**3.1 How good are we at diagnosing epithelial ovarian cancer?**

With the current tools available, clinicians are unable to reliably identify EOC early in its disease course, losing a valuable opportunity at early intervention and higher rates of cure. For patients who are diagnosed with EOC at stage I, disease that is confined to the ovaries, their five-year survival approaches 90% [2]. Survival drops precipitously for women with advanced stages of the disease, which is unfortunately the most

common presentation. Attempts at establishing screening systems have certainly been investigated, with the United Kingdom famously conducting a randomized controlled study in which women were screened for EOC with a combination of serum markers and ultrasound [12]. When compared to women who underwent no screening, no impact was observed on overall survival from EOC [12]. Based on the results of this trial, no screening is currently recommended for the general population because modern diagnostic tests do not help patients that are diagnosed with EOC live longer and these same tests lead more women with benign ovarian diseases to have unnecessary procedures because the testing does not distinguish well between benign ovarian disease and cancer [12]. EVs present a promising new frontier for EOC screening because they are detectable in the serum and urine of patients, providing a potential novel method for diagnosing this cancer at an early stage when the patient can be cured more easily.

#### **3.2 MicroRNA in EVs: how can they help to diagnose epithelial ovarian cancer?**

One promising method for early cancer detection involves the analysis of EVs carrying microRNA, or miRNA, in the blood of patients. As strands of non-coding RNA that are 19–25 nucleotides in length, miRNAs are transcription products of DNA that regulate genes, a process that can activate or suppress the expression of different factors that can promote the growth and metastasis of EOC [1]. While most miRNA found in body fluids is cell-free and easily degradable, miRNA that is present in EVs is more stable, amplifying the role of this information in intercellular communication because it reaches cells more effectively [13]. When normal cells, such as stem cells, receive the miRNA from EVs, they produce tumorigenic factors that enhance the cancer's ability to survive and promote invasion and dissemination [1].

When compared to healthy individuals, patients with EOC have levels of certain circulating miRNAs carried in EVs that are often elevated [13]. Numerous specific miRNAs have already been linked to EOC. For example, miR-222-3p, which Ying et al. showed promotes the conversion of normal macrophages into tumor-supporting macrophages through the activation of the SOCS3/STAT3 pathway, is elevated in patients with EOC. Once normal macrophages are transformed, they exert immunosuppressive effects that assist EOC cells in evading identification while also secreting factors that promote migration and growth. Since EV miR-222-3p levels are increased in this cancer, its detection in serum can serve as a diagnostic biomarker for early detection [14].

In another study Cappellesso et al. identified elevated levels of EVs with miR-21, a known regulator of the tumor suppressor gene programmed cell death 4 (PDCD4), in patients with EOC compared to patients with benign ovarian disease [15]. The gene PDCD4 typically prevents cancer through the regulation of apoptosis. However, in EOC, the increased expression of miR-21 directly inhibits PDCD4, allowing the cancer cell to further mutate and to invade other tissues. Similarly to miR-222-3p, EVs with miR-21 can enhance diagnostic testing and clinical staging of EOC.

While looking at individual EV miRNAs can provide clues for early detection of EOC, their true value will come from evaluating the miRNAs in large groups as diagnostic panels that together will provide screening with high sensitivity and specificity. In their study Taylor and Gercel-Taylor reviewed a panel of 8 miRNAs found in EVs miR-141, miR-214, miR-200a, miR-200b, miR-200c, miR-21, miR-205—that displayed distinct biological profiles between patients with benign ovarian disease and those with EOC [16]. By utilizing this panel of EV miRNAs and including other EV miRNAs, a simple blood sample may serve as a powerful test that can be employed by clinicians to apprehend EOC in asymptomatic populations before it lethally spreads.

### *Extracellular Vesicles and Ovarian Cancer DOI: http://dx.doi.org/10.5772/intechopen.101412*

Proteins are also transported in EVs and can potentially serve as biomarkers for early diagnosis of EOC. One example of these EV proteins is EpCAM, which is recognized for its role in tumorigenesis and tumor proliferation and is elevated in patients with EOC. However, the diagnostic utility of EpCAM and other proteins is limited because the proteins can be elevated in patients with benign ovarian disease, decreasing the specificity of these markers [16]. If such proteins are then implemented into screening protocols, patients may have false-positive test results and may subsequently undergo invasive procedures with their associated complications without any benefit.

However, some proteins carried by EVs appear to be specific to EOC. CD24, a known poor prognostic marker for EOC, can be detected within EVs in malignant ascites of EOC patients [17]. Additionally, about half of the blood samples from a cohort of EOC patients contained EV claudin-4, another protein that can potentially serve as a diagnostic marker [18]. With the development of new diagnostic panels that combine EV proteins and miRNAs, patients will 1 day obtain testing that identifies EOC early and gives them a better chance at a cure.

Even easier to obtain than blood, urine is another potentially rich source for EOC EVs. Studies have identified numerous EV miRNAs such as miR-92a and miR-30a-5p that are elevated in the urinary samples of patients with EOC when compared to healthy controls [19, 20]. Specifically, miR-30a-5p is elevated in EOC but decreased in other malignancies such as gastric and colon cancer, making it a potentially unique biomarker [20]. While EV miRNAs found in urine are a potentially exciting


#### **Table 1.**

*Potential panel of EV biomarkers for the diagnosis of epithelial ovarian cancer [1, 16, 17, 19, 20, 22].*

biomarker for diagnosing EOC, more research is required to further take advantage of this easily accessible opportunity.

While these many EV factors provide appealing options for future diagnostic applications, some barriers hinder the utilization of EVs in the clinical setting. Current methods for isolation and purification of EVs are still constrained, relying on identification of these vesicles by size, a non-specific criterion that does not distinguish EVs from large proteins and other types of vesicular structures. The purification process involves ultracentrifugation, a process that is inefficient and cumbersome, especially for serum samples [21]. Also, current methods of molecular identification are limited by the small size of EVs as well as by the difficulty in detecting the EV content [21]. Once scientists solve these issues and answer other questions regarding the viability and concentration of EVs in blood and urine samples, the detection of EOC EVs will bolster the strength of diagnostic tools (**Table 1**).
