**7. Conclusion: the future of lncRNAs in melanoma as biomarkers and targets for therapy**

Overall, lncRNAs serve as promising biomarkers for melanoma, though much more research needs to be done on them before they can be used clinically. The presence of lncRNA in blood and urine make them particularly valuable to the field of cancer diagnostics as presently, there is a dearth of early diagnostic measures for melanoma. Currently, potential melanomas must be detected by a patient or physician. The major shortcoming of this is that sometimes malignant melanomas do not appear obvious until it is at a late stage, and patients themselves often cannot identify harmful lesions at early stages. Additionally, it is difficult to keep track of potentially malignant nevus in certain areas of the body. Once a potentially tumorigenic nevus or lesion is clinically observed, the first line of diagnostics is the histopathology of biopsies, which are both invasive and expensive.

Diagnoses using circulating lncRNA could serve as an improvement to these biopsies. Not only are they minimally invasive, they are also less expensive and can be conducted at regular intervals for high-risk patients (those with a melanoma in the past 5 years, certain genes, phenotypic red hair, Irish-Scottish ancestry, high mole count, frequent sun exposure, etc.). Moreover, many lncRNAs can also provide valuable prognostic information, including progression, staging, and size to tumor.

between the lncRNA and DNA, RNA, and proteins, a variety of pull-down experiments are performed (**Figure 2A**). To determine RNA-DNA interactions, chromatin isolation by RNA purification (ChiRP) is most commonly used. Other methods include RNA antisense purification (RAP), which uses RNA antisense probes to map RNA interactions with chromatin

can then be analyzed via RT-qPCR to identify any lncRNAs of interest.

34 Human Skin Cancers - Pathways, Mechanisms, Targets and Treatments

**Figure 2.** Experimental techniques. (A) General workflow for immunoprecipitations. Many of these pull-down experiments to find RNA binding partners follow a similar protocol, including ChIRP, RAP RNA, and RIP. RNA-DNA: ChIRP, RAP, CHART. RNA-RNA: RAP RNA, CLASH. RNA-protein: RIP, CLIP. (B) Clinical detection of circulating lncRNA. lncRNA are found in serum exosomes. First, a blood sample collected from a patient is centrifuged to separate the plasma. Then, exosomes are precipitated from the plasma, lysed, and RNA collected and purified. This purified RNA Some concerns for the use of lncRNA as biomarkers do exist. The lncRNA must be present in sufficient quantity for it to be able to be detected and analyzed using standard methods. Additionally, as discussed previously, many of these lncRNAs are also upregulated in other cancers, lowering its specificity as a melanoma biomarker. However, this may not be a bad thing, as other malignancies may be able to be "accidentally" detected.

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Certain lncRNA can also be used as targets for novel therapies. LncRNAs like BANCR and MALAT-1 are responsible for cell migration and metastases. Targeting or knocking down these lncRNA in vivo may prevent further progression and invasion of early stage melanomas and limit the metastatic activity of late stage melanomas.

In conclusion, lncRNAs are likely to be suitable melanoma biomarkers for a variety of reasons: (1) They are secreted into the bloodstream and easily accessible for analysis using noninvasive and inexpensive methods. Because they are secreted within exosomes, they are also protected from various RNases within the bloodstream. (2) Various lncRNAs are secreted at different time-points of disease progression. Those secreted early on have valuable diagnostic potential while others may be useful in determining disease development and prognosis. (3) LncRNAs are generally highly specific for melanoma, a shortcoming of current protein biomarkers. (4) Noncoding RNAs are responsible for a variety of cellular functions and implicated in many important pathways, making them valuable prognosticators of disease. (5) LncRNA biology is still a relatively novel field, which holds a lot potential as more research is being conducted.

## **Acknowledgements**

Publication made possible in part by support from the Berkeley Research Impact Initiative (BRII) sponsored by the UC Berkeley Library.
