Thyroid transcription factor‐1 expression in lung neuroendocrine tumours: a gender-related biomarker?

Unambiguous evidence supports the relevance of gender difference in oncology, from the diagnosis to the response to treatments and treatments’ side-effects prolife [16‐18]. Overall, sex significantly influences the clinical and pathological features of cancer patients. These include disparities in incidence and mortality rates, clinical presentations including age, screening participation rates, site, stage and treatment utilisation, histopathology (including genetic and molecular features) and survival [19‐21]. Environmental and behavioural factors (e.g. smoking habit or metabolic syndrome onset) play a key role in this context [22‐24]. Moreover, biological (e.g. sex hormones) features have been showed to contribute to the differential risk in several tumour types [25, 26]. In the field of NET, indeed, the majority of available data arise from studies on gastro-entero-pancreatic (GEP) neuroendocrine neoplasms (NEN) [27]. A large population study, including 15,202 patients with pancreatic neuroendocrine tumours from The National Cancer Database (NCDB), suggested that men had more frequently tumours >2 cm, and poorly or undifferentiated tumours if compared to women [28]. Notably, no significant differences were found in the rates of lymph node involvement and metastatic recurrence after the surgical removal of the primary tumour. At the molecular level, MEN1 and DAXX mutations resulted more common in males and TP53 mutations in females, respectively. However, these data lacked to be confirmed at the multivariable analysis. Data of the Surveillance, Epidemiology and End Results Research (SEER) registry, based on 43,751 patients with GEP-NETs, demonstrated with multivariable analyses a prognostic value (in terms of OS) for sex with women associated to better outcomes (p < 0.001) [29]. In this analysis the 3-year survival rates resulted 84.6% and 87.7% and the 5-years survival rates of 80% vs. 84% for male and female, respectively. For lung NET a sex-difference has been described, with, even in this case, more favourable trends in female in a previous work by our group [13]. According to the Cox-univariate regression model a significant impact on patients’ outcome for sex was demonstrated, with male sex associated with dismal PFS (p < 0.0001) and OS (p < 0.0001). These data have been further confirmed by data coming from SEER registry, where female sex was associated with better OS compared with male sex (p < 0.001) [30]. Furthermore, few works suggest biological differences in terms of tumour aggressiveness in relation to patients’ sex [13]. Female sex has been associated with a more indolent disease, both considering a lower tumour stage (specifically, negative nodal status vs. positive) and also with regards to pathological features, as lower tumour grade (G1-2 vs. G3), lower Ki67 index and reduced mitotic count. In addition, a sex imbalance of the histological subtype (TC vs. AC) in males and females has been reported [31]. However, a sex-related distribution of the main IHC NET biomarkers (Chromogranin A, NSE and, specifically, TTF-1) has never been reported in literature. A retrospective study including 11 carcinoid tumorlets (TLs), 36 TC, 17 AC and 16 large cell neuroendocrine carcinomas (LCNECs) showed a more common positive TTF-1 IHC staining in LCNECs (5 of 6 positive cases), followed by TLs (4 of 8) respect to AC (1 of 4), and TC (0 of 10) [32]. Interestingly, in this study the percentage of female was higher in these two categories, LCNEC and TLs, whereas both TC and AC were well-balanced among male and female. In the current work we observed a statistically significant association between the IHC positivity for TTF-1 and female lung NET patients (p = 0.007). This result deserves further studies to confirm a potential biological significance of TTF-1 expression in connection with the sex of lung NET patients, with potentially relevant implications in the diagnostic work-up of these tumours.

TTF-1⁺ alveolar type II epithelial cells have been demonstrated to be the major source of vascular endothelial growth factor (VEGF) in the lung [33, 34]. At the molecular level, TTF-1 has been postulated to positively regulate VEGF expression and the major signalling receptor for VEGF as VEGFR2 lung cancer epithelial cells [34, 35]. TTF-1, indeed, has been suggested to reprogramme lung cancer secreted proteome into an antiangiogenic state. Interestingly, TTF-1 has been assessed as a potential predictive factor for antiangiogenic treatment in non-squamous NSCLC [36]. In this study, the 92 TTF-1-positive patients presented higher response rates (51.4% vs. 27.3%, p = 0.027) and PFS (216 days vs. 137 days, p = 0.012) in the group treated with the antiangiogenic bevacizumab to standard chemotherapy, whereas in TTF-1-negative patients no clinical benefit was obtained by the combination therapy (chemotherapy plus bevacizumab). Unfortunately, data about VEGF expression and TTF-1 in lung NET are lacking. In a previous work by our group, we demonstrated a significant association between the absence of expression of the TTF-1 and the presence of hypoxia (in 14/16, 87.5%, of TTF-1-negative cases, p = 0.012). Among hypoxia-negative cases, 11/13 (84.6%) were TTF-1 positive, whereas among hypoxia-positive cases, 10/24 expressed TTF-1 [14]. In the present study we detected a statistically significant correlation among TTF-1 positivity and the absence of necrosis (p = 0.018). Taken all together this data, it is possible to hypothesise that TTF-1 may be positively linked to increased angiogenesis, and associated with lower hypoxia and the absence of necrosis in lung neoplasms, potentially including lung NET.

Finally, in the present work we investigated the prognostic value of TTF-1 in our lung NET population. According to available evidences, TTF-1 positivity is considered an established positive prognostic factor for lung adenocarcinomas [10]. In lung NET field, more conflicting data have been reported. In a retrospective series of 370 lung NET, a difference in IHC positivity for TTF‐1 was found between patients with higher or lower Ki-67 [11]. Overall, a positive staining for TTF-1 was detected in 49 (17.1%) of the included lung NET, with TTF‐1 positivity in 30 (13.0%) of the low Ki‐67 group of patients and in 19 (34.5%) cases of the high Ki‐67 group. The second group (with higher Ki-67) was associated to a worse prognosis (p < 0.0001). Also, TTF-1 positivity correlated with a reduced survival outcome (p = 0.03). In a retrospective study of 34 lung NET treated with peptide receptor radionuclide therapy with (177) Lu-DOTATATE (Lu-PRRT), survival outcomes in terms of PFS were better in TTF-1 negative cases if compared to TTF-1 positive ones (26.3 vs. 7.2 months, respectively, p = 0.0009) [37]. However, these data have not been confirmed in subsequent works [12, 38]. A retrospective analysis of 108 lung NET lacked to demonstrate a correlation between TTF-1 positivity and patient outcomes [12,]. In another study, TTF-1 was positive in 78% of the 133 lung NET cases but was not associated with patients’ survival [38]. In the present study, in line with the available literature data, no significant association among TTF-1 expression and patient survival was observed (according to univariable and also multivariable model), despite a favourable trend for TTF-1 positive cases was noticed. Further prospective studies are encouraged to determine if this biomarker has a prognostic relevance for lung NET.