Advanced breast cancer diagnosis: Multiplex RT-qPCR for precise typing and angiogenesis profiling.

Breast cancer (BC) remains the foremost cause of cancer-related mortality, with an estimated 2.3 million new cases anticipated globally. The timely diagnosis of BC is pivotal for effective treatment. Currently, BC diagnosis predominantly relies on Immunohistochemistry (IHC), a method known for its sluggishness, expense, and dependence on proficient pathologists for confident cancer typing. In this study, we introduce a novel approach to enhance the accuracy, speed, and cost-effectiveness of BC diagnosis. We employ multiplex Reverse Transcription quantitative Polymerase Chain Reaction (RT-qPCR) with touch-down methods, which consistently yield significantly lower Cycle Threshold (CT) values. The study evaluates gene expression profiles of HER2, PGR, ESR, and Ki67 genes across 61 samples representing four BC subtypes, using RPL13A as the endogenous control gene. The results demonstrate that our method offers remarkable precision, nearly equivalent to IHC, in detecting gene expressions vital for BC diagnosis and subtyping. Moreover, we explore the gene expression of Hif1A, ANG, and VEGFR genes involved in angiogenesis, shedding light on the metastatic potential of the tested BC tumours. Notably, numerous samples exhibit elevated levels of Hif1A and VEGFR, indicating their potential as valuable biomarkers for assessing metastatic status. Collectively, our RT-qPCR methodology emerges as a powerful diagnostic tool for swiftly identifying BC subtypes and can be complemented with other essential tumorigenic biomarker assessments, such as angiogenesis, to further refine cancer characterisation and inform personalised therapeutic strategies for BC patients. This innovation holds the promise of revolutionising BC diagnosis and treatment, offering expedited and reliable insights for improved patient care.

Identification of SARS CoV-2 Omicron BA.1 and a novel Delta lineage by rapid methods and partial spike protein sequences in Sulaymaniyah Province, Iraq.

BACKGROUND: Five variants of concern (VOCs) of severe acute respiratory syndrome coronavirus 2 (SARS CoV-2) have been globally recorded including Alpha, Beta, Gamma, Delta, and Omicron. The Omicron variant has outcompeted the other variants including the Delta variant. Molecular screenings of VOCs are important for surveillance, treatment, and vaccination programs. This study aimed to identify VOCs by using rapid inexpensive methods and partial sequencing of the virus's spike gene. METHODS: Mutation-specific rRT PCR probes were used for both D614G and K417N mutations to potentially discriminate between Delta and Omicron variants. These were followed by sequencing of a fragment of spike gene (748 nucleotides), which covers the most notable VOC mutations in the receptor binding domain of SARS CoV-2. RESULTS: Rapid methods showed that out of 24 SARS CoV-2 positive samples, 19 carried the N417 mutation, which is present in the Omicron variant. Furthermore, 3 samples carried K417 wildtype, which is present in the Delta variant. Additionally, 2 samples containing both K417 and N417 suggested mixed infections between the two variants. The D614G mutation was present in all samples. Among the 4 samples sequenced, 3 samples carried 13 mutations, which are present in Omicron BA.1. The fourth sample contained the two common mutations (T478K and L452R) present in Delta, in addition to two more rare mutations (F456L and F490S), which are not commonly seen in Delta. Our data suggested that both Omicron variant BA.1 and a novel Delta variant might have circulated in this region that needs further investigations.