Deleterious and ethnic-related BRCA1/2 mutations in tissue and blood of Egyptian colorectal cancer patients and its correlation with human papillomavirus.

This study aimed to identify BRCA1/2 mutational patterns in the tissue and blood of Egyptian colorectal cancer (CRC) patients and to study the possible correlation of this mutational pattern with Human papillomavirus (HPV) infection. Eighty-two colonoscopic biopsies and forty-six blood samples were collected from Egyptian CRC patients, as well as blood samples of age and sex-matched healthy controls (n = 43) were enrolled. The libraries were performed using Qiaseq Human BRCA1 and BRCA2 targeted DNA panel and sequenced via Ion proton sequencer. Also, the CRC tissues were subjected to conventional PCR targeting the HPV Late 1 (L1) region. Our analysis revealed that the BRCA-DNA damage pathway had been altered in more than 65% of the CRC patients. Comparing tissue and blood samples from CRC patients, 25 somatic mutations were found exclusively in tissue, while 41 germline mutations were found exclusively in blood. Additionally, we identified 23 shared BRCA1/2 pathogenic (PVs) mutations in both blood and tissue samples, with a significantly higher frequency in blood samples compared to tissue samples. The most affected exon in BRCA1 was exon 10, while the most affected exons in BRCA2 were 11, 14, 18, 24, and 27 exons. Notably, we revealed an ethnic-related cluster of polymorphism variants in our population closely related to South Asian and African ethnicities. Novel PVs were identified and submitted to the ClinVar database. HPV was found in 23.8% of the CRC tissues, and 54% of HPV-positive cases had somatic BRCA1/2 PVs. The results of this research point to a possible connection between infection with HPV and BRCA1/2 mutations in the occurrence of colorectal cancer in the Egyptian population, which has a mixed ethnic background. Our data also indicate that liquid biopsy (blood samples) may be more representative than tissue samples for detecting BRCA1/2 mutations. These findings may have implications for cancer screening and the development of personalized, targeted therapies, such as PARP inhibitors, which can effectively target BRCA1/2 mutations.

Multigene Panel Sequencing Reveals Cancer-Specific and Common Somatic Mutations in Colorectal Cancer Patients: An Egyptian Experience.

This study aims at identifying common pathogenic somatic mutations at different stages of colorectal carcinogenesis in Egyptian patients. Our cohort included colonoscopic biopsies collected from 120 patients: 20 biopsies from patients with inflammatory bowel disease, 38 from colonic polyp patients, and 62 from patients with colorectal cancer. On top of this, the cohort included 20 biopsies from patients with non-specific mild to moderated colitis. Targeted DNA sequencing using a customized gene panel of 96 colorectal related genes running on the Ion Torrent NGS technology was used to process the samples. Our results revealed that 69% of all cases harbored at least one somatic mutation. Fifty-seven genes were found to carry 232 somatic non-synonymous variants. The most frequently pathogenic somatic mutations were localized in TP53, APC, KRAS, and PIK3CA. In total, 16 somatic mutations were detected in the CRC group and in either the IBD or CP group. In addition, our data showed that 51% of total somatic variants were CRC-specific variants. The average number of CRC-specific variants per sample is 2.4. The top genes carrying CRC-specific mutations are APC, TP53, PIK3CA, FBXW7, ATM, and SMAD4. It seems obvious that TP53 and APC genes were the most affected genes with somatic mutations in all groups. Of interest, 85% and 28% of the APC and TP53 deleterious somatic mutations were located in Exon 14 and Exon 3, respectively. Besides, 37% and 28% of the total somatic mutations identified in APC and TP53 were CRC-specific variants, respectively. Moreover, we identified that, in 29 somatic mutations in 21 genes, their association with CRC patients was unprecedented. Ten detected variants were likely to be novel: six in PIK3CA and four variants in FBXW7. The detected P53, Wnt/ßcatenin, Angiogenesis, EGFR, TGF-ß and Interleukin signaling pathways were the most altered pathways in 22%, 16%, 12%, 10%, 9% and 9% of the CRC patients, respectively. These results would contribute to a better understanding of the colorectal cancer and in introducing personalized therapies for Egyptian CRC patients.

Resource Oriented Decision Making for Treatment of Metastatic Colorectal Cancer (mCRC) in a Lower-Middle Income Country: Egyptian Foundation of Medical Sciences (EFMS) Consensus Recommendations 2020.

PURPOSE: Colorectal cancer (CRC) is the third most commonly diagnosed cancer worldwide and the second cause of cancer related mortality. Treatment options for patients with metastatic CRC (mCRC) expanded during the last two decades, with introduction of new chemotherapeutic and targeted agents. Egypt is a lower middle-income country; Egyptian health care system is fragmented with wide diversity in drug availability and reimbursement policies across different health care providing facilities. We report the results of consensus recommendations for treatment of patients with metastatic colorectal cancer developed by Egyptian Foundation of Medical Sciences (EFMS), aiming to harmonize clinical practice through structured expert consensus-based recommendations consistent with the national status. EFMS recommendations could be utilized in other countries with similar economic status. METHODS: EFMS recommendations were developed using a modified Delphi process, with three rounds of voting till the final recommendations were approved. A non-systematic review of literature was conducted before generating the provisional statements. Content experts were asked to vote on some recommendations in two different resource groups (restricted resources and non-restricted resources). External review board of experts from a low income and lower-middle countries voted on the applicability of EFMS recommendations in their countries. RESULTS: The current recommendations highlighted the discrepancy in health care between restricted and non-restricted resources with expected survival loss and quality of life deterioration. Access to targeted agents in first line is very limited in governmental institutions, and no access to agents approved for third line in patients who failed oxaliplatin and irinotecan containing regimens for patients treated in restricted resource settings. CONCLUSION: Management of mCRC in developing countries is a challenge. The currently available resource-stratified guidelines developed by international cancer societies represent a valuable decision-making tool, adaptation to national status in each country based on healthcare system status is required.

Laparoscopic gastrectomy with D2 lymphadenectomy for gastric cancer: initial Egyptian experience at the National Cancer Institute.

BACKGROUND: Laparoscopic gastrectomy has been used as a superior alternative to open gastrectomy for the treatment of early gastric cancer. However, the application of laparoscopic D2 lymphadenectomy remains controversial. This study aimed to evaluate the feasibility and outcomes of laparoscopic gastrectomy with D2 lymphadenectomy for gastric cancer. RESULTS: Between May 2016 and May 2018, twenty-five consecutive patients with gastric cancer underwent laparoscopic D2 gastrectomy: eighteen patients (72%) underwent distal gastrectomy, four patients (16%) underwent total gastrectomy, and three patients (12%) underwent proximal gastrectomy. The median number of lymph nodes retrieved was 18 (5-35). A positive proximal margin was detected in 2 patients (8%). The median operative time and amount of blood loss were 240 min (200-330) and 250 ml (200-450), respectively. Conversion to an open procedure was performed in seven patients (28%). The median hospital stay period was 8 days (6-30), and the median time to start oral fluids was 4 days (3-30). Postoperative complications were detected in 4 patients (16%). There were two cases of mortality (8%) in the postoperative period, and two patients required reoperation (8%). CONCLUSIONS: Laparoscopic gastrectomy with D2 lymphadenectomy can be carried out safely and in accordance with oncologic principles.

Removal of urea from dilute streams using RVC/nano-NiOx-modified electrode.

Reticulated vitreous carbon (RVC), a high surface area electrode (40 cm2/cm3), has been modified with nickel oxide nanoparticles (nano-NiOx) and used for electrochemical oxidation of urea from alkaline solution. For the cyclic voltammetry measurements, the used dimensions are 0.8 cm × 0.8 cm × 0.3 cm. The purpose was to offer high specific surface area using a porous open network structure to accelerate the electrochemical conversion. NiOx nanoparticles have been synthesized via an electrochemical route at some experimental conditions. The morphological, structural, and electrochemical properties of the RVC/nano-NiOx are characterized by using scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), cyclic voltammetry (CV), and potentiostatic measurements. The fabricated electrode, RVC/nano-NiOx, demonstrates high electrocatalytic activity towards urea oxidation in an alkaline electrolyte. The onset potential of the RVC/nano-NiOx compared to that of the planar GC/NiOx is shifted to more negative value with higher specific activity. The different loadings of the NiOx have a substantial influence on the conversion of urea which has been evaluated from concentration-time curves. The urea concentration decreases with time to a limit dependent on the loading extent. Maximum conversion is obtained at 0.86 mg of NiOx per cm3 of the RVC matrix.

Perineal reconstruction using pedicled vertical rectus abdominis myocutaneous flap (VRAM).

BACKGROUND: The perineal wound is a major source of morbidity after abdominoperineal resection (APR) for rectal cancer, particularly after preoperative irradiation. Myocutaneous flap reconstruction is a viable alternative to primary perineal wound repair providing well-vascularized non-irradiated tissue to fill the dead space, and improve wound healing. PATIENTS AND METHODS: Sixty patients with low rectal cancer receiving neoadjuvant radiotherapy were prospectively randomized to either primary perineal wound closure (Group B) or VRAM perineal reconstruction (Group A). The study was carried out in the surgical department, National Cancer Institute (NCI), Cairo University over a period of 4 years from August 2008 to July 2012. All patients were evaluated as regards incidence of perineal wound complications, abdominal wall complications, and added co-morbidity. RESULTS: Perineal wound complications were observed in 5 patients in group A (17.2%) and in 14 patients in group B (46.4%) (P = 0.015). There was no significant difference in the incidence of abdominal wound morbidity between both groups. Operative blood loss and time were lower in group B. CONCLUSION: The vertical rectus myocutaneous flap (VRAM) is a relatively simple and reliable technique for perineal wound reconstruction after abdominoperineal resection.

Biomimetic coating of bisphosphonate incorporated CDHA on Ti6Al4V.

Bi-functional coatings of carbonated calcium deficient hydroxyapatite (CDHA) on Ti alloys were developed by using a biomimetic coating process. The bi-functionality was achieved by loading alendonate sodium (AS), an approved bisphosphonate drug used for the treatment of osteoporosis, into the inner layers of CDHA coatings. Three possible methods of loading AS into CDHA coatings were systematically studied and compared. The results indicated that the co-precipitation method had greater benefits and can modify the release profile of AS by incorporating AS in the inner layers of the coatings. As a preliminary study, the influences of applied AS dosage to CDHA coatings were evaluated using XRD and SEM. In vitro tests indicated that the AS content on CDHA coatings played a significant role, and optimum AS content in local area is beneficial for osteoblast cells proliferation. It is expected that the CDHA-AS coatings via the co-precipitation approach have potential for bone tissue engineering applications.

Hydrolysis of monetite/chitosan composites in α-MEM and SBF solutions.

There are two objectives of this work. The first objective is to study the hydrolysis behavior of monetite cements formed in the presence and absence of the chitosan in cell culture media (α-MEM) and simulated body fluid (SBF) solutions at 37°C. During hydrolysis, monetite transformed to carbonated apatite. Therefore, the second objective is to examine how addition of chitosan affects on the formation of carbonated apatite phases. The changes in the phase structure of monetite after hydrolysis reactions were characterized using XRD, FTIR and SEM. Pure monetite and monetite/chitosan composite were soaked in α-MEM and SBF solution for 4 and 7 days. In α-MEM solution, the monetite particles started to transform into carbonated apatite with a slow rate. However, in SBF, the rate of monetite transformation to carbonated apatite was more rapid. The presence of the chitosan had no significant effect on the precipitation of carbonated apatite on the monetite particles.

Fabrication of novel PLA/CDHA bionanocomposite fibers for tissue engineering applications via electrospinning.

The main theme here is to fabricate PLA (poly lactic-acid)/CDHA (carbonated calcium deficient hydroxyapatite) bionanocomposites, where both the constituents are biocompatible and biodegradable with one dimension in nanometer scale. Such materials are important in tissue engineering applications. The bionanocomposite fibers were fabricated via electrospinning. There are two important signatures of this paper. First, CDHA, rather than HA, is added to PLA as the second phase. As opposed to HA, CDHA mimics the bone mineral composition better and is biodegradable. Therefore, PLA/CDHA fibers should have better biodegradability while maintaining a physiological pH during degradation. To the best of our knowledge, this is the first attempt of electrospinning of such a composite. Second, the CDHA nanoparticles were synthesized using the benign low temperature biomimetic technique, the only route available for the retention of carbonate ions in the HA lattice. The structural properties, degradation behavior, bioactivity, cell adhesion, and growth capability of as-fabricated PLA/CDHA bionanocomposites were investigated. The results show that the incorporation of CDHA decreased PLA fiber diameters, accelerated PLA degradation, buffered pH decrease caused by PLA degradation, improved the bioactivity and biocompatibility of the scaffold. These results prove that PLA/CDHA bionanocomposites have the potential in tissue regeneration applications.

Formation of calcium deficient HAp/collagen composites by hydrolysis of alpha-TCP.

Bone-like composites containing calcium deficient hydroxyapatite (CDHAp) were formed by the hydrolysis of alpha-tricalcium phosphate (alpha-TCP) in the presence of type I collagen. CDHAp-collagen composites were synthesized using two techniques. In one technique alpha-TCP was mixed with non-milled (as-received) collagen prior to the addition of the aqueous solution. In the second, the collagen was milled with alpha-TCP in heptane at room temperature prior to its conversion to CDHAp. The effect of milling strongly facilitates the formation of CDHAp at physiological temperature. The proportion of milled collagen between 5 and 20 wt% present in the alpha-TCP/collagen composites has no significant effect on the rate of CDHAp formation. Variations in pH and in calcium and phosphate concentrations were determined as a function of collagen processing and variations specific to the presence of collagen were discerned. Compared to CDHAp or to composites containing non-milled collagen, diametrical and compressive strengths of CDHAp increased in the presence of milled collagen. Lack of collagen dispersion and incomplete formation of CDHAp during 48 h were the bases for reduced strengths of composites containing non-milled collagen.

Formation of composites comprised of calcium deficient HAp and cross-linked gelatin.

Cross-linked gelatin/calcium deficient hydroxyapatite (CDHAp) composites were prepared at or near physiologic temperature. alpha-tricalcium phosphate (alpha-TCP) or a mixture of tetracalcium phosphate and dicalcium phosphate were used as CDHAp precursors. Glutaraldehyde was used to cross-link the gelatin fibers. CDHAp formation reached completion in the presence of cross-linked gelatin fibers. Effects of cross-linking concentrations, proportions of gelatin fiber, molecular weight of gelatin and the temperature of the hydration reaction on the formation of CDHAp were studied. Cross-linked gelatin acts as a nucleating agent for CDHAp formation. The pH variations during CDHAp formation are lower at the onset of the reactions in the presence of cross-linked gelatin fibers. Although cross-linked gelatin fibers enhance CDHAp formation their composites have low mechanical strengths. Swelling of gelatin appears to be a major factor that limits the strengths of the CDHAp/cross-linked gelatin composites.

Clinicopathologic and prognostic significance of overexpression of her-2/neu and p53 oncoproteins in gastric carcinoma using tissue microarray.

BACKGROUND: The aim of the study was to verify the frequency of the immunohistochemical overexpression of her-2/neu and p53 in gastric carcinoma and their relation to the other clinico-pathological features and the impact on survival rates. PATIENTS AND METHODS: A total of 93 patients of gastric carcinoma, who had a potential curative surgery in the period from 2001-2007 and with representative paraffin blocks, and sufficient follow-up data were included in this study. They were arrayed and evaluated for protein marker overexpression using tissue microarray (TMA). Patients, tumor and treatment characteristics were collected from the patients' files. The possible prognostic significance of p53 and her-2/neu over expression and different clinico-pathological features on survival rates were explored. RESULTS: Twenty four (25.8%) cases were her-2/neu and p53 positive. None of the examined clinico-pathologic factors had a significant relation to her-2/neu overexpression. p53 was overexpressed in intestinal type, 14/34 (41.2%), more than in diffuse type, 10/59 (16.9%), (p= 0.01). There was no relation between the overexpression of p53 and her-2/neu. The median survival period was 17.7 months. The survival rates at 12 months were 64.2%, 52.2%, 55.6% and 45.0% for overall (OS), local control (LC), metastasis free survival (MFS) and disease free survival (DFS) rates, respectively. Patients with advanced stages had a significantly lower OS and MFS. Age above 57 years was associated with significantly lower OS, LC, MFS and DFS. Patients who received radiotherapy had significantly higher OS, LC, MFS and DFS. None of the survival rates had been affected by the overexpression of p53, or her-2/neu. CONCLUSION: Although, this study failed to show any prognostic effect of p53 and her-2/neu on survival rates, we may suggest that p53 overexpression may play a role in the pathogenesis of intestinal gastric adenocarcinoma. It could also demonstrate the significantly improved survival rates with adjuvant chemoradiation. Also, TMA is a useful technique for rapid identification of protein expression profiles using minimal samples from archived tissues.

Clinico-Pathological and Prognostic Significance of p53, Bcl-2 and Her-2/neu Protein Markers in Colorectal Cancer Using Tissue Microarray.

BACKGROUND: The prognostic role of her-2/neu has been established in breast cancer but remains controversial in colorectal cancer (CRC). Widespread genetic mutations in colorectal carcinogenesis exist on chromosome 17. Her- 2/neu gene and the tumor suppressor gene p53 are both located on this chromosome. Bcl-2 protein prolongs survival of a variety of cells by blocking apoptosis. The aim of this study is to evaluate the relationship between the overexpression of p53, bcl-2 and her-2/neu protein markers and the clinico-pathologic characteristics of CRC, and their influence on survival rates. PATIENTS AND METHODS: One hundred and four cases of CRC had paraffin blocks with representative tissue, and sufficient follow-up data. They were arrayed and evaluated for protein marker expression using tissue microarray (TMA). RESULTS: Ten (9.6%), 35 (33.7%) and 27 (26%) of the patients were her-2/neu, p53 and bcl-2 positive, respectively. None of the examined clinico-pathologic factors had a significant relation with her-2/neu overexpression. Patients with +ve bcl-2 had a significantly higher mean age (52.4-/+13.3years) compared to 45.4-/+14.4 years for bcl-2 negative patients, p=0.03. Positive p53 was overexpressed in 20/44 (45.5%), 6/17 (35%), 9/43 (21%) cases of the colon, recto-sigmoid, and rectal sites, respectively, p=0.05. For the whole population, p53 overexpression had a significantly lower disease-free survival (DFS). For patients with Dukes' stage B, overexpression of p53 protein had a significant reduced overall survival (OS) p=0.04, metastasis free survival (MFS) p=0.004, and DFS p=0.01 rates. Expression of bcl-2 had a significantly better MFS p=0.001, while her-2/neu overexpression worsened the OS rate significantly, p=0.04. CONCLUSION: This study recommends the application of TMA technique for its economic importance and reliable quick throughput. The results from this study also suggest that overexpression of p53, bcl-2, and her-2/neu protein markers appear to be useful in selecting a group of CRC patients with a worse prognosis and constitute potential candidates for adjuvant therapy. Key Words: Her-2/neu , p53 , Bcl-2 , Colorectal cancer , Tissue microarray , Prognostic factors.