In this study, we investigated the association between STAS and clinicopathologic features, molecular alterations, the expression of immunohistochemical markers, and prognostic significance in both ADC and SQCC based on Chinese patients. We found that STAS was associated with aggressive clinicopathologic features, wild-type EGFR, rearranged ALK or ROS1, low expression of E-cadherin and high expression of vimentin, Ki67, and survivin. In the prognostic analysis, STAS was associated with poor DFS and OS in ADC by univariate analysis and was an independent risk factor for DFS by multivariate analysis. In addition, STAS was associated with poor DFS in single stage I, II, IA, IB, and IIA ADC patients according to the new 8th edition AJCC/UICC staging system.
Since 2018, a few studies have discussed the significance of STAS based on the 8th edition AJCC/UICC staging system, and the reported results mainly focused on ADC [7,8,9,10,11,12,13,14,15]. Some attention has been paid to the significance of STAS in stage I patients; however, few studies analyzed the significance of STAS in other stages exclusively. Recently, Terada and colleagues found that STAS was an independent predictor of recurrence in stage III (N2) ADC [15]. In the current study, STAS was found to be associated with poor DFS and OS in stage I-II patients but not in stage III-IV cases. This observation indicates that the prognostic significance of STAS mainly exists in early-stage ADC cases, and pathological evaluation of STAS should be performed more cautiously for these tumors. In the analysis of single-stage ADC, STAS was associated with poor DFS in stage I, II, IA, IB, and IIA patients, but not OS. These results reveal more details on the significance of STAS with respect to recurrence. When STAS is observed in these lymph node-negative ADCs, close follow-up should be implemented. Further studies are needed to discuss whether these patients need post-operative adjuvant therapy.
Only a few studies have analyzed STAS in SQCC. In SQCC, the incidence of STAS was generally lower than that in ADC, which was from 19.1% [16] to 40.3% [17]. Positive STAS was observed to be associated with larger tumor size, lymphovascular invasion, tumor necrosis, high-grade tumor budding, larger nuclear diameter, higher mitotic counts, and higher T, N, and p-stages [16,17,18]. In survival analyses, STAS was also reported to be a significant predictive factor of DFS and OS [16,17,18], especially in stage I patients [16]. In the current study, STAS was associated with shorter OS, although this trend was not statistically significant, and no correlation was found between STAS and DFS. This could be because the simple size of the current study was smaller than that of previous reports.
The association between STAS and molecular characteristics has not been clearly explicated. Molecular characteristics were exclusively studied in ADC. STAS was frequently observed in tumors with ALK and ROS1 rearrangements, BRAF mutations, or wild-type HER2 [6, 7, 19,20,21]. In the current study, 95.8% (23/24) cases with ALK rearrangements and all cases with ROS1 rearrangements demonstrated STAS, and this observation was similar to that of previous results. Three articles reported the association between STAS and KRAS mutations; one study concluded that STAS was frequently observed in tumors with KRAS mutations, whereas the other two reported no association [7, 19, 20]. Our results also concluded no association between STAS and KRAS mutations. However, as the KRAS mutation rate is quite low in Asian patients, more data are needed to clarify this issue. Regarding EGFR mutations, the reported results have varied among different studies. According to Hu and colleagues, STAS is frequently observed in tumors with EGFR mutations [7], whereas three other studies demonstrated that STAS was associated with wild-type EGFR [19,20,21]. In contrast, in some studies, no correlation was observed between STAS and EGFR [22,23,24]. In the current study, STAS was observed to be associated with wild-type EGFR. One possible explanation for the different frequencies of STAS based on different driver gene alterations could be that STAS is more frequently observed in poorly differentiated tumors including those with a solid/micropapillary pattern [25], and ALK or ROS1 rearrangements mainly exist in ADC with a predominant solid pattern [1, 26]. In contrast, STAS is also associated with a non-lepidic pattern [1, 7, 19, 20], whereas EGFR mutations were more frequently detected in ADC with lepidic growth [25].
The association between STAS and the expression of immunohistochemical markers was barely understood and the correlation between STAS and EMT has been poorly discussed. In ADC, positive STAS was reported to be significantly associated with tumor stroma metastasis-associated protein 1 expression levels [8] but was not significantly correlated with programmed death ligand 1, thyroid transcription factor 1, napsin, or CK7 expression, as well as Ki67 activity [19, 22, 23]. In the present study, STAS was found to be associated with lower E-cadherin and higher vimentin and Ki67 expression. In SQCC, previous reports concluded that STAS is associated with an increased tendency for high vimentin and Ki67 expression in comparison with levels in patients without STAS; however, the expression of p53 and E-cadherin was not associated with the status of STAS [16,17,18]. In the present study, STAS was found to be associated with lower E-cadherin and higher vimentin and survivin expression in the SQCC cohort. These results indicate that STAS might be more likely to be present in tumors exhibiting EMT features. EMT is a process by which epithelial cells transform into mesenchymal stem cells by losing their cell polarity and cell-to-cell adhesion and gaining migratory and invasive properties, and this process has been identified as an indicator of poor prognosis in non-small cell lung cancer [27]. Even though a relationship was found between the presence of STAS and EMT features, whether STAS cells underwent EMT remains unclear. According to Yagi and colleagues [28], the survival of STAS cells relies on blood vessel co-option, and these cells are E-cadherin-positive. This result, to some extent, challenged the opinion that STAS cells undergo EMT. In agreement with previous reports, the present results suggest that EMT might be a risk factor but not a mechanism for STAS, as tumors with EMT features were found to be more discohesive with fewer intercellular adhesions; this, it would be easier for the malignant cells to detach from the main tumor.
Our study had some limitations. On one hand, some early-stage patients in the present study received limited resection, and some patients with late-stage tumors received adjuvant therapy. These conditions might have influenced the prognosis and could affect the results of survival analysis. On the other hand, the sample size involved in the present study was small, especially for SQCC, and the patients were from one single institution.