The more common prevalence of HGSOC among all ovarian epithelial cancers associated with variable patient response to therapy and variable outcome should raise its concern in the scope of research. The Australian Ovarian Cancer Study Group defined four molecular subtypes of HGSOC by gene expression analysis  that was validated in the TCGA project [3, 6]. The correlation of these molecular subtypes with histopathological features was addressed in very few studies [7, 8].
Applying the histopathological features described in the literature to the current retrospective study allowed categorization of the cases into 4 groups including mesenchymal type represented by 29% of cases, the proliferative type which included 16.5%, immune reactive type that included other 16.5% of cases, and differentiated type represented by 32% cases; 15% of them had SET features and 22% had papillary architectural features. This study aimed to confirm this subtyping by IHC approach. However, since chemotherapy can alter the IHC expression of markers, cases were divided into 2 groups according to their treatment protocol (primary versus interval debulking) to allow accurate analysis of IHC markers.
In both protocol groups, proliferative subtype showed significant association with Ki67 > 25%. Similarly, the immunoreactive subtype showed a significant association with CD8 (≥20 TILs/HPF). On the contrary, no distinct immunoprofile was found for mesenchymal or differentiated subtypes. These findings were consistent with the study of Popa and colleagues who found that mean Ki-67 expression correlated with the high mitotic count >30/10HPF in their HGSC cases . Additionally, Darb-Esfahani et al. included cases of HGSC that received neoadjuvant chemotherapy and molecularly proved to be immunoreactive subtype. They found >50% of patients with this subtype were characterized by intratumoral enrichment of CD8 positive T cells . In concordance, Murakami et al. reported a significant correlation between CD8 lymphocytes and immunoreactive type .
Unexpectedly, ER showed a significant correlation with the proliferative subtype only in cases that underwent primary debulking. To some extent, this is similar to the reports of Popa et al. in their trial when most of their HGSOC cases showed high expression of ki67 and were positive for ER status, but they did not make a correlation between them .
Another study by Feng and colleagues applied a hormonal receptor-based classification of HGSC cases and found that cases that were ER-positive had the worst prognosis . In many reports, estrogen is proved to be the driver for ovarian cancer development, promoting proliferation as well as metastases through inhibition of cell to cell adhesion [19,20,21,22].
In the same treated patient group, a significant association was found between negativity for PR and E-cadherin in relation to low tumor progression. However, these results need to be validated on a wider scale since this significance may lose its weight due to the negativity of PR and E-cadherin that we found in most of our cases. Despite that, loss or decreased expression of E-cadherin is considered a feature of epithelial-mesenchymal transition (EMT) responsible for tumor dedifferentiation and invasiveness, which plays an important role in tumor progression in epithelial tissues .
Considering cases that received neoadjuvant chemotherapy (NACT), a significant correlation was found between high CD8 expression in lymphocytes within tumor cells and the differentiated subtype with a SET pattern. The Association of SET with diffusely dense TILs is known in most cases of BRCA1 mutation . However, the absence of this significant association in our cases that did not receive NACT was concerning. NACT proved their role in the induction of many local changes of the tumor microenvironment including activation of CD4+ and CD8+ T cells. This may be due to the presentation of degraded protein antigens by class I MHC molecules to stimulate CD8+ T lymphocytes to produce proinflammatory cytokines to kill tumor cells [24, 25].
In a trial to find an answer for the study question, whether there are prognostic differences between the histopathological groups, the current work found that most of the cases diagnosed at an advanced stage (III&IV) were mesenchymal and proliferative types but without statistically significant association. This is concordant to Murakami and coworkers who reported a statistically significant association between the mesenchymal type and advanced stage . The limited number of cases in the present study (85) compared to that of Murakami et al. (132) may be the cause for not approaching a statistically significant relation.
Moreover, the present study found a significant correlation between the histopathological subtype and progression-free survival where mesenchymal type had the least PFS and the differentiated type with SET features had the longest PFS. Similar findings were reported by the study of Murakami et al. for the mesenchymal type. On the other hand, they found the immunoreactive type to have the best PFS . In addition, the results of the current study are compatible with the study of Ohsuga et al. who suggested—by using CT—that cases with mesenchymal gene expression subtype had more mesenteric infiltration and wide peritoneal disease and shorter PFS .
In a more detailed analysis for factors significantly correlated with longer PFS, high PR expression, positive E-cadherin, absent LVI, and primary debulking were found. These findings are consistent with those reported by Faleiro-Rodrigues et al. (2004), Cho et al. (2006), Modugno et al. (2012), Feng et al. (2016), and Mohanty et al. (2019) [14, 17, 21, 26, 27]. While it could be logical for all these factors to prolong PFS, the effect of high PR expression is explained by the role of progesterone in promoting apoptosis of ovarian cancer cells as reported by Modugno et al. (2012) .
On the contrary, the current work did not find a significant correlation between the different histopathological subtypes and OS in contrast to the same previously mentioned studies that found that OS was better for immunoreactive type and worse in mesenchymal type cases [7, 8]. This may be due to the considerable number of cases that died because of the disease during the study period.
Longer OS in the current work is found to be significantly correlated with primary debulking surgery followed by adjuvant treatment as well as the use of combined taxol carboplatin-based chemotherapy. Similar findings are reported by Bristow and Chi (2006) and Murakami et al. (2016) [7, 28], but in contrast to the result of a recent study of Machida and colleagues .
Our work presented a detailed analysis for the histopathological features of HGSOC with evident histopathological findings that allowed categorization of cases into groups parallel to the molecular subtypes, with some of these features (the proliferative and immunoreactive subtype) are supported by IHC markers taking into consideration the management protocol that the patient received. These groups showed significant prognostic differences in terms of PFS that indicate the value of applying histopathological features during the evaluation of HGSOC and no longer considering it as a single tumor type.
On the other hand, the relatively small number of cases enrolled in the study limited us. This was due to the necessity for the selection of cases with the complete clinical and follow-up data. Additionally, the death of many cases during the study period hindered achieving a significant correlation with overall survival.