Welcome to a lecture on pathologic classification of non-small cell lung carcinomas. My name is Jeff Myers. I'm at the department of pathology at the University of Michigan, and I'm here to spend the next 40 minutes or so talking about lung cancer from the perspective of pathology. We have only a couple of objectives for our time together. The first is to learn to appropriately apply current terminology to specifically non-small cell lung carcinomas, which I'll abbreviate NSCLC. And as part of that conversation, I want to talk to you about how to apply the results of immunostains to a pathologic classification and sub-classification of non-small cell lung carcinomas. So that's our plan. Let's start with a question. A CT-guided core needle biopsy from a 4.3 centimeter spiculated peripheral lung mass in a 72 year old man demonstrated a poorly differentiated non-small cell lung carcinoma that could not be more specifically subclassified on the basis of routine histology. Immunostains were focally positive for TTF-1 and negative for p63. Which of the following is the most appropriate pathological diagnosis? Please respond now. We'll circle back to this question at the end of our time together. The language for lung carcinomas is driven in large part by the World Health Organization classification of tumors of the lung, pleura, thymus, and heart, the most recent edition having been published in the late fall of 2015. It divides lung carcinomas into these categories. Including adenocarcinoma, squamous cell carcinoma, adenosquamous carcinoma, and large cell carcinoma, which will be the focus of our conversation. Other categories that we'll not get to today include the neuroendocrine tumors, the category in which small cell carcinoma resides, pleomorphic carcinomas, a rare lesion referred to as pulmonary blastoma and less common, salivary gland-type carcinomas. In general, when pathologists and clinicians refer to non-small cell lung carcinoma, they're talking about these four categories. Adenocarcinoma, squamous cell carcinoma, adenosquamous carcinoma, and large cell carcinoma. And we're going to walk through each of these categories in turn. Adenocarcinoma, as currently defined, is based on a combination of histologic, cytologic, and immunohistochemical properties. Historically, adenocarcinomas have been referred to as malignant tumors that either show the ability to make glands or the ability to elaborate mucin. And in previous iterations of the WHO classification scheme, these features were the sole basis for recognizing adenocarcinoma as a discrete subset of non-small cell carcinomas. What's new in the 2015 classification, is that poorly differentiated carcinomas that failed to show these features, may also be designated adenocarcinomas based on their expression of one or two proteins, TTF-1 and napsin A. So today, the definition of adenocarcinoma includes the historical, histological, and cytologic features and, or immunostain positive for TTF-1 and, or napsin A. Adenocarcinoma is now the most common incident lung cancer in the United States. The lung cancer incident rate peaked in the early 1990s, but adenocarcinoma rates continued to rise through at least 2010. Adenocarcinoma rates uniformly higher than rates for squamous cell carcinomas across all demographic subcategories. The language applied to adenocarcinomas was dramatically shifted in 2011, with this publication that occurred in the February issue of the Journal of Thoracic Oncology. This was a large multidisciplinary effort lead by Bill Travis to redefine the language for adenocarcinomas of the lung. This is the classification scheme that emerged from this multidisciplinary international effort and was the language adopted by the most current WHO scheme. It divides lesions into those considered preinvasive, those that are minimally invasive, and fully invasive adenocarcinomas segregated by the predominant histologic growth pattern and I will illustrate each of them to you. This classification scheme collapses historical categories of bronchoalveolar carcinoma into preinvasive and minimally invasive categories. Key to understanding the difference between the preinvasive, minimally invasive, and invasive categories is familiarity with a new term, lepidic, l-e-p-i-d-i-c, which refers to this peculiar propensity for neoplastic cells in the lung to grow along pre existing scaffolds. That is, they grow along the ovular septa in the fashion illustrated at low magnification in this photomicrograph. If we look at a little higher magnification, you can see this psychologically atypical population of cells distributed along the surfaces of alveolar septa and you can see the sharp cutoff between the neoplastic population and the more normal appearing alveolar septa. This is a pattern of noninvasive growth, although this growth pattern is common in tumors that elsewhere show conventional invasive features. This is one of the common patterns of invasive carcinoma in the lung which is referred to as an acinar pattern because the tumor cells formed these tubules or acini that infiltrate what is in this case, a desmoplastic stroma. Lung cancers are sometimes papillary, meaning that the neoplastic cells are distributed along the surface of these papillary structures in which there are central connective tissue cores, often with small vessels, as illustrated at the center of this photomicrograph. This is different from the micropapillary pattern, in which tumor cells formed these aggregates that spill into the air spaces without a supporting connective tissue core. Adenocarcinomas occasionally form solid structures in which it is difficult to see any of the features that would have historically qualified a tumor to be designated adenocarcinoma. And it is in this subset, in particular that immunostains for TTF-1 and napsin A can be particularly helpful. In truth, most resected adenocarcinoma of the lung show some combination of these histologic patterns. In fact, in some studies, at least 90% of resected adenocarcinomas show some combination of these features. And adenocarcinomas are now segregated based on the predominant pattern illustrated in this slide. Squamous cell carcinoma is one of the second most common types of incident lung carcinoma, historically defined as a tumor that does one of two things. It either makes keratin, this acellular, densely eosinophilic material illustrated on the left or the cells connect to one another with intercellular ridges, as illustrated in the high magnification image on the right. Between the cells in this image Are very fine fibrils which represent the intercellular bridges corresponding to desmosomes when viewed with the electron microscope. Historically, one or both was required to qualify a tumor as a squamous cell carcinoma. As with adenocarcinoma, immunostains have now been added to the diagnostic criteria. And so solid tumors that show neither keratinization nor intercellular bridges can be considered squamous cell carcinomas if they express either p63 or p40 which are isophormes of the same protein. Or stained for antibodies directed against high molecular weight cytokeratins for which CK5/6 is the most commonly employed commercially available antibody. And by definition these tumors should be TTF-1 negative. There is a particular variant of squamous cell carcinoma that is worth knowing about and is new to the 2015 WHO classification scheme. Historically basaloid carcinomas were often lodged under the larger category of large cell carcinomas but it is now recognized that this is a subset of squamous cell carcinomas worth knowing about for two reasons. They are sometimes confused with small cell carcinomas and they often are associated with a particularly aggressive course. The reason they can be confused with small cell carcinomas is that by comparison to other squamous cell carcinomas, these comprise relatively small cells in the language of pathology. And the cells are composed primarily of nucleus, meaning they have a high ratio of nucleus to cytoplasm. That is a feature also characteristic of small cell carcinomas. The cells tend to be arranged in the lobular growth pattern illustrated in the top panel on the right. And as illustrated in the lower panel on the right, often show peripheral palisading in which the cells seem to be pointing towards the center of the lobule. That is another feature sometimes seen in small cell carcinomas. A useful feature that may differentiate these tumors from small cell carcinomas in appropriately sampled tissues, includes a hyalinized stroma and about half will show a clear cut squamous component, which is helpful in separating the lesion from some of the alternatives in the differential diagnosis. Squamous cell carcinoma, as you know, is one of the two types of lung carcinoma, the other being small cell carcinoma, that shows the strongest relationship to cigarette smoking. Currently, squamous cell carcinoma counts for about 25% of incident lung carcinoma cases in the United States. It is more common in men than women by a ratio of about 3 to 1. And it is more likely to occur centrally than peripherally, although peripheral squamous cell carcinomas certainly occur. Adenosquamous Carcinoma, as the term implies, is a category in which you see features of both adenocarcinoma and squamous cell carcinoma as discrete components in the same cancer. The WHO has arbitrarily suggested that the term only be applied to those tumors in which each component comprises at least 10% of the sampled tissue. At the left, you can see at a tumor in which there are that is glands which would qualify for an adenal carcinoma. And other more solid appearing components that in fact represent squamous cell carcinoma. Adenosquamous carcinoma is an uncommon sub set of non small cell carcinomas. Large cell carcinoma is our final category and is a category of lung carcinoma that is a category of exclusion. Meaning, that if it is not a adenocarcinoma, if it is not a squamous cell carcinoma, and therefore, not an adenosquamous carcinoma, Then it falls into this category. If we look to the WHO for the defining language, it is defined as an undifferentiated non-small cell carcinoma that lacks the cytological, architectural, and immunohistochemical features of small cell carcinoma, adenocarcinoma, or squamous cell carcinoma. So today, the diagnosis requires not only careful attention to histology, but also immunohistochemical stains to exclude particularly adenocarcinoma and squamous cell carcinoma as the alternatives. Because it's a diagnosis of exclusion, it is an inappropriate term to apply to small biopsies. And the WHO has suggested that the term only be applied to thoroughly sampled resected tumours, and that it should not be applied to small biopsies, or cytology specimens. Historical alliterations of the WHO classification scheme subcategorized large cell carcinoma into multiple groups, which have now been abandoned. In the 2015 large cell carcinoma has no subsets. For practical purposes, the approach to the diagnosis of large cell carcinoma is illustrated here. The first question, of course, for the pathologist is, are we are looking at a carcinoma at all? That refers to the fact that there are occasional tumors that can mimic the appearance of carcinoma. This applies to both primary and metastatic lesions like melanoma. If you are certain that it is a carcinoma, the first question is, is it a small cell carcinoma? And if the answer is, yes, obviously the diagnosis of large cell carcinoma is no longer relevant. If it is not small cell carcinoma, the next question might be is it an adenocarcinoma, which today requires more than review of the histology, but also immunostains for TTF-1 and napsin A. And those stains might reveal that in fact the tumor belongs in this category. If they are negative, the next question is, is it a squamous cell carcinoma? And again, immunostains are key to responding to this question. And they may show a phenotype that puts the tumor into this category. But if those stains are likewise negative then it becomes large cell carcinoma. And again, this pathway applies only to thoroughly sampled resected tumors. The WHO has suggested subtypes which really simply refer to the level of confidence that you've excluded the alternatives and these categories are not terribly useful in daily practice. Here is an example of a large cell carcinoma, seen at low magnification on the left, in which you see highly undifferentiated cells, disposed in this discohesive fashion, forming sheets without any architectural clues to its histogenesis. This is the sort of tumor for which the differential diagnosis might be broad and include not only carcinomas but also melanomas and even high grade lymphomas. As shown in the top panel on the right, the tumor cells at high magnification show none of the features that I illustrated for adenocarcinomas or squamous cell carcinomas. But the keratin stain, illustrated in the lower panel on the right, establishes the diagnosis of carcinoma and negative staining for TTF1 naps in a P63 or P40 and CK56 means that the tumor belongs in the category of large cell carcinoma. Because of the importance of immunostains in today's classification of lung carcinomas, the February 2011 consensus statement by our international multidisciplinary group recommended that for small biopsies and cytology samples, histology be augmented with immunohistochemistry. In sub-classifying non small cell lung cell carcinomas into adenocarcinoma or squamous cell carcinoma categories whenever possible. That means for pathologists, when we're confronted with small biopsies of lung carcinomas. The first question is whether the tumor can be sub-classified on the basis of histology and cytology alone. That is often the case, and in this circumstance, there is no rationale for doing additional studies. If however, on the basis of routine histology and cytology, the best you can do is to describe the tumor as a poorly differentiated non-small cell lung carcinoma. It is in this category that immuno stains can be helpful In further sub-categorizing the tumor. There are several proposed strategies for getting to this answer. Most would now recommend a relatively small, conservative panel of immunostaines conserving as much tissue as possible for other studies, particularly molecular studies that are increasingly useful. In triaging patients to therapeutic strategies. Theoretically, the idea is that if a tumor is positive for CK5/6 and P63 or p40, it belongs in the squamous cell carcinoma group. And if it's positive for TTF-1 and negative for the others, it belongs in the adenocarcinoma category. As is often the case, that simplistic approach belies the experience reported in the literature which tells us that there are variable sensitivities and specificities for all of these markers. So sometimes even after doing the stains it is not possible to confidently sub-categorize non small cell carcinomas into these two buckets. In that circumstance poorly differentiated non small cell lung carcinoma remains a category employed in daily practice. This summarizes that staining strategy to separate tumors into these categories and reminds us that there are some cases that cannot be resolved. In this example in the upper left hand panel is a highly undifferentiated non small cell carcinoma that would be difficult to classify on the basis of routine histology alone. As illustrated in the remaining panels the tumor cells were negative for CK 5 6 and p 63. The p63 decorating some non-neoplastic squamous epithelium in the field. The tumor cells themselves were strongly and diffusely positive for TTF-1. And therefore this tumor would be classified as a poorly differentiated adenocarcinoma. Again I reiterate that these immunostains will not solve all of these problems. And there remain cases that are immunohistochemically ambiguous, for which the term poorly differentiated non small cell lung carcinoma remains appropriate. So let's return to our question. A CT-guided core needle biopsy from a 4.3 centimeter spiculated peripheral lung mass in a 72-year old man demonstrated a poorly differentiated, non small lung carcinoma that could not be more specifically subclassified, on the basis of routine histopathology. Immunostains were focally positive for TTF-1, and negative for p63. Which of the following is the most appropriate pathological diagnosis? It is my hope that the answer is now clear to you on the basis of our conversation. The take home points are that first most biopsied and resected non-small-cell lung carcinomas can be subclassified as adenocarcinoma, squamous cell carcinoma, or adenosquamous carcinoma on the basis of routine histology alone. A limited panel of immunostains is usually sufficient to subclassified poorly differentiated Non Small Cell Lung Carcinomas that cannot be sub-classified on the basis of routine histology. Resected non-small cell lung carcinomas that cannot be sub-classified on the basis of either routine histology or immunostains should be classified as large cell carcinoma. Thank you for the time together. I hope that you have enjoyed this lecture.