The 3 common major histologic subtypes of renal cell carcinoma (RCC) are clear cell renal cell carcinoma (CCRCC), papillary renal cell carcinoma (PRCC), and suspicious cell renal cell carcinoma (CPRCC) in that order. Other rare RCCs include renal collecting duct carcinoma (CDC), mucinous tubule and spindle cell carcinoma (MTSCC), and RCC with MiTF/TFE3 gene translocation. With the increasing understanding of molecular genetics in RCC pathogenesis, many new RCC subtypes such as clear cell papillary renal cell carcinoma, tubular cystic renal cell carcinoma, and renal cell carcinoma associated with end-stage renal disease have been discovered in recent years, and each of these subtypes has unique clinicopathological features, prognostic significance, and molecular genetic background. Primary malignancies from other sites can also metastasize to the kidney. Accurate histologic differential diagnosis and subclassification of renal cell carcinoma are important to guide the clinical management of patients. However, there are obvious morphological overlaps between various histological subtypes of renal cell carcinoma, and certain renal tumors coming from distant metastases or direct spread from surrounding organs are extremely difficult to distinguish from primary renal tumors under light microscopy; therefore, ancillary diagnostic tools, especially immunohistochemical staining (IHC), have important utilization value in the differential diagnosis of renal tumors. (A) Immunohistochemical markers commonly used to diagnose renal tumors RCC Marker (Renal Cell Carcinoma Marker) RCC Ma is an antibody against glycoprotein antigen on the brush border of the proximal renal tubule in human kidney. CD10 is a type II cell surface glycoprotein molecule of approximately 100 kD size, and in normal renal tissue CD10 is expressed on the epithelial brush border of the proximal tubule and in the glomerular podocytes. e-cadherin and KSC (kidney-specific binding protein) : E-cadherin is a calcium-dependent cell adhesion molecule that regulates cell-cell junctions. ksc is KSC is a subunit of cadherin. Both are markers of the distal renal unit. PAX-2 is a nuclear transcription factor associated with the development and maturation process of the kidney, and most renal epithelial tumors express PAX-2 nuclearly. PAX-8 is another nuclear transcription factor closely related to PAX2, and its expression profile in renal tumors is similar to that of PAX2. Cytokeratin (CK) Different types of CK expression can be used to differentiate renal tumors, the most widely studied being CK7. Vimentin As a broad-spectrum mesenchymal marker, Vimentin can be expressed in many RCCs. CD117 is a proto-oncogene protein encoding a complexine transmembrane receptor and is widely used in the diagnosis of gastrointestinal mesenchymal tumors, most of which TFE3, TFEB, and Cathepsin-K (histone K) Xp11.2 translocated renal cell carcinoma immunohistochemically characterized by nuclear expression of the fusion gene transcription factor TFEB is another fusion gene transcription factor that is significantly overexpressed in renal cell carcinoma with t(6;11)(p21;q21) translocation, and nuclear expression of TFEB by immunohistochemistry is diagnostic for this subtype of RCC. cathepsin-K is The immunohistochemical staining for Cathepsin-K is useful for differentiating RCC with MiTF/TFE3 translocation from other RCCs. AMACR is a mitochondrial enzyme that regulates fatty acid synthesis and is expressed in normal kidney in the proximal Claudin-7, Claudin-8 Claudin-7 is a tight junction protein expressed in the distal renal unit of the human kidney. CD82 is a metastasis suppressor gene. Carbonic anhydrase (CA IX) is a complex transmembrane enzyme protein that regulates cell proliferation, adhesion and infiltration. (B) Immunohistochemical staining in the differential diagnosis of renal tumors Clear cell RCC vs. suspicious cell RCC Clear cell RCC and suspicious cell RCC are two completely different subtypes of renal cell carcinoma, and it is not difficult to distinguish them in the classical histological context. Sometimes the two tumors may show overlapping morphologic features that make differential diagnosis difficult, including clear cytoplasm, well-defined cell borders, and granular eosinophilic cytoplasm. However, in terms of prognosis, suspicious cell RCC is significantly better than clear cell RCC, including a 100% 5-year and 90% 10-year survival rate in the former, so it is important to accurately distinguish between these two subtypes. Suspicious cell RCC vs. eosinophilic adenoma Suspicious cell RCC and eosinophilic adenoma both originate from the distal renal tubules, and both have partial overlap in morphologic and immunophenotypic features. The eosinophilic subtype of RCC is difficult to differentiate from eosinophil adenoma, and both have a granular eosinophilic cytoplasm with poorly defined cellular contours. Some focal areas of typical eosinophilic adenomas may exhibit a perinuclear halo with a clear perinuclear appearance similar to that of RCC tumor cells. Eosinophilic adenoma is typically a benign tumor, whereas RCC is usually a low-grade malignant tumor with the potential for sarcomatoid dedifferentiation and distant metastasis. In some particularly difficult cases, morphologic features combined with appropriate immunohistochemical markers help us make a predictive diagnosis. Almost all eosinophilic adenomas express CK8 and CK18, characteristically presenting with a paranuclear and perimembranous globular granular staining pattern. As mentioned previously, the combined application of CK7, S-100A1 and CD82 may help to differentiate suspensory cell RCC from eosinophilic adenoma. Both express CD117, KSC and neither express Vimentin, which may reflect their common histogenetic origin, and the combined application of these markers may help to differentiate them from other renal tumors. Papillary RCC vs. mucinous tubular and spindle cell carcinoma (MTSCC) Papillary RCC characteristically presents with extensive papillae formation with or without interstitial foam cell aggregates in a ciliated vascular axis, with certain rare features including focal tubule formation and focal mucus secretion. The tumor cells are distributed in narrow tubular and extended spindle-shaped cells, and focal papillae formation and foam cell aggregates may be present. Recently, a mucus less subtype of MTCSS and papillary RCC with low grade spindle cell foci have been reported, while both papillary RCC and MSTCC may occasionally show sarcomatous differentiation with high grade spindle cells. In terms of immunophenotypic features, the two also have significant similarities, showing varying degrees of evidence of proximal and distal tubular differentiation. Indeed some authors have suggested that MTSCC may be a morphologic variant of papillary RCC, but genetic features do not support this hypothesis, and cytogenetic analysis has shown that papillary RCC typically presents with chromosome 7,17 gain and Y chromosome loss, which is rarely seen in MTSCC. Clear cell RCC vs. MiTF/TFE3 translocation RCC vs. epithelial MiTF/TFE3 translocation RCC is a recently described group of RCC with translocations of small eye transcription factors (including TFE3,TFEB, etc.) MiTF/TFE3 translocation RCC is morphologically easily confused with clear cell RCC, especially CCRCC with high Fuhrman nuclear grade, and immunohistochemical staining shows characteristic strong and diffuse nuclear TFE3 or TFEB expression is essential for the definitive diagnosis of genetically translocated RCC. As mentioned previously, the expression of Cathepsin-K, a target protein downstream of the small eye transcription factor, also contributes to its differential diagnosis from other RCCs. Unlike common RCCs, RCCs with MiTF/TFE3 translocation usually express less or only focally CK and EMA, especially certain rare TFE3 fusion types and α-TFEB fusion RCCs that barely express broad-spectrum keratins and abnormally express melanin markers such as HMB-45, which can cause differential diagnosis with renal vascular smooth muscle lipoma (AML) and especially its pure epithelioid subtype ( EAML presents morphologically as solid patches of epithelioid aggregates with abundant cytoplasm and expresses Cathepsin-K, HMB-45, and less myogenic markers (e.g., SMA, Desmin, etc.), and does not express CK or renal tumor markers. In contrast, RCC with MiTF/TFE3 translocation expresses PAX-8, CD10 and AMACR in addition to the characteristic nuclear expression of TFE3 or TFEB, which can be distinguished. RCC vs uroepithelial carcinoma These two types of tumors are mostly characterized by diffuse infiltration of patches of mesenchymal cells that often occupy a large portion of the renal parenchyma, and in the absence of certain histologic features that may suggest a tumor origin, the use of immunohistochemical packages can help distinguish between the two. UroplakinIII is a more specific marker of the uroepithelium staining approximately 57-100% of uroepithelial carcinomas, while high molecular weight keratin (HWCK) and thrombomodulin (thrombomodulin) appear to be more sensitive markers of uroepithelial tumor expression, and the majority of uroepithelial carcinomas express CK7 and P63, with varying degrees of CK5/6 and more than half expressing CK20. HWCK and thrombomodulin (thrombomodulin) appeared to be the more sensitive markers expressed in 80-90% and 68% of uroepithelial carcinomas, respectively. Almost none of these markers were expressed in RCC except CK7 and HWCK, whereas uroepithelial carcinomas typically do not express RCC markers such as RCC Ma, CD10, PAX-2, and PAX-8. Papillary RCC VS posterior renal adenoma (MA) Type I papillary RCC, especially the solid subtype, has significant overlap with posterior renal adenoma (MA) in terms of histopathological features. In addition to the typical papillae with a fibrovascular axis, papillary RCC can also appear as beams, tubules, glomeruli and solid growth patterns, whereas MA consists mainly of densely arranged tubular structures with sparse interstitium and focal areas of tubules that form papillary infoldings resembling immature glomeruli. A few studies have found that both papillary RCC and posterior renal adenoma can present with trisomy 7, 17 and Y chromosome loss suggesting a possible genetic link between the two, while most investigators do not agree with this view. Immunohistochemical staining can be helpful in cases where it is difficult to distinguish between the two, especially when papillary RCC shows more extensive tubular solid growth and MA has more papillary structures. Most MA expresses S-100, CD57, WT-1, less AMACR, and basically no CK7, whereas papillary RCC is the opposite, with 96-100% of papillary RCC expressing AMACR and 100% not expressing S-100. studies have been reported. Sarcomatoid RCC versus pure sarcoma Almost any subtype of RCC can present with a sarcomatoid transformation suggesting a poor prognosis. In the 2004 WHO classification, sarcomatoid RCC is not a tumor entity, but represents a common morphologic presentation of various subtypes of RCC presenting with high-grade transformation. Thus most sarcomatoid RCC will be accompanied by an identifiable low-grade or classic RCC component. Immunohistochemical staining provides some useful diagnostic clues to differentiate between the two. In contrast, true sarcomas often do not express CK and EMA, but may express CD34, SMA, MSADesmin, S-100, MyoD1, and other mesenchymal markers to varying degrees, depending on their direction of differentiation. Markers. Epithelioid vascular smooth muscle lipoma may show varying degrees of cellular atypia, and its characteristic co-expression of melanin marker HMB-45 and myogenic marker SMA can be distinguished from sarcomatoid carcinoma. Other renal tumor markers including CD10, KSC, and RCC Ma were not expressed in sarcomatoid RCC versus sarcoma, and therefore were not helpful in the differential diagnosis. In contrast, the expression of PAX-2 and PAX-8 in sarcomatoid RCC has been reported with inconsistent results; in general, positive expression of PAX-2 or PAX-8 is confirmatory for the diagnosis of sarcomatoid carcinoma in sarcomatoid tumors with apparent primary renal origin, while negative expression results do not exclude the sarcomatoid RCC component. Differentiation of collecting duct carcinoma (CDC) from other high-grade tumors CDC is a rare and highly aggressive tumor originating from the distal renal collecting duct, accounting for less than 1% of all renal tumors. Histologically, the tumor cells show adenoidal, tubular papillary infiltrative growth with varying degrees of spike-like cell differentiation and extensive interstitial pro-fibroproliferative responses with mixed chronic inflammatory cell infiltration. The main tumors that can be morphologically confused with CDC are other high-grade RCC, high-grade uroepithelial carcinoma, and metastatic carcinoma. Two recent studies demonstrated the significance of combined P63,PAX-8 expression in the differential diagnosis of CDC and high-grade uroepithelial carcinoma, with all collecting duct carcinomas (100%) expressing PAX-8 and the majority of uroepithelial carcinomas (83-91.2%) not expressing PAX-8, but 97-100% expressing P63. The PAX-8+/p63- immunophenotypic profile supports the diagnosis. CDC had 83-85.7% sensitivity and 100% specificity, in contrast to the immunophenotypic profile of PAX-8-/P63+ which had 88-88.2% sensitivity and 100% specificity for the diagnosis of uroepithelial carcinoma. Medullary carcinoma is a very distinctive and highly aggressive renal epithelial tumor with significant histologic overlap with collecting duct carcinoma. Other tumors associated with INI1 deletion include renal and extrarenal malignant rhabdomyosarcoma, atypical teratoid/rhabdomyosarcoma of the central nervous system, extraosseous mucinous chondrosarcoma etc. (iii) Differential diagnosis of metastatic RCC Compared with primary renal tumors, metastatic renal tumors have some unique clinical or pathological manifestations that may cause confusion. For example, certain metastatic renal cancers may appear before the tumor is found at the primary site; metastatic tumors may be more undifferentiated morphologically or exhibit histological features that are significantly different from those of the primary tumor, especially after receiving radiotherapy treatment. Alternatively, within the metastatic organ, renal cell carcinoma may closely resemble the morphology of certain primary tumors in that organ, and in addition metastatic renal tumors may show loss of their original immunophenotypic features, among others. Therefore, a wide range of immunomarkers are important for the identification of these tumors. Organ-specific immunomarkers are particularly useful in the diagnosis of metastatic renal cell carcinoma, such as PAX-2.PAX-8, but their expression in metastatic tumors usually appears to be lost or absent to varying degrees. It should be noted that other tumors of non-renal origin that co-express PAX-2,PAX-8 include parathyroid adenoma/carcinoma, ovarian carcinoma of Mullerian epithelial origin, Merkel cell carcinoma, adenoma of renal origin, and lymphoma. CD10 is significantly less sensitive and specific for the diagnosis of metastatic RCC with RCC Ma compared to PAX-2 or PAX-8, and most studies have shown that CD10 is only useful for identifying metastatic clear cell RCC; however, many tumors of non-renal origin express CD10, such as skin appendage tumors and endometrial mesenchymal tumors. A recent comprehensive immunohistochemical study has shown that the combination of PAX-2, PAX-8, CD10, RCC Maker and human kidney injury molecule-1 (hKIM) and CA IX is highly sensitive and specific for the diagnosis of metastatic clear cell RCC. kSC is basically useful only for the diagnosis of metastatic suspensory cell RCC, however, this subtype of RCC rarely metastasis (less than 2%). The vast majority of metastatic papillary RCCs express AMACR, but this marker is also expressed by tumors of non-renal origin, including those of gastrointestinal, liver, bladder, lung, breast, and ovarian origin, as well as certain neuroendocrine tumors. The diagnosis of metastatic sarcomatoid RCC is a difficult one, and immunohistochemistry currently appears to be of little use in identifying metastatic RCC, as studies have shown that some metastatic RCC do not express all of the above-mentioned markers associated with renal tumors. In summary, there is no single immunomarker that is absolutely reliable for the differential diagnosis of metastatic RCC. The combination of a range of immunohistochemical markers, complemented by certain metastatic site organ-specific markers or more specific markers for various non-renal tumors that need to be distinguished from metastatic RCC, is important for the differential diagnosis of metastatic RCC. However, it should be kept in mind that any evaluation of immunohistochemical staining results always needs to be combined with morphologic features and clinicopathologic background, and any diagnosis made by relying solely on immunohistochemical staining without analysis may ultimately have serious adverse consequences.