Publications | Ahmad Abdel-Azim

Statistical Genetics

Joint estimation of millions of genetic variant effects in Biobank-scale data for accurate early diagnosis and disease progression

A. Abdel-Azim, and X. Lin

In preparation

Description

We are developing novel robust genetic risk score paradigms for patient stratification and early disease diagnosis, leveraging a mixed effects model framework. The method developed is efficient, scalable, and can handle both longitudinal data and related samples, opening the door for genomics-enhanced treatment plans. We are validating the method via simulation and empirically testing the approach using UK Biobank data.

Cancer Genomics

Spectrum of Germline and Somatic Mitochondrial DNA Variants in Tuberous Sclerosis Complex. Frontiers in Genetics

K. Giannikou, K. Martin, A. Abdel-Azim, and 8 more authors

Frontiers in Genetics, Jan 2023

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Tuberous Sclerosis Complex (TSC) is caused by loss of function variants in either TSC1 or TSC2 and is characterized by broad phenotypic heterogeneity. Currently, there is limited knowledge regarding the role of the mitochondrial genome (mtDNA) in TSC pathogenesis. In this study, we aimed to determine the prevalence and spectrum of germline and somatic mtDNA variants in TSC and identify potential disease modifiers. Analysis of mtDNA amplicon massively parallel sequencing (aMPS) data, off-target mtDNA from whole-exome sequencing (WES), and/or qPCR, revealed mtDNA alterations in 270 diverse tissues (139 TSC-associated tumors and 131 normal tissue samples) from 199 patients and six healthy individuals. Correlation of clinical features to mtDNA variants and haplogroup analysis was done in 102 buccal swabs (age: 20-71 years). No correlation was found between clinical features and either mtDNA variants or haplogroups. No pathogenic variants were identified in the buccal swab samples. Using in silico analysis, we identified three predicted pathogenic variants in tumor samples: MT-ND4 (m.11742G>A, p. Cys328Tyr, VAF: 43%, kidney angiomyolipoma), MT-CYB (m.14775T>C, p. Leu10Pro, VAF: 43%, LAM abdominal tumor) and MT-CYB (m.15555C>T, p. Pro270Leu, VAF: 7%, renal cell carcinoma). Large deletions of the mitochondrial genome were not detected. Analysis of tumors from 23 patients with corresponding normal tissue did not reveal any recurrent tumor-associated somatic variants. The mtDNA/gDNA ratio between tumors and corresponding normal tissue was also unchanged. Overall, our findings demonstrate that the mitochondrial genome is highly stable across tissues and within TSC-associated tumors.
Comprehensive genomic and clinicopathologic characterization of advanced PEComa tumors with metastatic potential

K. Giannikou, A. Abdel-Azim, J. Tsuji, and 9 more authors

Jan In preparation

Description

Background: Malignant PEComa (Perivascular Epithelioid cell tumor) is a rare neoplasm that is predominantly seen in females. This tumor type affects multiple organs and demonstrates local invasion and/or distant metastases with poor prognosis. Approximately 35-40% of PEComa have mutations in TSC1/TSC2 genes. Aim: To identify other driver events beyond TSC1/TSC2 genetic alterations that might contribute to tumor development and to compare to The Cancer Genome Atlas (TCGA) Leiomyosarcoma (LMS), a similar sarcoma subtype with overlapping morphologic and immunophenotypic features which can make diagnosis challenging. Material- Methods: Exome sequencing was performed in 74 malignant PEComa tumors from 59 patients (median age 52). This is the first exome analysis conducted in the largest cohort of malignant PEComa tumors ever assembled. Results: We found that mutations in TP53 (27%), TSC2 (24%) and TSC1 (18%) were the most frequent alterations, whereas other somatic events were mainly non- recurrent missense alterations of uncertain clinical significance. C>T at CpG was the most common substitution, consistent with the aging mutation signature, similar to LMS. Copy number analysis showed that PEComa have multiple focal amplifications and deletions (e.g., RB1, TP53), also similar to LMS. Many of these PEComa patients (75%) had been treated with rapamycin or other mTORC1 inhibitors, as mutations in TSC1/TSC2 in tumors are associated with response to treatment with mTORC1 inhibitors (mTORi). Patients with TSC2 mutations had sustained clinical benefit in response to mTORi, median progression-free survival 1.4 years, and median overall survival 2.9 years. Conclusions: PEComa have a low somatic mutation rate, similar to LMS, and substantial TSC1/TSC2 mutations which are not seen in LMS. PEComa patients respond to rapalog therapy with favorable disease outcome and such responses are often durable (>1 year) and strongly correlate with alterations in TSC2.
Distinct oncogenic signatures in malignant PEComa and leiomyosarcoma identified by integrative RNA-seq and H3K27ac ChIP-seq analysis

K. Giannikou, A. Abdel-Azim, E. Adib, and 10 more authors

Jan In preparation

Description

Malignant Perivascular epithelioid cell tumor (PEComa) and leiomyosarcoma (LMS) are two sarcomas with overlapping morphologic and immunophenotypic features that can make them difficult to distinguish, leading to diagnostic uncertainty. We performed integrative RNA-seq and H3K27ac ChIP-seq analysis of PEComa and LMS and observed that PEComa and LMS co- cluster in comparison to six other sarcoma subtypes. Multiple differentially expressed and H3K27ac-marked genes were identified between groups, including DAPL1, MLANA, SULT1C2, GPR143, and CHI3L1 higher in PEComa; and MYOCD, WFDC2, DES, MYH11, and CNN1 higher in LMS. Gene set enrichment analysis showed higher expression of lysosome genes in PEComa; and myogenesis and smooth muscle contraction genes in LMS. Differentially expressed genes between these two sarcomas were assessed as candidate biomarkers by immunohistochemistry (IHC). PNL2, PMEL, and MLANA all demonstrated complete specificity for PEComa over LMS, but relatively low sensitivity, desmin positivity by IHC was enriched in LMS, while other select IHC markers could not discriminate between these two in an independent cohort. Given the success of mTORC1 inhibitors in the treatment of TSC1/TSC2- mutant PEComa and the potential misdiagnosis between LMS and PEComa, genetic analysis for TSC1/TSC2 mutations should be considered for all ambiguous LMS patients.

Microbiology

Characterization of the Gain and Loss of Resistance to Antibiotics versus Tolerance to Honey as an Antimutagenic and Antimicrobial Medium in Extended‑Time Serial Transfer Experiments

A. Abdel-Azim, S. Abdel-Azim, B. Piasecki, and 1 more author

Pharmacognosy Research, Apr 2019

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Background: Honey contains several substances with antimicrobial properties that appear more recalcitrant to generating bacterial‑evolved resistance than traditional antibiotics. Objectives: This study seeks to characterize the evolution of bacteria grown for successive generations in honey as an antimutagenic medium versus ampicillin. Materials and Methods: A naive strain of Staphylococcus aureus was serially cultured for 17 days in Lysogeny broth (LB) containing sublethal concentrations of medical‑grade Manuka honey, polyfloral honey, and ampicillin. Glucose as an osmotic control and pure LB were included for comparison. A portion of each culture was removed every 24 h to (a) determine the amount of growth that occurred during the previous 24 h and (b) use as a genetic stock in serially transferred tubes containing the same inhibitory compound as used previously. Results: As indicated by an increase in growth over sequential 24‑h period, bacteria rapidly gained resistance to ampicillin in a step‑wise pattern. However, bacteria grown in Manuka and polyfloral honey never exceeded their initial growth levels. Bacteria grown in relatively high concentrations of honey for a single 24‑h period consistently lost viability after one transfer, a phenomenon that has not been reported in the literature before and which indicates the inability of bacteria to adapt to honey as an antimutagenic medium. While bacteria grown in honey did not evolve the ability to grow at higher concentrations, a single isolate grown in Manuka honey gained tolerance to Manuka by successfully surviving and proliferating beyond second transfers. Bacteria that developed antibiotic resistance were found to remain sensitive to honey. Moreover, bacteria lost their resistance to ampicillin upon a single exposure to Manuka honey.
Determining the Contribution of Osmotic Stress to the Antibacterial Properties of Honey

A. Abdel-Azim, S. Abdel-Azim, and G. Abdel-Azim

Journal of Emerging Investigators, Mar 2019

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Researchers have repeatedly shown that honey possesses distinctive antimicrobial properties; however, there is uncertainty over which compounds in honey are responsible for these properties. In this research study, we sought to quantify the role of osmotic stress in honey and determine the efficacy of two types of honey: Manuka and raw pasture honey. Bacteria were sequentially cultured in sublethal concentrations of Manuka and raw pasture honey for five days. The role of osmotic stress as a contributor to the antibacterial properties of Manuka and raw pasture honey was quantified in the first culture and over five serial cultures. The growth levels of bacteria in honey were compared to growth levels in glucose, an osmotic control, to quantify the role of osmotic stress in the two types of honey. The results of this study indicate that in the first culture, the antibacterial impacts of Manuka and raw pasture honey were primarily attributable to osmotic stress. However, over five days of sequential transfers, both raw pasture honey and Manuka honey showed significant antibacterial properties beyond osmotic strength. It was established that the antibacterial properties of honey cannot be investigated based solely on the first culture. Serial transfers over several days should be employed to investigate the efficacy of honey as an antibacterial substance.
The Impact of Antibiotic Exposure and Concentration on Resistance in Bacteria

A. Abdel-Azim, and G. Abdel-Azim

Journal of Emerging Investigators, Oct 2016

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How many generations are needed for bacteria to gain antibiotic resistance and to what extent does antibiotic concentration make a difference? To address these questions, pure Escherichia coli was cultured on nutrient agar in the presence of low, medium, and high concentrations of amoxicillin. Surviving bacteria were repeatedly cultured for four consecutive generations, and the sensitivity of bacteria was measured within each generation. We measured antibiotic sensitivity by evaluating the diameter of the inhibition zone around antibiotic-saturated discs of all three amoxicillin concentrations. Although exposing bacteria to extremely heavy antibiotic concentrations resulted in greater inhibition zones around antibiotic-saturated discs, there were no major differences in sensitivity relative to the much lower concentrations studied. We also found that bacteria gained significant resistance upon first exposure to the antibiotic. In fact, the reduction in average inhibition zones was much greater in early generations than in late generations. We conclude that E. coli can gain major resistance upon surviving early exposure to amoxicillin, which suggests that antibiotic courses of medication should be prescribed to completely clear the body of bacterial infection upon first antibiotic exposure.
The characterization of quorum sensing trajectories of Vibrio fischeri using longitudinal data analytics

M. Abdel-Azim, and A. Abdel-Azim

Journal of Emerging Investigators, Dec 2023

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Quorum sensing (QS) is the process in which bacteria recognize and respond to the surrounding cell density, and it can be inhibited by certain antimicrobial substances. In Vibrio fischeri, bioluminescence intensifies as more cells utilize QS, making it an accurate biomarker for QS. In this study, we showed that observed illumination intensity data measured at multiple time points is insufficient for evaluating and comparing QS activity across different cultures without proper statistical modeling. We introduced a statistical approach to characterize illumination trajectories over time using longitudinal data analytics. The approach was tested on the naturally illuminating bacteria, V. fischeri, which emit light when QS is activated, but it can also be implemented with green fluorescent protein (GFP) and its numerous applications. To compare multiple QS trajectories, we suppressed QS activity at varying levels using honey, known for its antimicrobial and anti-QS properties. We characterized and compared illumination trajectories through time in bacterial cultures containing five Manuka honey concentrations. The longitudinal approach analyzed illumination data throughout the entire QS cycle and correctly ranked bacterial cultures according to their true level of QS activity. Conversely, the conventional cross-sectional approach was inconsistent in evaluating bacteria for their QS activity and trajectory. We conclude that modeling illumination intensity through time provides a more accurate evaluation of QS activity than conventional cross-sectional analysis.