CANCER 161 CYTOPATHOLOGY Fine-Needle Aspiration Biopsy of Chromophobe Renal Cell Carcinoma and Oncocytoma Comparison of Cytomorphologic Features Beata A. Wiatrowska, M.D. Maureen F. Zakowski, M.D. Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, New York. BACKGROUND. Chromophobe renal cell carcinoma (ChRCC) is a distinct tumor with a prognosis intermediate between renal oncocytoma (RO) and clear cell renal cell carcinoma. To our knowledge the cytologic features of only a limited number of ChRCC have been described to date. A retrospective review of the cytomorphologic features of ChRCC and a comparison with RO was performed. METHODS. Fine-needle aspiration biopsies (FNABs) of six cases of histopathologically proven ChRCC were reviewed. The material examined was comprised of smears, cytospins, Thin Prept Pap Test™ preparations, and cell block sections stained with Diff-Quikt, Papanicolaou, and hematoxylin and eosin. Six FNABs of ROs were examined similarly. The cytomorphology of each tumor was studied and particular attention was paid to features differentiating the two entities. RESULTS. The characteristic cytomorphologic features of ChRCC (present in all cases) included round/oval, occasionally polygonal, moderately pleomorphic large cells present singly and in small clusters. The abundant cytoplasm was variegated, ranging from dense to flocculent to vacuolated, with prominent cytoplasmic membranes. The nuclei were large and hyperchromatic, with nuclear membrane irregularities and grooves present at least focally. Frequent binucleation was observed. Small nucleoli were present in many cells, but rarely prominent. In contrast, RO showed large cells with homogenous granular cytoplasm. The nuclei showed minimal to no nuclear membrane irregularities, tiny nucleoli, mild pleomorphism, and only an occasional large, more hyperchromatic nucleus was observed. CONCLUSIONS. ChRCC has a distinct combination of cytomorphologic features. Careful attention to cytoplasmic and nuclear features allows for the distinction between ChRCC and RO in cytologic preparations. Cancer (Cancer Cytopathol) 1999;87:161–7. © 1999 American Cancer Society. KEYWORDS: chromophobe, renal cell carcinoma, oncocytoma, cytology, differentiating features. Presented in abstract form at the American Society of Cytopathology Annual Meeting, Nashville, Tennessee, November 3–7, 1998. Address for reprints: Maureen F. Zakowski, M.D., Cytology Service, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021. Received August 26, 1998; revision received December 28, 1998; accepted January 6, 1999. © 1999 American Cancer Society C hromophobe renal cell carcinoma (ChRCC) is a tumor with distinct morphologic, histochemical, ultrastructural, and genetic features. The lesions are well circumscribed and solitary, with a graybrown homogeneous cut surface. The tumor cells are large and polygonal with well defined borders and abundant cytoplasm that is pale (typical) or granular and strongly acidophilic (eosinophilic variant). The nuclei show slight pleomorphism and hyperchromasia with coarsely granular chromatin, irregular nuclear membranes, and occasional binucleation. There is diffuse cytoplasmic positivity of tumor cells for colloidal iron. The cells express cytokeratins but are vimentin negative. The characteristic ultrastructural feature is the presence of 162 CANCER (CANCER CYTOPATHOLOGY) June 25, 1999 / Volume 87 / Number 3 FIGURE 1. Cluster of cells of chromophobe renal cell carcinoma showing large round to oval cells with well defined cytoplasmic membranes. (DiffQuikT, 3200) round to oval cytoplasmic vesicles varying in size from 150 –300 nanometers that are concentrated around the nucleus.1 The distinct genetic features of ChRCC include allelic losses at chromosomes 1p, 2p, 6p, 10p, 13q, 17p, and 21q.2 The prognosis for ChRCC is intermediate between renal oncocytoma (RO) and clear cell renal carcinoma.3-5 The cancer staging system used for renal tumors including RO and ChRCC carcinoma is that of the American Joint Committee on Cancer.6 To our knowledge the cytologic features of only a limited number of cases of ChRCC have been described to date.7-9 A retrospective review of the cytomorphologic features of ChRCC and comparison with RO was performed in our laboratory. MATERIALS AND METHODS The files of the pathology department of Memorial Hospital were searched for cases of histologically confirmed ChRCC for which there was previous or concomitant cytologic material. Seven fine-needle aspiration biopsies (FNABs) (five renal and two extrarenal) were found. Material examined was comprised of smears, cytospin, ThinPrept Pap Test™ (Cytyc Corporation, Boxborough, MA) preparations, and cell block sections stained with Diff-Quikt, Papanicolaou, and hematoxylin and eosin (H & E). The histologic sections from nephrectomy specimens were prepared in a standard fashion, cut 4-mm thick, and stained with H & E and Hale’s colloidal iron. Six cases of RO were similarly identified. The cytomorphology of each tu- mor was studied, including the background, architecture, cell size and shape, cytoplasmic quantity and quality, nuclear shape and size, binucleation chromatin pattern, and presence and prominence of nucleoli. Particular attention was paid to features differentiating ChRCC from RO. No distinction was made between the typical and eosinophilic types of ChRCC. RESULTS FNABs from seven patients with ChRCC were reviewed. There were four females and three males, with an age range of 41–75 years (mean, 54 years). FNAB sites included the right kidney (five patients), the liver (one patient), and a left supraclavicular mass (one patient). In addition to the smears, two ThinPrept preparations, one cytospin preparation, and two cell block sections were available for review. FNABs from six patients with RO were reviewed similarly. There were two females and four males, with an age range of 63– 80 years (mean, 72 years). Four FNABs were from the right kidney and two were from the left kidney. In addition to air-dried, Diff-Quikt stained and alcohol fixed, Papanicolaou, and/or H & E stained smears, one ThinPrept and two cell block sections were available. All ChRCCs originally were recognized as malignant, with four diagnosed as RCC not further classified, one as RCC chromophobe cell type, one adenocarcinoma of renal versus adrenal origin, and one carcinoma favor transitional cell carcinoma. The cytomorphologic features of ChRCC (Figs. 1, 2, and 3) are detailed in Table 1. The cellularity was high with small clusters and single cells present in all Cytology of Chromophobe Carcinoma and Oncocytoma/Wiatrowska and Zakowski 163 FIGURE 2. Chromophobe renal cell carcinoma showing (A) variegated cytoplasm ranging from granular to flocculent to vacuolated (3200). (B) Single binucleate cell with vacuolated cytoplasm (DiffQuikT, 3400). FIGURE 3. Small cluster of chromophobe renal cell carcinoma demonstrating “raisinoid” nuclei, longitudinal nuclear grooves, and perinuclear halos (Papanicolaou stain, 3400). cases. The cells were large, round to oval, with abundant, variegated cytoplasm. A thick, prominent cytoplasmic membrane was observed in six of the seven cases. The cytoplasm ranged from granular to flocculent to vacuolated in all cases. Perinuclear clearing or halos were observed in three of seven cases in cyto- logic preparations and was appreciated better in cell block sections. There was moderate cellular pleomorphism, with greater variation in size than in shape of the cells. Although the nuclei were large or moderate to large in all cases the nuclear to cytoplasmic ratio (N:C) remained low due to the abundance of cyto- 164 CANCER (CANCER CYTOPATHOLOGY) June 25, 1999 / Volume 87 / Number 3 TABLE 1 Predominant Cytomorphologic Features of Chromophobe Renal Cell Carcinoma and Renal Oncocytoma Cytoplasm Cellular pleomorphism Nuclear features Nucleoli Fuhrman nuclear grade Other ChRCC RO Variegated, dense, granular, flocculent, vacuolated perinuclear clearing Thick/prominent cytoplasmic membrane Moderate Moderate pleomorphism Mildly irregular nuclear outline Grooves Intranuclear inclusions Raisinoid nuclei Rare 2–3 Uniform, granular, dense Well defined cytoplasmic membrane Subpopulation of larger cells with large, irregular hyperchromatic nuclei and occasional nucleoli Mild Mild pleomorphism Smooth contour No grooves Rare inclusions No raisinoid nuclei Occasional 1–2 Subpopulation of smaller cells with a higher N: C ratio ChRCC: chromophobe renal cell carcinoma; RO: renal oncocytoma; N:C: nuclear:cytoplasmic. plasm. Binucleation was frequent (present in five of seven cases). The large nuclei were round to oval with moderate pleomorphism. The nuclear membrane irregularities, including nuclear grooves, were present at least focally in all cases, intranuclear pseudoinclusions were present in three of seven cases, and raisinoid nuclei were present in four of seven cases. The nucleoli were present in many cells in five of seven cases; however, they were small and rarely prominent. The Fuhrman nuclear grade was 2–3 in all cases. Also noted in two of seven cases was a subpopulation of larger cells with large, irregular, hyperchromatic nuclei and occasional nucleoli. The background was clean (or bloody) with no necrosis and no inflammation in any case. FNABs from RO (Figures 4, 5, and 6) also were cellular with a similar architectural arrangement of small clusters and single cells (Table 1). The cells were large with a low N:C ratio but, in contrast to ChRCC, their cytoplasm was homogenous and granular in all cases. The nuclear features also were markedly different with only mild pleomorphism, absent or minimal nuclear membrane irregularities with no grooves, and no raisinoid nuclei. Very small nucleoli were observed in four of six cases. The Fuhrman nuclear index was 1-2 in all cases, with a grade of 2 granted mainly on the basis of the nucleolar presence. Only in one case were larger, more hyperchromatic nuclei observed occasionally among otherwise classic-appearing oncocytic cells. A subpopulation of similar cells with a higher N:C ratio and the same granular cytoplasm and low grade nuclei were observed in FNABs from two cases. The cytologic features differentiating ChRCC from RO are summarized in Table 2. DISCUSSION ChRCC first was described by Thoenes et al. in 1985.10 It is believed that ChRCC shows differentiation toward the intercalated cells of collecting ducts with numerous cytoplasmic vesicles containing carbonic anhydrases.11 It comprises between 2–5% of RCCs.1,12 Considered a clinicopathologic entity, it has distinct morphologic, histochemical, ultrastructural, and genetic2,3,10,11,13-15 features that are different from other types of RCC and RO. The characteristic cytomorphologic features noted in our series included large cells with variegated cytoplasm in all cases ranging from dense to granular to flocculent to vacuolated with perinuclear clearing in some cells. The characteristic nuclear features included moderate pleomorphism with irregular nuclear outlines, grooves, pseudoinclusions, and raisinoid nuclei. The nucleoli were relatively rare and smaller than expected compared with the overall appearance and pleomorphism of the nuclei. The cytologic features corresponded well with the histopathologic picture, including cytoplasmic and nuclear characteristics, particularly the admixture of pale and highly eosinophilic cells with many microvacuolated cells and nuclear hyperchromasia with an irregular nuclear membrane. These cytoplasmic features are similar to those in what to our knowledge are the relatively few cases described in the literature7-9 and correspond particularly well with the cases described most recently by Granter and Renshaw.9 It has been noted both in the histopathologic material and in the previous cytologic descriptions of ChRCC that the most difficult differential diagnosis for ChRCC most likely is that of an RO.7,9 Renshaw et al. reported that they were able to identify all four ROs and two ChRCC successfully in a retrospective review of 38 renal cell lesions.16 However, on the initial review, only one ChRCC was identified correctly by all authors and two of the ROs were misinterpreted as ChRCC. In our review of six cases of RO the most characteristic features included uniform granular cytoplasm, mild nuclear pleomorphism, round nuclei with smooth nuclear contour, frequent but not prominent nucleoli, and a subpopulation of smaller cells with a higher N:C ratio but the same cytoplasmic features. These match earlier reports.17-20 The differentiating features between the two entities include both cytoplasmic and nuclear characteristics. The most typical feature is the variegated appearance of Cytology of Chromophobe Carcinoma and Oncocytoma/Wiatrowska and Zakowski 165 FIGURES 4 AND 5. Renal oncocytoma showing small clusters and single cells. Note the uniformity of the cytoplasm (Figure 4: H & E, 3100) (Figure 5: DiffQuikT, 3200). cytoplasm in ChRCC and its monomorphism in RO, with very rare cytoplasmic vacuolation. Also the nuclear features are very different with typical pleomorphism, a very irregular nuclear membrane, and grooves of raisinoid nuclei with small nucleoli observed in RO. This observation contrasts somewhat with the report by Akhtar and Ali7 that found the nuclear morphology of RO and ChRCC to be essentially similar, with no nucleoli observed in RO. We observed nucleoli, albeit small, in many cells in RO. Because ChRCC is considered to have a prognosis intermediate between other types of RCC and RO3-5 the treatment usually is simple nephrectomy, although recently partial nephrectomy has been noted to be considered more often.21 RO is a benign entity and can be left untreated, although many physicians prefer to treat it surgically rather than leave the tumor in, fearing sampling error and the true diagnosis of RCC with oncocytic features. Nevertheless, in some patients (e.g., those with high surgical risk or known metastatic disease) nonsurgical treatment may be a viable and preferred option (one of the patients in the 166 CANCER (CANCER CYTOPATHOLOGY) June 25, 1999 / Volume 87 / Number 3 FIGURE 6. Low power and insert of renal oncocytoma demonstrating round, regular nuclei without grooves and homogenous granular cytoplasm (DiffQuikT, 3100). current study had biopsy proven metastatic melanoma at the time of the diagnosis of RO; the patient did not undergo surgery and died 2 years later of melanoma). Other important differential diagnostic considerations for ChRCC are clear and granular cell types of RCC. The features of RCC include large clusters of cells with abundant, clear to finely vacuolated, wispy, or granular cytoplasm; many stripped nuclei due to cytoplasmic fragility; centrally located round to slightly irregular nuclei with pale chromatin; and prominent nucleoli. Hyaline globules and phagocytized red blood cells may be observed in the cytoplasm of RCC. Renshaw et al.16 pointed out that in general the cells of clear cell RCC form larger groups and have more uniform nuclear size and round shape than cells of ChRCC. In comparing ChRCC and the granular cell variant of RCC, Akhtar and Ali7 noted indistinct cellular borders with irregular outlines and large nuclei with open chromatin and single, well developed nucleoli in the latter entity. With regard to other, less common differential diagnoses of ChRCC, the ones that arose from our initial diagnosis included adrenal cortical carcinoma and transitional cell carcinoma (TCC). The differential diagnosis between adrenal cortical carcinoma and RCC, particularly of the granular or clear cell variants, often is difficult or virtually impossible by morphology alone.22 FNAB of adrenal cortical carcinoma can vary in appearance from bland, lipid-laden cells to anaplastic malignant cells with dense acidophilic cytoplasm. Occasional giant bizarre as well as spindle cells can be observed. The nuclei also vary from small and round to large and anaplastic with irregularities of the nuclear membrane, abnormal chromatin, and prominent nucleoli. Occasionally oncocytic cells can be present.23 However, unlike ChRCC, perinuclear clearing with raisinoid nuclei (koilocyte-like) is not a feature of adrenal cortical carcinoma. In addition, an increase in the nuclear grade of this entity is accompanied by the presence of prominent nucleoli. With regard to the differential diagnosis between ChRCC and TCC, low grade TCC usually has papillary groups of nearly normal transitional cells with minimal nuclear atypia whereas high grade lesions show relatively dense cytoplasm with only scattered vacuoles with no flocculent or microvacuolated qualities. The nuclei are high grade with coarse chromatin and prominent nucleoli with no koilocyte-like appearance.24 Conclusions ChRCC has distinct cytologic features that allow for the correct diagnosis on FNAB. ChRCC can be differentiated from RO when attention is paid to cytoplasmic and particularly nuclear features. Although not usually a critical consideration, the distinction of ChRCC from RO may be important in certain clinical settings. Cytology of Chromophobe Carcinoma and Oncocytoma/Wiatrowska and Zakowski TABLE 2 Differentiating Cytologic Features between Chromophobe Renal Cell Carcinoma and Renal Oncocytoma ChRCC RO 7/7 7/7 7/7 5/6 6/6 6/6 7/7 7/7 0/7 6/7 6/7 7/7 3/7 6/7 — Mod 6/7 7/7 — 7/7 Mild 7/7 7/7 3/7 4/7 Mod 5/7 Mild/Mod 7/7 7/7 — 5/7 2–3 1/7 2/7 0/7 6/6 0/6 6/6 6/6 6/6 2/6 0/6 0/6 6/6 Mild 4/6 6/6 5/6 6/6 Mild 2/6 0/6 1/6 0/6 Mild 5/6 Mild 6/6 — Rare 3/6 4/6 1–2 0/6 0/6 0/6 6. 7. High/moderate cellularity Small clusters/single cells Large, round oval cells Cytoplasm Abundant Variegated Uniform Granular Flocculent Vacuolated Perinuclear clearing Thick/prominent cytoplasmic membrane Well defined cytoplasmic membrane Cellular pleomorphism Low N:C ratio Binucleation Round/oval, mod/large nuclei Irregular nuclear membrane Nuclear grooves Intranuclear inclusions Raisinoid nuclei Nuclear pleomorphism Hyperchromasia Chromocenters Clumpy chromatin Nucleoli Fuhrman nuclear grade Mitosis Cells with large hyperchromatic irreg nuclei Necrosis ChRCC: chromophobe renal cell carcinoma; RO: renal oncocytoma; mod: moderate; N:C: nuclear: cytoplasmic; irreg: irregular. REFERENCES 1. 2. 3. 4. 5. Murphy WM, Beckwith JB, Farrow GM. Tumors of the kidney, bladder, and related urinary structures. In: Atlas of tumor pathology. 3rd series. Fascicle 2. Rosai J, Sobin L, eds. Washington, DC: Armed Forces Institute of Pathology, 1994: 112–3. 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