Over the last two decades, a consistent body of literature has been accumulated demonstrating, both in vitro and in vivo, the potential antineoplastic efficacy of omega-3 PUFAs by using preclinical models of OvCa (see below for details). On the other hand, there are limited numbers of human observational studies, especially if compared to those available for BCa, and often their outcomes are also inconsistent [71,87]. Moreover, there are no clinical interventional trials to date [39]. This may have discouraged researchers from developing omega-3 PUFA-containing nanoformulations to improve OvCa therapy. For these reasons, it may be important to have a picture of the current knowledge in this field of research to better orientate future developments of the research both in the clinical setting and in the nanotherapy area. To this aim, we will describe and critically analyze the most relevant findings obtained with omega-3 PUFA treatments in preclinical in vivo models of OvCa, which are better than the in vitro ones that may mimic the human condition (Figure 1).

Recently, West et al. [88] reported the ability of DHA to inhibit the growth and proliferation of OvCa developing in a transgenic mouse model of OvCa in vivo (the KpB transgenic mouse model of high-grade serous epithelial OvCa mouse model (K18-gT121 ±; p53fl/fl; Brca1fl/fl; KpB)). In this work, DHA was intraperitoneally (IP) injected at the daily dose of 15 mg/kg for 28 days. The intraperitoneal way of administration appears very appropriate since the most common way of OvCa spreading is through the direct exfoliation of cancer cells into the peritoneal cavity [89]. It was also shown that the drug intraperitoneal (IP) injection increases their half-life and concentrations in the target of OvCa cells [90]. Nanoparticles encapsulating another nutraceutical, the epigallocatechin gallate, were recently administered IP in a peritoneal metastatic murine model of human OvCa [91]. The recent findings of West et al. [88] are extremely interesting, since their work is the first preclinical work where the anticancer effects of an omega-3 PUFA were investigated in an immunocompetent mammalian OvCa model, especially considering the crucial role exerted by the tumor microenvironment immune cells in the induction of the growth and progression of tumors. Instead, another early work was performed using SCID mice transplanted with paclitaxel-sensitive human ovarian carcinoma A121 cells [92]. That report also demonstrated that DHA covalently conjugated to the chemotherapeutic second-generation taxoid paclitaxel was able to potentiate the anticancer activity of this drug, inducing the complete regression of the A121 cell xenografts. Interestingly, when these conjugates were used by the same authors [92] against drug-resistant human colon cancer cells implanted in SCID mice, they were able to overcome the resistance and, also in this case, differently from paclitaxel alone and paclitaxel-DHA, induced the total regression of the tumor xenografts. A similar report was recently published by Udumula et al. [93], who implanted the human OvCa SKOV3ip or CaOV3 cells, in the peritoneal cavity of nude mice and then dietary supplemented the mice with 100 mg/kg EPA or DHA administered by gavage per 4–5 weeks. The antineoplastic efficacy of the two FAs was different, and dependent on the cell line implanted. In the SKOV3ip xenograft model, both the FAs resulted as less active than in the CaOV3 one. EPA resulted more efficacious than DHA, and increased the median mouse survival of control mice (23.5 days, corn oil-treated mice) by 97.8%, whereas DHA induced only a 23.4% increase. Moreover, just EPA reduced markedly and significantly the size of the excised tumors, as well as the Ki-67 proliferation index of the OvCa cells grown in vivo, as compared to the cells grown in control mice.

Instead, another work evaluated the ALA antineoplastic activity. In this case, another animal model was used, the xenograft model of zebrafish [94], which is farther than the xenograft mouse models in terms of evolutionary distance and phylogenetic trees from the human species. Anyway, this model was recently considered a helpful model for the study of metastasis in vivo [95], since it has the advantage of mirroring the degree of invasion/metastasis in vitro and in mice, and it is able to furnish information in a faster and less expensive way compared to mouse models of metastasis. In fact, in the zebrafish model, the human cancer cell lines are injected into the perivitelline space of two-days old larvae, where they can survive and metastasize, since the larvae will not develop an adaptive immune system until 14 days post-fertilization [96]. In Wang et al.’s work [94], performed on the zebrafish larvae (at 1-day post-fertilization) only DHA was able to suppress the development of cell metastasis. From the in vitro experiments, the authors suggested that this effect could be related to the DHA ability to suppress the NF-κB signaling.

Over the last two decades, a series of in vivo studies had been also conducted using a model of spontaneous OvCa frequent in incessantly ovulating hens (for a review see: [97]). This model, even though again not using mammalians, has the advantage of being the only known animal model of spontaneous OvCa, and it has been so far considered useful for pre-clinical and dietary intervention studies focused on OvCa prevention, since the OvCa developed in the hen is histologically similar and shares similar features with the human OvCa considering the ascites and metastasis formation in peritoneum [98,99]. The research developed over the years by using this model has demonstrated that a flaxseed-rich diet can decrease the severity and incidence of OvCa [99]. Moreover, it was observed that this diet promotes apoptosis and suppresses angiogenesis in the OvCa, but not in the normal avian ovaries [99]. Interestingly, besides containing high levels of ALA, which can then be metabolically converted into the long-chain EPA and DHA, flaxseed is also a source of the phytoestrogen lignan secoisolaricisresinol diglucoside, which is converted to enterodiol and enterolactone, both having anti-estrogenic and antioxidant properties [99]. Moreover, the enriched flaxseed diet was shown to induce the CYP1A1 pathway, and through it, the formation of 2-hydroxy estradiol metabolites, which are then converted into 2-methoxyestradiol (2MeOE₂) estrogen, which is the least powerful estrogenic metabolite as compared to the others (such as 4-hydroxy and 16-hydroxy metabolites) produced by the different CYP enzymes (CYP3A4 and CYP1B1, respectively) [100]. This will result in a higher 2-hydroxy/16-hydroxy estradiol ratio, previously shown to be protective against postmenopausal BCa [101]. On these bases, Pal et al. [99] recently also investigated the effects of individual treatments with DHA or 2MeOE₂ in vitro in human OvCa cells, and in an in vivo model that mimics the neo-angiogenetic process in vivo, i.e., the chorioallantoic membrane assay [102]. Using the two models, the authors demonstrated that, whereas only 2MeOE₂ induced apoptosis, both the compounds acted by inhibiting neo-angiogenesis, even though with different molecular mechanisms. These results suggest a possible 2MeOE₂/DHA combinational approach for the treatment of advanced OvCa. Therefore, the inclusion of both compounds in newly developed nanoformulations to be evaluated for their potential to contrast OvCa would deserve particular attention. Overall, the in vivo studies substantiate the hypothesis that omega-3 PUFAs may represent potential preventive/therapeutic agents against not only BCa, but also OvCa. Moreover, they suggest that their anticancer potential, when enclosed in nanoformulations in combination with other nutraceuticals or chemotherapeutic drugs, would be worth to be evaluated.

A considerable number of in vitro studies were also performed to evaluate the effects of omega-3 PUFAs on OvCa cells, but we will not analyze them here, both for the sake of brevity and being this issue beyond the scope of this review. However, it is interesting to underline that in a comparable number of these studies the cells were treated either with EPA or DHA, and just in two of them [93,103] both the two long-chain PUFAs were separately evaluated to make a comparison between them. In one case, DHA showed more pronounced anticancer properties [103], whereas in the other one EPA resulted in being the most active [93]. Therefore, robust indications are not yet available for choosing one or the other PUFA for further OvCa in vitro studies. In this context, also the hypothesis put forward very recently by Udumula et al. is of interest [93]. Based on their in vitro findings, these authors suggested that metformin, a “repurposed” drug for OvCa, previously reported to have anticancer effects in preclinical and human observational studies [104], could exert its powerful antineoplastic effects in OvCa by enhancing the endogenous production of EPA and DHA from the precursor ALA [93]. This stimulating hypothesis could represent the rationale for the evaluation of new treatments for OvCa both in vitro and in vivo, where the anticancer properties of metformin could be enhanced by the combined treatment with omega-3 PUFAs, furnishing all these antineoplastic agents (metformin and EPA or DHA) either in their free forms or co-delivered in newly developed nanoformulations. In our opinion, the reason why studies focused on the inclusion of omega-3 PUFAs in nanoformulations designed for the therapy of OvCa have not yet been performed, even despite all the robust and consistent data obtained in the preclinical setting, may be especially related to the inconsistency of the outcomes of the human observational studies performed in this field [71,87].