Carcinomas of Unknown Primary Site Kandalaft et al

(Arch Pathol Lab Med.doi: 10.5858/arpa.2015-0173-CP)

PRACTICAL APPLICATIONS IN

IMMUNOHISTOCHEMISTRY

Approximately 4% of all patients with cancer present as CUPs

IHC remains a gold standard at diagnosis

2 classes of antibody markers that can help- Antibodies to keratins- Antibodies to organ restricted markers

Determine the cell line of differentiation

Determine the CK type or types of distribution

Determine co expression of vimentin

Determine expression of supplemental Ags (CEA, EMA, PLAP)

Organ specific markers

KERATINSLow molecular

weight CK- “simple” epithelium- K8, K18- Glandular epithelium

of GIT, hepatocytes

High molecular weight CK

- “complex” epithelium

- K5, K14, K17- Stratified epithelium- Ductal and basal

cellsThe subclassification of carcinomas by HMW and LMW keratins has largely been superseded by subclassification using antibodies to K7 and K20 which is a far more powerful discriminator

CK 7 AND CK 20 - APPROACH

Cancer of Unknown PrimaryMD Anderson Cancer Center

Urothelial tumorsOvarian mucinous adenocarcinomaPancreatic adenocarcinomaCholangiocarcinomaGastric carcinoma

CK 7 POSITIVE/CK 20 POSITIVE

Lung adenocarcinomaBreast carcinomaThyroid carcinomaSalivary gland carcinomaEsophageal carcinoma

Endometrial carcinomaCervical carcinoma

CholangiocarcinomaPancreatic carcinomaGastric carcinoma

CK 7 POSITIVE/CK 20 NEGATIVE

As a rule, gastric

adenocarcinoma can show almost any

K7/K20 phenotype

Colorectal carcinomaMerkel cell carcinoma (dot like pattern)

CK 7 NEGATIVE/CK 20 POSITIVE

Hepatocellular carcinomaRenal cell carcinomaProstate carcinomaSquamous cell and small cell lung carcinomaHead and neck carcinoma

CK 7 NEGATIVE/CK 20 NEGATIVE

Keratin 5 (and its pair 14) – marker of squamous, transitional cell, myoepithelial and mesothelial differentiation

Keratin 17 – when expressed at high levels good marker for distinguishing between carcinomas of pancreatobiliary origin from gastric carcinomas

OTHER KERATINS

CK AND VIMENTIN COEXPRESSIONCARCINOMAS

Anaplastic thyroid carcinoma

Endometrial carcinomaMesotheliomaMetaplastic breast

carcinomaMyoepithelial carcinomaRenal cell carcinomaSarcomatoid carcinomaThyroid carcinomas

MESENCHYMALAdamantinomaChordomaDPSRCTEpithelioid

angiosarcomaEpithelioid sarcomaLeiomyosarcomaMalignant rhabdoid

tumorSynovial sarcoma

ORGAN SPECIFIC MARKERS

CLASSES OF TUMOR SPECIFIC ANTIBODIES

CYTOPLASMIC- Level of expression

and fraction Function of the state of differentiation of the tumor

NUCLEAR- When positive

entire tumor population

- Independent of the state of differentiation

BREAST CANCER MARKERS

2/3rd to 3/4th of primary breast Lower fraction of breast cancer in metastatic

sites

Primary in endometrium and ovaryPapillary carcinoma thyroidSkin adnexal tumors

10-20% of lung adenocarcinomas (focally)

Rare in adenocarcinoms of GIT

ESTROGEN RECEPTOR

23A3 monoclonal antibody- 80% sensitivityFunction of histological subtypeGreatest in lobular (particularly those with

signet ring cells) and those with apocrine features

Very small fraction of basal like carcinomas

Salivary gland carcinomas and sweat gland carcinomas

5-10% primary ovarian and endometrial carcinomas

5-6% lung adenocarcinomas

GCDFP 15

Sensitivity as a marker of breast cancer is LESS than that of GCDFP 15 (50-70%)

7% breast cancers are Mammaglobin A positive but GCDFP 15 negative

10% of ovarian and endometrial Skin adnexal and salivary gland tumors

MAMMAGLOBIN A

1 of 6 members of zinc finger transcription factor family

Very sensitive for breast and urothelial carcinoma

Ductal 91%, lobular 100% diffuse and strong nuclear staining

UNLIKE previous two seen in 43% of triple negative and 54% of metaplastic breast cancers

Maintained in metastatic breast cancer (>90%)

Skin adnexal, endometrial, pancreatic, salivary gland carcinomas

GATA BINDING PROTEIN 3

LUNG CANCER MARKERS

NKX2 family of DNA binding transcription factors

Selectively expressed during embryogenesis in the thyroid, diencephalon and respiratory epithelium

Expressed in both neuroendocrine and non neuroendocrine tumors of the lung

THYROID TRANSCRIPTION FACTOR-1

Sensitivity of TTF-1 is greatest among adenocarcinomas and nonmucinous bronchioloalveolar carcinomas

Lowest in mucinous adenocarcinomas and squamous cell carcinomas

Appears to retain similar sensitivity in metastatic sites

Small subset of ovarian, endometrial, and colorectal carcinomas, although the extent of positivity is usually focal, often in isolated clusters of cells

TTF-1 expression cannot be considered specific for high-grade neuroendocrine carcinomas of lung origin

Variable subset of small cell (neuroendocrine) carcinomas of the genitourinary and gynecologic (GYN) tract

Cell blocks of pleural fluids, which contain material that has been either fixed in alcohol or is nonfixed before creation of a formalin-fixed cell pellet, can manifest a profound loss of TTF-1 antigenicity

Aspartic protease that is crucial to the maturation of surfactant B and present in the cytoplasm of type 2 pneumocytes and alveolar macrophages

Very sensitive marker for detecting pulmonary adenocarcinomas

Subset of renal cell carcinomasMinority of endometrial adenocarcinomas and

papillary thyroid carcinomasVirtually all cases of clear cell carcinomas of the

ovary

NAPSIN A

GI TRACT MARKERS

Nuclear transcription factor controlling the proliferation and differentiation of intestinal epithelial cells

Virtually 100% of colorectal adenocarcinomas

MSI reduced or even absent expression

CRC uniform staining patternMost adenocarcinomas of the stomach,

pancreas, and biliary tract variegated or focal staining pattern

CDX2

½ gastric (more in intestinal type) and 1/3 of pancreatobiliary

Ovarian mucinous carcinomas, bladder adenocarcinomas, and sinonasal intestinal type adenocarcinomas

Limited subset of mucinous and nonmucinous pulmonary adenocarcinomas (enteric subtype)

Endocervical and endometrial mucinous differentiation

‘‘Squamous’’ morules of endometrioid hyperplasia and carcinoma

Germ cell tumors intestinal differentiation

GI neuroendocrine tumors, including those primary to the intestine (eg, carcinoid tumors) and, to a variable degree, the pancreas (islet cell tumors)

Actin-binding protein, found preferentially in microvilli

Expression is largely (but not entirely) restricted to glandular epithelium and corresponding adenocarcinomas of the GI tract

Expression is greatest and most reliably found in CRC

Lower levels of expression are found in adenocarcinomas primary to the pancreatobiliary tract and stomach

VILLIN

Scoring of the membranous or ‘‘brush border’’ signal is most significant

Cytoplasmic immunostaining can be seen in other types of tumors, particularly neuroendocrine carcinomas

Can also be seen in adenocarcinomas of other sites that display a GI-type histology and immunophenotype

Although the individual sensitivities of CDX2 and villin are each approximately 50%, their combined sensitivity is in excess of 75%.

HEPATOCELLULAR MARKERS

Detects a liver (hepatocyte)-specific marker, subsequently found to represent the enzyme carbamoyl phosphate synthase

Helps to distinguish metastatic carcinomas from primary HCCs

(1%–10%) subset of adenocarcinomas primary to the lung, pancreas, stomach, ovaries, and adrenal cortex hepatoid morphology

HEP-PAR 1 ANTIBODY (CPS1)

Enzyme involved in the urea cycle

Appears to represent the most-sensitive (and, perhaps, most-specific) marker of HCC to date

Cytoplasmic, granular pattern

High level of sensitivity even in the context of high-grade HCC

ARGINASE-1

It is not expressed in ‘‘hepatoid’’ and other non-HCCs (particularly carcinomas of the lung, stomach, and kidney)

This is the marker of choice for identifying HCC

Oncofetal protein

Proven useful in distinguishing HCC from nonneoplastic hepatic lesions and hepatic adenomas

In mets vs primary high level of sensitivity and specificity of arginase-1 to surpass the use of glypcian-3

GLYPICAN 3

GYN CANCER MARKERS

Nuclear transcription factor implicated in tumorigenesis and in specifying normal urogenital development

Mesothelial cells, ovarian surface epithelium, mesangial cells in the kidney, a subset of smooth muscle cells, and granulocytic cells and precursors

WILMS TUMOR ANTIBODY (WT1)

Marker of ovarian carcinomas in the context of adenocarcinomas

Mesothelioma distinguishing it from nonovarian adenocarcinomas

Desmoplastic small, round cell tumors

Ovarian serous carcinomas, primary peritoneal adenocarcinomas, and fallopian tube serous carcinomas

Very high sensitivity and specificity, both in excess of 90%.

In a poorly differentiated ovarian carcinoma, nuclear WT1 reactivity favors a serous neoplasm because endometrioid, clear cell and mucinous carcinomas are negative

In the breast, WT1 is expressed in around 6% of the cases, usually at low levels in pure mucinous (65%) and mixed mucinous (33%) subtypes

Subset of carcinomas arising within the female genital tract, exhibit nuclear expression for ER

In endometrial carcinomas of endometrioid type (type 1), ER antibodies are reactive

Whereas in uterine serous and clear cell carcinomas (type 2), they usually are not

ESTROGEN RECEPTOR

Can be part of a panel to differentiate endometrial adenocarcinoma from endocervical adenocarcinoma

No value in the distinction between a primary ovarian adenocarcinoma (mainly including endometrioid and serous carcinoma) and a metastasis from the breast or from elsewhere within the female genital tract

Transcription factor, which is critical to embryogenesis of the thyroid gland, kidney, and mullerian system

Nonciliated, mucosal cells of the fallopian tubes, endocervix, endometrium, and simple ovarian inclusion cysts BUT NOT on the surface of the epithelial cells of the ovary

90% to 100% of serous, endometrioid, clear cell, and transitional cell ovarian carcinomas

PAX 8

PAX8 is not expressed in mammary carcinomas, including ductal and lobular types

Because the ovary is a common site of involvement for metastasis by breast carcinoma, PAX8 can be a useful marker in the differential diagnosis of ovarian and breast carcinomas

Highly expressed in clear cell carcinomas of the ovary 100% of tumors

Clear cell carcinomas of the endometrium (82%)

Few endometrial serous carcinomas (8%)

NO endometrioid endometrial carcinoma

NAPSIN A

PROSTATE MARKERS

Very high sensitivity of this marker, apparently independent of Gleason score

Overall sensitivity in the range of 95% and specificity approaching 100%

Expressed by a subset of breast cancers

Also expressed focally in salivary gland and pancreatic carcinomas

PROSTATE SPECIFIC ANTIGEN

Antibody to the prostatic tumor suppressor gene NKX3.1

Recently reported to be an extremely sensitive marker for identifying metastatic prostatic adenocarcinoma (positive in 99%)

Level of sensitivity of NKX3.1 is maintained in high-grade prostatic carcinomas

NKX3.1

TCC MARKERS

More than 90% of urothelial carcinomas are positive

Useful marker in distinguishing TCC from other non–small cell carcinomas potentially in the differential diagnosis, such as, prostatic adenocarcinoma (especially high grade)

GATA3

Glycoprotein of the asymmetrical unit membrane, which forms plaques on the apical surfaces of urothelial umbrella cells

First, specific, urothelial-restricted marker described

High sensitivity in non invasive; low sensitivity in invasive

UROPLAKIN

RENAL CELL CARCINOMA MARKERS

Critical to the embryogenesis of the kidney, is identified in renal tubular epithelium and vas deferens, but not glomeruli

Most of the renal epithelial neoplasms

Clear cell > Papillary > Chromophobe = Sarcomatoid = Xp11 Translocation

Not expressed in bladder TCCSubset of renal pelvic urothelial carcinomas

PAX8

THYROID MARKERS

Specific and sensitive markers of both primary and metastatic carcinomas of the thyroid

Excellent marker of papillary and follicular carcinomas

Poor marker of anaplastic

NOT a marker for medullary

THYROGLOBULIN

Even more-sensitive marker of thyroid carcinomas than thyroglobulin

Medullary also

Anaplastic negative

TTF 1

Critical to the organogenesis of the thyroid gland and is highly expressed in the thyroid follicular epithelium

Papillary and follicular 100%

Anaplastic 80%

PAX8 is useful in discriminating between a TTF-1 + lung adenocarcinoma and a TTF-1+ thyroid carcinoma because PAX8 expression has not been identified in primary lung adenocarcinomas

PAX8

ADRENAL MARKERS

Expressed in a restricted subset of healthy cells, including ovarian granulosa cells, testicular Leydig cells, and adrenal cortical epithelium

Excellent marker for the identification of primary adrenal cortical tumors and their distinction from metastatic carcinomas to the adrenal gland

Ovarian and testicular stromal tumors

INHIBIN α

Alternative or supplementary marker of adrenal cortical differentiation

Sensitivity is comparable or even greater than that of Inhibin

MART 1 ANTIGEN

100% specificity at discriminating these neoplasms from other tumors with clear cell morphology, such as renal cell carcinoma, ovarian clear cell carcinoma, and chordomas

High levels in sex cord-stromal tumors of the ovary

Lower levels in testicular sex cord-stromal tumors

STEROIDOGENIC FACTOR 1

Unique among epithelial tumors very low level of keratins

Unique among nonneuroendocrine tumors synaptophysin

ANOMALOUS FINDINGS IN ADRENAL CORTICAL TUMORS

SQUAMOUS/ TRANSITIONAL CELL MARKERS (p63 and p40)

Uniformly and strongly p63 and p40 positive pure SCC (lung and cervix)

Thymomas can also be positive

SQUAMOUS DIFFERENTIATION

Uniform expression of p63 and p40, even in the setting of poorly differentiated tumors, such as spindle cell, bladder TCC

TRANSITIONAL DIFFERENTIATION

Carcinomas demonstrating myoepithelial differentiation (eg, adenoid cystic and other salivary gland carcinomas)

Carcinomas demonstrating trophoblastic differentiation.

NONSQUAMOUS, NONTRANSITIONAL CARCINOMAS

CASE 1

73 years old man

Long term smoker

Needle biopsy of single left lower lobe lung nodule

CK 20

CDX 2 TTF 1

Diagnosis Metastatic adenocarcinoma from rectosigmoid

CASE 2

63 years old male

Cervical lymph node needle biopsy

No known primary

Suspicious for lymphoma

CK 7

PSA VILLIN

Diagnosis Metastatic prostatic adenocarcinoma

CASE 3

55 years old woman

Right axillary lymph node biopsy

Ill defined density seen on mammogram

PET positive uptake in right parotid gland

GCDFP-15

GATA 3 HER 2

Diagnosis Metastatic ductal carcinoma from breast

CASE 4

73 years old woman

Needle biopsy of retroperitoneal lymph node

History of hysterectomy for unknown reasons many years back

Retroperitoneal lymphadenopathy, possible splenic metastasis

Left pelvic sidewall mass on CT scan - ?residual ovary

ER

WT 1 PAX8

Diagnosis Metastatic high grade serous carcinoma of ovary

CASE 5

58 years old man

Needle biopsy of mediastinal lymph node

Recent diagnosis of prostate adenocarcinoma (Gleason score 4, “hypernephroid”)

Nephrectomy 5 years ago for sarcomatoid renal cell carcinoma

Remote history of melanoma

Presented with mediastinal and lung masses

CK

PSA

PAX 8

Diagnosis PAX 8 + male thyroid and renal TTF negativeMetastatic renal cell carcinoma

Thank You