Human protozoa infection and dysplasia in ulcerative colitis: a neglected aspect in a prominent disease.

The chance of getting colorectal cancer (CRC) is higher in people with chronic ulcerative colitis (UC). The impact of parasitic infections on UC is underappreciated. The purpose of this study was to look into the effect of intestinal protozoal infections on the dysplastic changes generated by UC. The research included 152 adult patients with histologically confirmed UC and 152 healthy controls. Fecal samples were examined for the presence of parasites and fecal calprotectin (FC). The enzyme-linked immunosorbent assay measured serum anti-p53 antibodies (p53Abs) and metallothioneins (MTs). The advanced oxidation protein products (AOPPs) and reduced glutathione (GSH) levels were measured by a spectrophotometric method in all subjects. Serum C-reactive protein (CRP) and IL-6 were also measured. In addition, histopathological and immunohistochemical investigations of intestinal tissue were done. Our results exhibited significant increases in FC and CRP, IL-6, AOPPs, MTs, and p53Abs in ulcerative colitis patients with parasitic infections compared to those without parasites. In contrast, GSH levels showed a significant decrease in the same group compared with other groups. Histopathological and immunohistochemical assessments of intestinal tissue signified severe inflammation and strong expression of PD-L1 in patients with parasitic infections compared to others without parasitic infections. Our research indicated a greater frequency of intestinal protozoa in UC patients with elevated inflammatory and dysplastic biomarker levels. This suggests that these parasites may be involved in the etiology of chronic UC and the associated carcinogenetic process. This is the first report of a link between parasitic infections and dysplastic alterations in UC patients.

Clinicopathological significance of the immunologic signature (PDL1, FOXP3+ Tregs, TILs) in early stage triple-negative breast cancer treated with neoadjuvant chemotherapy.

Patients with breast cancer are appropriate candidates for neoadjuvant chemotherapy (NAC) to facilitate conservative surgery. The chemotherapeutic agents may exert their action by inducing the anti-tumor immune response. This study aimed to evaluate the tumor immune microenvironment including PD-L1, Foxp3+ Tregs, and TILs count in early stages TNBC patients (stage T1, T2) before and after neoadjuvant chemotherapy and their correlation with the clinical and pathological response. Fifty patients of TNBC patients were enrolled in this study; all of them received neoadjuvant chemotherapy. TILs count, Foxp3+ Tregs, and PD-L1 immunohistochemical expression were investigated in all cases before NAC and then evaluated in residual masses after surgery. Data on the clinical and pathological response to NAC were collected and then statistically analyzed. PDL1 expression was detected in 24% of all studied cases, all of them were node-positive (P < 0.002); while Foxp3+ Tregs expressed in 50% and high TILs in 28%. Pathological complete response (pCR) was achieved in 40% of patients and was associated with high TILs expression (P < 0.02) and absence of Foxp3+ Tregs and PDL1 (P < 0.001 for each). In conclusion, Pathologic complete response to NAC was associated with the immunological profile of TNBC. High TILs expression with concomitant decreased PD-L1 expression and low FOXP3+ Tregs is associated with favorable tumor prognosis. Combined therapeutic approaches aiming to PD-L1 block and Tregs depletion might improve treatment efficacy in TNBC.

Fatty acid synthase, Her2/neu, and E2F1 as prognostic markers of progression in non-muscle invasive bladder cancer.

Non-muscle-invasive bladder cancer (NMIBC) is a heterogeneous disease which has an unpredictable risk of progression to muscle-invasive bladder cancer (MIBC). The selection of patients who may benefit from early radical intervention is a challenge. To define the useful prognostic markers for progression, we analyzed the immunohistochemical expression of fatty acid synthase (FASN), Her2/neu, and E2F1 in 60 cases of NMIBC who underwent TURBT and adjuvant intravesical bacillus-Calmette-Guérin (BCG). Their predicting role for tumor recurrence, progression, recurrence-free survival (RFS) and progression-free survival (PFS) was analyzed. High FASN expression was observed in 56.7% (34/60) of NMIBC cases, and FASN expression was significantly associated with the tumor size, grade, and tumor stage (p = 0.003, p < 0.001, p < 0.0001 respectively). Positive Her2/neu was noted in 18.3% (11/60) of the cases, and its expression was significantly associated with the tumor size, histologic grade, and tumor stage (p = 0.001, p = 0.002, p = 0.011 respectively). High E2F1 expression was detected in 40% of the cases, and it was associated with tumor size, histologic grade, and tumor stage (p < 0.001 for each). Analysis of follow-up period revealed that NMIBC with high FASN, positive Her2/neu, and high E2F1 expression exhibited a potent relation with tumor progression, shorter RFS, and poor PFS. Conclusions: High FASN, Her2/neu, and E2F1 are considered as adverse prognostic factors of tumor recurrence and progression in NMIBC and these patients should be followed carefully. Therefore, we suggest that FASN, Her2/neu, and E2F1 should be considered and evaluated during the selection of the appropriate management strategy for NMIBC patients.

Molecular factors regulating E-cadherin expression in urothelial bladder cancer and their correlations with the clinicopathological features.

This study aimed to assess the expression of S100A4, Twist and E-cadherin (mRNA and protein) in urothelial bladder cancer, investigate the correlation between them and evaluate their association with the clinicopathological features of the disease. The study included 54 patients diagnosed as urothelial bladder cancer of different stages and grades. The expression levels of S100A4, Twist and E-cadherin (mRNA and protein) in tissue samples were determined by quantitative RT-PCR and immunohistochemistry. The expression of S100A4 and Twist was significantly upregulated while E- cadherin was significantly downregulated in urothelial bladder cancer tissues compared to the adjacent surrounding normal bladder tissues at both mRNA and protein levels (p < 0.001). Expression levels of S100A4 and Twist were significantly higher in recurrent tumor than in non-recurrent tumors (p < 0.001) while the expression level of E-cadherin was significantly lower in recurrent tumors than in non-recurrent tumors at both mRNA and protein levels (p < 0.001). There was a significant positive correlation between S100A4 and Twist expressions (r = 0.875, p < 0.001) while significant negative correlations were found between E- cadherin and S100A4 expressions(r=- 0.803, p < 0.001) and between E-cadherin and Twist (r = -0.809, p < 0.001). Up-regulation of S100A4 and Twist and down-regulation of E-cadherin in urothelial bladder cancer tissues compared to adjacent normal tissues were observed. There was a significant negative correlation between S100A4 and E- cadherin and between E- cadherin and Twist expression. However, there was a significant positive correlation between S100A4 and Twist expressions. Furthermore, the alterations in the gene expression were associated with disease stage and grade.

Prognostic impact of EGFR and cytokeratin 5/6 immunohistochemical expression in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) has an aggressive behavior and limited therapeutic options due to lack of targeted therapy. We aimed in this study to assess the immunohistochemical expression of EGFR and cytokeratin 5/6 and their ability to predict survival and response to neoadjuvant chemotherapy (NAC) among triple-negative breast cancer patients. Thirty-five cases with TNBC were studied by immunohistochemistry for EGFR and CK5/6 expression. Data on overall survival (OS), disease-free survival (DFS) and response to NAC were collected. The resulted data were statistically analyzed. Invasive carcinoma of no special type (NST) was the predominant histopathological type (80%). The commonest histological grade was grade II-III (88.6%). About 57.1% of TNBC cases were CK5/6-positive, and 71.4% were EGFR-positive. EGFR expression showed a significant association with tumor grade and axillary lymph node metastasis (p=0.006, 0.016 respectively). EGFR expression was related to unfavorable response to NAC (p=0.036), poor OS (p=0.002) and poor DFS (p=0.003). CK5/6 expression showed a significant association with tumor grade, unfavorable response to NAC, poor OS & DFS (p=0.007, 0.048, <0.001, 0.043 respectively). Immunohistochemical expression of EGFR and/or CK5/6 showed a high significant association with an unfavorable response to NAC, poor DFS &OS (p=0.010, 0.012, 0.030 respectively). CONCLUSIONS: EGFR and CK5/6 are adverse prognostic markers in TNBC. EGFR and CK5/6 expression could serve as biomarkers for identifying TNBC patients with poor survival that are unlikely to benefit from neoadjuvant chemotherapy. So, targeted therapy against EGFR may be a hopeful therapy for TNBC with NAC resistance.

Parthenolide repressed endometriosis induced surgically in rats: Role of PTEN/PI3Kinase/AKT/GSK-3ß/ß-catenin signaling in inhibition of epithelial mesenchymal transition.

AIM: PI3K/AKT/GSK-3ß/ß-catenin signaling pathway is a triggering factor for epithelial to mesenchymal transition (EMT) which plays a pivotal role in the pathogenesis of endometriosis. Parthenolide is a sesquiterpene lactone extract that has anti-inflammatory, analgesic and anticancer properties. Hence, we investigated the effect of parthenolide against EMT in the endometrial tissue implants and immortalized epithelial endometriotic cell lines 12Z. MAIN METHODS: Twenty- four female Rats with surgically induced endometriosis were treated with parthenolide (2, 4 mg/kg), for 4 weeks. Endometriotic cell line 12Z was used to identify the effect of parthenolide on the wound healing, cellular migration and invasion properties of endometriotic cells. KEY FINDINGS: Parthenolide decreased the endometriotic implant tissue expression of total PI3K, PI3K-p85, p-AKT, p/total AKT, p-GSK-3ß, P/total GSK-3ß, and nß-catenin, as well as increased E-cadherin and decreased vimentin mRNA expression. Parthenolide upregulated PTEN immunoreactivity as well as the endometriotic tissue caspase-3, caspase-9, BAX levels while reducing Bcl2 level. Additionally, parthenolide decreased endometriotic tissue implants surface area and histopathological score of the epithelial growth. SIGNIFICANCE: Our findings showed that parthenolide in a dose dependent manner inhibited PI3K/AKT/GSK-3ß/nß-catenin cascade via enhancement of PTEN with subsequent inhibition of EMT evidenced by elevation of the epithelial marker, E-cadherin and reduction of mesenchymal marker, vimentin, of the endometriotic implants in addition to reversal of invasion and migration properties of epithelial endometriotic cell lines. These findings provide a valuable therapeutic approach for treatment of endometriosis.

A Promising Diagnostic Role of Immunohistochemical Expression of Insulin-Like Growth Factor II mRNA Binding Protein 3 (IMP3) in Pancreatic Lesions Using Endoscopic Ultrasound-Guided Fine Needle Aspiration (EUS-FNA) Cytology.

BACKGROUND: Poor prognosis and short survival of patients harboring pancreatic cancer emerge how advanced disease it is. In a trial to achieve the earliest and most accurate diagnosis to manage this progressive disease, we proposed that using endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) with an adjuvant diagnostic immunohistochemical marker would give better diagnostic results. IMP3 has gained recently wide attention, as many studies found that IMP3 has not only diagnostic but also prognostic role in different types of malignancies. AIM OF THE STUDY: This prospective work is to assess the diagnostic role of EUS-FNA combined with the immunohistochemical expression of IMP3 on different benign and malignant pancreatic lesions. MATERIAL AND METHOD: The included pancreatic lesions (n = 140) were obtained by EUS-FNA technique and stained for IMP3 immunohistochemically. Paraffin blocks from patients who underwent excision (n = 92) or core biopsies (n = 48) were performed for confirming diagnosis. RESULTS: The combined method for diagnosis showed that IMP3 was positive in 78.7%, 91.7%, 100% PAC, mucinous neoplasm with high grade dysplasia, and IPMN with high grade dysplasia, respectively, while almost all benign lesions showed negative IMP3. Also, this method showed sensitivity (78.26%), specificity (95.83%), and accuracy (84.3%). CONCLUSION: EUS-FNA cytology with IMP3 could be a reliable diagnostic tool especially for assessment of malignant pancreatic lesions.

Predictive and Prognostic Value of Tumor- Infiltrating Lymphocytes for Pathological Response to Neoadjuvant Chemotherapy in Triple Negative Breast Cancer.

OBJECTIVES: Background: Tumor-infiltrating lymphocytes (TILs) reflect the antitumor response of the host. This study aimed to assess the value of TILs in predicting pathological response after neoadjuvant chemotherapy (NAC) and survival outcomes in patients with triple-negative breast cancer (TNBC). METHODS: A retrospective analysis conducted between February 2012 and December 2015. Patients with stage I, II, and III TNBC patients were enrolled. TILs were assessed in haematoxylin and eosin-stained sections from true cut needle biopsies before NAC. According to international TILs working group, we had three groups; low (0-10%), intermediate (11-59%), and high TILs (= 60%). RESULTS: A total of 159 patients was included, 56% were premenopausal and 76.1% were less than 60 years. The main bulk of patients had histological grade III, high Ki 67, and high TILs (74.2%, 84.3%, and 72.3%), respectively. The pre-treatment high TILs was significantly correlated with high Ki-67 (p = 0.001), pCR (p<0.001), and late relapse (p<0.001). Other clinico-pathological features such as age, menopausal status, tumor size, histological grade, lymph node involvement and lympho-vascular invasion weren't significantly correlated with TILs levels. 71.3% of enrolled patients having high TILs achieved pCR, vs 27.8% in the intermediate group and 30.8% in low group. After a median follow-up of 45.3 months, patients with high TILs were significantly associated with longer DFS and OS as compared to intermediate and low TILs (27.2 vs 15.9 vs11.4 months for DFS and 70.2 vs 34.3 vs 27.6 months for OS)p<0.001). CONCLUSIONS: Pre-treatment level of TILs had a predictive and prognostic value in TNBC patients receiving NAC. TILs may be integrated into the basic laboratory for TNBC prognostication as a credible biomarker.

Effect of spiramycin versus aminoguanidine and their combined use in experimental toxoplasmosis.

Toxoplasmosis is one of the widest spread parasitic infections which is caused by Toxoplasma gondii protozoon. Many experimental studies have evaluated the effect of aminoguanidine upon parasitic load and inflammatory process. However, few reports have illustrated the impact of combining aminoguanidine with spiramycin in the treatment of toxoplasmosis. Therefore, our study aimed to explore the possible effects of spiramycin used alone and combined with aminoguanidine against the avirulent (ME49) Toxoplasma gondii strain in experimental toxoplasmosis. Fifty-five Swiss albino mice were included in the study and were divided into five groups: (GI): non-infected control group; (GII): infected untreated control group; (GIII): infected- spiramycin treated group; (GIV): infected-aminoguanidine treated group; (GV): infected and received combination of spiramycin and aminoguanidine. Obtained results exhibited a significant increase in brain cysts numbers in aminoguanidine treated groups compared to infected untreated control groups. Histopathological studies denoted that combination between spiramycin and aminoguanidine improved the pathological features only in liver and heart tissues of the studied groups. Moreover, it was noticed that spiramycin administered alone had no effect on nitric oxide expression, whereas its combination with aminoguanidine had an inhibitory effect on inducible nitric oxide synthase enzyme in brain, liver and heart tissues of different study groups. In conclusion, the combination of spiramycin and aminoguanidine significantly reduced the parasitic burden, yet, it failed to resolve the pathological sequels in brain tissues of Toxoplasma gondii infected mice.

Neoadjuvant Chemotherapy in Triple Negative Breast Cancer: Correlation between Androgen Receptor Expression and Pathological Response.

BACKGROUND: There is growing evidence that the response to chemotherapy may be affected by Androgen Receptor (AR) expression suggesting that triple-negative breast cancers (TNBC) AR+ and quadruple negative breast cancer (QNBC) subtypes may have different diseases behavior. METHODOLOGY: We retrospectively estimated the predictive value of the AR expression in stage II and stage III TNBC patients treated with neoadjuvant chemotherapy (NAC) and correlated with the rate of pathological response (pCR). RESULTS: Of 89 TNBC patients, 29 patients (32.6%) were TNBC AR+ and 60 patients (67.4) were QNBC. Most of the patients were less than 60 years old. Of note, approximately 62% in the QNBC group were less than 40 years old compared with 39 % in the TNBC AR+ group. The Ki-67 expression was higher in the QNBC in comparison with TNBC AR+ being 86.7% and 65.5%, respectively. QNBC subgroup showed higher rates of pCR compared with TNBC; 60% and 24%, respectively. Higher Ki-67 expression, higher grade, and lymph node involvement were statistically significantly correlated with the rate of pCR in the QNBC group (p=0.02, p=0.04, and p=0.03, respectively). In contrast, no significant association was observed between pCR and clinical-pathological features in the TNBC AR+ group. CONCLUSION: Our results suggested that the AR expression in TNBC may be applied as a predictive marker for NAC. TNBC AR+ had a lower rate of pCR compared with QNBC, suggesting that this subtype may have a partial chemoresistance.

C-MYC and BCL2: Correlation between Protein Over-Expression and Gene Translocation and Impact on Outcome in Diffuse Large B Cell Lymphoma

Background: Due to lack of availability of gene expression profiling (GEP) for most developing countries and clinicians; the immunohistochemistry (IHC) is mostly used in the clinical application. The aim of our study is to check the possibility of using IHC to detect MYC and BCL2 in our patients with diffuse large B-cell lymphoma (DLBCL) instead of GEP to stratify them into high and low-risk groups. This will help in a proper treatment choice of subsequent improvement in the survival outcome. Method: During the study period, 90 DLBCL patients were eligible. MYC and BCL2 evaluated by IHC and gene rearrangement by real-time PCR (RT-PCR) and correlated with clinical-pathological features and survival. Results: Through IHC, the expression of MYC, BCL2, and double expression was detected in 35.6%, 46.7% and 30% of patients, respectively. While by RT-PCR, it was 4.53±0.74 for MYC compared with 2.18±0.78 for BCL-2. Most patients with BCL2+/MYC+; double-expressor and double-hit lymphomas (DEL and DHL) had high stage (III, IV), more extra-nodal involvement, (P value <0.001) and intermediate to high International Prognostic Index (IPI) risk profile (P-value <0.001). The median overall survival was 14 months and 6 months for DEL and DHL, respectively. While all patients with DHL died during the follow-up period, the median PFS were only 2 months for DEL. There was a statistically significant correlation between mRNA of MYC and BCL2 with their protein expression (p<0.001). Conclusion: Our results confirmed the unique characters and poor outcome associated with DEL and DHL mandated the need for more intense therapy and not the standard protocol. Moreover, the significant correlation between protein overexpression and gene rearrangement may open the door for the possibility to use IHC instead of RT-PCR in developing countries.

Up-regulated miR-221 expression as a molecular diagnostic marker in laryngeal squamous cell carcinoma and its correlation with Apaf-1 expression.

BACKGROUND: Despite the improvement in the diagnosis and the management of laryngeal squamous cell carcinoma (LSCC), many patients with advanced-stage have poor prognosis in the form of recurrence, metastasis or death. So, recognition of new molecular markers would facilitate the development of targeted therapies. OBJECTIVES: To investigate miR-221 expression in LSCC and its possible correlation to apoptotic protease activating factor-1 (Apaf-1). Also, we aimed to investigate the association between miR-221 and Apaf-1 expressions and the clinicopathological features of LSCC. METHODS: We investigated the expression of miR-221 (by qRT-PCR) and Apaf-1 (by qRT-PCR and immunohistochemistry) in primary LSCC and adjacent normal tissues. RESULTS: We found significant up-regulation of miR-221 and significant down-regulation of Apaf-1 expression in LSCC tissues compared to normal nearby laryngeal tissues. In addition, significant associations between up-regulated miR-221 and down-regulated Apaf-1 expressions and clinical stage and lymph node (LN) metastasis (P< 0.001 for each) were found. Furthermore, there was a negative correlation between miR-221 gene expression and Apaf-1 gene expression (r=-0.73, P< 0.001). CONCLUSION: miR-221 can be considered as a diagnostic marker in LSCC and Apaf-1 may be considered as a possible target of miR-221.

Prognostic Significance of Programmed Cell Death Ligand 1 (PD-L1), CD8+ Tumor-Infiltrating Lymphocytes and p53 in Non-Small Cell Lung Cancer: An Immunohistochemical Study.

OBJECTIVE: Programmed cell death ligand-1 interacts with the immune receptors on the surface of CD8+ tumor infiltrating lymphocytes and PD-1, thereby blocking its anti-tumor activity. Therapeutics suppression of this interaction will show a promise in the treatment of non-small cell lung cancer by restoring the functional anti-tumor T-cell activity. We aimed to evaluate the association between the immunohistochemical expression of PD-L1, stromal CD8+ tumor infiltrating lymphocytes and p53 with the clinicopathological characteristics, response to chemotherapy, progression-free-survival, and overall survival. MATERIAL AND METHOD: We examined the immunohistochemical expression of PD-L1, stromal CD8+ TILs, and p53 expression in 50 patients with advanced stage (III&IV) non-small cell lung cancer. RESULTS: PD-L1 was expressed in 56% of the studied cases. PD-L1 expression was related to unfavorable response to the therapy without significant difference. PD-L1 expression was significantly associated with disease progression, poor progression-free-survival & overall survival. CD8+ TILs were high in 32% of the cases. Tumors with high CD8+ TILs showed a partial response to therapy and had a better progression-free-survival and overall survival. p53 expressed in 82% of the studied cases. There was a significant negative association between PD-L1 and CD8+ TILs (p=0.009), while a non-significant association was found between p53 and PD-L1 (p=0.183). CONCLUSION: PD-L1 overexpression is an unfavorable prognostic marker, while the high CD8 + TILs is a good prognostic marker in non-small cell lung cancer. PD-L1 immunohistochemical assessment may be used for the selection of patients legible for treatment with anti-PD-L1 therapy.

Prognostic significance of the genetic and the immunohistochemical expression of epithelial-mesenchymal-related markers in colon cancer.

BACKGROUND: Epithelial-mesenchymal transition (EMT) is one of the main events in colorectal cancer (CRC) spread. Snail-1 is a zinc transcription factor that mediates EMT in tumor cells probably by down-regulation of E-cadherin and claudin-1. OBJECTIVES: To detect the expression of epithelial markers (claudin-1 and E-cadherin) and mesenchymal markers (snail-1 and vimentin) in primary cancer colon. Also, to select stage II cancer patients of a high risk that can benefit from postoperative adjuvant chemotherapy. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemical analysis were performed to investigate snail-1, claudin-1, E-cadherin and vimentin expressions at mRNA and protein levels in fresh tissues of cancer colon and normal colonic mucosa. The correlations between the expression of these markers and clinicopathological parameters were performed. RESULTS: Normal colonic mucosa revealed complete membranous expression of claudin-1, preserved E-cadherin and negative snail-1 and vimentin expressions. Compared to control, the expression of snail-1 and vimentin mRNA in cancer colon was significantly up-regulated while the expression of claudin-1 and E-cadherin mRNA was significantly down-regulated. These changes were significantly associated with stage and lymph node involvement at both mRNA and protein levels (p< 0.05). There were significant negative correlations between vimentin and each of E-cadherin and claudin-1 gene expression and between snail-1 and each of E-cadherin and claudin-1 gene expression. Moreover, these changes were independent predictors of recurrence of stage II cancer colon cases. CONCLUSION: There is a clinical significance of snail-1, claudin-1, E-cadherin and vimentin as possible markers for recognizing patients with lymph node involvement, advanced stage and high incidence of tumor recurrence in cancer colon.

IMMUNOGENICITY ASSESSMENT FOR LUNG-STAGE AND BIOMPHALARIA ALEXANDRINA COCKTAIIVACCINE IN SCHISTOSOMA MANSONI INFECTED MICE.

The availability of a new vaccine is usually needed as an additional component to chemotherapy for control of schistosomiasis. Different strategies of different types of vaccines were assessed to decrease morbidity but did not give the best protection. The study assessed the efficacy of BAAP, SLAP and their combined preparations together with BCG adjuvant as an effective anti-schistosomal vaccine. METHODOLOGY: Six groups of Swiss albino mice were used; (Gl) as a control, (G2) infected non immunized; (G3) infected and supported by Adj.; (G4) infected; vaccinated with BAAP and supported by Adj.; (G5) infected, vaccinated with SLAP and supported by Adj. and the target group (G6) infected, vaccinated with combined antigens (BAAP + SLAP) and supported by Adjuvant. Mice were sacrificed 8 weeks post infection for assessment the effect of our vaccine through parasitological, histopathological, serological and immunohisto- chemical study. The vaccination of mice with BAAP, SLAP and Adjuvant followed by challenge S. mansoni infection resulted in highest reduction percentages (92% & 86%) for mean numbers of adult burdens and fecal egg counts respectively,(82.4%, 81%) for granuloma number and diameter respectively compared with other groups. The improvement % of all measured enzymes was higher in G6 than other groups.IL1O was significantly increased in G6 than other groups; also, TNF was significantly decreased in G6 than other groups.