microRNAs are small, single-stranded, non-coding RNAs which are 22-25 nucleotides long. They target mRNAs and either cleave or deadenylate the mRNAs or repress the translation. By these ways, they facilitate gene silencing and they are one of the main regulators of biological processes. They have role in every cellular activity and are very important in maintaining cell homeostasis, differentiation and development. They also play a vital role in development of many diseases including cancers, autoimmune diseases such as rheumatoid arthritis, cardiovascular diseases, neurological disorders and liver diseases.
Biogenesis of microRNAs
There are 2 types of miRNAs; intragenic and intergenic miRNAs. Half of the currently discovered miRNAs are intergenic which means that they are found and processed from either introns or exons of the gene; but mostly from the introns. The other half of the miRNAs are intergenic and they are transcribed from a different gene regulated by a different promoter. The most common pathway for biogenesis of miRNAs is the canonical pathway. In the canonical pathway, primary microRNAs (pri-miRNAs) transcribed by RNA Polymerase II are capped and polyadenylated. Mature miRNA sequences are found in the stem of a hairpin on the pri-miRNA. Then, they are processed to single hairpins; pre-miRNAs, by the microprocessor complex. Microprocessor complex consists of two components; DiGeorge Syndrome Critical Region 8 (DGCR8) and Drosha which is a ribonuclease III enzyme. Pre-miRNAs are transported to the cytoplasm by Exportin 5/Ran GTP complex and the complex cleaved by RNase III endonuclease, Dicer and RNA Binding Protein, TRBP. The length of the miRNA is the mature miRNA length after this step. Argonaute (Ago2) proteins load functional strand of the miRNA into the RNA-induced silencing complex (RISC) and lead it to the target mRNA. Other biogenesis pathways are Drosha- independent and Dicer-independent pathways but they are not as common as the canonical pathway.
microRNAs and Cancer
MicroRNAs are shown to be good biomarkers and therapeutic agents for cancer. Besides, dysregulation of some miRNAs are found to be the key player in progression of several cancer types. Cellular processes such as proliferation, growth, cell death, activation of invasion and metastasis are regulated by microRNAs. miRNAs can function as oncogenes or tumor-suppressors. If a miRNA functions as an oncomiR, it targets a tumor suppressor gene and decreases its expression for inducing cell proliferation and repressing apoptosis. On the other hand, if it acts as a tumor suppressor, it targets an oncogene and reduces its expression. The first study that investigated role of microRNAs in human cancers is from Calin et al. They revealed that miR-15 and miR-16 were downregulated in chronic lymphocytic leukemia. After this study in 2002, many researches about miRNAs as cancer hallmarks have been conducted.
Studying microRNAs in cancer researches for diagnostic or therapeutic reasons has high potential and many advantages. First of all, microRNAs can be isolated from many biological fluids such as blood and urine, saliva which makes the noninvasive biomarker testing methods possible. Secondly, miRNAs are very stable due to their small size so they make the technically challenging samples such as formalin fixed samples easier. Additionally, microRNAs are found free in the circulation as well as encapsulated in extracellular vesicles. EV miRNAs give more information about tumor such as the tumor microenvironment. Moreover, microRNAs serve as biomarkers even for the grade of cancers. Finally, miRNAs are tissue-specific so that every tumor has different microRNA profile depending on the tissue type which make the profile specific for each tissue and more reliable for comparison with healthy profile.
microRNAs as Cancer Biomarkers
Tumor-suppressor and oncogenic properties of microRNAs make them good biomarker candidates. Differential expression of miRNAs results in changes of gene expression. There are hundreds of different targets for miRNAs and miRNA expression differs in each tissue; which means that different cancer types have different miRNA profiles. Especially due to the advances in high-throughput studies in last years, miRNA biomarker panel researches for almost every cancer type has increased tremendously.
There are many promising researches and evidences about discovery of miRNA biomarkers for several cancers by examination of blood specimens. Heneghan and colleagues conducted a study for determining miRNA biomarkers of breast cancer. They reported that miR-195 expression was increased in cancer cohort. Moreover, miR- 215, miR-299-5p, and miR-411 expression levels was found to be downregulated by van Schooneveld et al. Additionally, Konishi et al revealed that miR-21 concentration was significantly higher and miR-375 expression was significantly lower in esophageal squamous cell carcinoma when compared to healthy controls. Besides, miR-486 was found to be a potential biomarker for non-small cell lung cancer and its expression was significantly increased in patients. For gastric cancer, miR-204 level was significantly lower in the cancer cohort when compared to healthy donors. Further, it has been documented that miR-145 levels were lower than the healthy controls as well as miR-204-5p levels.
Although miRNA biomarker discovery from blood is more common, urinary biomarker researches gathered speed because it is a non-invasive method and has high potential for being used as biomarker specimen. Erbes et al stated that miR-21-5p, miR-125b-5p, miR- 155-5p, and miR-451-5p were differentially expressed in urine of breast cancer patients. Indeed, in bladder cancer patients, it was observed that miR-10b and miR-34b were upregulated compared to controls. Furthermore, Andreu et al revealed that miR-375 level was significantly lower in high-grade bladder cancer patients. On the other hand, miR-146a was significantly upregulated in low-grade bladder cancer patients. For prostate cancer, a study pointed up diagnostic power of miR-222-3p*miR-24-3p/miR- 30c-5p biomarkers in cell free urine for prostate cancer diagnosis