Case 2, 3A, 3B, 3C -

An Approach to the Diagnosis of Limited Cancer with Prudent Utilization of Immunohistochemical Markers to Resolve the Diagnosis Rajal Shah and Ming Zhou

Case 2: A 65 year-old man with serum PSA 4.5 ng/ml, found on routine health check-up. He underwent a sextant prostate biopsy. Diagnosis: Minute (< 5%) focus of adenocarcinoma of the prostate, Gleason score 3+3=6. Case 3: A 72 year-old man with urinary urgency and an elevated serum PSA. He underwent a 10-core prostate biopsy. Diagnosis: Minute (<5%) focus of adenocarcinoma of the prostate, Gleason score 3+3=6. Case 3B: A 68 year-old man with elevated serum PSA and underwent prostate biopsy. Diagnosis: Pseudohyperplastic prostate adenocarcinoma. Case 3C: A 62 year-old man with a serum PSA 5.2 ng/ml. He underwent a prostate biopsy. Diagnosis: Minute focus of adenocarcinoma of the prostate, Gleason score 3+3=6. Diagnosis of Limited Prostate Cancer in Prostate Biopsy Limited, or minimal volume, prostate cancer is defined as involving <5% of biopsy tissue. Diagnosis of limited prostate cancer is often challenging in prostate biopsy. There are two main issues. The first one is the recognition of limited cancer in needle biopsies to avoid under-diagnosis (false negative). The second one is the recognition of benign mimics of prostate cancer to avoid over-diagnosis (false positive). In this handout, we will summarize the histological features as well as ancillary immunohistochemical markers that are important for the diagnosis of limited cancer in prostate biopsies. However, we understand that no two pathologists have the same threshold for diagnosing limited cancer in biopsy, and even one's own threshold changes with time and experience. It is expected that not everyone will feel equally comfortable in establishing a cancer diagnosis on the cases we present. However, it is critical to recognize these as being atypical and suspicious for carcinoma; therefore, further work-up may lead to a more definitive diagnosis. In this sense, the most critical issue in prostate biopsy interpretation is to identify prostate glands that are morphologically atypical and suspicious for cancer. Histological Features Diagnostic of Prostate Cancer There are many histological features that are important for the diagnosis of limited prostate cancer in biopsy. Only three histological features are diagnostic of prostate cancer, as they have not been found in benign prostatic glands to date [1, 2]. They are collagenous micronodules (mucinous fibroplasia), glomerulation and perineural invasion. Collagenous micronodules are loose collagenous stroma within the glandular lumina or adjacent to cancer glands, probably as the result of hyalinization of extravasated mucin. Glomerulation results from intraluminal proliferation of cancer cells that form balls or tufts within the cancer glands. Perineural invasion represents circumferential or near-circumferential and tight encircling of a nerve by cancer cells. Occasionally, intraneural invasion of cancer cells can also be seen. The cancer cells in perineural or intraneural invasion may paradoxically have less nuclear atypia and mimic benign glands. In addition, benign prostatic glands can occasionally lie adjacent to and indent, but do not circumferentially encircle, a nerve, a finding termed perineural indentation by benign prostatic glands [3]. Therefore, it is critical to differentiate perineural invasion by cancer cells from perineural indentation by benign glands. In difficult cases, basal cell markers would help to identify them as benign glands. These 3 histological findings are specific and diagnostic for prostate cancer; therefore, a cancer diagnosis can be established with confidence in a prostate biopsy when any one of these features is present, even when only limited amount of cancer is present. However, the diagnostic value of these findings is limited by their low prevalence [1, 4], present only in <10% of the prostate cancer in biopsies. Other Histological Features of Prostate Cancer Since the histological features diagnostic for prostate cancer (collagenous micronodule, glomerulation, and perineural invasion) are infrequently found in prostate cancer in needle biopsies, diagnosis of limited cancer in most cases relies on other histological features. For conceptual purpose, these histological features are categorized as major and minor diagnostic criteria (Table 1). The major diagnostic criteria are present in the majority of prostate cancer and minor criteria are present only in minority of cases [2]. None of these features, however, is specific and diagnostic for prostate cancer. Therefore, a constellation of major and minor criteria is required to establish a cancer diagnosis. Table 1: Histological criteria for diagnosis of limited cancer in prostate needle biopsies Major Architectural: infiltrative growth, small crowded glands, confluent/irregular cribriform glands, single/cords of cells Loss of basal cells Nuclear atypia: Nuclear and nucleolar enlargement, hyperchromasia Minor Intraluminal blue mucin Pink amorphous secretions Mitotic figures and apoptotic bodies Crystalloids Adjacent HGPIN Amphophilic cytoplasm Evaluation of a prostate biopsy should start with evaluation of benign prostatic glands. Such exercise is important since fixation, section thickness and staining all affect the appearance of prostate glands. By looking at benign glands first, one can calibrate his eyes. At lower magnification, benign glands typically are medium to large in size, and have irregular contour, papillary infoldings and undulating luminal borders. There is moderate amount of stroma between the glands. In well-fixed tissue section, secretory cells have dark, reddish and slightly coarse chromatin, while basal cells have smooth, grayish chromatin. There may be inconspicuous nucleoli in basal cells; however, there shouldn't be any significant nuclear atypia, including prominent nucleoli, in both secretory cells and basal cells. One can then scan the biopsy cores to identify any glands that significantly deviate from, or do not fit in with, the rest of the benign glands. Always compare the glands in question to the benign glands that you are certain about for architectural and cytological features. Architectural Patterns. The first histological feature one should look for is the architectural growth patterns (Table 2). The pattern most characteristic for prostate cancer is so called "infiltrative" pattern with cancer cells situated between larger and paler benign glands and/or on both sides of benign glands [5]. This growth pattern is indicative of an invasive nature of prostate cancer as the prostate cancer cells in general do not induce desmoplastic stromal response characteristic of other types of cancer. On the other hand, benign processes usually maintain their lobular architecture. Cancer glands can also form a focus with closely packed, pale and small glands. The focus is often well-circumscribed without infiltrative pattern. The differential diagnosis should always include a prostate cancer that arises in the anterior or transition zone, and adenosis (atypical adenomatous hyperplasia). The cancer glands can also be exhibit a haphazard growth and dissect stroma and smooth muscle bundles without accompanying benign glands. They can also form large cribriform structures with irregular and infiltrative border. Finally they can grow in single cell fashion or cords of cells. All these growth patterns are in general inconsistent with a benign process; therefore, should raise the strong suspicion for prostate cancer. Table 2. Abnormal architectural patterns of prostate cancer Infiltrative Well-circumscribed small crowded glands Haphazard arrangement of cancer glands that dissect stroma and smooth muscles Large cribriform glands Single or cords of cells Absence of basal cells. Absence of basal cells is the hallmark of prostate cancer and is the second major diagnostic criteria. However, it is not always easy to identify basal cells on H & E slides as basal cells can assume a range of appearance [6, 7]. In addition, crushed cancer cells and stromal cells can mimic basal cells. It is also important to note that basal cells are not always present in benign glands, as noted by Dr. Totten almost half a century ago [8]. Therefore, absence of basal cells by itself is not diagnostic of prostate cancer. If a focus is otherwise diagnostic of prostate cancer, one need not go down to high power to look for basal cells. Such practice may be counter-productive. Nuclear atypia. The third major criterion is nuclear atypia in the forms of nuclear enlargement, prominent nucleoli and hyperchromasia. Nuclear enlargement, as compared to adjacent benign glands, is virtually seen in all the cases [4]. Prominent nucleoli are also present in majority of prostate cancer [4]. The definition of prominent nucleoli is always a subject of debate, and the presence of nucleoli is affected by many factors, including tissue fixation, thickness of sections and staining. There is no need to measure the nucleoli with a micrometer. Nucleoli that are distinct and significantly larger than those in adjacent benign glands are considered prominent. The importance of prominent nucleoli in the diagnosis of prostate cancer is well known and probably over emphasized in the past to the point that it is almost synonymous to prostate cancer. The presence of prominent nucleoli is very helpful in establishing a cancer diagnosis. However, some prostate cancers, especially the low grade cancers in the transition zone, may appear bland and with prominent nucleoli [9]. A prostate cancer diagnosis can also be established in the absence of prominent nucleoli. When prominent nucleoli are absent, there should be significant nuclear enlargement with or without hyperchromasia to diagnose prostate cancer. Minor diagnostic criteria. Other histological features are considered minor, or soft, criteria as they are present in lower percentage of prostate cancer [2, 5]. Same as the major diagnostic criteria, they are not specific for prostate cancer either. However, they are very useful in alerting pathologists to the atypical glands that harbor these features and perform in depth study of these atypical glands. Amphophilic, or dark, cytoplasm is one of the features one can often be appreciated even at low power when comparing atypical glands to adjacent benign glands with clear cytoplasm. Cancer glands also lack lipofuscin pigment that can be seen in benign glands. The presence of pigments in the focus that one is considering a cancer diagnosis should prompt one to reconsider the diagnosis. The intraluminal contents may also shed light on the nature of the prostate glands in question. Amorphous, pink secretions and crystalloids are more common in cancer glands, and only infrequently seen in benign conditions with the exception of adenosis. Blue mucin is also commonly seen in cancer glands, but rarely in benign glands. Detection of blue mucin depends to a large degree on the histological staining. If the tissue section is over-stained with the colonic mucosa that is often present in the prostate biopsy stained intensely blue, one then should not put too much weight on the diagnostic utility of blue mucin. Corpora amylacea are usually seen in benign glands, but not in cancer glands. The presence of corpora amylacea in glands for which one is considering a cancer diagnosis should also raise the concern about the diagnosis and prompt further study. Exclusion of benign mimics of prostate cancer. The final step towards establishing a cancer diagnosis is to rule out any conditions that may cause architectural and cytological atypia (Table 3). Benign glands with intense inflammation, especially acute inflammation, can cause both architectural and cytological atypia seen in cancer. Benign glands with atrophic cytoplasm, including partial atrophy [10] and post-atrophic hyperplasia [11], can present as small crowded glands with nuclear atypia, although the degree of the nuclear atypia is often mild and prominent nucleoli should be absent. Adenosis should always be considered and ruled out when the atypical glands form a relatively well-circumscribed focus and have cytological features similar to the adjacent benign glands [12, 13]. Lastly, a small focus of atypical glands immediately adjacent to HGPIN may represent a focus of microinvasive cancer, or tangential/out-pouching from the HGPIN glands [14]. Only after all these conditions are ruled out, one can then establish a cancer diagnosis with confidence. Table 3: Histological features against a diagnosis of prostate cancer Atypia associated with inflammation Atrophic cytoplasm Small glands merging with benign glands with indistinct cytoplasm and cytology Presence or mixture of adjacent PIN Quantitative threshold for diagnosis of prostate cancer. The major diagnostic criteria do not include the quantitative threshold for the number of glands required to make a cancer diagnosis. Such a number depends on one's diagnostic confidence and the degree of architectural and cytological atypia. The more architectural and cytological atypia, the fewer glands are required to make a definitive diagnosis. However, most urological pathologists require at least three glands to make a cancer diagnosis with confidence, although a cancer diagnosis can be made by the presence of fewer than 3 neoplastic glands provided other characteristic architectural and cytological features of prostate cancer are present [2]. In summary, there are only very few histological features that are diagnostic of prostate cancer. Therefore, diagnosis of limited prostate cancer in the majority of needle biopsies cannot rely upon any single histological feature; rather it requires a compilation of many features and exclusion of other features against such a diagnosis. A practical approach that I use is shown in the Figure 1. If there is a focus of glands that are both architecturally and cytologically atypical, and conditions that may cause architectural/cytological atypia can be ruled out, a cancer diagnosis can then be established with confidence. If, however, any one of these three criteria is not met, additional studies, including immunohistochemistry and deeper sections, should be performed, and hopefully, will lead to a more definitive diagnosis. Figure 1: Diagnosis of limited cancer in prostate needle biopsies: a practical approach. References: Baisden, B.L., H. Kahane, and J.I. 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