMS-derived exosomes results in a significant increase in proliferation, migration, and invasion, these effects are reversed when using exosomes from CD147-downregulated RMS cells. In human RMS tissue, CD147 was expressed exclusively in metastatic tumors. Altogether, our results demonstrate that CD147 contributes to RMS tumor cell aggressiveness, and is involved in modulating the microenvironment through RMS-secreted exosomes. Targeted inhibition of CD147 reduces its expression levels within the isolated exosomes and reduces the capacity of these exosomes to enhance cellular invasive properties.

Thymoquinone Radiosensitizes Human Colorectal Cancer Cells in 2D and 3D Culture Models.

Resistance of cancer cells and normal tissue toxicity of ionizing radiation (IR) are known to limit the success of radiotherapy. There is growing interest in using IR with natural compounds to sensitize cancer cells and spare healthy tissues. Thymoquinone (TQ) was shown to radiosensitize several cancers, yet no studies have investigated its radiosensitizing effects on colorectal cancer (CRC). Here, we combined TQ with IR and determined its effects in two-dimensional (2D) and three-dimensional (3D) culture models derived from HCT116 and HT29 CRC cells, and in patient-derived organoids (PDOs). TQ sensitized CRC cells to IR and reduced cell viability and clonogenic survival and was non-toxic to non-tumorigenic intestinal cells. TQ sensitizing effects were associated with G2/M arrest and DNA damage as well as changes in key signaling molecules involved in this process. Combining a low dose of TQ (3 µM) with IR (2 Gy) inhibited sphere formation by 100% at generation 5 and this was associated with inhibition of stemness and DNA repair. These doses also led to ~1.4- to ~3.4-fold decrease in organoid forming ability of PDOs. Our findings show that combining TQ and IR could be a promising therapeutic strategy for eradicating CRC cells.

Role of Ultrastructural Study in the Diagnosis of Medical Renal Diseases: A Tertiary Medical Center Experience and Review of the Literature.

The 20th century unfolded many mysteries regarding renal pathology with the advent of ancillary techniques such as immunofluorescence (IF) and electron microscopy (EM) studies. EM plays a major role in confirming the routine and IF findings or may uncover new and unsuspected features. The aim of the study is to elucidate the role of ultrastructural findings in patients with medical kidney diseases on whom kidney biopsy was performed at the American University of Beirut Medical Center, between November 2018 and June 2019. A total of 188 renal biopsies were examined during the study period. EM confirmed the light microscopy diagnosis in 54% of the cases while completely changed the diagnosis in 23% of the cases. In 23% of the sample, EM provided additional features and a secondary diagnosis. Our study emphasizes the important diagnostic role of EM and its significance, particularly in minimal change disease, basement membrane abnormalities, and glomerulopathies.

Establishment and characterization of prostate organoids from treatment-naïve patients with prostate cancer.

Three-dimensional (3D) organoid culture systems are emerging as potential reliable tools to investigate basic developmental processes of human disease, especially cancer. The present study used established and modified culture conditions to report successful generation and characterization of patient-derived organoids from fresh primary tissue specimens of patients with treatment-naïve prostate cancer (PCa). Fresh tissue specimens were collected, digested enzymatically and the resulting cell suspensions were plated in a 3D environment using Matrigel as an extracellular matrix. Previously established 12-factor medium for organoid culturing was modified to create a minimal 5-factor medium. Organoids and corresponding tissue specimens were characterized using transcriptomic analysis, immunofluorescent analysis, and immunohistochemistry. Furthermore, patient-derived organoids were used to assess the drug response. Treatment-naïve patient-derived PCa organoids were obtained from fresh radical prostatectomy specimens. These PCa organoids mimicked the heterogeneity of corresponding parental tumor tissue. Histopathological analysis demonstrated similar tissue architecture and cellular morphology, as well as consistent immunohistochemical marker expression. Also, the results confirmed the potential of organoids as an in vitro model to assess potential personalized treatment responses as there was a differential drug response between different patient samples. In conclusion, the present study investigated patient-derived organoids from a cohort of treatment-naïve patients. Derived organoids mimicked the histological features and prostate lineage profiles of their corresponding parental tissue and may present a potential model to predict patient-specific treatment response in a pre-clinical setting.

An Unusual Gastric Polyp: Brunneroma.

Brunneromas or polypoid hamartomas are benign lesions arising from Brunner glands. They are usually benign lesions with low potential for malignancy. They are usually located in the duodenum and manifest with different unspecific symptoms, such as abdominal pain, nausea, or bloating. Other more serious manifestations are also reported in the literature that are related to the size of the lesion. Usually, they are treated with endoscopic resection, with some lesions requiring surgical intervention. We present a case of a gastric antral polypoid lesion that was consistent with Brunneroma on histology.

Prevalence and characteristics of post-gastroscopy gastric cancer: A retrospective study from an academic medical center.

BACKGROUND AND STUDY AIMS: Gastric cancer is diagnosed by endoscopy but false negative rates of up to 10% in the west and 40% in Asia have been reported. In Lebanon, little is known about the rates of post-gastroscopy gastric cancer (PGGC), defined as the proportion of patients diagnosed with gastric cancer with a negative previous examination within 2 years of diagnosis. We aimed to examine the rate of PGGC and its risk factors, clinico-pathologic and endoscopic characteristics at a University medical Center. PATIENTS AND METHODS: Retrospective analysis of patients with histologically proven gastric malignancy over the last 14 years. Patients with history of upper endoscopy preceding the index diagnostic endoscopy by 6 to 24 months were included. RESULTS: 18,976 patients underwent upper endoscopy and gastric cancer was diagnosed in 323 (1.7%). Of those, only 4 (1.2%) had a preceding endoscopy within 6 to 24 months of diagnosis: 3 adenocarcinoma and one MALT lymphoma. Upon review of the initial endoscopy, a mucosal abnormality had been noted in all 4 patients and biopsies taken in 3 were negative for cancer. The mean time to cancer diagnosis was 8 months (range 6-13 months). CONCLUSION: A small proportion of gastric carcinomas are missed on endoscopy in this study. Patients with endoscopic evidence of mucosal abnormalities and negative biopsies should undergo repeat examination with multiple biopsies. Proper endoscopic technique, lesion recognition and adoption of performance improvement measures are important to optimize endoscopic practice.

Positive predictive value of fecal immunochemical test for high-risk colonic adenomas and carcinoma: A health maintenance organization cohort screening study in Lebanon.

BACKGROUND AND STUDY AIMS: Fecal Immunochemical Test (FIT) is one of the leading modalities for colorectal cancer screening. Studies show that FIT is highly sensitive for the detection of colorectal cancer (CRC) but not similarly accurate for detection of pre-cancerous advanced adenomas (AA). We studied the performance metrics of FIT for the detection of CRC and AA in ahealth maintenance organization (HMO) cohort screening program. PATIENTS AND METHODS: Retrospective cohort study of asymptomatic persons of screening age belonging to a HMO. Endoscopy and pathology reports of those who tested positive were used to calculate the positive predictive value (PPV) of FIT, and characterize endoscopic findings on colonoscopy. RESULTS: Between 1995 and 2017, 3000 persons had screening fecal occult testing as part of their Employee Health Care plan. Of those, 150 had a positive qualitative FIT (cutoff 10 Âµg hemoglobin/g feces). All underwentcolonoscopy, and median time to colonoscopy was 27 days. 4 (2.6%) had carcinoma(2 stage IIIA and 2 stage IIIB), 106 (70.6%) had adenomas of which 40 (26.6% of the total cohort) had advanced adenomas (≥1 cm, villous features, or high-grade dysplasia) giving a PPV for AA and carcinoma of 29% and 3% respectively. When stratified by age, the PPV of AA; carcinoma was [50-59 (21.7%; 0.0%)], [60-69 (14.6%; 4.2%)], [70-79 (42.6%; 2.1%)], [80-89 (33.3%; 11.1%)]. CONCLUSION: The performance characteristics of FIT testing are acceptable for population screening in resource-limited settings. The resultsof this study are helpful when discussing expectations prior to colonoscopy in people with positive FIT.

Extraskeletal Ewing sarcoma: Diagnosis, management and prognosis.

Extraskeletal Ewing sarcoma (EES) is a relatively uncommon primary tumor of the soft tissues, which accounts for 20-30% of all reported cases of ES. Being uncommon, all members of the ES family tumors are treated following the same general protocol of sarcoma tumors. The present review summarizes the diagnosis, management and prognosis of EES, focusing on the differences between the subtypes of ESS. The clinical features and imaging of EES are also discussed. Magnetic resonance imaging is the modality of choice for diagnostic imaging and local staging, while core-needle biopsy with pathological testing is used to obtain a definitive diagnosis. Although several oncology groups endorse that ES family of tumors should be treated with similar algorithm and protocols, some studies have demonstrated that surgery and radiotherapy may be used as a form of local control. However, further studies are required to conclude the optimum treatment option for EES.

Unique Presentation and Management Approach of Pleural Solitary Fibrous Tumor.

Solitary fibrous tumor (SFT) of the pleura is an uncommon tumor that is often discovered incidentally on a routine chest X-ray. We report a case of a young female with a large, sessile, hypervascularized SFT of the pleura presenting with cardiopulmonary shock to a rural hospital with limited therapeutic interventions. We propose, in this case report, a unique multidisciplinary approach for the management of such a critical patient and the safe resection of the tumor.

The Impact of Partial Weak Staining in Normal Breast Epithelium on the Reliability of Immunohistochemistry Results in HercepTest-positive Breast Cancer.

INTRODUCTION: Although normal epithelial cells do not show human epidermal growth factor receptor-2 (HER2) gene amplification and should lack membrane staining by HER2 immunohistochemistry (IHC), HER2 staining in benign breast epithelium is occasionally encountered. The significance of this occurrence has not yet been adequately studied, and its associated American Society of Clinical Oncology/College of American Pathologists recommendations are vague. Our objective is to assess the correlation between HER2 IHC 3+ breast cancer cases with normal epithelium staining (NES) and their corresponding fluorescence in situ hybridization (FISH) results, and to suggest recommendations for interpretation. MATERIALS AND METHODS: A total of 154 breast cancer cases with HER2 IHC 3+ were reviewed. NES, along with other clinicopathologic characteristics, were recorded. NES was scored as present or absent. All study cases were sent for FISH testing. All cases, and particularly those that showed false positivity for IHC (positive IHC, negative FISH) were examined for NES. RESULTS: Of the 154 cases, 146 cases were FISH-positive (94.8%) and 2 failed FISH testing (1.3%). Conversely, 22% (34/154) of the cases showed NES for HER2. Of these 34 cases, 23 (67%) were FISH-amplified, 9 (26%) were FISH not amplified, and 2 failed FISH testing. Notably, all of the false-positive (FISH-negative) breast cancer cases showed some degree of positivity in normal breast epithelium. CONCLUSIONS: Our findings, though descriptive, show a very strong association between NES and false-positive HER2 IHC. This confirms the need to carefully evaluate IHC-positive breast cancers for NES, and to have a low threshold for confirmatory testing by FISH.

EMT Markers in Locally-Advanced Prostate Cancer: Predicting Recurrence?

Background: Prostate cancer (PCa) is the second most frequent cause of cancer-related death in men worldwide. It is a heterogeneous disease at molecular and clinical levels which makes its prognosis and treatment outcome hard to predict. The epithelial-to-mesenchymal transition (EMT) marks a key step in the invasion and malignant progression of PCa. We sought to assess the co-expression of epithelial cytokeratin 8 (CK8) and mesenchymal vimentin (Vim) in locally-advanced PCa as indicators of EMT and consequently predictors of the progression status of the disease. Methods: Co-expression of CK8 and Vim was evaluated by immunofluorescence (IF) on paraffin-embedded tissue sections of 122 patients with PCa who underwent radical prostatectomies between 1998 and 2016 at the American University of Beirut Medical Center (AUBMC). EMT score was calculated accordingly and then correlated with the patients' clinicopathological parameters and PSA failure. Results: The co-expression of CK8/Vim (EMT score), was associated with increasing Gleason group. A highly significant linear association was detected wherein higher Gleason group was associated with higher mean EMT score. In addition, the median estimated biochemical recurrence-free survival for patients with < 25% EMT score was almost double that of patients with more than 25%. The validity of this score for prediction of prognosis was further demonstrated using cox regression model. Our data also confirmed that the EMT score can predict PSA failure irrespective of Gleason group, pathological stage, or surgical margins. Conclusion: This study suggests that assessment of molecular markers of EMT, particularly CK8 and Vim, in radical prostatectomy specimens, in addition to conventional clinicopathological prognostic parameters, can aid in the development of a novel system for predicting the prognosis of locally-advanced PCa.

Adrenal Cavernous Hemangioma: A Rarely Perceived Pathology-Case Illustration and Review of Literature.

Cavernous hemangiomas are endothelial tumors that rarely affect the adrenal glands. Most of these tumors remain silent and are incidentally found on abdominal imaging. Hardly ever, these tumors are endocrinologically functional. They may present as vague abdominal pain. Surgical resection remains the mainstay for large masses. In this paper, we are presenting a case of adrenal cavernous hemangioma in a 83-year-old male patient who initially presented for workup of vague abdominal and bilateral flank pain. A computed tomography scan of the abdomen showed an 8 cm right adrenal adenoma which was metabolically nonfunctional. The mass was completely resected through an open subcostal incision, with no encountered postoperative complications. A highlight of all published cases of adrenal hemangiomas since 1955 is also presented and reviewed.

Large Tick (Ixodes) Infestation of the Upper Eyelid Presenting as Eyelid Mass and Preseptal Cellulitis.

A child was referred for removal of an eyelid mass. She had preseptal cellulitis and a large tick deeply embedded in the tarsus of the upper eyelid necessitating antibiotic therapy and en-bloc excision of the tick with the attached eyelid portion. Large ticks that are embedded in the eyelid are best treated surgically with en-bloc excision of the tick and its attached lid. On the contrary, for small ticks involving the very superficial skin, fine-tipped tweezers can be used to grasp the insect. Ticks are vectors of several diseases like Lyme borreliosis, hence prophylactic antibiotic treatment and close observation are recommended.

Pulmonary Sarcoidosis Activation following Neoadjuvant Pembrolizumab plus Chemotherapy Combination Therapy in a Patient with Non-Small Cell Lung Cancer: A Case Report.

BACKGROUND: Pembrolizumab is a humanized monoclonal antibody which serves to enhance the antitumor immune response by targeting programmed cell death 1 receptor. The use of pembrolizumab plus carboplatin/pemetrexed combination therapy results in improvement in overall survival and progression-free survival rates for non-small cell lung cancer (NSCLC) patients as compared to chemotherapy alone. However, numerous immune-mediated toxicities of pembrolizumab have been reported. CASE PRESENTATION: We report the case of a 74-year-old male patient diagnosed with stage IIIA programmed death-ligand 1-positive non-small cell lung adenocarcinoma treated with 4 cycles of carboplatin/pemetrexed plus pembrolizumab combination therapy followed by 2 cycles of pembrolizumab treatment. Follow-up PET-CT scanning showed a very good response at the level of the tumor but new-onset activity in bilateral hilar and mediastinal lymph nodes. Biopsy of these lymph nodes revealed a benign pathology with noncaseating granulomas consistent with immune-mediated sarcoidosis. CONCLUSION: The pathogenesis of immunotherapy-induced sarcoidosis is not yet known but has been reported in different cancers and using different checkpoint inhibitors. To our knowledge, this case is the first in the literature displaying pulmonary sarcoidosis in a patient with NSCLC 4 months after having initiated chemotherapy plus pembrolizumab combination therapy.

Phosphorus Supplementation Mitigated Food Intake and Growth of Rats Fed a Low-Protein Diet.

Background: Low protein intake is associated with various negative health outcomes at any life stage. When diets do not contain sufficient protein, phosphorus availability is compromised because proteins are the major sources of phosphorus. However, whether mineral phosphorus supplementation mitigates this problem is unknown, to our knowledge. Objective: Our goal was to determine the impact of dietary phosphorus supplementation on food intake, weight gain, energy efficiency, body composition, blood metabolites, and liver histology in rats fed a low-protein diet for 9 wk. Methods: Forty-nine 6-wk-old male Sprague-Dawley rats were randomly allocated to 5 groups and consumed 5 isocaloric diets ad libitum that varied only in protein (egg white) and phosphorus concentrations for 9 wk. The control group received a 20% protein diet with 0.3% P (NP-0.3P). The 4 other groups were fed a low-protein (10%) diet with a phosphorus concentration of 0.015%, 0.056%, 0.1%, or 0.3% (LP-0.3P). The rats' weight, body and liver composition, and plasma biomarkers were then assessed. Results: The addition of phosphorus to the low-protein diet significantly increased food intake, weight gain, and energy efficiency, which were similar among the groups that received 0.3% P (LP-0.3P and NP-0.3P) regardless of dietary protein content. In addition, phosphorus supplementation of low-protein diets reduced plasma urea nitrogen and increased total body protein content (defatted). Changes in food intake and efficiency, body weight and composition, and plasma urea concentration were highly pronounced at a dietary phosphorus content <0.1%, which may represent a critical threshold. Conclusions: The addition of phosphorus to low-protein diets improved growth measures in rats, mainly as a result of enhanced energy efficiency. A dietary phosphorus concentration of 0.3% mitigated detrimental effects of low-protein diets on growth parameters.

Programmed Death-Ligand 1 Expression in Muscle-Invasive Bladder Cancer Cystectomy Specimens and Lymph Node Metastasis: A Reliable Treatment Selection Biomarker?

BACKGROUND: The programmed death-1 (PD-1) pathway negatively regulates T-cell activation and has an important role in regulating antitumor host immunity. Monoclonal antibodies directed against PD-1 or the PD-1 ligand (PD-L1) have shown activity in several tumor types with preliminary data suggesting a relationship between PD-L1 expression and response. The aim of this study was to establish the frequency of PD-L1 expression in muscle-invasive bladder cancer and associated lymph node metastasis using immunohistochemistry and to investigate the feasibility of using PD-L1 expression as a biomarker to select patients for PD-1-directed therapy. PATIENTS AND METHODS: Cases of radical cystectomy for muscle-invasive bladder cancer with no exposure to previous chemotherapy were identified and representative slides from archived paraffin-embedded blocks stained with anti-PD-L1 antibody (5H1 clone) were identified. PD-L1 positivity was defined by a 5% expression threshold. RESULTS: Fifty-two radical cystectomy specimens were reviewed. PD-L1 was overexpressed in the tumor cells of 5/52 (9.6%) of cystectomy specimens in this cohort with 17/52 (32.7%) of cases showing PD-L1 overexpression in tumor-infiltrating immune cells. Discordance was observed between PD-L1 expression in lymph node metastasis and the primary tumor. CONCLUSION: Standard assays for PD-L1 expression have yet to be established. The observation of discordance between PD-L1 expression in metastatic sites and primary tumors suggests that prospective biomarker studies should aim to acquire material immediately before treatment initiation rather than archived tissue from resected specimens that might not reflect the current immune-active microenvironment.

p53 Restoration in Induction and Maintenance of Senescence: Differential Effects in Premalignant and Malignant Tumor Cells.

The restoration of p53 has been suggested as a therapeutic approach in tumors. However, the timing of p53 restoration in relation to its efficacy during tumor progression still is unclear. We now show that the restoration of p53 in murine premalignant proliferating pineal lesions resulted in cellular senescence, while p53 restoration in invasive pineal tumors did not. The effectiveness of p53 restoration was not dependent on p19(Arf) expression but showed an inverse correlation with Mdm2 expression. In tumor cells, p53 restoration became effective when paired with either DNA-damaging therapy or with nutlin, an inhibitor of p53-Mdm2 interaction. Interestingly, the inactivation of p53 after senescence resulted in reentry into the cell cycle and rapid tumor progression. The evaluation of a panel of human supratentorial primitive neuroectodermal tumors (sPNET) showed low activity of the p53 pathway. Together, these data suggest that the restoration of the p53 pathway has different effects in premalignant versus invasive pineal tumors, and that p53 activation needs to be continually sustained, as reversion from senescence occurs rapidly with aggressive tumor growth when p53 is lost again. Finally, p53 restoration approaches may be worth exploring in sPNET, where the p53 gene is intact but the pathway is inactive in the majority of examined tumors.