Biology subjects

Lam, A. K.

Publications and source records attributed to Lam, A. K..

4 recordsLinked to original sources

bioRxiv2025

Mutation profiling of KRAS and BRAF in primary tumors and circulating tumor cells of colorectal cancer patients using PNA-LNA molecular switch

Identifying KRAS and BRAF mutation status is essential for guiding targeted therapies and enhancing treatment outcomes in colorectal cancer (CRC). This study employs the "PNA-LNA molecular switch" to detect mutations in KRAS codon 12 (c.35G>T/G12V) and BRAF codon 600 (c.1799T>A/V600E) from primary tumours and circulating tumour cells (CTCs) in CRC patients, correlating mutation status with clinicopathological parameters. DNA was isolated from 71 primary tumours and 37 CTC samples. Mutation profiles were generated using the PNA-LNA molecular switch. KRAS mutations were detected in 26 primary tumours (36.6%) and 13 CTCs (26.8%), while BRAF mutations were observed in 19 primary tumours (26.8%) and 7 CTCs (19%). No significant correlation was observed between mutation status and clinicopathological parameters in primary tumours. However, KRAS G12V mutations in CTCs significantly correlated with lymph node metastasis (p=0.002), overall pathological stage (p=0.005), and lymphovascular invasion (p=0.034). BRAF V600E mutation status showed no significant clinicopathological associations. Validation of the PNA-LNA molecular switch against Next-Generation Sequencing (NGS) showed 89% concordance with p-values < 0.001 for both genes. This method is highly comparable to NGS for detecting KRAS and BRAF mutations and shows promise as a point-of-care diagnostic tool. Larger patient cohorts are required to confirm its clinical utility.

cancer biology↗

bioRxiv2023

β Cell microRNAs Function as Molecular Hubs of Type 1 Diabetes Pathogenesis and as Biomarkers of Diabetes Risk

MicroRNAs (miRNAs) are small non-coding RNAs that play a crucial role in modulating gene expression and are enriched in cell-derived extracellular vesicles (EVs). We investigated whether miRNAs from human islets and islet-derived EVs could provide insight into {beta} cell stress pathways activated during type 1 diabetes (T1D) evolution, therefore serving as potential disease biomarkers. We treated human islets from 10 cadaveric donors with IL-1{beta} and IFN-{gamma} to model T1D ex vivo. MicroRNAs were isolated from islets and islet-derived EVs, and small RNA sequencing was performed. We found 20 and 14 differentially expressed (DE) miRNAs in cytokine-versus control-treated islets and EVs, respectively. Interestingly, the miRNAs found in EVs were mostly different from those found in islets. Only two miRNAs, miR-155-5p and miR-146a-5p, were upregulated in both islets and EVs, suggesting selective sorting of miRNAs into EVs. We used machine learning algorithms to rank DE EV-associated miRNAs, and developed custom label-free Localized Surface Plasmon Resonance-based biosensors to measure top ranked EVs in human plasma. Results from this analysis revealed that miR-155, miR-146, miR-30c, and miR-802 were upregulated and miR-124-3p was downregulated in plasma-derived EVs from children with recent-onset T1D. In addition, miR-146 and miR-30c were upregulated in plasma-derived EVs of autoantibody positive (AAb+) children compared to matched non-diabetic controls, while miR-124 was downregulated in both T1D and AAb+ groups. Furthermore, single-molecule fluorescence in situ hybridization confirmed increased expression of the most highly upregulated islet miRNA, miR-155, in pancreatic sections from organ donors with AAb+ and T1D. One Sentence SummarymiRNA expression patterns in human pancreatic islets and EVs change under inflammatory conditions and can be leveraged to inform biomarkers strategies for T1D.

physiology↗

bioRxiv2022

Treg cells drive MYCN-mediated immunosuppression and tumor aggressiveness in high-risk neuroblastoma

Solid tumors, especially those with aberrant MYCN activation, harbor an immunosuppressive microenvironment to fuel malignant growth and trigger treatment resistance1,2, yet the underlying mechanisms are elusive and effective strategies to tackle this challenge are lacking. Here we demonstrated the crucial role of T regulatory (Treg) cells in MYCN-mediated immune repression and tumor aggression using high-risk neuroblastoma (NB) as a model system. Human MYCN-activated NB attracts CD4+ Treg cells, which are also found enriched in MYCN-high primary patient samples. Zebrafish MYCN-overexpressing neural crests recruit Cd4+ cells before tumor formation and induce an immunosuppressive microenvironment, thereby promoting tumor onset and progression. Strikingly, disruption of Treg cells through depletion of forkhead box protein 3a restores anti-tumor immunity and impairs NB development. Together, our studies establish Treg cells as the key driver of MYCN-mediated immunosuppression and tumor aggressiveness, providing mechanistic insights and therapeutic implications.

cancer biology↗

bioRxiv2020

Deletion of immune evasion genes provides an effective vaccine design for tumor-associated herpesviruses

Vaccines based on live attenuated viruses often induce broad, multifaceted immune responses. However, they also usually sacrifice immunogenicity for attenuation. It is particularly difficult to elicit an effective vaccine for herpesviruses due to an armament of immune evasion genes and a latent phase. Here, to overcome the limitation of attenuation, we developed a rational herpesvirus vaccine in which viral immune evasion genes were deleted to enhance immunogenicity while also attaining safety. To test this vaccine strategy, we utilized murine gammaherpesvirus-68 (MHV-68) as a proof-of-concept model for the cancer-associated human {gamma}-herpesviruses, Epstein-Barr virus and Kaposi sarcoma-associated herpesvirus. We engineered a recombinant MHV-68 virus by targeted inactivation of viral antagonists of type I interferon (IFN-I) pathway and deletion of the latency locus responsible for persistent infection. This recombinant virus is highly attenuated with no measurable capacity for replication, latency, or persistence in immunocompetent hosts. It stimulates robust innate immunity, differentiates virus-specific memory T cells, and elicits neutralizing antibodies. A single vaccination affords durable protection that blocks the establishment of latency following challenge with the wild type MHV-68 for at least six months post-vaccination. These results provide a novel approach to effective vaccination against cancer-associated herpesviruses through the elimination of latency and key immune evasion mechanisms from the pathogen.

microbiology↗