Immunohistochemical Expression of p16 in Melanocytic Lesions: An Updated Review and Meta-analysis.
Misdiagnosis of melanoma, mainly underdiagnosis, constitutes 13% of total pathology-related medical malpractice lawsuits, the second largest group of malpractice claims. (2) Recurrent problems include melanoma misdiagnosed as a melanocytic nevus (without disclosure of diagnostic doubt), chronically inflamed nevus, Spitz nevus, and dysplastic/ atypical nevus. (2-4) These lesions are often difficult to diagnose, and interpretation may be subjective, leading to a range of diagnoses from different pathologists, including expert dermatopathologists. In a study (5) wherein 11 expert pathologists reviewed 37 classic melanocytic lesions, there was total agreement in only 30% of cases. The potentially high rates of misdiagnosis and the possibility that they may contribute to an unfavorable outcome are of great concern to dermatologists and pathologists. The problem is that distinction between benign and malignant melanocytic lesions is often quite difficult for general pathologists, and even for skilled dermatopathologists. (6)
Additionally, a large study surveying 207 dermatopathologists interpreting melanocytic lesions in 10 US states revealed that the majority of dermatopathologists' interpretive behavior was altered because of concerns about malpractice. One of the alterations in behavior included ordering additional tests. (7) Immunohistochemistry is likely the most common additional test used by pathologists to assist in the distinction between benign and malignant melanocytic lesions. Unfortunately, there is no single marker or panel of markers that can achieve this desired end with absolute certainty in all cases. Immunohistochemistry is used by general pathologists and dermatopathologists as an adjunctive diagnostic tool primarily because of accessibility, turnaround time, and familiarity of interpretation. The emerging field of molecular diagnostics is promising; however, it is currently impractical for daily use because it may require a molecular pathologist to interpret the findings, the test is typically expensive, and turnaround time is longer. Therefore, acquisition of molecular tests for diagnostic purposes is mostly reserved for the most difficult cases.
A few immunohistochemical markers have been proposed and used to assist the pathologist with melanocytic lesions. (8-13) Currently, a panel comprising HMB45, Ki-67, and p16 is commonly used to seek assistance in distinguishing benign from malignant lesions. (8) Although the panel may be helpful in some cases, it variably lacks the confidence to provide definitive answers sought by the pathologist. Each marker has its own advantages and disadvantages, dependent on the case; for example, Ki-67 is difficult to interpret in settings with extensive lymphocytic inflammation, although dual labeling with a cytoplasmic marker, such as MART-1, with contrasting chromogen has been proposed to alleviate this problem. (14,15) Additionally, HMB45 may not be as useful in blue nevus-related lesions and malignant blue nevus cases. HMB45 and Ki-67, nevertheless, have been used for some time, and there is substantial literature supportive of their use with difficult melanocytic lesions. However, p16 has been less evaluated and reviewed for its efficacy and practicality for adjunctive diagnostic use in melanocytic lesions. Therefore, herein we review p16 immunohistochemistry as a marker for diagnostic use, particularly for the distinction of nevi from melanoma.
Cyclin-dependent kinase inhibitors (CDKIs) are essential proteins involved in withdrawing from cell cycle progression. Lack of CDKIs allows activation of cyclin-dependent kinases, which may then phosphorylate retinoblastoma protein (Rb). Unregulated hypophosphorylated Rb initially sequesters E2F transcription factors, thereby acting as a tumor suppressor. The phosphorylation of Rb releases the bound E2F. Free E2F then acts to progress cell cycle progression by the transcription of genes required for mitotic S phase entry and DNA synthesis. (16-18)
There are 2 families of CDKIs. (17) The CIP/KIP family of kinase inhibitors includes p21, p27, and p57. The INK4 family of CDKIs include [p15.sup.INK4b], [p16.sup.INK4a] (referred to herein as p16), [p18.sup.INK4c], and [p19.sup.INK4d]. The most studied among these are p21 and p16. (16) It is thought that induction of cellular senescence is established and/or maintained by either or both of the p53/p21 and p16/pRb pathways. (19-22) The upregulation of p16 has been shown to respond to hypermitogenic/oncogenic signals such as overexpression of RAS, MAP kinases, or Myc. Multiple other stimuli for upregulation of p16 have also been proposed, such as response to DNA damage. (16-19)
Among the CDKIs, there has been great interest in studying the relation of p16 with melanoma. This may partially be due to the linkage of chromosome 9p21, which includes the p16 gene, to familial melanoma. Although germline mutations of p16 were strongly correlated with familial melanoma, there was no definitive proof that p16 mutations or deletions were absolute requirements for melanoma, because not all melanomas have p16 mutations. Interestingly, few nondiseased controls were also detected to have p16 mutations. Atypical/dysplastic nevi were also analyzed and had a lesser rate of p16 mutations compared with melanomas. (23)
The role of p16 in cellular senescence, particularly in melanocytes, is also of great importance, because senescence is considered to be a critical barrier for tumorigenesis or progression of melanoma. (24) In vitro cell culture studies further support this view. Analysis of human melanocytes revealed that cells depleted of p16 displayed enhanced proliferation and an extended replicative lifespan in the presence of replication-associated DNA damage. (25) Additionally, similar findings were seen with mouse melanocyte culture studies. Mouse Ink4a-[Arf.sup.-/-] melanocytes were shown to readily become immortal upon culture, whereas wild-type Ink4a-[Arf.sup.+/+] cells senesced within 4 to 5 weeks of culture. Restoration of p16 by retroviral vectors containing normal p16 complementary DNAs into Ink4a-[Arf.sup.-/-] melanocytes stopped growth of the cells. (26)
Within melanocytes, the loss of p16 (by mutation, deletion, or silencing) is thought to be a possible contributing factor for tumorigenesis, consistent with our current understanding of the molecular biology of tumor suppressor genes/products. This is in contrast to malignant human papillomavirus-driven tumors (which include mostly anogenital, gynecologic, and oropharyngeal tumors) that show increased p16 expression by immunohistochemistry. This is thought to be due to the integration and expression of E6 and E7 viral oncoproteins. E7 inactivates pRb, preventing the sequestration of E2F transcription factor. The increase in E2F then leads to upregulation of p16 by a feedback mechanism. E6, on the other hand, acts by degrading p53, preventing apoptosis. (27,28)
Observation of germline mutations within melanoma cases and also through experimental basic science has led to the implication of p16 in the tumorigenesis of melanoma. Many independent groups have therefore studied the possible incorporation of p16 immunohistochemistry for the purpose of distinguishing benign from malignant melanocytic lesions, particularly in sporadic melanoma cases. A review and meta-analysis of studies analyzing nevi, atypical melanocytic lesions, and melanoma is presented.
p16 IMMUNOHISTOCHEMISTRY AND ITS USE IN MELANOCYTIC LESIONS
Multiple studies (29-71) evaluating p16 expression by immunohistochemistry of melanocytic lesions were reviewed, analyzed, and tabulated. Essential information (number of samples, antibody used, collection dates, overall p16 immunohistochemistry results, and general method of interpretation) from each study are listed in the Table. Most studies showed a high proportion of benign cutaneous nevi to stain positively for p16 immunohistochemistry. The reported percentage of benign nevus cases staining positive ranged from 61% to 100%. * Most cases, however, reported p16 positivity close to 100% for benign nevi. For primary cutaneous invasive melanomas, ([dagger]) the percentage of cases with positive p16 staining reported by multiple studies ranged from 12% to 93%. Most studies (10) reported percentages within the 40s to 60s. There were 4 studies that reported less than 40% and 2 studies that reported greater than 80%. Metastatic melanoma cases, ([double dagger]) on the other hand, ranged from 0% to 71% of cases staining for p16 immunohistochemistry. Because each group had differing methodologies and reporting styles, not all studies were included, and the aforementioned range of percentages was derived from studies that defined p16 immunohistochemistry simply as either positive or negative. Still, the definition of what was interpreted as positive versus negative varied from study to study. Excluded were studies ([section]) that used a scale or grading-type system in reporting their p16 immunohistochemistry results, because they did not clearly distinguish a positive versus negative result but rather a gradient of staining patterns or relative expression. It is evident that the range of reported percentages is wide and appears insignificant. In general, however, there does appear to be a decreasing trend of p16 staining going from benign to advanced melanoma (especially metastatic lesions). This may also be highlighted by one of the studies (Ghiorzo et al (45)) that showed 1 subset of primary invasive melanoma with smaller than 0.8-mm Breslow thickness to have 51% relative p16 protein expression, and another subset larger than 0.8 mm to have 21% relative p16 protein expression. Relative expression within this unique study was based on staining intensity derived from using benign nevi as reference for comparison. Unfortunately, however, the wide variation in the numerous studies appears to undermine the possible value of p16 immunohistochemistry in its application for melanocytic lesions. It is uncertain, though, whether this wide variation could be due to subjective interpretation, unstandardized laboratory techniques, source of the antibody, or other factors. There is too much variability to consider; however, it is possible that digital imaging, as used by Ghiorzo et al, (45) may assist in the reduction of intraobserver variability, although variations could also exist among imaging hardware and software.
Figure 1, a through h, illustrates the variability of p16 immunohistochemistry staining patterns experienced with melanocytic lesions. For baseline comparison, a nevus with corresponding p16 immunohistochemistry characteristically shows strong and diffuse staining pattern of nevocytes (Figure 1, b). A more variable staining pattern is illustrated in primary invasive cutaneous melanoma, in which there is mostly cytoplasmic staining of the invasive cells (Figure 1, d), versus negative staining (Figure 1, f), versus cases in which there is heterogenous positive and negative staining of the invasive cells (Figure 1, h).
Multiple studies have also incorporated specialized types of melanocytic lesions, such as Spitz lesions, (58-61,64,66,68) desmoplastic melanomas, (58,67,71) blue nevus-related lesions, (69,71) and mucosal melanocytic lesions (41,49,62,63); 2 studies (43,70) focused on nodal metastases, and a recent study (S. S. Koh, unpublished data, 2018) compared nevi of pregnancy and nevoid melanomas. The wide range of variable results is further highlighted when analyzing studies from Spitz nevi. The percentage of Spitz nevi cases positive for p16 immunohistochemistry reported by multiple groups (30,34,50,61,64) ranged from 0% to 100%. Thus, the entire range of staining characteristics disfavors the use of p16 immunohistochemistry for spitzoid lesions. Mason et al (64) showed no difference in p16 staining in Spitz nevi and spitzoid melanomas, concluding that p16 was not a useful marker to distinguish the two. Additionally, George et al (60) had similar conclusions for atypical Spitz nevi, having shown that their p16 immunoreactivity is intermediate between those of Spitz nevi and melanoma, although they claimed that p16 was useful for discriminating Spitz nevus from melanoma. In contrast, Hilliard et al (58) (desmoplastic Spitz nevi versus desmoplastic melanoma) and Al Dhaybi et al (61) (Spitz nevi versus spitzoid melanoma) reported profound differences in expression in benign versus malignant cases and proposed p16 immunohistochemistry expression as an aid for differentiation. Although it is inconclusive for diagnostic purposes in spitzoid lesions, Horst et al (66) and Yazdan et al (68) proposed p16 immunohistochemistry as a possible screening tool for 9p21 cytogenetic abnormalities. Similar to spitzoid lesions, multiple studies analyzing nevi and primary invasive melanomas have also had mixed conclusions: some studies (52,62) support p16 immunohistochemistry for diagnostic purposes (differentiating nevi from primary invasive melanoma), and some studies (29,65) do not show convincing evidence. Interestingly, in the 2 studies (43,70) analyzing nodal metastases, both supported the use of p16 immunohistochemistry as a diagnostic utility to distinguish nodal nevi from metastatic melanoma.
From the molecular level, some authors (29,54) have suggested that p16 loss is not necessary for tumor initiation, perhaps because it is retained mostly in melanoma in situ and primary invasive melanoma. Others (31,34,63) have, in contrast, supported the possibility that p16 may be contributory as a primary event. Some (35) have proposed that the gradual loss of p16 correlates with the progression of melanoma, but is not an initiating event. Most studies did, however, show gradual loss of p16 with melanoma progression (see Table). Furthermore, 2 studies (31,39) correlated loss of p16 with increased Ki-67, supporting its role in the loss of regulating and inhibiting cell cycle entry.
Although some studies incorporated atypical nevi in their studies, most did not analyze p16 immunohistochemistry staining characteristics based on grade of atypia. Chang and Cassarino (69) evaluated atypical cellular blue nevi, including cases with mild, moderate, and severe atypia, versus melanoma (including cases of malignant cellular blue nevi). They found that mildly and moderately atypical cellular blue nevi could be distinguished from severely atypical cellular blue nevi and melanomas when p16 was used within a panel of markers. Unfortunately, however, severely atypical cases showed loss of p16 similar to melanomas, and were therefore not distinguishable from melanoma based on p16 staining. It may be reasonable to raise the possibility that the severely atypical cases could have been underdiagnosed. However, the authors reported low Ki-67 staining pattern for the severely atypical cases that were similar to mild and moderately atypical cellular blue nevi and distinctly different from the high Ki-67 present in melanoma.
Apart from its use in diagnostics, p16 immunohistochemistry expression has also been explored as a potential prognostic marker. Similar to its use for diagnostics, p16 was found to have mixed results as to its role in prognostication. Some studies proposed lack of p16 to be associated with recurrent disease, (32) predicting decreased patient survival, (39,44) and as a predictor of lymph node metastasis. (48) Sanki et al, (51) in contrast, proposed that p16 immunohistochemistry expression did not reliably predict recurrence or survival. Additionally, for sinonasal melanomas, loss of p16 did not correlate with prognosis. (49)
Analysis of multiple studies shows a wide range of results, as previously discussed. The variation could be due to how the observer is interpreting the results. One of the main differences seen among studies is whether positive staining was considered to be nuclear and/or cytoplasmic. Reanalysis was performed by separating studies depending on whether nuclear alone or nuclear and cytoplasmic staining was considered positive for p16 staining (Table). For benign nevi, 89% to 100% of cases positive for p16 was the range reported (31,34,50,54) if only nuclear staining was considered positive, compared with 61% to 100% for those studies (29,31,43,52,61) that used both nuclear and cytoplasmic staining for positivity. For primary invasive melanomas, studies (31,33,34,50,54) considering only nuclear staining had a range of 50% to 68% compared with studies (29,30,37,42,51,52) considering both nuclear and cytoplasmic staining showing a range of 12% to 91%. For metastatic melanomas, studies (33,34,37,56) considering only nuclear staining had a range of 0% to 64% compared with studies (29,43,51) considering both nuclear and cytoplasmic staining showing a range of 2% to 56%. By analyzing the studies as 2 groups (one group interpreting only nuclear staining, and a second group interpreting both nuclear and cytoplasmic staining), it appears that the group considering only nuclear staining characteristics had a shorter range, or less variability, from study to study. These results may suggest that the use of only nuclear staining for interpreting p16 immunohistochemistry for melanocytic lesions may be more optimal. However, it is unclear at the moment whether only nuclear staining is biologically relevant and whether cytoplasmic staining should be considered.
Figure 2 illustrates the staining patterns encountered with p16 immunohistochemistry in melanocytic lesions. Strong staining is usually encountered with benign nevi. Here we see an example of a nevus showing strong cytoplasmic and nuclear staining in many cells (Figure 2, a). In contrast, an example of primary invasive melanoma with the majority of cells staining is present, but the staining pattern is mostly cytoplasmic, with only a few cells showing nuclear staining (Figure 2, b). In this example, if only nuclear staining is considered within this illustrated field, then positivity may be interpreted as being around 5%. However, if cytoplasmic staining is considered, then overall positivity may be interpreted as being 50% to 60%. Depending on the methodology one uses, results may be very different, as illustrated in this example. As experienced by many pathologists, variability of staining is inevitable, as seen in another example (Figure 2, c) of a primary invasive melanoma showing many cells with weak cytoplasmic staining and no nuclear staining (yellow arrow), cytoplasmic staining without nuclear staining (red arrow), and an adjacent cell showing strong nuclear and cytoplasmic staining (green arrow). The point at which to call a positive-staining cell is quite arbitrary, and unfortunately is left to the discretion of the observer.
The use of p16 for diagnostic purposes in melanocytic lesions appears limited. For the differentiation of metastatic lesions, such as in nodal metastasis versus nodal nevi (although based only on 2 reports), there appears to be some evidence supporting its diagnostic utility. However, for the purposes of distinguishing primary cutaneous melanoma from benign lesions, there is currently a lack of substantial evidence to support its use, especially when it is used alone. When it is used in a panel of other melanocytic markers, however, the possibility for diagnostic utility is likely increased. (8) Perhaps other potential markers similar to p16 might provide better use for distinguishing benign from malignant melanocytic lesions in the future, such as with the recent report (72) of p15 immunohistochemistry in melanocytic lesions. Otherwise, currently, because of the variability of results encountered by analyzing multiple studies, p16 appears to have many limitations, especially for differentiating benign from malignant primary lesions. Nevertheless, if interpretation methods and techniques for its use were better defined, perhaps its role for melanocytic lesions might become more acceptable. Because studies considering only nuclear immunohistochemical p16 staining as positive appeared to show more consistent results, limiting the interpretation to nuclear p16 staining may prove to be more accurate, and may improve interobserver variability, thus potentially making it more useful in the routine distinction of benign versus malignant melanocytic lesions.
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Stephen S. Koh, MD, PhD; David S. Cassarino, MD, PhD
Accepted for publication December 13, 2017.
From the Department of Pathology and Dermatopathology, Kaiser Permanente Anaheim Medical Center, Anaheim, California.
The authors have no relevant financial interest in the products or companies described in this article.
Presented at the 16th Spring Seminar of the Korean Pathologists Association of North America (KOPANA); March 3, 2017; San Antonio, Texas.
Corresponding author: Stephen S. Koh, MD, PhD, Department of Pathology and Dermatopathology, Kaiser Permanente Anaheim Medical Center, 3430 E La Palma Ave, Anaheim, CA 92806 (email: email@example.com).
* References 29-31, 34, 35, 43, 44, 46, 50, 52-54, 61.
([dagger]) References 29-31, 33-35, 37, 42, 44, 50-54.
([double dagger]) References 29, 33-35, 37, 43-44, 51, 53, 56.
([section]) References 32, 36, 38-40, 45, 47-48, 55, 57, 60, 62-63, 65-67, 69.
Caption: Figure 1. a and b, Nevus with corresponding p 16 staining. Higher magnification of p16 staining provided as inset on lower right corner of b illustrates a clearer view of strong and diffuse nuclear and cytoplasmic staining of dermal nevus cells, c and d, Primary invasive melanoma with corresponding p16 staining. Higher magnification of p16 staining provided as inset on lower right corner of d illustrates a clearer view of partial and mostly cytoplasmic staining of dermal invasive cells, e and f, Primary invasive melanoma with corresponding negative p 16 staining, g and h, Primary invasive melanoma with corresponding heterogenous p16 staining (hematoxylin-eosin, original magnifications x 10 objective [a, c, e, and gl; p16 immunohistochemistry, original magnifications x 10 objective [b, d, f, and h] and X60 objective [b and d, insets]).
Caption: Figure 2. a, Nevus with many cells showing both cytoplasmic and nuclear staining. b, Primary invasive melanoma with mostly cytoplasmic staining. c, Primary invasive melanoma showing weak cytoplasmic staining with negative nuclear staining (yellow arrow), cytoplasmic staining with negative nuclear staining (red arrow), and strong cytoplasmic and strong nuclear staining (green arrow) (p16 immunohistochemistry, original magnification X60 objective).
Results and Highlighted Findings From Each Reviewed Study Study Source, y Period Samples Reed et al, NA 20 BN (29) 1995 18 AN 14 MIS 37 PIM 14 MM Wang et al, NA 28 BN (6 Spitz nevi) (30) 1996 19 PIM Talve et al, 1973-1982 10 BN (31) 1997 79 PIM Straume and 1981-1989 91 PIM, NM Akslen, (32) 1997 Piccinin et NA 8 PIM al, (33) 1997 8 MM Sparrow et 5 yrs or less 35 BN (5 Spitz nevi) al, (34) 1998 12 MIS 38 PIM 25 MM Keller- NA 51 BN + AN Melchior et al, (35) 1998 7 MIS 17 PIM 33 MM Funk et NA 24 BN al, (36) 1998 2 lentigo simplex 4 MIS, LM 26 PIM, SSM 28 PIM, NM 3 PIM, LMM 5 MM Morita et NA 14 PIM, ALM + al, (37) 1998 SSM + NM 14 MM, corresponding cases Radhi, NA 20 PIM with (38) 1999 residual dermal nevi Straume et 1981-1997 190 PIM al, (39) 2000 58 MM Vuhahula et January 1997- 28 PIM, 50% ALM al, (40) 2000 December 1998 Tanaka et NA 13 PIM, oral al, (41) 2001 Pavey et July 1993- 108 PIM, SSM + al, (42) 2002 June 1994 NM Mihic-Probst et NA 1 5 BN, dermal al, (43) 2003 1 8 nodal nevi 26 MM, SLN 10 MM, NSLN 13 MM, organs Alonso et 1995-2000 9 BN al, (44) 2004 26 PIM, radial 66 PIM, vertical 68 MM Ghiorzo et NA 33 BN al, (45) 2004 12 MIS 31 PIM <0.8 mm 9 PIM >0.8 mm 11 MM Herron et 1994-2001 30 BN al, (46) 2004 Gray-Schopfer NA 11 BN et al, (47) 2006 9 AN 14 PIM, RGP 14 PIM, VGP Mihic-Probst et NA 34 BN al, (48) 2006 64 PIM Franchi et NA 37 PIM, sinonasal al, (49) 2006 Stefanaki et NA 20 BN al, (50) 2007 10 Spitz nevi 16 PIM Sanki et al, NA 19 BN + AN (51) 2007 39 PIM 34 MM Demirkan et NA 28 BN al, (52) 2007 24 PIM, NM + LMM + ALM + SSM Sini et al, NA 20 BN (53) 2008 22 AN 14 PIM, T1-T2 18 PIM, T3-T4 8 MM Stefanaki et NA 41 BN, congenital al, (54) 2008 20 BN, acquired 16 PIM Richmond- January 1998- 109 PIM, LMM + Sinclair et December SSM al, (55) 2008 1999 De Sa et 1994-2000 125 MIS + PIM al, (56) 2009 29 MM Karim et 1997-1999 8 BN, dermal al, (57) 2009 7 BN, compound 15 MIS 25 PIM, thin 29 PIM, thick 28 MM Hilliard et NA 15 desmoplastic al, (58)2009 Spitz 11 desmoplastic melanoma Hsieh et al, NA 32 ALM, 3 MIS + (59) 2009 29 PIM George et al, 1983-2000 27 Spitz nevi (60) 2010 19 atypical Spitz 42 PIM, 6 with spitzoid features Al Dhaybi et 1998-2010 12 BN al, (61) 2011 18 Spitz nevi 6 spitzoid melanoma Zoroquiain et 1993-2009 19 BN al, (62) 2012 15 PAM w/o atypia 2 PAM w atypia 9 PIM de Andrade et NA 38 BN, intramucosal al, (63) 2012 13 PIM Mason et al, NA 18 BN, Spitz nevi (64) 2012 19 PIM, spitzoid Tran et al, 1993-2010 19 BN (65) 2012 18 MM 20 BN, tissue array 21 PIM, tissue array Horst et al, July 2000- 22 Spitz nevi, (66) 2013 June 2010 patient age >50 y 11 Spitz nevi, patient age <50 y Blokhin et al, NA 5 BN, desmoplastic (67) 2013 22 PIM, desmoplastic Yazdan et al, NA 12 atypical Spitz, (68) 2014 heterozygous 9p21 deletion 13 atypical Spitz, homozygous 9p21 deletion 16 PIM, with homozygous 9p21 deletion Chang and NA 6 CBN, Cassarino, (69) conventional and 2014 mildly atypical 4 CBN, moderately atypical 5 CBN, severely atypical 12 melanomas, MIS + superficial spreading + in transit + spitzoid + nevoid 5 CBN, malignant Piana et al, January 2000- 22 nodal nevi with (70) 2015 January 2014 5 coexistent metastatic deposit Donigan et 2007-2015 9 common blue al, (71) 2017 nevi 8 cellular blue nevi 12 PIM, spitzoid + desmoplastic + nevoid + nodular 3 PIM, blue nevuslike p16 I mmunohistochemistry Source, y Results Interpretation Reed et al, 100% of cases homogeneously N and C (29) 1995 positive 100% of cases homogeneously positive 100% of cases homogeneously positive 48% of cases homogeneously positive, 43% variable proportion positive, 9% negative 28% of cases homogenously positive, 28% variable proportion positive, 44% negative Wang et al, 17 cases (61%) positive, all 6 N or C (30) 1996 Spitz reported as negative 3 cases (16%) positive Talve et al, 10 cases (100%) positive N (31) 1997 46 cases (58%) positive Straume and 8 cases (9%) negative or very N and C Akslen, (32) weak, 4 cases (4%) very strong, 1997 and all remaining cases had intermediate staining Piccinin et 5/8 cases positive N al, (33) 1997 0/8 cases positive Sparrow et 89% of cases positive, 1/5 Spitz N, <5% N al, (34) 1998 reported negative staining 92% of cases positive considered 50% of cases positive negative 64% of cases positive Keller- 44 cases (86%) with uniform Unspecified Melchior et labeling al, (35) 1998 5 cases (72%) with uniform labeling 10 cases (59%) with uniform labeling 16 cases (49%) with uniform labeling Funk et 6 (25%) 0%-10% positive, 6 N + C "mixed al, (36) (25%) 10%-50% positive, 12 pattern" 1998 (50%) >50% positive 1 (50%) 0%-l 0% positive, 1 (50%) 10%-50% positive, 0 (0%) >50% positive 2 (50%) 0%-l 0% positive, 1 (25%) 10%-50% positive, 1 (25%) >50% positive 1 8 (69%) 0%-l 0% positive, 5 (19%) 10%-50% positive, 3 (12%) >50% positive 17 (61%) 0%-10% positive, 6 (21%) 10%-50% positive, 5 (18%) >50% positive 3 (100%) 0%-10% positive, 0 (0%) 10%-50% positive, 0 (0%) >50% positive 5 (100%) 0%-10% positive, 0 (0%) 10%-50% positive, 0 (0%) >50% positive Morita et 9/14 cases positive N al, (37) 1998 7/14 cases positive Radhi, 14 cases (70%) strong N and 6 N or C; see (38) 1999 (30%) weak N positivity in comment residual nevi 1 8 cases (90%) weak N and no reported strong N positivity in malignant melanocytes Straume et Nuclear scores: 28% = 0, 1 7% = N or C al, (39) 2000 1, 21% = 2, 4% = 3, 14% = 4, 13% = 6, 3% = 9 Cytoplasmic scores: 15% = 0, 6% = 1,12% = 2, 34% = 3, 2% = 4, 25% = 6, 6% = 9 Nuclear scores: 74% = 0, 3% = 1, 7%-2, 0%-3, 12%-4, Cytoplasmic scores: 36% = 0, 22% = 1,14% = 2, 3% = 3, 12% = 4, 9% = 6, 3% = 9 Vuhahula et 18% = 0, 32% = 1,18% = 2, N al, (40) 2000 18% = 4, 11 % = [greater than or equal to] 6 Tanaka et 7/13 cases positive (4/8 stage N > C; see al, (41) 2001 II, 2/ 3 stage III, 1/2 stage comment IV) Pavey et 52 (48%) of tumors with some N or C al, (42) 2002 degree of immunoreactivity, as defined by a lesion having more than a rare cell with staining. Mihic-Probst et All cases N or C positive N or C al, (43) 2003 All cases N or C positive All cases N negative, 6 cases C staining All cases N negative, 3 cases C staining 12 cases N negative, 4 cases C staining Alonso et 9 cases (100%) positive Unspecified al, (44) 2004 23 cases (88%) positive 59 cases (89%) positive 48 cases (71%) positive Ghiorzo et 100% relative p16 protein N and C al, (45) 2004 expression 43% relative p16 protein expression 51% relative p16 protein expression 21% relative p16 protein expression 18% relative p16 protein expression Herron et 100% positive Unspecified al, (46) 2004 Gray-Schopfer 10/11 cases ++ positive, 1/11 + N and C et al, positive (47) 2006 5/9 cases ++ positive, 4/9 + positive 7/14 cases ++ positive, 7/14 + positive 3/14 cases ++ positive, 11/14 + positive Mihic-Probst et 67% [+ or -] 22% of cells N or C al, (48) 2006 positive (N + C), 9% [+ or -] 16% of cells positive (C only) 16% [+ or -] 24% of cells positive (N + C), 34% [+ or -] 34% of cells positive (C only) Franchi et 27 cases (72.9%) showed loss of N and C al, (49) 2006 p16 expression Stefanaki et 20/20 cases positive, moderate N al, (50) 2007 to strong staining in >60% of nuclei 10/10 cases positive, strong N expression in 40%-70% of nuclei 11/16 cases positive, heterogeneous N weak expression from <5% to 40% of nuclei Sanki et al, 73.7% (N) and 94.7% (C) of cases N or C (51) 2007 positive 28.2% (N) and 38.5% (C) of cases positive 14.7% (N) and 17.6% (C) of cases Demirkan et 79% of cases positive N and C al, (52) 2007 12% of cases positive Sini et al, 0 cases (0%) showing loss of p16 Unspecified (53) 2008 0 cases (0%) showing loss of p16 1 case (7%) showing loss of p16 3 cases (1 7%) showing loss of p16 5 cases (62%) showing loss of p16 Stefanaki et 41/41 cases positive, N al, (54) 2008 heterogeneous intense >70% nuclei staining 20/20 cases positive, diffuse moderate to strong >60% N staining 11/20 cases positive, heterogeneous weak expression of <5%-40% nuclei Richmond- 0% (64 cases), <1% (5 cases), N and/or C Sinclair et 1 %-10% (12 cases), 10%-30% al, (55) 2008 (10 cases), 30%-50% (2 cases), >50% (16 cases) De Sa et 4% of cases positive N, <5% N al, (56) 2009 0% of cases positive staining considered negative Karim et 6.1 (mean), 7 (median), 3-7 N al, (57) 2009 (range) 5.4 (mean), 5 (median), 2-6 (range) 3.6 (mean); 4 (median); 0-6 (range) 3.9 (mean), 4 (median), 0-7 (range) 2.2 (mean), 2 (median), 0-7 (range) 1.2 (mean), 0 (median), 0-7 (range) Hilliard et 15 cases (100%) positive N and C al, (58)2009 2 cases (1 8.2%) positive Hsieh et al, 25 cases (78%) positive, 1/3 MIS N (59) 2009 cases positive George et al, 68% (N) and 80% (C) mean N or C (60) 2010 positive dermal tumor cells 52% (N) and 65% (C) mean positive dermal tumor cells 40% (N) and 57% (C) mean positive dermal tumor cells Al Dhaybi et All positive N and C al, (61) 2011 All positive All negative Zoroquiain et 7.63 [+ or -] 3.24 N al, (62) 2012 11 [+ or -]1.69 12 [+ or -] 0 3.3 [+ or -] 1.8 de Andrade et Strongly expressed in more than N al, (63) 2012 70% of cells in all cases Only one oral melanoma was positive in 35% of neoplastic cells. Other 12 cases negative. Mason et al, 15 cases (83%) positive N or C (64) 2012 15 cases (79%) positive Tran et al, 40% median percentage of cells N (65) 2012 positive 0% median percentage of cells positive 45% median percentage of cells positive 20% median percentage of cells Horst et al, 6.1 (mean), 7 (median) N (66) 2013 6.0 (mean), 6 (median) Blokhin et al, All positive Unspecified (67) 2013 6 cases negative, 6 cases with <25% of cells staining, 4 cases with 25%-50% of cells staining, and 6 cases with diffuse positive staining Yazdan et al, 8 cases (67%) positive N (68) 2014 0 cases (0%) positive 0 cases (0%) positive Chang and 69.58% [+ or -] 8.23% positive N and C Cassarino, (69) melanocytes 2014 66.88% [+ or -] 12.56% positive melanocytes 20.2% [+ or -] 11.96% positive melanocytes 13.92% [+ or -] 6.32% positive melanocytes 6% [+ or -] 3.24% positive melanocytes Piana et al, All 22 nodal nevi components N and C (70) 2015 positive and all 5 metastatic components negative Donigan et 0 cases positive Unspecified al, (71) 2017 3 cases positive 7 cases positive 1 case positive Source, y P16 Antibody Reed et al, Pharmingen Inc, San (29) 1995 Diego, California; Santa Cruz Biotechnology Inc, Santa Cruz, California, Wang et al, Pharmingen (30) 1996 Talve et al, Pharmingen (31) 1997 Straume and SC-469, Santa Cruz Akslen, (32) 1997 Piccinin et 151262, Pharmingen; al, (33) 1997 sc468, Santa Cruz Sparrow et Pharmingen; Santa Cruz al, (34) 1998 Keller- G175-405 Pharmingen Melchior et al, (35) 1998 Funk et G1 75-405, Pharmingen al, (36) 1998 Morita et Pharmingen al, (37) 1998 Radhi, Pharmingen Canada (38) 1999 Straume et SC-468, Santa Cruz al, (39) 2000 Vuhahula et SC-468, Santa Cruz al, (40) 2000 Tanaka et G1 75-405, Pharmingen al, (41) 2001 Pavey et G1 75-405, Pharmingen al, (42) 2002 Mihic-Probst et Neomarkers al, (43) 2003 Alonso et F12, Santa Cruz al, (44) 2004 Ghiorzo et G1 75-405, Pharmingen; al, (45) 2004 SC-468 C-20 and SC-467 N-20, Santa Cruz Herron et Santa Cruz al, (46) 2004 Gray-Schopfer Novocastra et al, (47) 2006 Mihic-Probst et Neomarkers al, (48) 2006 Franchi et E6H4, Dakocytomation al, (49) 2006 Stefanaki et 1F4, Santa Cruz al, (50) 2007 Sanki et al, Neomarkers (51) 2007 Demirkan et 6H12, Novocastra, al, (52) 2007 Norwell, Massachusetts Sini et al, 16P04JC-2, Lab Vision/ (53) 2008 Neomarkers, Fremont, California Stefanaki et 1F4, Santa Cruz al, (54) 2008 Richmond- G175-405, BD Sinclair et Pharmingen, San Diego, al, (55) 2008 California De Sa et SCI 661-F12, Santa Cruz al, (56) 2009 Karim et AB-4 16p04, Neomarkers al, (57) 2009 Hilliard et Cell Marque, Hot al, (58)2009 Springs, Arkansas Hsieh et al, 16P04, Labvision, (59) 2009 Fremont, California George et al, G175-405, Becton (60) 2010 Dickinson, Franklin Lakes, New Jersey Al Dhaybi et Mtm Laboratories, al, (61) 2011 Heidelberg, Germany Zoroquiain et Abcam, Cambridge, United al, (62) 2012 Kingdom de Andrade et SC-1661, Santa Cruz al, (63) 2012 Mason et al, E6H4, mtm Laboratories, (64) 2012 Heidelberg, Germany Tran et al, N20 and C20, Santa Cruz (65) 2012 Horst et al, E6H4, Ventana, Tucson, (66) 2013 Arizona Blokhin et al, E6H4, Dakocytomation, (67) 2013 mtm laboratories, Germany Yazdan et al, JC8, Santa Cruz (68) 2014 Chang and E6H4, Roche, Tucson, Cassarino, (69) Arizona 2014 Piana et al, E6H4, Ventana (70) 2015 Donigan et Santa Cruz al, (71) 2017 Source, y Comments Reed et al, (29) 1995 Wang et al, (30) 1996 Talve et al, (31) 1997 Straume and Akslen, (32) 1997 Piccinin et al, (33) 1997 Sparrow et al, (34) 1998 Keller- Uniform labeling Melchior et considered when at least al, (35) 1998 two-thirds of melanocytic cells positive. Funk et al, (36) 1998 Morita et al, (37) 1998 Radhi, Author analyzed both C (38) 1999 and N staining. However, only N immunoreactivity was considered as a sign of p16 positivity. Straume et MMs are corresponding al, (39) 2000 cases. p16 staining was reported through a semiquantitative scoring system. A 3-point percentage of stained cells score (0 = absent, 1 = <10%, 2 = 10%- 50%, 3 = >50%) was multiplied by a 3-point N intensity score (0 = negative, 1 = weak, 2 = moderate, 3 strong) for highest value of 9. Vuhahula et p16 staining was reported al, (40) 2000 through a semiquantitative scoring system. A 3-point percentage of stained cells score (0 = absent, 1 = <10%, 2 = 10%- 50%, 3 = >50%) was multiplied by a 3-point N intensity score (0 to 3 = no staining to strong staining) for highest value of 9. Tanaka et Positive if N was stronger al, (41) 2001 than C. Pavey et al, (42) 2002 Mihic-Probst et al, (43) 2003 Alonso et al, (44) 2004 Ghiorzo et Expression is relative al, (45) 2004 compared with nevi (100%) analyzed by digital imaging techniques. PIM also stratified by superficial spreading melanoma versus nodular melanoma and by Clark levels. Herron et al, (46) 2004 Gray-Schopfer Immunostaining scored as et al, ++ (51 %-100% of cells (47) 2006 immunostained), + (up to 50%), or 0. Mihic-Probst et al, (48) 2006 Franchi et al, (49) 2006 Stefanaki et al, (50) 2007 Sanki et al, Also additional data (51) 2007 analysis based on type of nevi, thick versus thin melanoma, type of metastasis, and pathologic variables of melanoma. Demirkan et al, (52) 2007 Sini et al, (53) 2008 Stefanaki et al, (54) 2008 Richmond- % of cells with Sinclair et immunopositivity. al, (55) 2008 De Sa et MIS and PIM were grouped al, (56) 2009 in their global assessment as primary tumor. Separated by tumor thickness; [less than or equal to] 1.0 mm had 7.9% positive p16 staining and >1.0 mm had 0% positive p16 staining. Karim et p16 staining was reported al, (57) 2009 through a semiquantitative scoring system. A 3-point N intensity score (0 = no staining, 1 = weak staining, 2 = moderate staining, 3 = strong staining) was added to a 4-point percentage of stained cells score (0 = 0%, 1 = 1 %-5%, 2 = 6%-10%, 3 = 11 %- 30%, 4 = >30%) for a value ranging from 0 to 7. Hilliard et Intensity and percentage of al, (58)2009 positive tumor cells also analyzed separately. Hsieh et al, (59) 2009 George et al, Epidermal component also (60) 2010 evaluated separately. Al Dhaybi et al, (61) 2011 Zoroquiain et Conjunctival lesions. al, (62) 2012 p16 staining was reported through a semiquantitative scoring system. A 4-point percentage of stained cells score (0 = absent, 1 = 1%-10%, 2 = 11%- 50%, 3 = 51 %-80%, 4 = 81 %-100%) was multiplied by a 3-point N intensity score (0 = negative, 1 = weak, 2 = moderate, 3 strong) for a value ranging from 0 to 12. de Andrade et Oral lesions. al, (63) 2012 Mason et al, (64) 2012 Tran et al, (65) 2012 Horst et al, p16 staining was reported (66) 2013 through a semiquantitative scoring system. A 3-point N intensity score (0 = negative, 1 = weak, 2 = moderate, 3 = strong) was added to a 4-point percentage of stained cells score (0 = 0%, 1 = 1 %-25%, 2 = 2 6%- 50%, 3 = 51 %-75%, 4 = >76%-100%) for a value ranging from 0 to 7. Blokhin et al, (67) 2013 Yazdan et al, Complete absence of (68) 2014 staining and aggregates of tumor cells with complete loss were considered negative. Chang and p16 intensity was also Cassarino, (69) separately graded. 2014 Piana et al, (70) 2015 Donigan et al, (71) 2017 Abbreviations: ALM, acral lentiginous melanoma; AN, atypical/ dysplastic nevi; BN, benign nevi/melanocytic lesion; C, cytoplasmic; CBN, cellular blue nevus; LM, lentigo maligna; LMM, lentigo maligna melanoma; MIS, melanoma in situ; MM, metastatic melanoma; N, nuclear; NA, not available; NM, nodular melanoma; NSLN, nonsentinel lymph node; PAM, primary acquired melanosis (conjunctival); PIM, primary invasive melanoma; RGP, radial growth phase; SLN, sentinel lymph node; SSM, superficial spreading melanoma; VGP, vertical growth phase.
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|Author:||Koh, Stephen S.; Cassarino, David S.|
|Publication:||Archives of Pathology & Laboratory Medicine|
|Date:||Jul 1, 2018|
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