Immunohistochemistry in the Differential Diagnosis of Cutaneous Basal Cell Carcinoma

Basal cell carcinoma and squamous cell carcinoma are two of the most common cutaneous tumors seen by pathologists. In the large majority of cases, the distinction between these two tumors is readily made on the basis of standard H&E morphology. However, many of us see cases from time to time that for one reason or another (minuscule biopsy, mishandled specimen, crushed beyond recognition, dryed out, poorly fixed, etc., etc.), it is difficult to know for certain whether one is dealing with a squamous carcinoma or a basal cell carcinoma. This month, we discuss several immunostains that can be of utility in approaching this problem. 
It is worth mentioning that both basal cell carcinoma and cutaneous squamous cell carcinoma characteristically express strong and diffuse high molecular weight cytokeratin, cytokeratin 5 (or cytokeratin5/6) and nuclear p63, so the absence of staining with these markers (assuming adequate tissue and technique of course) should lead you to consider another diagnosis. 
EMA is a useful antibody for this problem, since basal cell carcinomas are negative for EMA, although occasionally lumina associated with sebaceous differentiation in these tumors may show EMA positivity. In contrast, most squamous cell carcinomas of the skin will have substantial EMA immunoreactivity.

Ber-EP4 is also a useful marker, as basal cell carcinomas are typically positive for this marker, unlike cutaneous squamous carcinoma. Interestingly, non-cutaneous squamous carcinomas (e.g., pulmonary squamous carcinoma) may express Ber-EP4, so conceivably reactivity of Ber-EP4 in a known cutaneous squamous tumor might suggest the possibility of metastatic squamous carcinoma, although I do not know of any published reports that have specifically addressed that question. 
Interestingly, smooth muscle actin (SMA) has been found to be expressed in a significant number of basal cell carcinomas of the skin (13 of 17 cases in one study). Indeed, we have observed strong SMA reactivity in a number of basal cell carcinomas that we have stained, although the frequency of reactivity is not as high in our hands as in some published series. Cutaneous squamous carcinomas are negative for SMA.
BCL-2 has been reported by some authors to be useful in this situation, since basal cell carcinomas are typically diffusely positive for this marker. Cutaneous squamous cell carcinomas are generally negative, although some authors describe focal positivity enough to 26% of cutaneous squamous carcinoms.
In summary, when faced with the differential diagnosis of cutaneous basal cell carcinoma versus cutaneous squamous carcinoma, a reasonable first approach would be to employ immunostains for EMA and Ber-EP4. If these results are not diagnostic, immunostains for SMA and BCL-2 would be worth a try. Again, if the tumor in question does not show strong high molecular weight cytokeratin, cytokeratin 5, cytokeratin 5/6, and nuclear p63, consideration of another diagnosis would be prudent. Results of expected staining in these tumors are listed in table below.

1. Wick MR: Practical immunohistology of cutaneous neoplasms: an update. Presentation at the American Society of Dermatopathology Companion Meeting, 2004 Annual Meeting of the United States and Canadian Academy of Patholgy, Vancouver, BC, March 7, 2004.
2. Jimenez FJ et al: Ber-EP4 immunoreactivity  innormal skin and cutaneous neoplasms. Mod Pathol8(8): 854-858, Oct 1995.
3. Peterdy G et al: Immunohistochemical separationof microcystic adnexal carcinoma from basal cellcarcinoma and squamous cell carcinoma. ModPathol 14(1):72A (abstract # 407), Jan 2001.
4. Varma M et al: Expression of smooth muscle antigensin basal cell carcinomas of skin. Mod Pathol12(1):65A (abstract # 365), Jan 1999.
5. Williams GA et al: Immunoreactivity for alphasmoothmuscle actin aids in the separation of basalcell carcinoma from both squamous cell carcinomaand trichoepithelioma. Lab Investig 78(1):54A(abstract # 303), Jan 1998.
6.Rodney T. Miller, M.D., Director of Immunohistochemistry


Tumor Size (Size of Invasive Carcinoma) in Breast Carcinoma

Breast Carcinoma reporting;

Tumor Size (Size of Invasive Carcinoma)

  • The size of an invasive carcinoma is an important prognostic factor. 
  • The single greatest dimension of the largest invasive carcinoma is used to determine T classification 
  • The best size for AJCC T classification should use information from imaging, gross examination, and microscopic evaluation. 
  • Visual determination of size is often unreliable, as carcinomas often blend into adjacent fibrous tissue. 
  • The size by palpation of a hard mass correlates better with invasion of tumor cells into stroma with a desmoplastic response. 
  • Sizes should be measured to the nearest millimeter. 
  • In some cases, the size may be difficult to determine.
How to measure size of breast invasive carcinoma:

A. Invasive carcinoma with surrounding ductal carcinoma in situ (DCIS). The size only includes the area of the invasive carcinoma and does not include the adjacent DCIS. The size should be measured to the closest 1 mm.

Invasive carcinoma and DCIS: The size measurement includes only the largest area of contiguous invasion of stroma. Surrounding DCIS is not included in the size measurement.


B. Small invasive carcinoma with prior core needle biopsy. The size of the carcinoma in the core needle biopsy should not be added to the size of the carcinoma in the excisional specimen, as this will generally overestimate the true size. The best size for classification must take into consideration the largest dimension of the carcinoma in both specimens as well as the size by imaging before the core needle biopsy.

C. Small invasive carcinomas with adjacent biopsy site changes. In some excisional specimens, a small carcinoma will be present adjacent to a relatively large area of biopsy site changes. The actual size cannot be determined with certainty. The size in the core needle biopsy, in the excisional specimen, and by imaging should be considered to determine the best size for classification.


D. Multiple invasive carcinomas. If multiple carcinomas are present, the size of the largest invasive carcinoma is used for T classification. The modifier “m” is used to indicate that multiple invasive carcinomas are present.


E. Multiple invasive carcinomas in close proximity. It may be difficult to distinguish multiple adjacent carcinomas from one large invasive carcinoma. Careful examination of the specimen with submission of tissue between grossly evident carcinomas is essential. Correlation with imaging findings can be helpful. Generally, microscopic size confirmation of the largest grossly identified invasive carcinoma is used for T classification.

F. Invasive carcinomas that have been transected. If an invasive carcinoma has been transected and is present in more than 1 tissue fragment, the sizes in each fragment should not be added together, as this may overestimate the true size. In many cases, correlation with the size on breast imaging will be helpful to choose the best size for classification. In other cases, the pathologist will need to use his or her judgment in assigning an AJCC T category.


DCIS with microinvasion: 
  • Microinvasion is defined by the AJCC as invasion measuring 1 mm or less in size. 
  • If more than 1 focus of microinvasion is present, the number of foci present, an estimate of the number, or a note that the number of foci is too numerous to quantify should be reported.
  •  In some cases, immunoperoxidase studies for myoepithelial cells may be helpful to document areas of invasion and the size of the invasive foci.
  •  Invasive tumors that are larger than 1.0 mm but less than 2.0 mm are rounded up to 2.0 mm.


Grade Group in reporting of  prostate acinar adenocarcinoma:

  • The 9 Gleason scores (2-10) have been variably lumped into different groups for prognosis and patient management purposes. Epstein and associates proposed grouping scores into 5 prognostic categories, grade groups 1-5. 
  • This grade grouping, shown below in the table, strongly correlate with biochemical recurrence and have been incorporated into the new Partin tables.
  •  At the 2014 ISUP Consensus Conference, details of this prognostic system were clarified and it was recommended for usage together with the Gleason system.
  • This grade grouping has also been subsequently validated by other independent studies in surgical and radiation cohorts show significant correlation with survival.
  • The new grade grouping has been endorsed in the 2016 WHO classification.
  • The grade grouping has also been endorsed by ISUP and is referred to as ISUP grade in some publications. Like Gleason scoring in needle biopsies, the grade group can be applied at core, specimen, or case levels.


Methods for estimating the size/extent of DCIS in specimen.

Why to measure size of DCIS in specimen:

  • Higher rates of invasive cancer detected according to DCIS size. 
  • Progression to invasive cancer occurred in 10% of DCIS patients with a  DCIS tumor size between 2.5 to 3.5 cms, 57% for tumor size 3.6 to 4.5 cms and 71% for tumors between 4.5 and 6 cms.
  • Tumors over 2.5 cms have a higher risk of progressing to invasive cancers.

J Exp Clin Cancer Res. 2006 Jun;25(2):223-7.

There are multiple methods for estimating the extent of DCIS (see Figure):

  • DCIS in 1 block: The area involved by DCIS can be measured from a single slide, if DCIS is present in only 1 block. If separate foci are present, the largest distance between foci should be reported. This method will underestimate the extent of DCIS when multiple blocks are involved and should not be used in such cases.
  •  Serial sequential sampling: The entire specimen is blocked out in such a way that the location of each block can be determined. The extent of the DCIS can be calculated by using a diagram of the specimen, the thickness of the slices, and the location of the involved blocks.7-9 This method is recommended for all excisions likely to harbor DCIS or with previously diagnosed DCIS (eg, by diagnosis on a prior core needle biopsy).
  •  Nonsequential sampling: The number of blocks involved by DCIS is correlated with the extent of DCIS up to 40 mm.8 Multiplying the number of blocks involved by DCIS by the approximate width of a tissue section gives an estimate of the extent. In 2 studies, multiplying by 3 mm underestimated the extent of DCIS, and multiplying by 5 mm may overestimate the extent.8,9 Therefore, multiplying by 4 mm is recommended unless there is additional information that a different number would yield a more accurate result. This method may underestimate extent if not all areas of DCIS are sampled. Therefore, it is recommended that all tissue likely to be involved by DCIS be sampled (eg, all grossly abnormal tissue and all tissue with radiologically suspicious calcifications). When feasible, the entire specimen should be examined microscopically.
  • This method may result in a larger estimation of extent than the serial sequential sampling method when DCIS is present in a large volume of tissue in 3 dimensions rather than in a predominantly linear distribution. The best estimate for correlation with outcomes (eg, residual disease or recurrence) will require further studies.
  • This method can be applied to any specimen and will give a better estimation of extent than measuring extent on a single slide when multiple blocks contain DCIS.
  • • Margins: If DCIS involves or is close to 2 opposing margins, the distance between the margins can be used as the extent of the DCIS within the specimen.
  • • Gross lesions: In some cases of high-grade DCIS, there may be a gross lesion that can be measured. Confirmation of the gross size must be confirmed by microscopic evaluation.
  • The largest estimate obtained using any of these methods should be used to report the estimated size (extent) of the DCIS.

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