To the best of our knowledge, we are the first to compare FSA and PSA results from the URM of PUC cases and to demonstrate two separate patterns of tumor infiltration. In this study, we found a notable 50% discrepancy rate between FSA and PSA results for the URM. Moreover, we observed a consistent periureteral fat infiltrative pattern (adventitia invasion) in all the discordant cases, with sectioning errors and misinterpretation identified as the primary causal factors.

Submission of URM for intraoperative pathology consultation is common for bladder cancer patients undergoing RC [11]. However, studies about the significance of FSA of the URM in PUC are rare. Kaimakliotis et al. [5] reported that a 32% URM-positive status in PUC, which aligns closely with our results of a 40% URM positivity rate. In the study of Kaimakliotis et al. [5], the URM positivity rate in PUC was 8.6 times higher than that in conventional UC (2.3%). Furthermore, PUC showed a much higher URM positivity rate than that (17.9%) in micropapillary UC (MPC), another aggressive subtype [5]. Notably, all + URM in conventional UC and MPC involved only carcinoma in situ (CIS) on the endoluminal URM, whereas 5 of the 8 + URM cases in PUC showed tumor cell invasion without concurrent CIS [5]. In accordance with the literature, all + URM cases in our cohort exhibited only invasion of PUC tumor cells. These findings are consistent with previous studies indicating that PUC has a propensity to manifest disease on peritoneal surfaces, leading to common + URM and surgical upstaging [6, 7]. These results suggest a significant likelihood that residual tumor will remain at the operative site even after radical surgery. These observations are associated with the independent effect of plasmacytoid subtype histology on survival outcomes. It remains unknown whether + URM in PUC is independently associated with disease-specific or overall survival, but our findings in this study and those in the literature imply that a thorough assessment of intraoperative URMs is essential in PUC.

We found that + URM cases in PUC showed 2 distinct patterns of tumor infiltration. In our cohort, PUC tumor cells infiltrate through the submucosa and muscularis propria layer of the distal ureter, or tumor cells were noted along the adventitia over a longitudinal section of the ureter in the RC specimen. Kaimakliotis et al. [5] also reported that all + URM involving invasive tumor cells in PUC showed an infiltrative pattern of the subserosal and adventitia ureteral plane, and PUC tumor cells were also observed along the subserosal surface and ureteral adventitia on multiple, sequential surgical excisions. These findings explain that PUC tumor cells that penetrate beyond the detrusor muscle could potentially move along the bladder serosa beneath the peritoneal reflection or the endopelvic fascia. Once within Waldeyer’s space, they can be tracked along the distal ureteral subserosa and adventitia [5]. On the other hand, the mechanism of intramuscular invasion was elucidated by the study of Beunk et al. [12]. They explained that smooth muscle tumor invasion in the bladder involves multiple steps, beginning with epithelial cancer cells infiltrating the connective tissue that separates the epithelial and muscle layers, followed by muscle invasion. Here, bladder tumor cells exhibit chain-like invasion into smooth muscle tissue, showing characteristics of jammed or collective migration aligned with the linear endomysium [12]. They also reported that muscle invasion in cancer patients is frequently correlated with destructive growth and a worse prognosis [12]. Anatomically, the inner smooth muscle layer of the ureter merges with that of the contralateral ureter to form the superficial trigone [13]. The distal ureter is accompanied by a separate external smooth muscle layer (Waldeyer sheath) to the bladder [13]. As it traverses the bladder wall, the Waldeyer sheath merges with detrusor muscle fibers to form the deep trigone of the bladder, which is enclosed by the 2 ureter orifices [13]. These distinctive anatomical structures between the ureter and bladder can provide additional support for the 2 PUC tumor cell infiltration patterns highlighted in this study, as well as the observation that these 2 infiltration patterns align with the longitudinal ureter muscle. These explanations are in line with the findings in our cohort that PUC with + URM was significantly more common in the trigone or bladder neck, and all PUC cases with + URM showed tumor involvement in the ureteral orifice, tended to exhibit higher tumor and LN stages, and showed slightly more progressive behavior. The aggressiveness of PUC can be explained by the loss of E-cadherin. Several studies have demonstrated that decreased expression of E-cadherin, a membrane connective tissue protein, is associated with more aggressive pathology, loss of cell differentiation, increased cell invasion, and poor prognosis in UC [14,15,16]. Loss of membranous E-cadherin expression and its nuclear accumulation are highly associated with PUC [14, 15] and could allow PUC tumor cells to easily invade surrounding tissues, ultimately leading to an advanced stage and rapid progression.

In this study, discordance between FSA and PSA was observed in 3 of the 6 (50%) + URM cases in the histopathological examination of the permanent URM tissues. Interestingly, all the discordant cases exhibited the periureteral fat infiltrative pattern (adventitia invasion), and the causes of the discordance between FSA and PSA were sectioning errors and misinterpretation by the pathologist of tumor cells as inflammatory cells. These findings indicate that a thorough evaluation of the ureteral margin should be conducted, with a focus on the periureteral fat tissue. The freezing and thawing processes used for frozen sectioning can cause tissue damage, with fatty tissues being especially vulnerable, potentially resulting in tissue loss and sectioning challenges. When preparing frozen sections, it is important to include periureteral fat tissue in the embedding process. Additionally, if the periureteral fat tissue is not visible on the slide or in any suspicious cases, additional frozen section examinations should be conducted. Pathologists should also pay close attention to the periureteral fat tissue when interpreting frozen slides. Ultimately, this thorough assessment of intraoperative URM should be applicable to all UCs that include PUC. In addition to this, it is recommended that pathologists include detailed information on PUC in every pathological report, as lack of awareness regarding the plasmacytoid subtype during FSA appears to pose greater challenges. Given that PUC tumor cells show morphological resemblance to plasma cells, it can be suggested that including detailed information about PUC in pathological reports may assist pathologists in minimizing the risk of misinterpretation during FSA.

This study has several limitations. It had a retrospective design that could not avoid inevitable limitations such as selection bias. The small sample size caused by the rarity of this entity and our focus on PUC with a plasmacytoid subtype component greater than 25%, and inclusion of patients who received neoadjuvant chemotherapy could affect the overall data quality. Also, statistically significant differences were not found in recurrence and survival between + URM patients and -URM patients, although + URM patients showed slightly more progressive behavior in the present study. Larger cohorts and further well-designed multicenter studies are needed to clarify the clinical significance of frozen assessment in PUC and to establish strong recommendations. Despite those shortcomings, this study offers valuable insights based on currently available data. These findings can aid clinicians and pathologists in shared decision-making discussions about the management and prognosis of patients with PUC.