Dual color multiplex TTF-1 + Napsin A and p63 + CK5 immunostaining for subcategorizing of poorly differentiated pulmonary non-small carcinomas into adenocarcinoma and squamous cell carcinoma in fine needle aspiration specimens

Abstract

Background:The distinction of lung adenocarcinoma (ADC) from squamous cell carcinoma (SCC) has important therapeutic implications. Napsin A is a recently developed marker, which has shown high specificity for lung tissue in the surgical pathology specimens. In this study, we have evaluated whether the use of a panel of novel multiplex cocktails of TTF-1 + Napsin A and p63 + CK5 for dual color immunostaining will improve the diagnostic accuracy of lung adenocarcinoma and squamous cell carcinoma in fine needle aspiration (FNA) specimens, usually with relatively scant microfragments of diagnostic material.Materials and Methods:Formalin-fixed, paraffin-embedded, adequately cellular FNA cell blocks with a confirmed diagnosis of either ADC (n = 22), SCC (n = 20) or poorly differentiated carcinoma (PDC; n = 7), from a total of 49 consecutive cases, were studied. All these cases had subsequently confirmed diagnosis in biopsies or resection specimens. The sections were immunostained with two color methods of TTF-1 + Napsin A and p63 + CK5 multiplex cocktails. The presence of one or more unequivocal individual tumor cells with convincing brown nuclear TTF-1 and red cytoplasmic Napsin A staining, and cells with brown nuclear p63 and membranous / cytoplasmic CK5 staining were interpreted as ‘positive’.Results: All 20 FNA cell blocks from SCC cases were positive for dual stain p63 + CK5 and negative for dual stain TTF-1 + Napsin A. The sensitivity and specificity of the dual immunoexpressions of p63 + CK5 for SCC of lung FNAs were both 100%. All 22 ADC cases were positive with dual stain of TTF-1 + Napsin A and negative for dual stain of p63 + CK5. On follow-up of the surgical pathology specimens, 22 cases were confirmed as ADC. The sensitivity of the dual immunoexpression of TTF-1 + Napsin A for ADC of lung FNAs was 100% and the specificity was also 100%. Of the seven PDC cases, five cases that were positive for dual stain p63 + CK5 and negative for dual stain TTF-1 + Napsin A could be categorized as SCC. Two of the seven (2 / 7) PDC cases were positive for dual stain TTF-1 + Napsin A and negative for dual stain p63 + CK5, consistent with ADC.Conclusions:Simultaneous coordinate or individual immunostaining for Napsin A / TTF-1 in ADC and p63 / CK5 in SCC demonstrated high sensitivity and specificity. The panel with multiplex Napsin A / TTF-1 and p63 / CK5 dual color immunostains could specifically subcategorize PDC into ADC and SCC in lung FNA specimens. Multiplex dual color Napsin A / TTF-1 and p63 / CK5 immunostaining is especially recommended for evaluation of FNA specimens with relatively scant cellularity.

The utility of a novel triple marker (combination of TTF1, napsin A, and p40) in the subclassification of non–small cell lung cancer

“…Several recent studies have proposed to combine individual markers as a dual or triple marker for subtyping NSCLC; and showed variable sensitivities and specificities [25], [26], [27] and [28]. Fatima et al have studied the combination of TTF1 and Napsin A in diagnosing lung ADC using FNA cell block material, and they showed a sensitivity and specificity of 74% and 87%, respectively [25].…”

“…Although Brown’s study demonstrated an optimal performance of the desmoglein 3 plus CK5 in subtyping SqCC, a recent study by Sakai et al showed that desmoglein 3 was also variably expressed by a subset of lung ADCs [30]. Sethi et al have reported that the combination of TTF1 and Napsin A had 100% sensitivity and specificity in ADC, and the combination of p63 and CK5 had 100% sensitivity and specificity in SqCC [27]. Even though a few studies suggested 100% sensitivity and 100% specificity of certain markers, sampling error and bias should be considered when interpreting the data, particularly when studies were performed using a small set of selected samples.…”

“…Recently, several studies have addressed this issue by investigating the potential utility of the combination of several IHC markers as a single marker for NSCLC subclassification [25], [26], [27] and [28]. A dual marker combining TTF1 and Napsin A showed 74% sensitivity and 87% specificity [25], and 74% sensitivity and 88% to 96% specificity [26] in the identification of lung ADC using FNA material.…”

“…A dual marker combining TTF1 and Napsin A showed 74% sensitivity and 87% specificity [25], and 74% sensitivity and 88% to 96% specificity [26] in the identification of lung ADC using FNA material. A dual marker of p63 and CK5 showed 100% sensitivity and 100% specificity in the identification of lung SqCC using FNA material [27]. Lung tumor tissue microarray (TMA) data showed that a dual marker of TTF1 and p40 had 93% sensitivity and 92% specificity in diagnosing SqCC [28].…”

Classification of lung adenocarcinoma and squamous cell carcinoma samples based on their gene expression profile in the sbv IMPROVER Diagnostic Signature Challenge

“…Interestingly, although no published prior knowledge was used for the identification of this gene, KRT5 is a well-known marker used to distinguish between AC and SCC. 18 It has also been identified in a number of the published signatures and by the team with the third best performance in the sbv IMPROVER LC Challenge. 19 The empirical method used in the challenge included visual inspection of the profiles of a set of candidates in the test set: profiles were identified according to their ability to separate samples in two clusters (assuming these two clusters would be AC and SCC samples).…”