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Association of small foci of diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH) with adenocarcinoma of the lung
Pathology – Research and Practice 209 (2013) 578–584
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Original article
Association of small foci of diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH) with adenocarcinoma of the lung Masoud Mireskandari a , Azita Abdirad a , Qing Zhang a , Manfred Dietel b , Iver Petersen a,∗ a b
Institute of Pathology, Jena University Hospital, Friedrich Schiller University, Jena, Germany Institute of Pathology, Charité – University Medicine Berlin, Campus Mitte, Berlin, Germany
a r t i c l e
i n f o
Article history: Received 29 January 2013 Received in revised form 27 May 2013 Accepted 25 June 2013 Keywords: Diffuse idiopathic pulmonary neueroendocrine cell hyperplasia (DIPNECH) Adenocarcinoma of lung Lung preneoplasia Grading of lung adenocarcinoma
a b s t r a c t DIPNECH is regarded as a precursor lesion of neuroendocrine lung tumors, specifically carcinoids. A relationship with lung adenocarcinomas has not been clearly established so far. We present a series of four cases with a concomitant presence of adenocarcinoma and DIPNECH in the lung. The cases were retrieved from the archives of the Institutes of Pathology of the Jena University Hospital and the Charité, Berlin. The clinical data were collected from the hospital information system. The microscopic findings of adenocarcinoma and DIPNECH were reviewed. A panel of neuroendocrine and epithelial markers was analyzed immunohistochemically. In addition, the H&E slides of a series of 82 lung carcinomas were reevaluated for the presence of DIPNECH foci and the parameters of the IASLC/ATS/ERS classification for lung adenocarcinoma. DIPNECH foci were composed of small intramucosal nests of proliferating pulmonary neuroendocrine cells alongside or at the periphery of terminal airways. All detected foci measured less than 5 mm in maximal diameter and showed a consistent reactivity against Synaptophysin. They did not express epithelial markers of squamous cell carcinoma and adenocarcinoma. In three cases, the DIPNECH foci were clearly associated with the adenocarcinoma, while in one case, they were observed in the non-neoplastic lung tissue. The adenocarcinoma with DIPNECH inside mainly showed low grade histology, while the fourth case was intermediate to high grade. The histologic evaluation of the HE slides of the other 82 lung cancer cases showed no suspected or definite DIPNECH foci. Within this series, we could confirm the prognostic significance of the IASLC/ATS/ERS classification of lung adenocarcinoma. Our series suggest that a subset of lung adenocarcinoma is characterized by the concomitant presence of DIPNECH within the tumor, suggesting a causal relationship. These adenocarcinomas seem to be low grade ones, and may have a particular tumorigenesis and clinical behavior. These observations need to be confirmed in larger tumor collectives. We could confirm the prognostic relevance of the new adenocarcinoma classification. © 2013 Elsevier GmbH. All rights reserved.
Introduction
Definition and clinical symptoms of DIPNECH
Recently, during the histologic evaluation of two cases of adenocarcinoma (ADC) of the lung in our institute, we noticed the presence of small DIPNECH foci in the lung tissue. Together with two other cases from a collection of lung tumors obtained from the Charite Institute of Pathology in Berlin, they constitute a series of four cases with simultaneous involvement of lung tissue with adenocarcinoma and DIPNECH.
DIPNECH is an exceedingly rare lesion. It was not completely recognized until the early 1990s, although before that time, cases with similar clinical and pathologic findings but with different nomenclature had been reported [1]. To date, fewer than one hundred cases have been documented in the literature [2]. WHO defines the DIPNECH as follows: a generalized proliferation of scattered single cells, small nodules (neuroendocrine bodies), or linear proliferation of pulmonary neuroendocrine cells (PNCs) that may be confined to the bronchial and bronchiolar epithelium [1]. Practically, it is very difficult to separate the reactive proliferations of neuroendocrine cells and formation of tumorlets from DIPNECH. Gosney and colleagues believe that DIPNECH is usually more florid. It accompanies in many cases the carcinoid tumor of the lung, often
∗ Corresponding author at: Institute of pathology, Universitätsklinikum Jena, Friedrich-Schiller-Universität, Ziegelmühlenweg 1, D-07743 Jena, Germany. Tel.: +49 3641 933120; fax: +49 3641 933111. E-mail address: [email protected] (I. Petersen). 0344-0338/$ – see front matter © 2013 Elsevier GmbH. All rights reserved. http://dx.doi.org/10.1016/j.prp.2013.06.019
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leads to obliteration of small airways and, in contrast to reactive proliferations, shows no significant pathologic alteration in the surrounding lung tissue apart from mild fibrosis and inflammation [3]. They could also show a meaningful difference in the expression pattern of cell proliferation and death antigens (e.g. p53, p16 and Ki67), as well as CD10 (CALLA), between reactive proliferations and DIPNECH. In many case reports and studies was DIPNECH histologically defined as an intramucosal proliferation of PNCs that is confined to the basement membrane. The proliferations extending beyond the basement membrane were called tumorlets when they were smaller than 5 mm in maximal diameter and carcinoid when they are larger than five millimeters [4–7]. DIPNECH, as a diagnostic term, is reserved only for cases in which the hyperplasia is diffusely primary in nature [8], which was also reflected in the nomenclature of the lesion and implication of the term “idiopathic” to describe it. In some case reports, the different clinical symptoms have been attributed to the presence of DIPNECH. Some of these clinical findings are recurrent pneumonia [4], unexplained bronchial asthma [5], and hyper-adrenocorticotropic symptoms [6]. Pulmonary preneoplasia Classically, three lesions are considered preneoplastic lesions of the lung. According to the WHO-classification of lung tumors [1], as well as many other studies [7,9–12], epithelial dysplasia, atypical adenomatous hyperplasia (AAH), and DIPNECH are considered preneoplastic lesions for squamous cell carcinoma (SCC), adenocarcinoma (ADC), and carcinoid tumor of the lung, respectively. Until now, no definite preneoplastic lesion for small cell carcinoma of the lung has been detected. New classification of adenocarcinoma of the lung In 2011, a group of experts proposed and published a new classification for adenocarcinoma of the lung [13]. The classification was supported by the International Association for the Study of Lung Cancer (IASLC), the American Thoracic Society (ATC), and the European Respiratory Society (ERS), and is expected to be integrated in the new edition of the WHO classification of lung tumors. According to this new classification, three parameters play an important role in the diagnosis and classification of lung adenocarcinomas. They include the presence or absence of invasion, size of invasion, and tumor size, as well as the predominant histological growth pattern. The following categories were generated: preinvasive lesions (atypical adenomatous hyperplasia/AAH, adenocarcinoma in situ/AIS), minimally invasive adenocarcinoma/MIA (≤3 cm in total tumor diameter, lepidic predominant growth pattern, and with ≤5 mm invasion), and invasive adenocarcinomas (with >5 mm invasion or predominant non-lepidic growth). Furthermore, the term/entity bronchioloalveolar adenocarcinoma was eliminated because it was previously associated and thus confused with entirely distinct lesions, such as AAH, AIS, MIA, lepidic predominant adenocarcinoma/LPA, and invasive mucinous adenocarcinoma. Since most of the lung ADCs are heterogenous in microscopic appearance and display a spectrum of histologic patterns, the new classification suggests the application of the term “predominant” to describe the most extensive histologic growth pattern and then to describe all other patterns in percentage in 5% increments. This form of reporting provides a basis for the grading of lung ADCs. It has been shown that among different morphologic growth patterns, solid and micropapillary patterns have the worst prognosis and the non-mucinous lepidic growth pattern the most favorable one. Other patterns, including papillary and acinar/cribriform, constitute the ADCs with intermediate prognostic association [14–16].
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Aims of present study The questions which we are going to address in this study are as follows: – Is DIPNECH a precursor lesion in a subset of lung ADCs? – Is there any characteristic feature for the DIPNECH-associated lung adenocarcinomas which can be discriminated from other ADCs of the lung regarding tumor phenotype (e.g. grading according to the new classification of lung adenocarcinoma or tumor-morphologic characteristics)? – Is there any DIPNECH-associated clinical sign or symptom which can lead the physician to suspect the possible presence of DIPNECH in a patient with lung adenocarcinoma? Materials and methods Cases All four DIPNECH cases of this study had undergone partial lung resection because of ADC of the lung. Detection of DIPNECH was an incidental finding during the microscopic evaluation of resection specimens. The hospital information systems were used to retrieve any potentially DIPNECH-related clinical symptoms (dyspnea, wheezing, cough, hemoptysis and chest tightness), as well as demographic data. Immunohistochemitry and molecular analysis Both tumoral tissue and DIPNECH foci were evaluated immunohistochemically using antibodies against TTF1, Napsin, CK7, CK 5/6, CD56, Synaptophysin, Chromogranin, and proliferation marker Ki67 according to the provider guidelines, as well as an automatic stainer (Dako, Germany). Microscopic re-evaluation of carcinomas of the lung To look for any presumably undetected DIPNECH foci during the first microscopic evaluation, we reviewed the HE slides of 81 cases of non small cell and one small cell lung cancer, which had been collected for other studies [17,18]. Only the slides containing both neoplastic and non-neoplastic lung tissue were reviewed. In total, 247 slides were reviewed. In 45 cases (about 55%) only one slide, in 29 cases (about 35%) between 2 and 5 slides, in 5 cases (about 6%) between 6 and 8 slides, and in only three cases (about 4%) 10 or more than ten slides were available for review. The mean number of slides per each case was about 3. The adenocarcinomas were graded according to the above described new lung ADC classification. Results Patients’ clinical data The age, sex, and other demographic attributes of four DIPNECH cases are summarized in Table 1. Briefly, three of four cases are female. The mean age of the patients was 66.5 years (55–72). The average diameter of ADC was 2.5 cm (2.0–3.0). At the time of diagnosis, all tumors were in early stages (pT1 to pT2 without lymph node metastasis). The search of the hospital information system for the clinical data of the two recently diagnosed cases (cases 1 and 2) revealed no clinical sign or symptom, which is attributable to neuroendocrine secretions. Interestingly, both cases had a history of other malignancies. Case number one experienced a stage Ib malignant melanoma of the skin two years before the present
S R R R 5 4 >10 1
Relation to ADC No. Of DIPNECH foci
Alive (10) Alive (10) Alive (30) Dead (13)
Fig. 1. Microscopic features of proliferating neuroendocrine cells.
admission without any sign of recurrence. Case number two had a papillary thyroid carcinoma which had been diagnosed 4 years before the diagnosis of the present lung tumor. The cases 3 and 4 are from another Institute, and there was no access to their clinical information. Histologic findings of lung adenocarcinoms with DIPNECH
Abbreviations: M, male; F, female; S, separate; R, related. a Architectural and cytologic grade, respectively [see the Refs. 14,15,18,19]. b Survival time after follow up in month.
pT2a, pN0 (0/38) pT1b, pN0 (0/10) pT2, pN0 (0/28) pT1, pN0 (0/14) 2/high 1/low 1/Low 2/Low 3.0 × 1.5 × 1.4 2.5 × 2.4 × 1.5 2.0 × 2.0 × 1.5 2.8 Right upper lobe Left lower lobe Right middle lobe Right upper lobe 72 69 70 55 1 2 3 4
M F F F
Tumor gradea Tumor size Tumor location Sex Age Case no.
Table 1 Clinical, demographic and pathologic findings of cases with ADC and DIPNECH.
Tumor stage
Survivalb
0.274–0.600 0.223–0.753 0.177–0.864 0.441
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Diameter Range (mm)
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Two ADCs were low grade with a predominantly lepidic growth pattern, and two were intermediate grade according to the histological growth pattern with a predominantly acinar growth pattern. One of two recent cases showed low grade features according to cytological criteria [18,19]. In all cases, the number of foci of PNC proliferation was more than one. In most cases, the foci of proliferating PNCs were seen as small solid syncytial nests of cells with uniform round to oval nuclei, a homogenous chromatin pattern without chromatin clumping, and a small to moderate amount of amphophilic cytoplasm (Fig. 1). The cell borders were not prominent. In most locations, the aggregates of proliferating cells grew alongside the airspaces or were located just at the periphery of the terminal airways (Fig. 2). The maximal diameter of each focus of DIPNECH was measured using a digital camera and “microscopic discussion” software from JVC. It did not exceed the threshold of 5 millimeters in all cases (Table 1). In some locations, they formed aggregates of variably and usually pinpoint sized nests with intervening fibrous stroma (Fig. 3a and b). The relationship between tumoral tissue and DIPNECH foci was different (Table 1). While in case one, the DIPNECH foci were detected completely out of the confines of tumor in the neighboring non-neoplastic lung tissue, in other three cases, the DIPNECH foci were located right in the middle (case number 3) or at the border but inside the tumor area (case number 4). Immunohistochemistry The immunohistochemical findings of DIPNECH foci and adenocarcinomas are summarized in Table 2. The most consistent positive immunohistochemical finding in the foci with PNC proliferation (DIPNECH) was a strong cytoplasmic reactivity for Synaptophysin (Fig. 3b). In fact, all the PNC-proliferations except for one focus in case number 2 (Fig. 4) showed a strong and uniform reaction for Synaptophysin. This reaction eases the identification of small and microscopic proliferations and can be used as a reliable indicator of DIPNECH. The reaction against other neuroendocrine markers,
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Fig. 2. One of the typical growth patterns of DIPNECH foci was proliferation of neuroendorine cells alongside the basement membrane of small airways (left). Note the strong cytoplasmic reactivity against synaptophysin (right).
Fig. 3. (Left) A focus of DIPNECH composed of small nests of proliferating pulmonary neuroendocrine cells. (Right) Immunohistochemistry for synaptophysin in the same focus.
Fig. 4. Negative reaction of a focus of DIPNECH against synaptophysin in case number 2 (left). Note the positive reaction in the same focus against CD56 (right).
such as Chromogranin and CD56, was more variable and ranged from a completely negative to a strongly and diffusely cytoplasmic reaction with some examples of intermediate or equivocal reactions. In all examined cases, the proliferating PNCs were negative
for epithelial markers of carcinomas, such as CK7, Napsin, CK5/6, and p63. In general, the proliferation index of proliferating PNCs was very low and did not exceed 2% in all cases. In case number 2, we detected a strong cytoplasmic reactivity against CD56 (Fig. 5). In
Table 2 Immunohistochemical characteristics of adenocarcinomas and DIPNECH foci. Case no.
Syn.
Chr. A
CD56
TTF1
CK5/6
Napsin
p63
1 (tumor) 1 (DIPNECH) 2 (tumor) 2 (DIPNECH) 3 (tumor) 3 (DIPNECH)
N P N P/Nb N P
N P/Na N P N P
N P P P/Nc N P
P N P N P N
P N N N P N
P N P N P N
N N N N N N
Abbreviations: Syn., synaptophysin; Chr. A, chromogranin A; P, positive; N, negative. a In two foci, the reaction against chromogranin was strongly positive, in two foci faintly positive and in one focus completely negative. b A single focus was negative. The other foci were positive. c In three foci moderately to strongly positive. In one focus negative.
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Discussion
Fig. 5. Strong cytoplasmic reactivity against CD56 in cells of adenocarcinoma in case 2.
another two cases, there was no reactivity against neuroendocrine markers.
Microscopic review of lung carcinomas The reviewed lung carcinomas encompassed 38 cases of ADC and 42 cases of squamous cell carcinoma (SCC) or large cell lung carcinoma (LCC). In this series, there was only one case of small cell lung cancer. A single case showed both adenomatous and squamous components together (adeno-squamous carcinoma). The adenocarcinomas (38 cases) were graded according to the new, previously described classification. They included 11 low grade, 19 intermediate grade, and 8 high grade adenocarcinomas. In only three cases was recognized a single microscopic growth pattern. In 24 cases two growth patterns, in 6 cases three growth patterns, and in 4 cases four different growth patterns were demonstrable. In a single case, we could see 5 different microscopic growth patterns. The most frequent microscopic growth pattern was acinar and/or cribriform. It was encountered in 36 cases (94.7%,) and was the dominant growth pattern in 19 cases (50%). The other growth patterns, in decreasing order of frequency, were solid (22 cases/57.9%), lepidic (19 cases/50%), papillary (7 cases/18.4%), and micropapillary (6 cases/15.8%). In four cases, we recognized clear cell changes in different proportions which ranged from 20% to 70% of the bulk of tumor.
Correlation of grading and survival time Among the 38 cases with adenocarcinoma, the survival data of 16 cases were available. The follow-up time ranged from 2 to 66 months (mean: 32.5). In order to analyze the correlation between tumor grade and the survival of the patients, we allocated the cases to two groups. Group one includes the cases of good and intermediate differentiated tumors (G1 and G2). Group two is composed of the cases with low differentiation (G3). The number of patients in group one and two was 11 and 5, and their mean survival time was 52.6 (39.3–65.9) and 14.4 (5.9–22.8) months, respectively. We performed the Kaplan–Meier and the chi-square analysis for overall and disease-free survival (SPSS software, SPSS Inc., Chicago, IL, USA). There was a statistically meaningful difference between these two groups with a P-value of 0.013.
We have seen four cases of lung adenocarcinoma with simultaneous PNC proliferation(s). Such simultaneity has been recently documented in a single case report [2]. DIPNECH is a rarely encountered lesion, and the number of documented cases is less than 100 [2]. Rizivi and colleagues reviewed a series of cases with neuroendocrine and non-neuroendocrine carcinomas of the lung and reported immunohistochemical evidence of neuroendocrine cell hyperplasia in 4 of 26 cases (15%) of lung adenocarcinoma [21]. In this study, the border between reactive and pathologic proliferations of PNCs was not clearly defined. On the other hand, a spectrum of lesions with different levels of proliferation was gathered into a group of neuroendocrine cell hyperplasia. The relationship between the foci of neuroendocrine cell hyperplasia and lung adenocarcinoma, as well as the number and dimension of these foci, was not discussed in the article. Many factors (for instance full attention on the evidently more important background lesion, e.g. adenocarcinoma of lung) can lead to under-detection of DIPNECH. To challenge this assumption, we reviewed the available HE slides of a series of lung tumors which have been used for other studies [17]. Our careful histologic examination revealed that there were no foci of PNC proliferation inside or around the tumoral tissue and in the neighboring non-neoplastic lung parenchyma. Arguably, the number of reviewed slides per each case was generally low. In more than 50% of cases, only one slide was available for microscopic review. Although there was a correlation between histologic grade of adenocarcinoma by the new classification and the survival of patients, the number of cases in this series was not large enough to reach a definitive conclusion. In a large series of 1090 lung resection specimens, Ruffini and colleagues could detect preinvasive lesions of lung carcinoma in 73 cases [7]. Among them, in only three of all 1090 cases, including 5.4% (3/55) of cases with carcinoid tumor, DIPNECH was detectable. In all these 3 cases, DIPNECH was associated with typical or atypical carcinoid of the lung. In fact, in none of the cases with ADC or SCC of the lung were the lesions of DIPNECH detectable. To our knowledge, there is no case series or single case report which could show an incidental finding of DIPNECH in an otherwise normal or non-neoplastic lung tissue, as well as concurrent synchronous or metachronous detection of SCC or other infrequent forms of lung tumors and DIPNECH. Taking these facts into account, we think that the concurrent presence of DIPNECH and ADC of the lung in our case series is not a pure coincidence. The ambiguities and impreciseness in the standard definition of DIPNECH can also affect the detection rate of this lesion. In the latest edition of the WHO classification of lung and mediastinal tumors, no size and location attribute has been applied for the definition of DIPNECH [1]. However, in many case reports and studies published recently, the nomenclature of DIPNECH is reserved only for intramucosal proliferations of PNCs which are confined to the basement membrane, and their size does not exceed 5 mm in maximal dimension [4–7]. Tumorlet was used to name the lesions that are located out of the confines of basement membrane and measure less than 5 mm, and carcinoid was applied for similar lesions which are larger than 5 mm. Consideration of the threshold of 5 mm seems to be rather arbitrary and not evidence-based. Interestingly, Gosney et al. considered tumorlet as a reactive process in continuum with reactive proliferations of PNCs in reaction to the lung injury without any malignant potential [1]. He did not give any detailed diagnostic criteria to discriminate the reactive processes from preneoplastic (DIPNECH) lesions. All the proliferations of PNCs in our cases were less than 5 mm in maximal diameter and were confined histologically to the basement membrane. The largest focus has been encountered in case 1, with a maximal diameter of about 4.2 mm. It was composed, in turn, from many small
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intramucosal aggregates and solid nests of PNCs. In three of the present cases, we could detect 4 or more separate DIPNECH foci with similar cytologic and immunohistochemical characteristics. One can argue that the neuroendocrine proliferations in these cases can be a reaction to hypoxic effects of a slow-growing tumor on adjacent non-neoplastic lung tissue. The multiplicity and, in at least one case, the presence of foci of neuroendocrine proliferations far from tumor support a rather diffuse and idiopathic process that is compatible with the basic definition of DIPNECH. The immunohistochemical evaluation revealed that the most frequent and strongest reaction against neuroendocrine markers in foci of DIPNECH belonged to Synaptophysin. Except for one case, in which the focus of DIPNECH was not present in the new section used for immunohistochemical staining, as well as one single focus in patient number 2, all foci of DIPNECH showed strong cytoplasmic reactivity against Synaptophysin. Other neuroendocrine markers, such as Chromogranin and CD56, were completely negative or either partly or faintly positive. As expected, they showed a very low proliferative index using Ki67 immunostaining. To assess any similarity between the immunohistochemical profile of adenocarcinomas and DIPNECH, we performed a similar block of immunohistochemical staining on both lesions using epithelial and neuroendocrine markers, including Synaptophysin, Chromogranin, CD56, TTF1, Napsin, CK5/6, and p63. None of the DIPNECH foci showed a reaction against epithelial markers. Other than case number two with cytoplasmic CD56 reactivity (Fig. 5), none of the adenocarcinomas expressed neuroendocrine markers. Because of the rarity of DIPNECH, its association with any specific sets of clinical or paraclinical findings cannot be proved. On the other hand, the presence of accompanying lesions can mask or dominate the symptoms of DIPNECH. Falkenstern and colleagues reported a case of DIPNECH that was initially evaluated for recurrent pneumonia [4]. Stenzinger and colleagues noticed diffusely disseminated DIPNECH lesions in both lungs during the autopsy of a female patient under treatment of urosepsis with a history of an unexplained bronchial asthma about 25 years before the present admission [5]. Cameron et al. have discussed a case with a spectrum of PNC proliferative lesions, including DIPNECH with patchy immunohistochemical presentation of ACTH as an explanation for cyclic hyper-adrenocorticotropic symptoms [6]. In a recently published article, Gorshtein et al. collected and summarized the clinical data of 100 published DIPNECH cases, including their new 11 cases [12]. They showed that 89% of all cases were female. The mean age at the time of diagnosis was 57.8 years, and the duration of clinical symptoms before the diagnosis varied between 3 up to more than 30 years. In 94 cases with available data, 59 showed evidence of obstructive lung disease determined by pulmonary function tests. Only in one of 100 cases with DIPNECH has a disease-related death occurred 7 years after the onset of symptoms. In comparison to this case collection, the mean age of our cases is higher (66.5). Similar to the case collection of Gorshtein, three of our cases are female. We could not find any clinical or laboratory findings that are attributable to neuroendocrine secretions. The tumor location, size, grade, and stage are summarized in Table 1. We used the new proposed system for tumor grading to show if there is any correlation between a specific tumor growth pattern and presence of PNCs proliferation. Two of four adenocarcinomas were well differentiated and showed a predominantly lepidic growth pattern. In two other cases, the acinar and cribriform growth patterns were the predominant ones. In general, no correlation between the predominant growth pattern of adenocarcinoma and detection of DIPNECH was detectable. However, the cytological features of the ADC cells and most of the DIPNECH foci were quite similar. In case 2, in particular, both cell proliferations seemed to have morphological transition, arguing for a causal relationship between both manifestations. It is important to note that all ADC cases were either low grade (except
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for case number 1) or low stage tumors, suggesting an indolent or at least less aggressive clinical course than conventional ADC. In addition, in this series and in other reported series of DIPNECH cases [13], a female predominance is evident. Female gender is generally considered a positive prognostic factor in lung cancer. All these data support the hypothesis that lung ADC cases with DIPNECH are associated with a more favorable prognosis. Thus, pulmonary cell hyperplasia with neuroendocrine differentiation seems to represent a relevant morphologic feature to be looked for in lung cancer specimens. We would like to stress that neuroendocrine cell differentiation in such low grade/stage lung adenocarcinoma is not a marker for tumor progression in contrast to high grade/stage lung cancer [22]. Summary We described in this study four cases with a concomitant presence of adenocarcinoma and DIPNECH in the lung tissue. Even though such concomitance seems to be a rare event, we feel that it does not represent a pure incidental finding but might instead define a subgroup of lung adenocarcinomas with a specific clinical behavior with a better prognosis than conventional lung ADC, as well as a potentially distinct tumorigenesis being related to neuroendocrine cell hyperplasia. Thus, DIPNECH may not only represent a precursor lesion for carcinoids but also lung ADC [23]. This issue would be verified in larger series of cases. Considering the rarity of DIPNECH, a routine immunohistochemical evaluation of neuroendocrine differentiation in all cases with lung adenocarcinoma is in our view not necessary. Conflict of interest The authors declare that they have no conflict of interest. Acknowledgments We acknowledge the very kind and professional technical support of Mrs. Barbara Bergholz and other colleagues from the immunohistochemistry laboratory of the Jena Institute of Pathology. References [1] J.r. Gosney, W.D. Travis, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia, in: Tumors of the Lung, Pleura, Thymus and Heart, IARC Press, 2004, pp. 76–77. [2] A. Warth, E. Herpel, A. Schmähl, K. Storz, P.A. Schnabel, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH) in association with an adenocarcinoma: a case report, J. Med. Case Rep. 2 (January) (2008) 21. [3] J.R. Gosney, I.J. Williams, A.R. Dodson, C.S. Foster, Morphology and antigen expression profile of pulmonary neuroendocrine cells in reactive proliferations and diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH), Histopathology 59 (2011) 751–762. [4] R.F. Falkenstern, M. Kimmich, G. Friedel, A. Tannapfel, V. Neumann, M.M. Kohlhaeufl, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia: 7year follow-up of a rare clinicopathologic syndrome, J. Cancer Res. Clin. Oncol. 137 (2011) 1495–1498. [5] A. Stenzinger, W. Weichert, M. Hensel, H. Bruns, M. Dietel, A. Erbersdobler, Incidental postmortem diagnosis of Dipnech in a patient with previously unexplained asthma bronchiale, Pathol. Res. Pract. 206 (November (11)) (2010) 785–787. [6] C.M. Cameron, F. Roberts, J. Connel, M.W. Sproule, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia: an unusual cause of cyclical ectopic adrenocorticotrophic syndrome, Br. J. Radiol. 84 (January (997)) (2011) e14–e17, http://dx.doi.org/10.1259/bjr/91375895. [7] E. Ruffini, M. Bongiovanni, A. Cavallo, P.I. Filosso, R. Giobbe, M. Mancuso, M. Molinatti, A. Oliaro, The significance of associated pre-invasive lesions in the patients resected for primary lung neoplasms, Eur. J. Cardiothorac. Surg. 26 (July (1)) (2004) 165–172. [8] S. Irshad, E. McLean, S. Rankin, S. Barrington, G. Santis, J. Spicer, L. Lang-Lazdunski, Unilateral diffuse idiopathic pulmonary neuroendocrine cell
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Pathology – Research and Practice journal homepage: www.elsevier.com/locate/prp
Original article
Association of small foci of diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH) with adenocarcinoma of the lung Masoud Mireskandari a , Azita Abdirad a , Qing Zhang a , Manfred Dietel b , Iver Petersen a,∗ a b
Institute of Pathology, Jena University Hospital, Friedrich Schiller University, Jena, Germany Institute of Pathology, Charité – University Medicine Berlin, Campus Mitte, Berlin, Germany
a r t i c l e
i n f o
Article history: Received 29 January 2013 Received in revised form 27 May 2013 Accepted 25 June 2013 Keywords: Diffuse idiopathic pulmonary neueroendocrine cell hyperplasia (DIPNECH) Adenocarcinoma of lung Lung preneoplasia Grading of lung adenocarcinoma
a b s t r a c t DIPNECH is regarded as a precursor lesion of neuroendocrine lung tumors, specifically carcinoids. A relationship with lung adenocarcinomas has not been clearly established so far. We present a series of four cases with a concomitant presence of adenocarcinoma and DIPNECH in the lung. The cases were retrieved from the archives of the Institutes of Pathology of the Jena University Hospital and the Charité, Berlin. The clinical data were collected from the hospital information system. The microscopic findings of adenocarcinoma and DIPNECH were reviewed. A panel of neuroendocrine and epithelial markers was analyzed immunohistochemically. In addition, the H&E slides of a series of 82 lung carcinomas were reevaluated for the presence of DIPNECH foci and the parameters of the IASLC/ATS/ERS classification for lung adenocarcinoma. DIPNECH foci were composed of small intramucosal nests of proliferating pulmonary neuroendocrine cells alongside or at the periphery of terminal airways. All detected foci measured less than 5 mm in maximal diameter and showed a consistent reactivity against Synaptophysin. They did not express epithelial markers of squamous cell carcinoma and adenocarcinoma. In three cases, the DIPNECH foci were clearly associated with the adenocarcinoma, while in one case, they were observed in the non-neoplastic lung tissue. The adenocarcinoma with DIPNECH inside mainly showed low grade histology, while the fourth case was intermediate to high grade. The histologic evaluation of the HE slides of the other 82 lung cancer cases showed no suspected or definite DIPNECH foci. Within this series, we could confirm the prognostic significance of the IASLC/ATS/ERS classification of lung adenocarcinoma. Our series suggest that a subset of lung adenocarcinoma is characterized by the concomitant presence of DIPNECH within the tumor, suggesting a causal relationship. These adenocarcinomas seem to be low grade ones, and may have a particular tumorigenesis and clinical behavior. These observations need to be confirmed in larger tumor collectives. We could confirm the prognostic relevance of the new adenocarcinoma classification. © 2013 Elsevier GmbH. All rights reserved.
Introduction
Definition and clinical symptoms of DIPNECH
Recently, during the histologic evaluation of two cases of adenocarcinoma (ADC) of the lung in our institute, we noticed the presence of small DIPNECH foci in the lung tissue. Together with two other cases from a collection of lung tumors obtained from the Charite Institute of Pathology in Berlin, they constitute a series of four cases with simultaneous involvement of lung tissue with adenocarcinoma and DIPNECH.
DIPNECH is an exceedingly rare lesion. It was not completely recognized until the early 1990s, although before that time, cases with similar clinical and pathologic findings but with different nomenclature had been reported [1]. To date, fewer than one hundred cases have been documented in the literature [2]. WHO defines the DIPNECH as follows: a generalized proliferation of scattered single cells, small nodules (neuroendocrine bodies), or linear proliferation of pulmonary neuroendocrine cells (PNCs) that may be confined to the bronchial and bronchiolar epithelium [1]. Practically, it is very difficult to separate the reactive proliferations of neuroendocrine cells and formation of tumorlets from DIPNECH. Gosney and colleagues believe that DIPNECH is usually more florid. It accompanies in many cases the carcinoid tumor of the lung, often
∗ Corresponding author at: Institute of pathology, Universitätsklinikum Jena, Friedrich-Schiller-Universität, Ziegelmühlenweg 1, D-07743 Jena, Germany. Tel.: +49 3641 933120; fax: +49 3641 933111. E-mail address: [email protected] (I. Petersen). 0344-0338/$ – see front matter © 2013 Elsevier GmbH. All rights reserved. http://dx.doi.org/10.1016/j.prp.2013.06.019
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leads to obliteration of small airways and, in contrast to reactive proliferations, shows no significant pathologic alteration in the surrounding lung tissue apart from mild fibrosis and inflammation [3]. They could also show a meaningful difference in the expression pattern of cell proliferation and death antigens (e.g. p53, p16 and Ki67), as well as CD10 (CALLA), between reactive proliferations and DIPNECH. In many case reports and studies was DIPNECH histologically defined as an intramucosal proliferation of PNCs that is confined to the basement membrane. The proliferations extending beyond the basement membrane were called tumorlets when they were smaller than 5 mm in maximal diameter and carcinoid when they are larger than five millimeters [4–7]. DIPNECH, as a diagnostic term, is reserved only for cases in which the hyperplasia is diffusely primary in nature [8], which was also reflected in the nomenclature of the lesion and implication of the term “idiopathic” to describe it. In some case reports, the different clinical symptoms have been attributed to the presence of DIPNECH. Some of these clinical findings are recurrent pneumonia [4], unexplained bronchial asthma [5], and hyper-adrenocorticotropic symptoms [6]. Pulmonary preneoplasia Classically, three lesions are considered preneoplastic lesions of the lung. According to the WHO-classification of lung tumors [1], as well as many other studies [7,9–12], epithelial dysplasia, atypical adenomatous hyperplasia (AAH), and DIPNECH are considered preneoplastic lesions for squamous cell carcinoma (SCC), adenocarcinoma (ADC), and carcinoid tumor of the lung, respectively. Until now, no definite preneoplastic lesion for small cell carcinoma of the lung has been detected. New classification of adenocarcinoma of the lung In 2011, a group of experts proposed and published a new classification for adenocarcinoma of the lung [13]. The classification was supported by the International Association for the Study of Lung Cancer (IASLC), the American Thoracic Society (ATC), and the European Respiratory Society (ERS), and is expected to be integrated in the new edition of the WHO classification of lung tumors. According to this new classification, three parameters play an important role in the diagnosis and classification of lung adenocarcinomas. They include the presence or absence of invasion, size of invasion, and tumor size, as well as the predominant histological growth pattern. The following categories were generated: preinvasive lesions (atypical adenomatous hyperplasia/AAH, adenocarcinoma in situ/AIS), minimally invasive adenocarcinoma/MIA (≤3 cm in total tumor diameter, lepidic predominant growth pattern, and with ≤5 mm invasion), and invasive adenocarcinomas (with >5 mm invasion or predominant non-lepidic growth). Furthermore, the term/entity bronchioloalveolar adenocarcinoma was eliminated because it was previously associated and thus confused with entirely distinct lesions, such as AAH, AIS, MIA, lepidic predominant adenocarcinoma/LPA, and invasive mucinous adenocarcinoma. Since most of the lung ADCs are heterogenous in microscopic appearance and display a spectrum of histologic patterns, the new classification suggests the application of the term “predominant” to describe the most extensive histologic growth pattern and then to describe all other patterns in percentage in 5% increments. This form of reporting provides a basis for the grading of lung ADCs. It has been shown that among different morphologic growth patterns, solid and micropapillary patterns have the worst prognosis and the non-mucinous lepidic growth pattern the most favorable one. Other patterns, including papillary and acinar/cribriform, constitute the ADCs with intermediate prognostic association [14–16].
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Aims of present study The questions which we are going to address in this study are as follows: – Is DIPNECH a precursor lesion in a subset of lung ADCs? – Is there any characteristic feature for the DIPNECH-associated lung adenocarcinomas which can be discriminated from other ADCs of the lung regarding tumor phenotype (e.g. grading according to the new classification of lung adenocarcinoma or tumor-morphologic characteristics)? – Is there any DIPNECH-associated clinical sign or symptom which can lead the physician to suspect the possible presence of DIPNECH in a patient with lung adenocarcinoma? Materials and methods Cases All four DIPNECH cases of this study had undergone partial lung resection because of ADC of the lung. Detection of DIPNECH was an incidental finding during the microscopic evaluation of resection specimens. The hospital information systems were used to retrieve any potentially DIPNECH-related clinical symptoms (dyspnea, wheezing, cough, hemoptysis and chest tightness), as well as demographic data. Immunohistochemitry and molecular analysis Both tumoral tissue and DIPNECH foci were evaluated immunohistochemically using antibodies against TTF1, Napsin, CK7, CK 5/6, CD56, Synaptophysin, Chromogranin, and proliferation marker Ki67 according to the provider guidelines, as well as an automatic stainer (Dako, Germany). Microscopic re-evaluation of carcinomas of the lung To look for any presumably undetected DIPNECH foci during the first microscopic evaluation, we reviewed the HE slides of 81 cases of non small cell and one small cell lung cancer, which had been collected for other studies [17,18]. Only the slides containing both neoplastic and non-neoplastic lung tissue were reviewed. In total, 247 slides were reviewed. In 45 cases (about 55%) only one slide, in 29 cases (about 35%) between 2 and 5 slides, in 5 cases (about 6%) between 6 and 8 slides, and in only three cases (about 4%) 10 or more than ten slides were available for review. The mean number of slides per each case was about 3. The adenocarcinomas were graded according to the above described new lung ADC classification. Results Patients’ clinical data The age, sex, and other demographic attributes of four DIPNECH cases are summarized in Table 1. Briefly, three of four cases are female. The mean age of the patients was 66.5 years (55–72). The average diameter of ADC was 2.5 cm (2.0–3.0). At the time of diagnosis, all tumors were in early stages (pT1 to pT2 without lymph node metastasis). The search of the hospital information system for the clinical data of the two recently diagnosed cases (cases 1 and 2) revealed no clinical sign or symptom, which is attributable to neuroendocrine secretions. Interestingly, both cases had a history of other malignancies. Case number one experienced a stage Ib malignant melanoma of the skin two years before the present
S R R R 5 4 >10 1
Relation to ADC No. Of DIPNECH foci
Alive (10) Alive (10) Alive (30) Dead (13)
Fig. 1. Microscopic features of proliferating neuroendocrine cells.
admission without any sign of recurrence. Case number two had a papillary thyroid carcinoma which had been diagnosed 4 years before the diagnosis of the present lung tumor. The cases 3 and 4 are from another Institute, and there was no access to their clinical information. Histologic findings of lung adenocarcinoms with DIPNECH
Abbreviations: M, male; F, female; S, separate; R, related. a Architectural and cytologic grade, respectively [see the Refs. 14,15,18,19]. b Survival time after follow up in month.
pT2a, pN0 (0/38) pT1b, pN0 (0/10) pT2, pN0 (0/28) pT1, pN0 (0/14) 2/high 1/low 1/Low 2/Low 3.0 × 1.5 × 1.4 2.5 × 2.4 × 1.5 2.0 × 2.0 × 1.5 2.8 Right upper lobe Left lower lobe Right middle lobe Right upper lobe 72 69 70 55 1 2 3 4
M F F F
Tumor gradea Tumor size Tumor location Sex Age Case no.
Table 1 Clinical, demographic and pathologic findings of cases with ADC and DIPNECH.
Tumor stage
Survivalb
0.274–0.600 0.223–0.753 0.177–0.864 0.441
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Diameter Range (mm)
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Two ADCs were low grade with a predominantly lepidic growth pattern, and two were intermediate grade according to the histological growth pattern with a predominantly acinar growth pattern. One of two recent cases showed low grade features according to cytological criteria [18,19]. In all cases, the number of foci of PNC proliferation was more than one. In most cases, the foci of proliferating PNCs were seen as small solid syncytial nests of cells with uniform round to oval nuclei, a homogenous chromatin pattern without chromatin clumping, and a small to moderate amount of amphophilic cytoplasm (Fig. 1). The cell borders were not prominent. In most locations, the aggregates of proliferating cells grew alongside the airspaces or were located just at the periphery of the terminal airways (Fig. 2). The maximal diameter of each focus of DIPNECH was measured using a digital camera and “microscopic discussion” software from JVC. It did not exceed the threshold of 5 millimeters in all cases (Table 1). In some locations, they formed aggregates of variably and usually pinpoint sized nests with intervening fibrous stroma (Fig. 3a and b). The relationship between tumoral tissue and DIPNECH foci was different (Table 1). While in case one, the DIPNECH foci were detected completely out of the confines of tumor in the neighboring non-neoplastic lung tissue, in other three cases, the DIPNECH foci were located right in the middle (case number 3) or at the border but inside the tumor area (case number 4). Immunohistochemistry The immunohistochemical findings of DIPNECH foci and adenocarcinomas are summarized in Table 2. The most consistent positive immunohistochemical finding in the foci with PNC proliferation (DIPNECH) was a strong cytoplasmic reactivity for Synaptophysin (Fig. 3b). In fact, all the PNC-proliferations except for one focus in case number 2 (Fig. 4) showed a strong and uniform reaction for Synaptophysin. This reaction eases the identification of small and microscopic proliferations and can be used as a reliable indicator of DIPNECH. The reaction against other neuroendocrine markers,
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Fig. 2. One of the typical growth patterns of DIPNECH foci was proliferation of neuroendorine cells alongside the basement membrane of small airways (left). Note the strong cytoplasmic reactivity against synaptophysin (right).
Fig. 3. (Left) A focus of DIPNECH composed of small nests of proliferating pulmonary neuroendocrine cells. (Right) Immunohistochemistry for synaptophysin in the same focus.
Fig. 4. Negative reaction of a focus of DIPNECH against synaptophysin in case number 2 (left). Note the positive reaction in the same focus against CD56 (right).
such as Chromogranin and CD56, was more variable and ranged from a completely negative to a strongly and diffusely cytoplasmic reaction with some examples of intermediate or equivocal reactions. In all examined cases, the proliferating PNCs were negative
for epithelial markers of carcinomas, such as CK7, Napsin, CK5/6, and p63. In general, the proliferation index of proliferating PNCs was very low and did not exceed 2% in all cases. In case number 2, we detected a strong cytoplasmic reactivity against CD56 (Fig. 5). In
Table 2 Immunohistochemical characteristics of adenocarcinomas and DIPNECH foci. Case no.
Syn.
Chr. A
CD56
TTF1
CK5/6
Napsin
p63
1 (tumor) 1 (DIPNECH) 2 (tumor) 2 (DIPNECH) 3 (tumor) 3 (DIPNECH)
N P N P/Nb N P
N P/Na N P N P
N P P P/Nc N P
P N P N P N
P N N N P N
P N P N P N
N N N N N N
Abbreviations: Syn., synaptophysin; Chr. A, chromogranin A; P, positive; N, negative. a In two foci, the reaction against chromogranin was strongly positive, in two foci faintly positive and in one focus completely negative. b A single focus was negative. The other foci were positive. c In three foci moderately to strongly positive. In one focus negative.
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Discussion
Fig. 5. Strong cytoplasmic reactivity against CD56 in cells of adenocarcinoma in case 2.
another two cases, there was no reactivity against neuroendocrine markers.
Microscopic review of lung carcinomas The reviewed lung carcinomas encompassed 38 cases of ADC and 42 cases of squamous cell carcinoma (SCC) or large cell lung carcinoma (LCC). In this series, there was only one case of small cell lung cancer. A single case showed both adenomatous and squamous components together (adeno-squamous carcinoma). The adenocarcinomas (38 cases) were graded according to the new, previously described classification. They included 11 low grade, 19 intermediate grade, and 8 high grade adenocarcinomas. In only three cases was recognized a single microscopic growth pattern. In 24 cases two growth patterns, in 6 cases three growth patterns, and in 4 cases four different growth patterns were demonstrable. In a single case, we could see 5 different microscopic growth patterns. The most frequent microscopic growth pattern was acinar and/or cribriform. It was encountered in 36 cases (94.7%,) and was the dominant growth pattern in 19 cases (50%). The other growth patterns, in decreasing order of frequency, were solid (22 cases/57.9%), lepidic (19 cases/50%), papillary (7 cases/18.4%), and micropapillary (6 cases/15.8%). In four cases, we recognized clear cell changes in different proportions which ranged from 20% to 70% of the bulk of tumor.
Correlation of grading and survival time Among the 38 cases with adenocarcinoma, the survival data of 16 cases were available. The follow-up time ranged from 2 to 66 months (mean: 32.5). In order to analyze the correlation between tumor grade and the survival of the patients, we allocated the cases to two groups. Group one includes the cases of good and intermediate differentiated tumors (G1 and G2). Group two is composed of the cases with low differentiation (G3). The number of patients in group one and two was 11 and 5, and their mean survival time was 52.6 (39.3–65.9) and 14.4 (5.9–22.8) months, respectively. We performed the Kaplan–Meier and the chi-square analysis for overall and disease-free survival (SPSS software, SPSS Inc., Chicago, IL, USA). There was a statistically meaningful difference between these two groups with a P-value of 0.013.
We have seen four cases of lung adenocarcinoma with simultaneous PNC proliferation(s). Such simultaneity has been recently documented in a single case report [2]. DIPNECH is a rarely encountered lesion, and the number of documented cases is less than 100 [2]. Rizivi and colleagues reviewed a series of cases with neuroendocrine and non-neuroendocrine carcinomas of the lung and reported immunohistochemical evidence of neuroendocrine cell hyperplasia in 4 of 26 cases (15%) of lung adenocarcinoma [21]. In this study, the border between reactive and pathologic proliferations of PNCs was not clearly defined. On the other hand, a spectrum of lesions with different levels of proliferation was gathered into a group of neuroendocrine cell hyperplasia. The relationship between the foci of neuroendocrine cell hyperplasia and lung adenocarcinoma, as well as the number and dimension of these foci, was not discussed in the article. Many factors (for instance full attention on the evidently more important background lesion, e.g. adenocarcinoma of lung) can lead to under-detection of DIPNECH. To challenge this assumption, we reviewed the available HE slides of a series of lung tumors which have been used for other studies [17]. Our careful histologic examination revealed that there were no foci of PNC proliferation inside or around the tumoral tissue and in the neighboring non-neoplastic lung parenchyma. Arguably, the number of reviewed slides per each case was generally low. In more than 50% of cases, only one slide was available for microscopic review. Although there was a correlation between histologic grade of adenocarcinoma by the new classification and the survival of patients, the number of cases in this series was not large enough to reach a definitive conclusion. In a large series of 1090 lung resection specimens, Ruffini and colleagues could detect preinvasive lesions of lung carcinoma in 73 cases [7]. Among them, in only three of all 1090 cases, including 5.4% (3/55) of cases with carcinoid tumor, DIPNECH was detectable. In all these 3 cases, DIPNECH was associated with typical or atypical carcinoid of the lung. In fact, in none of the cases with ADC or SCC of the lung were the lesions of DIPNECH detectable. To our knowledge, there is no case series or single case report which could show an incidental finding of DIPNECH in an otherwise normal or non-neoplastic lung tissue, as well as concurrent synchronous or metachronous detection of SCC or other infrequent forms of lung tumors and DIPNECH. Taking these facts into account, we think that the concurrent presence of DIPNECH and ADC of the lung in our case series is not a pure coincidence. The ambiguities and impreciseness in the standard definition of DIPNECH can also affect the detection rate of this lesion. In the latest edition of the WHO classification of lung and mediastinal tumors, no size and location attribute has been applied for the definition of DIPNECH [1]. However, in many case reports and studies published recently, the nomenclature of DIPNECH is reserved only for intramucosal proliferations of PNCs which are confined to the basement membrane, and their size does not exceed 5 mm in maximal dimension [4–7]. Tumorlet was used to name the lesions that are located out of the confines of basement membrane and measure less than 5 mm, and carcinoid was applied for similar lesions which are larger than 5 mm. Consideration of the threshold of 5 mm seems to be rather arbitrary and not evidence-based. Interestingly, Gosney et al. considered tumorlet as a reactive process in continuum with reactive proliferations of PNCs in reaction to the lung injury without any malignant potential [1]. He did not give any detailed diagnostic criteria to discriminate the reactive processes from preneoplastic (DIPNECH) lesions. All the proliferations of PNCs in our cases were less than 5 mm in maximal diameter and were confined histologically to the basement membrane. The largest focus has been encountered in case 1, with a maximal diameter of about 4.2 mm. It was composed, in turn, from many small
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intramucosal aggregates and solid nests of PNCs. In three of the present cases, we could detect 4 or more separate DIPNECH foci with similar cytologic and immunohistochemical characteristics. One can argue that the neuroendocrine proliferations in these cases can be a reaction to hypoxic effects of a slow-growing tumor on adjacent non-neoplastic lung tissue. The multiplicity and, in at least one case, the presence of foci of neuroendocrine proliferations far from tumor support a rather diffuse and idiopathic process that is compatible with the basic definition of DIPNECH. The immunohistochemical evaluation revealed that the most frequent and strongest reaction against neuroendocrine markers in foci of DIPNECH belonged to Synaptophysin. Except for one case, in which the focus of DIPNECH was not present in the new section used for immunohistochemical staining, as well as one single focus in patient number 2, all foci of DIPNECH showed strong cytoplasmic reactivity against Synaptophysin. Other neuroendocrine markers, such as Chromogranin and CD56, were completely negative or either partly or faintly positive. As expected, they showed a very low proliferative index using Ki67 immunostaining. To assess any similarity between the immunohistochemical profile of adenocarcinomas and DIPNECH, we performed a similar block of immunohistochemical staining on both lesions using epithelial and neuroendocrine markers, including Synaptophysin, Chromogranin, CD56, TTF1, Napsin, CK5/6, and p63. None of the DIPNECH foci showed a reaction against epithelial markers. Other than case number two with cytoplasmic CD56 reactivity (Fig. 5), none of the adenocarcinomas expressed neuroendocrine markers. Because of the rarity of DIPNECH, its association with any specific sets of clinical or paraclinical findings cannot be proved. On the other hand, the presence of accompanying lesions can mask or dominate the symptoms of DIPNECH. Falkenstern and colleagues reported a case of DIPNECH that was initially evaluated for recurrent pneumonia [4]. Stenzinger and colleagues noticed diffusely disseminated DIPNECH lesions in both lungs during the autopsy of a female patient under treatment of urosepsis with a history of an unexplained bronchial asthma about 25 years before the present admission [5]. Cameron et al. have discussed a case with a spectrum of PNC proliferative lesions, including DIPNECH with patchy immunohistochemical presentation of ACTH as an explanation for cyclic hyper-adrenocorticotropic symptoms [6]. In a recently published article, Gorshtein et al. collected and summarized the clinical data of 100 published DIPNECH cases, including their new 11 cases [12]. They showed that 89% of all cases were female. The mean age at the time of diagnosis was 57.8 years, and the duration of clinical symptoms before the diagnosis varied between 3 up to more than 30 years. In 94 cases with available data, 59 showed evidence of obstructive lung disease determined by pulmonary function tests. Only in one of 100 cases with DIPNECH has a disease-related death occurred 7 years after the onset of symptoms. In comparison to this case collection, the mean age of our cases is higher (66.5). Similar to the case collection of Gorshtein, three of our cases are female. We could not find any clinical or laboratory findings that are attributable to neuroendocrine secretions. The tumor location, size, grade, and stage are summarized in Table 1. We used the new proposed system for tumor grading to show if there is any correlation between a specific tumor growth pattern and presence of PNCs proliferation. Two of four adenocarcinomas were well differentiated and showed a predominantly lepidic growth pattern. In two other cases, the acinar and cribriform growth patterns were the predominant ones. In general, no correlation between the predominant growth pattern of adenocarcinoma and detection of DIPNECH was detectable. However, the cytological features of the ADC cells and most of the DIPNECH foci were quite similar. In case 2, in particular, both cell proliferations seemed to have morphological transition, arguing for a causal relationship between both manifestations. It is important to note that all ADC cases were either low grade (except
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for case number 1) or low stage tumors, suggesting an indolent or at least less aggressive clinical course than conventional ADC. In addition, in this series and in other reported series of DIPNECH cases [13], a female predominance is evident. Female gender is generally considered a positive prognostic factor in lung cancer. All these data support the hypothesis that lung ADC cases with DIPNECH are associated with a more favorable prognosis. Thus, pulmonary cell hyperplasia with neuroendocrine differentiation seems to represent a relevant morphologic feature to be looked for in lung cancer specimens. We would like to stress that neuroendocrine cell differentiation in such low grade/stage lung adenocarcinoma is not a marker for tumor progression in contrast to high grade/stage lung cancer [22]. Summary We described in this study four cases with a concomitant presence of adenocarcinoma and DIPNECH in the lung tissue. Even though such concomitance seems to be a rare event, we feel that it does not represent a pure incidental finding but might instead define a subgroup of lung adenocarcinomas with a specific clinical behavior with a better prognosis than conventional lung ADC, as well as a potentially distinct tumorigenesis being related to neuroendocrine cell hyperplasia. Thus, DIPNECH may not only represent a precursor lesion for carcinoids but also lung ADC [23]. This issue would be verified in larger series of cases. Considering the rarity of DIPNECH, a routine immunohistochemical evaluation of neuroendocrine differentiation in all cases with lung adenocarcinoma is in our view not necessary. Conflict of interest The authors declare that they have no conflict of interest. Acknowledgments We acknowledge the very kind and professional technical support of Mrs. Barbara Bergholz and other colleagues from the immunohistochemistry laboratory of the Jena Institute of Pathology. References [1] J.r. Gosney, W.D. Travis, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia, in: Tumors of the Lung, Pleura, Thymus and Heart, IARC Press, 2004, pp. 76–77. [2] A. Warth, E. Herpel, A. Schmähl, K. Storz, P.A. Schnabel, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH) in association with an adenocarcinoma: a case report, J. Med. Case Rep. 2 (January) (2008) 21. [3] J.R. Gosney, I.J. Williams, A.R. Dodson, C.S. Foster, Morphology and antigen expression profile of pulmonary neuroendocrine cells in reactive proliferations and diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH), Histopathology 59 (2011) 751–762. [4] R.F. Falkenstern, M. Kimmich, G. Friedel, A. Tannapfel, V. Neumann, M.M. Kohlhaeufl, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia: 7year follow-up of a rare clinicopathologic syndrome, J. Cancer Res. Clin. Oncol. 137 (2011) 1495–1498. [5] A. Stenzinger, W. Weichert, M. Hensel, H. Bruns, M. Dietel, A. Erbersdobler, Incidental postmortem diagnosis of Dipnech in a patient with previously unexplained asthma bronchiale, Pathol. Res. Pract. 206 (November (11)) (2010) 785–787. [6] C.M. Cameron, F. Roberts, J. Connel, M.W. Sproule, Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia: an unusual cause of cyclical ectopic adrenocorticotrophic syndrome, Br. J. Radiol. 84 (January (997)) (2011) e14–e17, http://dx.doi.org/10.1259/bjr/91375895. [7] E. Ruffini, M. Bongiovanni, A. Cavallo, P.I. Filosso, R. Giobbe, M. Mancuso, M. Molinatti, A. Oliaro, The significance of associated pre-invasive lesions in the patients resected for primary lung neoplasms, Eur. J. Cardiothorac. Surg. 26 (July (1)) (2004) 165–172. [8] S. Irshad, E. McLean, S. Rankin, S. Barrington, G. Santis, J. Spicer, L. Lang-Lazdunski, Unilateral diffuse idiopathic pulmonary neuroendocrine cell
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