Hybrid Oncocytic/Chromophobe Tumor Of The Kidney Associated With Sporadic Renal Oncocytosis And Chronic B-Cell Lymphocytic Leukemia

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Miguel A.Idoate, Inmaculada Trigo, Jesus Saenz de, Zaitiguib Manuel, Pérez-Péreza, Juan Joseé Riosa

Abstract

Hybrid oncocytic/chromophobe tumor (HOCT) of the kidney arising from a precursor oncocytosis not associated with the Birt-Hogg-Dubé (BHD) syndrome is unusual and highly interesting neoplasm. Immunohistochemical and molecular findings suggest that HOCT(Hybrid oncocytic/chromophobe tumor) is an entity distinct from both oncocytoma and chromophobe carcinoma. Although uncertainty persists regarding the factors predisposing to the development of HOCT(Hybrid oncocytic/chromophobe tumor), experimental findings suggest that it may arise due to the effect of toxins or in association with chronic kidney failure. The potential role of prior renal lymphoma in the development of oncocytosis has not hitherto been examined. We present a morphological, immunohistochemical, and molecular analysis of a HOCT(Hybrid oncocytic/chromophobe tumor) arising from renal oncocytosis in conjunction with CLL affecting the kidney. The findings suggest that this tumor belongs to a family of similar neoplasms including oncocytoma, the eosinophilic variant of chromophobe renal cell carcinoma (CRCC), and low-grade oncocytic tumor, even though these neoplasms may arise from different precursor lesions. HOCT(Hybrid oncocytic/chromophobe tumor) and oncocytosis revealed the same immunohistochemical profile consistent on positivity for epithelial membrane antigen (EMA), cytokeratin 7 (Ck7), E-cadherin, CAM 5.2, and negativity for Pax-8, vimentin, renal cell carcinoma (RCC) antigen, CD117, racemase, progesterone receptor, and CD10. The Ki-67 proliferation index was <1%. Molecular analysis of the tumor revealed the AKT3 gene mutation variant, classified as probably pathogenic, together with FOS1 gene amplification and no copy number variations (CNVs). Finally, we present a case of HOCT(Hybrid oncocytic/chromophobe tumor) arising from a nonhereditary renal oncocytosis in conjunction with B lymphoma that raises interesting questions regarding pathogenesis.

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Introduction

Hybrid oncocytic/chromophobe tumor (HOCT) is a rare renal neoplasm currently classified as a variant of chromophobe renal cell carcinoma (CRCC) [1]. In morphological terms, HOCT(Hybrid oncocytic/chromophobe tumor) strongly resembles oncocytoma, from which it differs essentially in the presence of perinuclear halos and frequent binucleation [2, 3]. Though the sporadic form is the most common, HOCT(Hybrid oncocytic/chromophobe tumor) has also been reported in a hereditary setting, in association with Birt-Hogg-Dubé (BHD) syndrome [4] Sporadic HOCT(Hybrid oncocytic/chromophobe tumor) may arise from renal oncocytosis, that is, the diffuse proliferation in the kidney of oncocytic epithelial cells originating in the renal collecting tubules [5]. The precise causes of renal oncocytosis are yet unknown, although it has been associated both with chemical carcinogens and with chronic kidney failure [6, 7]. We report on an unusual sporadic HOCT(Hybrid oncocytic/chromophobe tumor) associated with oncocytosis in conjunction with a previously diagnosed B cell lymphoma of the kidney. Morphological and molecular findings are discussed, together with the potential role of B-cell lymphoma as a factor predisposing to the development of HOCT(Hybrid oncocytic/chromophobe tumor).

Case Report/Case Presentation

A 72-year-old male was diagnosed with B-cell CLL(Binet Stage A) in 2009 and is currently undergoing standard cyclophosphamide fludarabine rituximab chemotherapy, with continued clinical follow-up.MRI follow-up showed no changes in the abdominal CT in 2012. In a CT scan of the left kidney, in 2014, a small image appeared in the cortex of less than 5 mm. In the retrospective review of the CT scan of 2012, there was no visible injury. In 2015, the patient displayed multiple swollen lymph nodes(retroperitoneal, mesenteric, mediastinal, pelvic, hilar, and axillary).In addition, a CT scan revealed a solid cortical nodule 20 mm in the left kidney in diameter, isodense relative to the renal parenchyma, and showing low contrast uptake. A further CT scan in February 2019 disclosed the solid nodule measuring 24 mm. In 2020,2 solid nodules were identified,1 in each renal pole, the higher of 27 mm in diameter.No significant radiological changes were observed in the right kidney. Contrast-enhanced MRI revealed 2 solid masses showing heterogeneous enhancement on the administration of i.v. contrast; the masses, measuring 28 and 25 mm, respectively, were hypointense on T1 weighted and is hypointense on T2 weighted images and showed increased contrast uptake in the portal phase (shown in Fig. 1). Renal function was conserved. Small splenic nodules were also observed, together with numerous swollen retroperitoneal, mesenteric, and pelvic lymph nodes. Patient history included psoriasis and colon carcinoma in 2000. The patient was a nonsmoker with no relevant family history. No skin or lung lesions were apparent. A left radical nephrectomy was performed. No lymph nodes were removed from the surgical specimen.

Fig. 1. Evolution of the left kidney tumoral nodule by CT images (period 2012–2017): In the first image of 2012, there was no visible injury; in 2014, a small image appears in the cortex of less than 5 mm, which changes to 24 mm in 2017.

The nephrectomy specimen comprised a kidney measuring 10 ×6×5cm, together with adjacent pericapsular fat and an a2-cm segment of the ureter. There were no grossly apparent changes on the kidney surface. Examination of serial sections revealed 3 separate well-circumscribed nodular lesions of elastic consistency, light brown, and of maximum width 3.5,1.5, and 0.3 cm; these nodules did not extend to the capsule or renal sinus(shown in Fig.2).

Fig. 2. a. Macroscopy of highest nodular tumor expansive and well-circumscribed light brown. b. A large expansive, well-circumscribed, and oncocytic tumor nodule partially encapsulated is observed. H and E, panoramic. c Tumor is separated from the rest of the parenchyma by a fibrous tract. The dense lymphoma is observed around and inside the tumor. H&E. ×100.

Histopathological examination disclosed 3 tumors of similar morphological appearance, together with multiple nodular lesions not discernible at the gross examination, and numerous nests of cells sharing similar features. These proliferating oncocyte-like cells were surrounded by an atypical lymphoid infiltrate (shown in Fig. 3). The 3 large tumor-like nodules were expansive, well-circumscribed but not encapsulated, and bore a strong histological resemblance to oncocytomas. Nodules were composed of cells with eosinophilic granular cytoplasm and rounded nuclei with finely dispersed chromatin, inconspicuous nucleoli, and clearer-nuclear halos. Irregular"resinoid" nuclei characteristic of CRCC were not detected. Binucleation was common. There was no evidence of atypia, necrosis, or mitotic activity, nor any invasion of blood or lymph vessels or sarcomatoid transformation. Despite the multifocal partial transformation of the renal tubular epithelium into oncocytic epithelium with a metastatic appearance, the general architecture of renal tubules was conserved. Strikingly, oncocytic nests and incipient oncocytic lesions were embedded in an atypical lymphoid infiltrate. Other findings included cortical cysts, some lined by a single layer of oncocytic cells, several small papillary adenomas displayingcalcifications, and occasional cortical fibromas.

Fig. 3. Histological view of the 1 oncocytic nodule. The dense aggregate of oncocytic cells with more or less evident luces is seen. The dense atypical infiltrate is observed inside the nodule. No atypia, necrosis, or mitotic activity is observed (a), H, and E. ×100. Tumoral nodules were composed of cells with eosinophilic granular cytoplasm and rounded nuclei with finely dispersed chromatin, inconspicuous nucleoli, and clear perinuclear halos. Irregular “raisaid” nuclei are not observed. Binucleation was common. (b). H&E. ×200. Multifocal infiltrative lymphoma with the associated oncocytic transformation of renal tubular epithelium with preservation of the general architecture of adjacent renal parenchyma (c), H&E. ×100. Incipient oncocytic lesions were embedded in an atypical lymphoid infiltrate (d), H&E. ×200.

Immunohistochemical analysis of tumors and oncocytic epithelial proliferations revealed positive staining for epithelial membrane antigen (EMA), with an apically reinforced cytoplasmic expression pattern. Staining for cytokeratin 7(Ck7) was positive in some areas and negative in others. In positive staining areas, Ck7 expression was not homogeneous; negative cells were intermingled with positive cells displaying varying cytoplasmic staining intensity. Expression of E-cadherin and CAM 5.2 was also observed (shown in Fig.4). The Ki-67 proliferation index was<1%. Tumor cells were negative for Pax-8, vimentin, renal cell carcinoma (RCC) antigen, CD117, racemase, progesterone receptors, and CD10.

Fig. 4. Immunohistochemical analysis revealed positive staining for EMA, with an apically reinforced cytoplasmic expression pattern. Staining for Ck7 in positive-staining areas was not homogeneous; negative cells were intermingled with positive cells displaying varying cytoplasmic staining intensity. The expression of E-cadherin and CAM 5.2 was both intense and diffuse. EMA, epithelial membrane antigen; Ck7, cytokeratin 7.

Atypical lymphoid infiltrate was identified throughout the renal parenchyma in a multifocal pattern, extending beyond the capsule into perirenal adipose tissue. Infiltrate was composed of relatively small monomorphic atypical lymphoid cells with occasional blasts forming discrete proliferative clusters. Tumor lymphoid cells stained positive for BCL-2, CD20,CD5, and CD79alpha; and negative force-6, CD10,MUM-1,and cyclin D1.Ki-67 expression in proliferative clusters was focally 60%(shown in Fig.5). The immunohistochemical profile of the lymphoid infiltrates matched that observed in a previously mphnode biopsy. The diagnosis was kidney involvement of a B cell chronic lymphoid leukemia/small cell lymphocytic lymphoma.

Fig. 5. Tumor lymphoid cells corresponding to a B-CLL/B lymphoma well-differentiated inside and outside the epithelial tumoral nodule stained positive for CD20, ×100.

Molecular analysis based on high throughput sequencing revealed the variant c.973C>T(p.Arg325Ter)at exon 10 of the AKT3 gene, classified as probably pathogenic, together with FOS1 gene amplification. No copy number variations(CNVs) were observed, except for again on chromosome 12 (shown in Fig.6). The final diagnosis was multiple HOCT(Hybrid oncocytic/chromophobe tumor) of the kidney associated with oncocytosis (impala, Nx) in conjunction with renal involvement by a chronic lymphoid leukemia/well-differentiated B cell lymphoma.

Fig. 6. No CNVs were observed, except for again on chromosome 12 in the molecular analysis. CNVs, copy number variations.

Discussion/Conclusion

HOCT(Hybrid oncocytic/chromophobe tumor) has been reported in3 clinical settings: as a sporadic tumor; in association with renal oncocytosis; and as a component of BHDsyndrome, an autosomal dominant inherited condition caused by mutations in the gene coding for folliculin(FLCN) and characterized by the lung and skin lesions(fibrofolliculomas). Sporadic oncocytoma disassociated with HOCT(Hybrid oncocytic/chromophobe tumor) and its BHD-related counterpart share certain common features: both arise from a precursor lesion termed renal oncocytosis, and in both cases, neoplasms tend to be multifocal and bilateral [4]. Sporadic and hereditary HOCTs(Hybrid oncocytic/chromophobe tumor) are invariably benign[1, 4, 5]. Renal oncocytosis often affects both kidneys and is generally asymptomatic. HOCT(Hybrid oncocytic/chromophobe tumor) arising from renal oncocytosis is a rare neoplasm mostly reported in adults non-BHD oncocytosis usually affects elderly patients and gender distribution is almost equal.

Renal oncocytosis has been defined as diffuse involvement of the kidney by a gradual proliferation of oncocytic epithelial cells originating in the collecting ducts, leading to the development of oncocytic tumors. The process thus spans a lesional spectrum ranging from initial oncocytic changes in renal tubules with scarcely any alteration of nephron architecture, through the presence of hyperplasticlike nests of oncocytic cells to the formation of dominant nodules of neoplastic appearance [8]. Lesions may be innumerable and affect the entire renal parenchyma. Tumors arising in this context are mostly HOCTs(Hybrid oncocytic/chromophobe tumor), although oncocytomas and CRCCs are also reported in this setting. Several authors have even suggested that oncocytomas may progress to CRCs [9]. It would seem more likely, as Tickoo et al. [5]have noted that HOCTs(Hybrid oncocytic/chromophobe tumor)-like oncocytomas and CRCCs arise through the progressive transformation of oncocytic nests, as was apparent in this case. It is thus unsurprising that oncocytic tumors should present with another overlapping spectrum of lesions with varying morphologies.

The kidney affected by oncocytosis also tends to display other, non-oncocytic, lesions such as the papillary adenomas observed here although, as other authors have reported, these are probably chance findings. One interesting feature of the present case was the appearance of cortical cysts partially lined by an oncocytic epithelium.

hosts(Hybrid oncocytic/chromophobe tumor) are currently classified as a subset of tumors with overlapping histology between renal oncocytoma (RO)and chromophobe RCC[1]. Yet, the morphology of HOCT(Hybrid oncocytic/chromophobe tumor) closely resembles that of oncocytoma, from which it differs only in minor aspects such as the appearance of perinuclear halos. Cytological differences concerning CRCC are more marked. One proof of the singular status of HOCT(Hybrid oncocytic/chromophobe tumor) is the joint presence of Ck7 positive and Ck7 negative cells.Ck7 expression tends to be negative in RO and positive in CRCC. Here, positive staining areas alternated with negative areas; even within Ck7 positive areas, staining was by no means homogeneous, since negative cells intermingled with positive cells displaying varying cytoplasmic staining intensity.

However, this intermediate expression was not recorded for other markers(shown in Table 1): immunostaining for cKit and Pax8 was negative for HOCT(Hybrid oncocytic/chromophobe tumor) but is positive in both CRCCand RO[10-12], while for the other markers tested, HOCT(Hybrid oncocytic/chromophobe tumor) displayed staining similar to that of the other tumor types [13]. The expression of certain markers provides pointers to the histogenetic origin of these oncocytic tumors. Interestingly, E-cadherin and racemase expression would suggest that tumor cells in HOCT(Hybrid oncocytic/chromophobe tumor), RO, and CRCCshare a common origin in the renal collecting ducts. Indeed, positive staining in renal leucocytosis would support the view that RO also originates in the collecting ducts[14-18].In the present study, positive staining for Ck7 and EMA indicated a combination of the expression profile of the principal cells in the collecting ducts with that of the intercalated cells. Expression of Ck7, with apical membranous staining for EMA but no cytoplasmic staining, would appear to be characteristic of collecting duct principal cells, whereas negative staining for Ck7 and intense positive cytoplasmic staining for EMA is characteristic of intercalated cells [19]. No previously published reports have highlighted this singular apically enhanced cytoplasmic staining pattern for EMA in HOCTs, which may well be a distinctive feature enabling the hybrid tumor to be differentiated from both RO and chromophobe RCC.

Since HOCTs(Hybrid oncocytic/chromophobe tumor) arise in 3 different clinical settings, molecular changes might be expected to differ in each case, even though morphological features may be similar. HOCT(Hybrid oncocytic/chromophobe tumor) associated with BHD syndrome [20] is characterized by germline mutations in the FLCN gene which are not observed in HOCTs (Hybrid oncocytic/chromophobe tumor) arising in the other settings. Similarly, this mutation is not found in sporadic oncocytomas or CRCCs, whether or not associated with oncocytosis, but has been reported in those arising within the context of BHD syndrome [20].

A further interesting consideration is that mitochondrial DNA mutations are common in oncocytoma, in CRCC[21], and focal oncocytosis; they are likely to be present in HOCT(Hybrid oncocytic/chromophobe tumor), too, although this has yet to be determined. Mitochondrial DNA mutations may account for mitochondrial proliferation as a compensation mechanism in all 3 tumor types. No studies have yet compared cytogenetic changes in the 2 groups of sporadic HOCTs(Hybrid oncocytic/chromophobe tumor). In those not associated with oncocytosis, cytogenetic anomalies such as monosomies and polysomies have been observed, particularly in chromosomes 1,14,20-1 of the most widely affected 21, and Y[3,12]; by contrast, little research has focused on cytogenetic changes in HOCTs(Hybrid oncocytic/chromophobe tumor) associated with renal oncocytosis [22]. The gain on chromosome 12 observed here has not been reported previously.

The molecular expression profile of sporadic HOCT(Hybrid oncocytic/chromophobe tumor) differs from that of both oncocytoma and CRCC [23]. Though closer to that of oncocytoma in terms of CNVs, low mutational load, absence of mutations in driver genes, and cytogenetic aberrations[23,24], its molecular expression profile is different. HOCT(Hybrid oncocytic/chromophobe tumor) also differs considerably from CRCC not only in molecular expression but also in the molecular and chromosomal changes indicated earlier[23,25]. In molecular terms, therefore, HOCT(Hybrid oncocytic/chromophobe tumor) may be regarded as closer to oncocytoma than to CRCC [3,12,24,26].

The AKT3 gene mutation observed here is known to induce cell growth and proliferation and enhance apoptosis resistance. Totheauthors'knowledge, this is the first report of the mutation in HOCT(Hybrid oncocytic/chromophobe tumor) of the kidney. Interestingly, the PTEN, TERT, and TP53 gene mutations typical of CRCC and the ERCC2 mutation characteristic of RO [23] were not observed in the present case. The absence of CNVs and the low mutational load indicate a genetic profile closer to that of RO.

About the pathogenesis of HOCT(Hybrid oncocytic/chromophobe tumor), renal oncocytosis is the precursor lesion in a large number of cases [26]. Oncocytosis in patients with BHD syn-drome may be attributed to germline mutations in the FLCN gene [4] encoding FLCN protein, which is expressed in the distal tubule and collecting ducts. It has been argued that FLCN acts as a tumor suppressor gene and that its mutation may induce expression of the potent angiogenic trigger hypoxia-inducible factor 1-alpha, thus favoring tumorigenesis. However, the FLCN gene mutation is not observed in renal oncocytosis arising in a nonhereditary context. Although the cause of non-BHD oncocytosis is unknown, oncocytomas have been experimentally induced in rats treated with certain chemical carcinogens, including N-nitrosomorpholine [6]. In a clinical setting, oncocytosis has been reported in patients with chronic kidney failure and those undergoing dialysis [7,27]. The patient under study here had no history either of contact with carcinogens or of chronic kidney failure.

A striking feature of the case presented here was the concurrent presentation of oncocytosis and infiltration of the renal parenchyma by B-cell lymphoma. Whether the lymphoma may have triggered the development of oncocytosis remains a matter of speculation. Reports of histopathological findings in lymphoma-affected kidneys make no mention of oncocytosis, although sample sizes were very small. In 2 clinical series comprising a total of 54 patients with lymphoma B diagnosed by percutaneous kidney biopsy, 18 of whom had CLL/well-differentiated B-lymphocyte lymphoma, no evidence was reported of oncocytic changes inrenaltubules[28,29].In an additional case report of a RO associated with oncocytosis in a patient previously diagnosed with B-cell small lymphocytic lymphoma, no atypical lymphoid infiltration was observed in the nephrectomy surgical specimen [30]. Even so,2 findings in the present case point to the possible contribution of lymphoma to oncocytosis. First, a close topographical relationship was observed between initial oncocytic lesions and atypical lymphoid infiltrate, although it was not possible to determine whether this was a causal relationship or simply reflected a particular tropism of atypical lymphoid cells for previously developing areas of oncocytosis. Second, radiological evidence shows that the kidney involvement developed progressively between 6 and 10 years after the diagnosis of chronic lymphoid leukemia. However, it is impossible to determine whether oncocytosis developed naturally and independently of renal lymphoma.

The differential diagnosis of HOCT(Hybrid oncocytic/chromophobe tumor) should include other eosinophilic kidney tumors, particularly the eosinophilic variant of clear cell carcinoma, angiomyolipoma, the eosinophilic variant of CRCC[25], the oncocytic variant of CRCC[22], and low-grade oncocytic tumor [31]. The first 2 differ considerably from HOCT(Hybrid oncocytic/chromophobe tumor) in terms of morphology and immunohistochemical features, and should thus pose no particular diagnostic challenge. In most cases, HOCT(Hybrid oncocytic/chromophobe tumor) displays slight morphological, immunophenotypic, and molecular differences concerning eosinophilic CRCC [32], itself a heterogeneous variant not associated with any single morphological pattern[23,26]. The main differences are their regular nuclei and intense cKit and Ck7 expression observed in eosinophilic CRCC, although these findings would not alone justify their classification as distinct entities. The oncocytic variant of CRCC shares certain morphological and immunophenotypic features with HOCT(Hybrid oncocytic/chromophobe tumor). To add to the confusion, Trpkov et al. (2019)[31] recently reported on a "low-grade oncocytic tumor of the kidney, "not associated with either the BHDsyndrome, which displayed morphological and immunohistochemical features very similar to those of HOCT(Hybrid oncocytic/chromophobe tumor), particularly in terms of intense diffuse positive staining for Ck7. The eosinophilic variant of CRCC, this low-grade oncocytic tumor of the kidney, and HOCT(Hybrid oncocytic/chromophobe tumor) would appear to be, in essence, the same entity, forming part of a spectrum of tumors probably arising from a common precursor lesion and with overlapping histological characteristics.

To conclude, this article reports on a hybrid tumor arising from a nonhereditary renal oncocytosis in conjunction with Blymphoma that raises interesting questions regarding pathogenesis. This tumor bore a clear morphological resemblance to the eosinophilic and oncocytic variants of CRCC and a recently reported low-grade oncocytic tumor of the kidney, suggesting that all these tumors may form part of a spectrum belonging to the same family of neoplasms, deriving from renal oncocytic epithelial proliferation arising in different clinical contexts.

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