http://hdl.handle.net/123456789/255 2026-04-04T10:43:09Z http://hdl.handle.net/123456789/20711 DIFFERENTIAL EXPRESSION OF LGR5 AND ITS DIAGNOSTIC SIGNIFICANCE IN NEOPLASTIC AND NON –NEOPLASTIC COLORECTAL LESIONS DR MARVI ZIA (06-114232-003) Cancer stem cells (CSCs) play a major role in development, progression and in aggressive chemo-resistant behavior as they provide a continuous pool for clonal proliferation of malignant cells. LGR5, one of the most important stem cell marker is implicated very early in metamorphosis of malignant transformation. LGR5 is directly involved in cell proliferation through its effects on Wnt- β catenin pathway. In this cross- sectional study we used 109 formalin fixed paraffin embedded (FFPE) tissue samples and evaluated immuno-reactive score (IRS) of LGR5 expression in normal colonic mucosa, adenomas and colorectal cancer tissue samples. Association between LGR5 expression and various clinico-pathological parameters including grade, stage and lymph-node metastasis was assessed .The comparison of mean LGR5 IRS scores across non-neoplastic tissue, adenomas, and carcinomas was done . A binary logistic regression was performed to examine the association of LGR5 expression (high vs. low) with lesion type, T stage, N stage, and histological grade. Results: Histological type and tumour grade significantly influence LGR5 expression. Conventional Adenocarcinomas and well differentiated Grade 1 tumours exhibited strong LGR5 expression followed by moderately differentiated (Grade 2) tumors, as compared to mucinous and signet ring cell variants and poorly differentiated tumours Grade 3(p <0.001). There was a statistically significant difference in mean IRS scores among the three groups (p < 0.001). The highest mean IRS score was observed in adenomas (mean = 9.42 ± 3.06), followed by carcinomas (6.12 ± 3.62), whereas non-neoplastic tissues exhibited the lowest mean IRS score (2.15 ± 2.02). The patients with Non-Neoplastic Lesions were less likely to have high LGR5 expression (OR=0.077, p<0.001) in comparison to patients with neoplastic lesions. The study identified higher IRS score for LGR5 expression in colorectal cancers and adenoma as compared to normal mucosa, thus identifying it as a marker of neoplastic transformation. Increased LGR5 expression in adenomas and grade 1 CRC identified this stem cell marker as a significant diagnostic marker useful for early neoplastic lesion. Decline of LGR5 expression in poorly differentiated tumours verified it as a marker of poor prognosis. Supervised by Prof. Dr. Sumayyah Shawana 2025-12-01T00:00:00Z http://hdl.handle.net/123456789/20712 IDENTIFICATION OF BRCA1 VARIANTS RS2053936273 AND RS80357670 IN TRIPLE NEGATIVE BREAST CANCER (TNBC) IN THE PAKISTANI POPULATION AT A TERTIARY CARE HOSPITAL IN KARACHI: A SANGER SEQUENCING STUDY DR HIRA SHOAIB (06-114232-001) Triple-negative breast cancer (TNBC) is an aggressive subtype of breast carcinoma characterized by the absence of estrogen receptor (ER), progesterone receptor (PR), and HER2 expression, commonly associated with germline Breast cancer Gene 1 (BRCA1) mutations. These mutations compromise homologous recombination DNA repair and increase cancer susceptibility. Despite the high burden of Triple Negative Breast Cancer in Pakistan, limited genetic data exists on the distribution of BRCA1 variants. Two BRCA1 variants, rs2053936273 and rs80357670, have been reported in major Pakistani studies as pathogenic frameshift mutations leading to truncated protein products; however, their prevalence and clinical relevance in remains unclear. The present study aimed to detect these BRCA1 variants in Triple Negative Breast cancer patients and assess their correlation with clinico-pathological parameters. A cross-sectional study was conducted at Pakistan Navy Station Shifa Hospital in Karachi from November 2024 to October 2025. Fifty female Triple Negative Breast Cancer patients aged 28–60 years were enrolled after biopsy and immunohistochemical confirmation. Genomic DNA was extracted using Qiagen® kits, and Polymerase chain reaction (PCR) amplification was performed at Ziauddin Medical University, Karachi. Sanger sequencing was outsourced to Eurofins, United States of America. Sequence alignment and variant annotation were performed using Molecular Evolutionary Genetic Analysis (MEGA X), BioEdit, and Database Single Nucleotide Polymorphism (dbSNP). Descriptive and associative statistics were analyzed using SPSS v23. Of 50 samples, 39 yielded high-quality sequences suitable for variant analysis. Neither rs2053936273 nor rs80357670 was detected in the study participants. However, two variants of uncertain significance (VUS) were observed in 5.1% of samples. The majority of cases occurred in patients under 50 years, with a subset showing a positive family history. The absence of the targeted BRCA1 variants suggests potential ethnic specificity and highlights the need for expanded genomic studies in the Pakistani TNBC population. VIII In conclusion, the findings emphasize the importance of population-based BRCA1 variant profiling to develop localized genetic testing panels and inform early detection, prevention, and counseling strategies for hereditary breast cancer in Pakistan. Supervised by Prof. Dr. Summayyah Shawana 2025-11-01T00:00:00Z http://hdl.handle.net/123456789/20710 ANTIBIOFILM FORMING ACTIVITY OF NATURAL PRODUCTS AND PROBIOTIC AGAINST STAPHYLOCOCCUS AUREUS AND STREPTOCOCCUS PYOGENES BIOFILM DR. MUHAMMAD MOHSIN SAEED (06-114212-003) Staphylococcus aureus and Streptococcus pyogenes are part of normal flora of human body. Both of them can form a biofilm. Biofilm is described as an adhered collection of immobile groups of microbes fenced within extracellular polymeric substance (EPS). Biofilm will provide microbes safe shelter and ideal environment for maximum growth even under unfavorable conditions like extreme temperature, dehydration and radiations. Biofilm associated infection of both microbes are becoming life-threatening problem due to emerging antibiotic resistance, which necessitates discovery of new alternatives. Many probiotics and natural products have advantageous anti-biofilm properties. Objectives of the study were to assess the biofilm forming activity of Staphylococcus aureus and Streptococcus pyogenes, to evaluate the anti-biofilm role of natural products i.e. Allium cepa (Onion), Mentha piperita (mint) and Zingiber officinale (ginger) and also evaluate the anti-biofilm activity of Probiotics (Lactobacillus). Another objective was to assess the synergistic action of natural product and probiotics against biofilm formation. It is a cross-sectional study. The samples collection is from different labs. The sample size is of 246 participants that belong to 1 - 75 years of age. Informed consent from patients taken. Specimens were inoculated in lab on culture plates, and put for incubation at 37 °C for 24 to 48 hours. Staphylococcus aureus and Streptococcus pyogenes are identified on the basis of colony morphology, gram staining and biochemical test. Post identification ally we use micro-titer plate assay for observing biofilm producing action of Staphylococcus aureus and Streptococcus pyogenes. Then we employ natural product and probiotics on these biofilms and observe the anti-biofilm potential by performing 96 well Plate methods for biofilm assessment. 3 plant {Allium cepa (Onion), Mentha piperita (mint) and (ginger) Zingiber officinale} and probiotic (Lactobacilli acidophilus) possess anti biofilm activity individually and in combination. This anti-biofilm capability is further enhanced when natural products and probiotics used in combination because they act synergistically. Supervised by Prof,Dr.Naveed Faraz 2025-11-01T00:00:00Z http://hdl.handle.net/123456789/18859 P16 AND TOP2A IMMUNOHISTOCHEMICAL EXPRESSION FOR DETECTION OF CERVICAL INTRAEPITHELIAL AND EARLY MALIGNANT LESIONS DR. ARIBA NASREEN 06-114222-001 Cervical Cancer (CC) is the fourth most common type of cancer worldwide. In vast majority of cases, it is caused by Human Papillomavirus (HPV) - a Sexually Transmitted Disease (STD). In Pakistan, the incidence of cervical cancer is 4.9% while its morbidity and mortality rates are considerably higher due to ignorance in terms of screening, prevention and vaccination. As a result more than 70% women with CC are diagnosed at very advanced stage of malignancy. In this time of vaccines, systematic screening for CC may be considered as one of the most valuable tools for reducing the disease burden and death rates. Despite the characteristic advantages of Liquid Based Cytology (LBC) in cervical screening, LBC has low sensitivity (40-50%) which usually leads to under diagnosis. While HPV testing in cervical screening has been proven effective to some extent but its low specificity may result in unnecessary treatment and high-risk HPV testing alone cannot differentiate between transient and persistent infections. Hence, applying different markers in CC screening is necessary for its control, early-diagnosis and prevention. P16 is a cyclin-dependant kinase inhibitor and plays a crucial role as a cell cycle regulator by decelerating cell progression from G1 to S phase. While Topoisomerase IIA is an enzyme responsible for DNA strands uncoupling during its replication and expressed only in cycling cells. The primary objective of this study was to assess immunohistochemical expression of P16 and TOP2A in normal, pre cancerous and cancerous cervical cytology cell blocks and to correlate their expression level with cytological and various clinicopathological parameters. It was a cross-sectional study conducted at PNS Shifa Hospital Karachi and Dr. Ruth K. M. Pfau, Civil Hospital, Karachi over a time period of eight months. A total of 60 cervical cytology samples were analyzed for immunohistochemical staining using CDKN2A/P16-INK4a Mouse (IgG) and Topoisomerase II alpha Rabbit (IgG) Monoclonal antibodies. Demographic data and various clinicopathological parameters were recorded using the designed Performa and the results of immunostaining were analyzed and correlated with the documented parameters. The results revealed that there is a significant association of P16 immunohistochemical expression with age group, ethnicity, menopausal status, cytological diagnosis, and cancer status. While TOP2A was only associated with menopausal status, cytological diagnosis, and cancer status of the patients. P16 ix immunostaining was directly related with the increasing severity of cervical cytological abnormalities. The sensitivity and specificity of P16 for detecting pre-cancerous and cancerous lesion was significantly higher, whereas TOP2A demonstrated higher specificity for detecting cancerous lesions. Therefore, P16 can be a reliable immunohistochemical marker for diagnosing early as well as late cancerous lesion while TOP2A is highly specific marker for ruling out non-cancerous lesion Supervised by Prof. Dr. Summayya Shawana 2024-10-01T00:00:00Z