Descriptions of primary pulmonary NMC are rare and have only been reported in a few case reports and case series; thus, it is necessary to summarise the clinical features and treatment of this entity. Herein, we summarised the clinical and pathological processes of seven patients with primary pulmonary NMC and explored the efficacy of different treatment options. Furthermore, WES was used to explore the TMB, gene fusions, and potential coexisting mutations. We also explored the relationship between TMB and immunotherapy efficacy.
The median ages of diagnosis for patients with lung cancer and NMC overall were 70 and 24 years, respectively, while the median age of this group of patients was 31 years (mean 42 years) 7,22. All patients were diagnosed in an advanced stage with a poor PS score. Before diagnosis, all patients complained of a cough lasting for more than a month and some patients had difficulty breathing, chest pain, fever, or haemoptysis. Four patients had a significant smoking history while three patients had no or negligible tobacco exposure. Consistent with previous studies, all patients had no EBV exposure, suggesting that EBV was not associated with NMC 5. No longer restricted to paediatric patients and midline structures, with the development of diagnostic techniques, this entity has been reported in patients of all ages (0–81.7 years) and in non-midline structures 19. Therefore, regardless of the primary lung mass in young or elderly patients, even those without a smoking history, NMC should always be included in the differential diagnosis of malignant lung tumours when the clinical course appears to be aggressive.
Similar to the resulted reported by Lynette, the imaging results lacked specificity but had some distinct features 9. Chest CT showed a large primary pulmonary mass (diameter ≥ 5 cm) fused with the hilar and mediastinal lymph nodes, with obstructive atelectasis and ipsilateral pleural involvement, while the contralateral lungs were essentially unaffected. Furthermore, local invasion involved the oesophagus and back, with distant metastases mainly involving the adrenal glands and bone. In our study, bone metastases were the most common (3/7) extrathoracic metastases assessed by CT and PET/CT. Brain metastases were most common site in lung cancer compared to other cancers with cumulative 1-years incidence rates of 3.0% in patients with Stage III non-small cell lung cancer (NSCLC) and 10.8% in patients with Stage IV NSCLC 23,24. Brain metastases from lung NMC have not been reported and it is often believed that the tumours progress too rapidly to cause brain metastasis. Consistent with previous studies, no brain metastases were observed by brain CT or MR, including in the three patients who survived for more than 1 year.
The cytomorphology of NMC originating in the lungs and other sites has been extensively described 5,9,25. The histological manifestations of this entity can be confused with other high-grade lung malignancies and lead to the misdiagnosis of NMC 9. In our series, two patients were initially diagnosed with mucinous epithelial carcinoma and squamous cell carcinoma before undergoing NUT IHC. Therefore, the possibility of NMC lung cancer cannot be ruled out in cases manifesting as other poorly differentiated tumours. Consistent with previous studies, cases with pulmonary NMC mainly expressed keratin, P63, and/or P40 in IHC, which suggests that NMC cells are derived from squamous cell carcinoma 9,25,26. Expression of neuroendocrine cancer markers was not observed by IHC in our patients. Two patients (Cases 2 and 7) were positive for TTF-1 expression, which favoured the diagnosis of lung adenocarcinoma 27. The significance of TTF-1 expression in this entity is unclear. From a diagnostic point of view, the co-expression of TTF-1 with extensive squamous differentiation markers such as p63/p40 is very rare in lung cancer and NMC should be considered 9,28.
The diagnosis of NMC relies on NUMT1-positive IHC (≥ 50%) findings because NUMT1 protein expression indicates the presence of a NUTM1 gene translocation. FISH using NUTM1 split-apart probes are a sensitive approach for detecting NUTM1 rearrangement 5. However, in our case series, only one patient was positive by FISH. Given that Haack et al. reported a new probe spanning NUT rather than two flanking probes to confirm the presence of cryptic BRD4-NUT rearrangements in two NUTM1-IHC-positive but standard FISH-negative patients, a possible explanation for our result is that the NUTM1 fusion gene break sites of these patients were not covered by the probe we used. 12 In addition, although a small number of fusion genes do not meet the diagnostic criteria for FISH, they may result in high expression of oncoproteins under other regulatory mechanisms at the level of gene transcription and translation. Therefore, the small number of fusion genes or the limitations of the probe may have caused the negative FISH results in our NUTM1-IHC-positive specimens.
Four patients in our series underwent WES due to fewer specimens and unqualified quality assessment. Among them, only Case 7 (with the lowest TMB) was confirmed to have a rare fusion of CHRM5-NUTM1 (intron1-5UTR, 12.47%). We did not identify a NUTM1 fusion partner in the remaining three patients with high TMB. This may be caused by the significant limitations of WES as it does not capture intronic regions where fusion breakpoints often occur 29. In addition, samples from three patients with high TMB had significant DNA degradation, which may have led to false-positive mutations, resulting in significantly higher TMB. ATXN3 and ZNF429 are co-mutated genes detected in all samples but were not found to be associated with cancer pathways in functional enrichment analysis. In short, while WES can be used to detect the fusion partners of NUTM1, it has limitations and specimen preservation significantly impacts TMB detection.
The published mOS of the largest series of NMCs originating from all sites was 6.7 months 6. In our series, other than Case 5, patients with lung masses died of the disease within 5 months despite receiving multiple treatments. The mOS of patients with lung masses was only 2.75 months, while that of patients with tracheal masses was significantly longer than 1 year. These findings suggest that NMC patients with tracheal masses have a longer OS than that in patients with lung masses.
There is currently no standard protocol for the treatment of primary pulmonary NMC, and exploration of effective treatment options is urgently needed. While surgery is the most effective treatment for patients with early-stage primary pulmonary NMC, it may not prolong survival in patients with advanced disease. In our series, Case 1 underwent surgery but eventually died of surgical complications. Although patients with primary pulmonary NMC can benefit from BETi even after multi-line treatment, the difficulty in obtaining the drug limits its application 21. Therefore, chemotherapy or radiation therapy is inevitable in patients with advanced disease. Consistent with previous studies, chemotherapy did not significantly prolong PFS and OS in the patients in our series, whereas concurrent chemoradiotherapy significantly prolonged the OS in Case 6. 6 Therefore, concurrent chemoradiotherapy may play an important role in prolonging patient survival.
Rapid squamous differentiation caused by knockdown of NUT fusion oncogenes in NMC cell lines suggests that NMCs are derived from squamous cells 30. PD-1/PD-L1 inhibitors prolong OS in NSCLC patients and, therefore, may be effective against primary pulmonary NMC 31. Immunotherapy is more effective in patients with high TMB and/or PD-L1 expression32,33. Among the patients in our study with pulmonary masses administered immunotherapy, only Case 5 received radiotherapy as a first-line treatment; this patient showed significant efficacy, with an OS of 19.5 months. In contrast, Case 3 received chemotherapy as a first-line treatment and showed a very poor prognosis. Although Case 1 showed high TMB and received immunotherapy, he died of severe postoperative complications one month after surgery. These findings suggest that highly lethal radiotherapy may improve the efficacy of immunotherapy and increase patient survival. The high TMB may better explain the long survival of Case 2, who was administered immunotherapy. In contrast, although Cases 6 and 7 had high and low TMB, respectively, patients with tracheal masses administered second- or third-line immunotherapy showed significantly long PFS. However, given the small number of cases and the long survival time of patients with primary tracheal tumours, it is difficult to prove that high TMB predicts better immunotherapy efficacy in these patients. Radiotherapy plus immunotherapy may prolong survival in patients with NMC originating from the lungs; although the effect of first-line immunotherapy cannot be assessed in patients with NMC originating from the trachea, immunotherapy may significantly prolong their survival.
Our study detailed the clinical and pathological features of lung NC and treatment experience and explored the diagnosis of WES in NC. Our case series summarised the largest number of patients administered checkpoint immunotherapy in lung NC and explored the efficacy of immunotherapy. The results suggested that immunotherapy can benefit patients with these deadly tumours. However, our study has certain limitations. This retrospective study of rare lung NC included a limited number of patients. The initial diagnosis of all patients was based on positive NUT-IHC results and the commercial probe did not detect the presence of NUT rupture in all patients. Based on these limitations, the results of exploration of immunotherapy in lung NC remain controversial. More studies on lung NC are needed to confirm our findings.