An amassing body of literature supports the role of immune cells in ovarian cancer proliferation. In 2003, Zhang et al reported a seminal study in which tissue from 186 patients was assessed for the degree of CD3⁺ tumor-infiltrating T-cells. ^[1] The presence of these cells was found to independently predict delayed recurrence and prolonged survival, with a large magnitude of effect. Several other studies have yielded the same conclusion, summarized recently in a meta-analysis by Hwang et al.^[2] With a cumulative experience of 1,815 patients across 10 studies, it was demonstrated that a lack of tumor-infiltrating lymphocytes (TILs) was associated with poorer survival (hazard ratio [HR] 2.24; 95% CI 1.71–2.91). In these studies, TILs were characterized by expression of CD3 and/or CD8. ^[1], ^[3], ^[4], ^[5], ^[6], ^[7], ^[8], ^[9], ^[10], ^[11] The observed correlations between T-cell infiltration and clinical outcome is a biologically plausible one – specifically, CD8⁺ T-cells may cause a direct cytolytic antitumor effect.

With a greater understanding of T-cell biology, it has become clear that certain subsets of T-cells may actually have a procancer effect. Specifically, Barnett et al have assessed the extent of regulatory T-cell (Treg) infiltration in a series of 232 primary serous ovarian cancer specimens. ^[12] In this series, the extent of Treg infiltration was associated with higher cancer grade and advanced clinical stage. Corroborating this data, peripheral blood assessments from patients with ovarian cancer suggest and increasing number of circulating Tregs accompanying disease progression. ^[13]

The role of B-cells in ovarian cancer is more challenging to discern. ^[14] In early studies using murine models of B-cell deficiency, it was suggested that B-cells may actually inhibit the antitumor effect of tumor infiltrating T-cells. ^[15], ^[16] As a potential mechanism, B-cells have been demonstrated to inhibit the priming effect of CD4⁺ T-cells on CD8⁺ T-cells. ^[17] Despite these observations, clinical evidence points to a link between CD20⁺ tumor infiltrating B-cells and a favorable outcome in the setting of ovarian cancer. Specifically, Milne et al examined tissue microarray (TMA) data derived from 199 patients with high-grade serous epithelial ovarian cancer (EOC). ^[18] In this report, the presence of intraepithelial CD20+ cells was associated with an improvement in disease-specific survival (DSS). However, it is important to note that this report was accompanied by other paradoxical observations – for instance, the presence of FoxP3⁺ regulatory T-cells (Tregs) were also associated with a favorable DSS. This finding contradicts the preponderance of evidence suggesting that Tregs may have a pro-cancer effect.

Clearly, the relationship between B-cells and clinical outcome warrants further assessment. In the current manuscript, we assess the extent of B-cell infiltration in omental tissue derived from patients with ovarian cancer, and attempt to correlate the extent of infiltration with overall survival (OS). We have previously demonstrated that phosphorylated signal transducer and activator of transcription-3 (pSTAT3) plays a critical role in cancer-related inflammation and tumor progression through recruitment of myeloid cells and Tregs. ^[19], ^[20] More recently, we have demonstrated a critical role of STAT3 in promoting B-cell mediated tumor angiogenesis in mouse models (unpublished data). To this end, we further explored the association of between pSTAT3 and B-cell infiltration.

Several unique observations emerge from the current study. A greater extent of B-cell infiltration in omental tissue appears to correlate with poorer survival. B-cell infiltration is associated strongly with pSTAT3 expression in omental tissue, and in fact, increased pSTAT3 expression in omental tissue is also associated with poorer survival. On multivariate analysis, B-cell infiltration entered the final multivariate model along with clinical stage, and a strong association was noted between pSTAT3 expression and B-cell infiltration.

As already noted, the impact of T-cell infiltration is relatively well documented in the setting of ovarian cancer, now summarized in a meta-analytic dataset. ^[2] These correlative studies have spurned efforts to employ adoptive transfer of T-cells, both alone and in combination with cytotoxic therapy. ^[21], ^[22], ^[23] The effects of relevant T-cell subsets, such as the procancer effects of Tregs, have also been discerned. The role of B-cells remains more controversial – although traditionally thought to have an antagonistic effect on cytotoxic T-cell responses, the aforementioned dataset from Milne et al suggests that tumor infiltrating B-cells may be a positive prognostic marker. However, it is important to note that our study utilized a distinct B-cell marker (CD19), and also assessed a more heterogeneous group of patients (patients who received secondary debulking and prior systemic therapy). With respect to the latter, we found no significant difference in outcome based on these clinical features, and surprisingly, the distribution of biomarkers did not vary in these subgroups. Our study also assessed individual sections derived from paraffin-embedded specimens; in contrast to other studies evaluating TMAs, this approach allowed for assessment of multiple fields (10 per specimen) to provide a more accurate biomarker assessment.

The close association between B-cells and pSTAT3 in this study supports preclinical data we have previously reported delineating the role of pSTAT3 in immune cell recruitment. ^[24] Our preclinical data also suggest that abrogation of STAT3 activity may have an antitumor effect, consistent with our clinical observation that higher pSTAT3 may be associated with poorer clinical outcome. ^[25], ^[26] In combination, if these clinical observations are confirmed in larger series, this may serve as rationale to pursue therapeutic trials of STAT3 antagonists to target B cells in ovarian cancer. As one example, we have developed a CpG-conjugated siRNA directed at STAT3 in myeloid cells and B cells. ^[26] While the delivery of siRNA-based therapies has been a formidable challenge, ovarian cancer may be an ideal disease entity to approach with these agents. Intraperitoneal chemotherapy has a well-established survival benefit in phase III studies, and intraperitoneal delivery of RNA-based therapeutics may circumvent certain stability and solubility issues. ^[27] In addition to directly antagonizing STAT3, an approach targeting upstream moieties such as JAK2 could be considered. Several JAK2 inhibitors have been developed for the treatment of myeloproliferative diseases bearing aberrations in this protein; it has been demonstrated that one of these small molecules, AZD1480, has an antitumor effect in the MDAH2774 ovarian cancer cell line. ^[28], ^[29].

Several limitations should be acknowledged. Firstly, our sample size was relatively small in contrast to other published reports. ^[1], ^[30] Secondly, we did not assess T-cell populations (i.e., CD8⁺ T-cells and FOXP3⁺ regulatory T-cells) to confirm previously reported findings. ^[30] Third, it remains unclear what effect certain treatment variables (i.e., prior chemotherapy) may have had on the observed B-cell or pSTAT3 distribution – this warrants further study in a larger series. Finally, our study exclusively focuses on tissue-based markers. Although conventional serum markers such as the CA-125 have fallen out of favor after recent prospective assessments, there are a multitude of new serum-based assays on the horizon. ^[31] For instance, autoantibody profiling studies have identified signatures that differ between ovarian cancer patients and healthy controls. ^[32] It will be curious to see whether a combination of the biomarkers we have proposed herein with others may yield superior personalized prognostic tools. Despite these limitations, our data adds to a very limited body of literature related to the impact of B-cell infiltration on clinical outcome in advanced ovarian cancer, and provides a plausible mechanism for this to occur (namely, B cell recruitment via STAT3-dependent pathways). Clearly, larger efforts will be needed to validate the observations made herein. With a shadow cast over traditional biomarkers such as the CA-125, immune markers hold immense promise in ovarian cancer. ^[33] Given a burgeoning pipeline of JAK/STAT pathway inhibitors, our data linking pSTAT3 expression and B-cell infiltration to clinical outcome may ultimately support a novel therapeutic avenue for this disease.