The National Institutes of Health this month awarded grants worth more than $400,000 to two researchers investigating the role of microRNAs in cancer, and one who is developing a new method to detect the small, non-coding RNAs.
The first grant went to Jeffrey Petty of Furman University to help fund his development of an oligonucleotide-sensing technology, which is being optimized for miRNAs.
While fluorescence is useful for biological analysis, background interference often requires extensive purification for in vitro analysis, Petty wrote in his grant’s abstract.
To address this limitation, he and his colleagues developed small silver clusters of around 10 atoms that exhibit strong emission in the “near-infrared spectral region where biological samples are relatively transparent.” The metallic ligands associate with oligonucleotide sensors, forming specific silver clusters through nucleobase coordination within certain sequences and allowing the recognition of target oligonucleotides through complementary base pairing.
“Highly sensitive detection is accomplished when hybridization of the analyte with this sensor transforms a cluster from a non-emissive state with a violet absorption to a highly emissive state with near-infrared absorption,” according to the abstract.
With the help of a three-year NIH grant, which began on May 1, Petty and his lab will examine the use of the silver clusters to detect miRNA sequences with potential as disease biomarkers. The grant is worth $323,150 in the first year.
The second grant was awarded to Yale University’s Christopher Cheng to support his studies into the inhibition of miR-21 and miR-155 to combat lymphoma.
Both miRNAs show differential expression levels in cancers, acting as oncogenes to impact cellular transformation and carcinogenesis. “In fact, cancers can become addicted to miR-21 and miR-155” such that oncogenesis is dependent on their overexpression, and their removal triggers cancer cell apoptosis or a reversal of the malignant phenotype, according to the grant’s abstract.
To gain a better understanding into how these two miRNAs influence cancer, specifically lymphoma, Cheng aims to uncover their downstream targets in order to “reveal the mechanisms involved in the onset and maintenance” of the disease.
“In vivo pathways impacted by miR-21 and 155 in lymphoma will be identified using unbiased screens in [miRNA]-addicted transgenic mouse models and human B cell lines,” he wrote in the abstract.
The miRNAs’ targets will then be identified by comparing the populations of cross-linked mRNA from B cells over-expressing miR-21 or miR-155 to normal B-cells. Cheng and colleagues will then examine the resulting RNA sequences for small indels and point mutations found in the tumors but not in normal spleen or lymph tissue, and see whether the downstream targets they identify are also altered in human pre-B-cell lymphoma cells.
Also with the support of the NIH grant, Cheng plans to test whether miR-21 and miR-155 antagonists have therapeutic effects in mouse models and human lymphoma cell lines.
Drug-delivery nanoparticles will be coated with undisclosed, novel molecules that “improve tumor accumulation via low pH-induced targeting,” and then loaded with the miRNA inhibitors, according to the abstract. The formulations will be evaluated for biodistribution and ability to effect tumor regression in the mouse models, with the most effective ones tested in in human lymphoma cells and xenograft models.
Cheng’s two-year grant began on May 1 and is worth $49,214 in its first year.
Also receiving a grant from the NIH is University of Pennsylvania researcher Amy Demicco, who is investigating the mechanisms used by developing lymphocytes to coordinate DNA double-strand break repair and cell cycle progression, which are key to maintaining genomic stability and preventing cancer.
As lymphocytes develop, they induce and repair several DNA double strand breaks, or DSBs, during somatic and class switch recombination. A key aspect of these processes is the upregulation of cyclin D3, which induces cellular proliferation.
“D3 protein and mRNA expression is downregulated in primary mouse lymphocytes in response to DSBs induced by [ionizing radiation] through a mechanism that requires the tumor suppressors ATM and p53,” as well as the miRNA-processing enzyme Dicer, Demicco noted in her grant’s abstract.
Preliminary data indicate that the D3 repression occurs as a result of increased mRNA turnover, which is “consistent with several known pathways by which ATM and p53 regulate the expression of [miRNAs] involved in the DSB response,” she added.
To further explore this, she aims to uncover the ATM and p53-dependent mechanisms through which DSBs downregulate D3 expression, focusing on miRNA-mediated pathways, according to the grant’s abstract. To do so, mutational analysis of the 3' UTR will be combined with miRNA overexpression and knockdown assays to pinpoint those miRNAs that specifically target the D3 mRNA. The potential of ATM and p53 to enhance miRNA biogenesis will also be examined.
Demicco’s NIH grant, which also includes in vivo studies of the DSB-induced D3 repression, began on May 1 and runs for three years. It is worth $42,232 in the first year.