: Breast cancer remains the most common malignant disease in women worldwide. Despite advances in detection and therapies, studies are still needed to understand the mechanisms underlying this cancer. Cancer stem cells (CSC) play an important role in tumor formation, growth, drug resistance, and recurrence. Here, it is demonstrated that the transcription factor RUNX1, well known as essential for hematopoietic differentiation, represses the breast cancer stem cell (BCSC) phenotype and suppresses tumor growth . The current studies show that BCSCs sorted from premalignant breast cancer cells exhibit decreased RUNX1 levels, whereas ectopic expression of RUNX1 suppresses tumorsphere formation and reduces the BCSC population. RUNX1 ectopic expression in breast cancer cells reduces migration, invasion, and tumor growth (57%) in mouse mammary fat pad. Mechanistically, RUNX1 functions to suppress breast cancer tumor growth through repression of CSC activity and direct inhibition of ZEB1 expression. Consistent with these cellular and biochemical results, clinical findings using patient specimens reveal that the highest RUNX1 levels occur in normal mammary epithelial cells and that low RUNX1 expression in tumors is associated with poor patient survival. IMPLICATIONS: The key finding that RUNX1 represses stemness in several breast cancer cell lines points to the importance of RUNX1 in other solid tumors where RUNX1 may regulate CSC properties.

Genetically engineered mouse models of prostate cancer allow for study of disease progression from localized tumor formation through distal metastasis. The anatomy of the mouse prostate differs dramatically from the human prostate, being composed of four lobe pairs (anterior, dorsal, lateral, and ventral), making the identification and dissection technically challenging. Although the entire murine prostate and surrounding tissue, including urethra, bladder, seminal vesicles, and associated adipose tissue, can be quickly dissected for en bloc analysis, it is necessary to isolate individual prostate lobes for gene expression studies elucidating the molecular mechanisms of prostate cancer. The procedure as described here includes full color images, allowing the researcher to appreciate the unique prostate morphology and tissue manipulation required to harvest individual prostate lobes. Along with removing all extraneous tissue, the procedure allows for direct comparison of the different prostate lobes by established downstream techniques. Importantly, high quality RNA required for next-generation gene expression analysis can only consistently be obtained from ventral and lateral lobes. Finally, preclinical studies using prostate targeted therapies can be monitored specifically in individual prostate lobes on the histological and gene expression studies.

Loop-mediated isothermal amplification (LAMP) is a power tool for the amplification of specific RNA and DNA targets. Much like PCR, LAMP requires primers that surround a target amplification region and generates exponential product through a unique highly specific daisy-chain single-temperature amplification reaction. However, until recently, attempts to amplify targets of greater than 200 base pairs (bp) have been mostly unsuccessful and limited to short amplicon targets of less than 150 bp. Although short amplicons have the benefit of a rapid detection (,40 min), they do not allow for the prediction of RNA integrity based on RNA length and possible intactness. In this study, 8 primer sets were developed using 2 LAMP primer-specific software packages against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid gene with insert lengths ranging from 262 to 945 bp in order to amplify and infer the integrity of viral RNA. Because these amplification lengths are greater than the current methods that use an insert length of 130 or less, they require a longer incubation, modified primer and temperature strategies, and the addition of specific adjuncts to prevent nonspecific amplification. This proof of concept study resulted in successful reverse transcription LAMP reactions for amplicon targets of 262, 687, 693, and 945 bp using a clinical nasopharyngeal NP sample, purified SARS-CoV-2 RNA, and crude lysate containing inactivated virus.

Cover: The cover image, by Areg Zingiryan et al., is based on the From the Bench Paper Dissection of Individual Prostate Lobes in Mouse Models of Prostate Cancer to Obtain High Quality RNA, DOI: .

<p>S1. RUNX1 mRNA is decreased during breast cancer progression. S2. RUNX1 does not change cell proliferation. S3. RUNX1 represses tumor growth in mammary fat pad. S4. Gate for MCF10AT1 sorting. S5. CD24high Cells have high RUNX1 expression in MCF10AT1 cells. S6. Loss of RUNX1 promotes stemness in MCF10A and MCF7 cells. S7. Overexpression RUNX1 in MCF10CA1a cells does not change BCSC population. S8. Zeb1 is expressed at low level in MCF10CA1a cells. S9. Schematic diagram of ChIP qPCR primers and amplicons over Zeb1 for ChIP-qPCR. S10. Knockdown Zeb1 in MCF10AT1 ns cells does not change cancer stem cell phenotypes.</p>