Where mutation does not become cancer: RNA and neurons
Neuro-inflammation can worsen the outcomes of stroke, traumatic brain injury or spinal cord injury, as well as accelerate neurodegenerative diseases such as ALS, Parkinson’s or Alzheimer’s. This suggests that limiting neuroinflammation may represent a promising new approach for the treatment of neurological diseases and neuropathic pain. Glial cells are the non-neuronal cells of the central nervous system, which help support and protect neurons. One of the types, microglia, are brain macrophages that respond to injury or infection. Microglia and astroglia are key cells of the central nervous system that, when activated, drive neuroinflammation by secreting toxic inflammatory mediators, including cytokines and chemokines. In a preclinical study published in the journal Glia, professors Peter King and Burt Nabors, both of the University of Alabama at Birmingham’s Department of Neurology, show that their drug, SRI-42127, can potently attenuate neuroinflammation triggers.
These experiments in glial cell cultures and mice now open the door to testing SRI-42127 in models of acute and chronic neurological damage. After 25 years of cooperation, their study builds on previous findings that microglial and astroglial cells rely on a key RNA-binding protein called HuR that protects messenger RNAs encoding inflammatory mediators from degradation and promotes their translation into proteins. Neuroinflammation occurs when activated microglia and astrocytes in the brain or spine secrete cytokines and chemokines such as IL1β, IL-6, TNF-α, CXCL1, and CCL2. The messenger RNAs for those pro-inflammatory proteins contain adenine- and uridine-rich elements, or AUREs, that regulate their expression. Scientists have previously shown that HuR plays an important positive role in regulating the production of inflammatory cytokines. HuR normally concentrates in the nuclei of glial cells.
However, when glial cells are activated, HuR moves from the nucleus to the cytoplasm, where it can increase the production of cytokines and chemokines. Previously, UAB researchers have shown that HuR translocates out of the nucleus of astrocytes in acute CNS diseases, spinal injuries, and stroke. They also showed that it translocates out of the nucleus into microglia in chronic CNS disease ALS or amyotrophic lateral sclerosis. Importantly, the HuR monomer cannot pass through the nuclear envelope which serves as a regulatory membrane barrier between the nucleus and the cytoplasm. Only HuR dimers (consisting of two single HuR molecules) are able to cross the nuclear membrane. This knowledge allowed collaborative research by Southern Research, Birmingham, Alabama, and UAB, using high-throughput screening, to identify a small molecule called SRI-42127 that inhibits HuR dimerization. The scientists used bacterial LPS to activate glial cells and initiate the inflammatory cascade.
They then found that treatment with SRI-42127 suppressed the translocation of HuR from the nucleus to the cytoplasm in LPS-activated glial cells, both in tissue culture and in mice. SRI-42127 also significantly attenuated the production of inflammatory mediators, such as IL-1β, IL-6, TNF-α, and the chemokines CXCL1 and CCL2. Furthermore, the compound suppressed the activation of microglia in the brains of mice and attenuated the recruitment of other immune cells outside the CNS. Such blood-brain barrier (BBB)-mediated entry of neutrophils and monocytes can exacerbate inflammation in the brain or spinal cord. In summary, SRI-42127 penetrated the blood-brain barrier and rapidly suppressed neuroinflammatory responses. These findings highlight the critical role of HuR in promoting glial activation and the potential for SRI-42127 and other HuR inhibitors to treat neurological diseases driven by this activation.
In unpublished work in collaboration with Robert Sorge, PhD, associate professor in the Department of Psychology, UAB College of Arts and Sciences, Professors King and Nabors discovered potential beneficial effects of SRI-42127 for reducing neuropathic pain, a condition that it is triggered by microglia-induced neuro-inflammation. This would be a non-opioid approach to treating pain. Overall, this discovery opens universal doors towards the treatment of neuroinflammation that underlies almost all modern neurodegenerative processes, such as Alzheimer’s, ischemic stroke, multiple sclerosis, ALS and all known neuropathies.
FrateRNAl alliance: from neuroscience to malignancy
In their long collaboration, King and Nabors used glioblastoma, a primary brain tumor, as a disease model to study HuR because many of the factors that drive neuroinflammation also promote glioblastoma growth. And this is where the link with tumors lies: Nabors focused on the tumor suppressive properties of SRI-42127 and its potential use in the treatment of glioblastoma and other tumors. One of the hallmarks of cancer is genomic instability, or the tendency to accumulate mutations and DNA damage that leads to alterations of the genome during cell division. DNA mutations can result from exposure to ultraviolet radiation or X-rays or from certain chemicals known to cause cancer; however, our cells have developed mechanisms to monitor and repair damaged DNA. Genome stability may also be threatened by the translation of some messenger RNAs (mRNAs). Some mRNAs are known to be associated with cancer metastasis.
To counter this threat, a specific protein, heterogeneous nuclear ribonucleoprotein E1 (hnRNP E1), binds these mRNAs and prevents them from producing proteins. Researchers at the Medical University of South Carolina have previously demonstrated how hnRNP E1 binds to metastasis-associated RNAs to inhibit their translation. hnRNP E1 binds RNA in the cytoplasm of the cell, but the protein is also found in the cell nucleus. This led researchers to hypothesize that hnRNP E1 might also interact with DNA. Their findings describe a new role for hnRNP E1 in binding DNA in the nucleus. How hnRNP E1 binds and interacts with RNA has been extensively studied, but the discovery that hnRNP E1 also binds to DNA has opened up new research avenues to explore. The DNA binding of hnRNP E1 is not limited to a few sites, but rather the protein has a plethora of potential binding sites, allowing it to detect or prevent DNA damage throughout the genome (a kind of sensor).
The team also found that hnRNP E1 binds to a specific structure that can form on DNA known as an I-motif, which forms in regions enriched in the cytosine nucleotide and acts as regulators of gene expression. Since DNA is formed by specific bonds between nucleotides, numerous guanine bases are located in front of cytosine-rich I-motifs. These guanine-rich regions have the potential to form their own structure known as the G-quadruplex (G4). G4s are present at the beginning of several oncogenes. However, it is not known whether I-motifs and G4 can exist at the same time or whether they are mutually exclusive. Therefore, binding of hnRNP to motif I regions could suppress the formation of G4 structures in order to protect the cell. The scientists hypothesized that hnRNP E1 would protect against genomic instability by maintaining I-motifs and suppressing G4s. Indeed, experiments using cells that lack hnRNP E1 simultaneously showed fewer I-motifs and more G4, DNA damage signals and mutations.
Treating these cells with additional DNA-damaging agents, such as UV light and hydroxyurea, resulted in an intensification of the cells’ DNA damage response which caused them to stop progressing through the cell cycle. These findings have great relevance in the field of cancer genetics and biology. Researchers have studied the contribution of G4s to cancer biology for decades. Due to its association with oncogenes, these regions have been the target for drug design and anticancer therapies. Understanding the protein-DNA interactions that occur at sites opposite G4s may contribute to the efficacy of these drugs, thus facilitating better drug targeting and specificity. And the work becomes more complicated, in light of the latest discovery made by researchers at UC San Francisco, which offers clues to overcoming resistance to drugs, such as tamoxifen which is used against breast cancer. New research has found that in addition to its well-known activity in the nucleus, it can also help malignant cells overcome innate anti-tumor mechanisms and develop resistance to treatment.
Estrogen receptor α (ERα) drives over 70% of breast cancers. In the nucleus, ERα regulates the conversion of DNA to messenger mRNA. Once formed, the mRNA strand travels from the nucleus into the cytoplasm, where it instructs ribosomes to make proteins, a process known as translation. To their surprise, the researchers found that ERα also plays a role in this process by binding to newly formed mRNA. Using breast cancer cell lines, the research team saw how ERα tends to bind to RNAs, particularly mRNAs involved in cancer progression. Some of these mRNAs prevent cells from committing suicide when they accumulate too many harmful mutations. Others help them thrive in harsh conditions, such as lack of oxygen or nutrients or the presence of chemotherapy drugs. Endocrine therapies, such as tamoxifen, block the transcription activity of ERα in the nucleus of a cancer cell. While they may initially be highly effective for most patients with ERα-positive breast cancer, a significant number develop resistance to the drug.
To understand ERα’s role in this, the team analyzed tumor cells from 14 patients diagnosed with ERα-positive breast cancer and found that they had elevated levels of ERα mRNA. Then they experimented with breast cancer cell lines that had acquired resistance to tamoxifen, both in tissue culture and in mouse xenografts. Inhibition of the RNA-ERα interaction restored tamoxifen’s potency against tumors in mice, making cultured cells more sensitive to stress and apoptosis (programmed cell death). A better understanding of the many functions of ERα, therefore, could help optimize current treatments, such as tamoxifen, as well as lead to new therapeutic targets. Compounds targeting translational control in cancer are already in the clinic and can now be tested for potency against breast tumors associated with ERα expression.
- edited by Dr. Gianfrancesco Cormaci, PhD, specialist in Clinical Biochemistry.
Scientific references
Huang XT et al. Cancer Lett. 2021 Oct 10; 518:196-206.
Mohanty BK et al. Life Sci Allian 2021; 4(9):e202000995
Xu Y et al. Cell 2021 Sep 18: S0092-8674(21)01047-3.
Varshney D et al. Nat Rev Mol Cell Biol 2020; 21:459–74.
Giaginis C et al. Pathol Oncol Res. 2018; 24(3):631-40.
Tan S et al. J Biol Chem. 2017; 292(33):13551-13564.