Clear Cell Chondrosarcoma With Somatic VHL Inactivation: A Case Report With Integrated Genomic and Transcriptomic Analysis

Abstract

Background:

Clear cell chondrosarcoma (CCC) is a rare, low-grade malignant bone tumor and a specific subtype of chondrosarcoma. In contrast to conventional central chondrosarcoma, which often exhibits recurrent IDH1/2 mutations, CCC is IDH-wildtype, and its genomic profile remains uncharacterized. Sporadic cases have been described in association with von Hippel–Lindau (VHL) disease, but somatic VHL alterations have not been demonstrated in this tumor.

Methods:

A 35-year-old man presented with local recurrence of a distal femoral tumor 5 years after curettage and allograft reconstruction for a tumor that was initially diagnosed as clear cell chondrosarcoma. The recurrent tumor was resected and analyzed using a comprehensive DNA/RNA next-generation sequencing assay with matched germline testing to distinguish somatic from germline alterations.

Results:

The recurrent tumor measured 5.0 cm and showed classic CCC morphology with focal areas of conventional chondrosarcoma-like differentiation. No IDH1/2 mutations were identified. Two pathogenic loss-of-function variants were detected: COL2A1 c.1262_1266+2delinsTCC (variant allele frequency [VAF], 34%) and VHL c.341–2A>C (VAF, 45%). The tumor had a low tumor mutational burden, low loss of heterozygosity, and a complex karyotype with monosomy of chromosome 3. Gene expression profiling revealed decreased VHL mRNA expression. Germline testing showed no germline VHL alteration, confirming somatic VHL inactivation.

Conclusion:

To our knowledge, this report is the first to describe a molecularly profiled CCC with somatic VHL inactivation. These results expand the evolving genomic spectrum of CCC beyond its previously known IDH-wildtype status and suggest that VHL pathway dysregulation may contribute to tumorigenesis in a subset of cases.

1 |. Introduction

Clear cell chondrosarcoma (CCC) is a rare distinct subtype of chondrosarcoma that typically arises in adults, most often in the third to fifth decades, and shows a predilection for the epiphyses of long bones, most commonly the femur and humerus [1, 2]. Histologically, it is composed of sheets and lobules of polygonal cells with abundant clear cytoplasm and distinct cell borders, often admixed with woven bone and osteoclast-like giant cells. Foci resembling low-grade conventional chondrosarcoma are commonly present [2].

At the molecular level, CCC is distinct from conventional central chondrosarcoma, which harbors recurrent hotspot mutations in IDH1 and IDH2 in approximately half of cases [3]. In contrast, CCC is consistently IDH-wildtype [4, 5]. Integrative genomic studies of chondrosarcoma have demonstrated genomic heterogeneity and identified potential therapeutic vulnerabilities, but data specific to CCC remain limited, and no recurrent pathognomonic driver alteration has been established [3–7].

Isolated case reports of CCC in the setting of von Hippel–Lindau (VHL) disease have raised the possibility of VHL pathway involvement in tumorigenesis [8, 9]. However, somatic VHL inactivation has not been reported in CCC arising outside the context of VHL disease. Here, we report a case of CCC with biallelic somatic VHL inactivation identified by integrated genomic and transcriptomic profiling.

2 |. Case Report

All procedures were conducted in accordance with the ethical standards of the institutional research committee and with the 1975 Helsinki Declaration and its later amendments. In accordance with institutional and federal guidelines, this work did not require Institutional Review Board oversight. Informed consent for publication was obtained from the patient.

A 35-year-old man was seen for right knee pain and referred for a bone lesion in the medial femoral condyle detected on radiographic imaging. The patient was otherwise asymptomatic and had no significant past medical history or family history of cancer. On physical examination, he had focal tenderness to the anteromedial aspect of the distal femur. An open biopsy and saucerization with bone grafting of the right distal femur were performed. Histologic examination revealed a lobular proliferation of chondrocytes with abundant clear cytoplasm and well-defined cell borders, admixed with woven bone and scattered osteoclast-like multinucleated giant cells; features diagnostic of CCC. Staging studies including computed tomography (CT) scan of the chest and magnetic resonance imaging (MRI) of the brain showed no evidence of metastatic disease. He subsequently underwent partial resection of the right distal femur with allograft reconstruction and internal fixation. Histologic examination of the resection specimen demonstrated a few microscopic residual foci of CCC, the largest being 0.6 cm, with negative surgical margins. He underwent hardware removal within a year and remained clinically disease-free with stable surveillance imaging studies.

Approximately 5 years after the initial diagnosis, the patient returned with a 1 to 2 year history of progressive right knee pain. Imaging studies revealed a new bony abnormality in the region of the medial femoral condyle on radiographs (Figure 1A), and MRI demonstrated a well-demarcated epiphyseal lesion consistent with local recurrence (Figure 1B). Satellite lesions were also identified along the posteromedial femoral condyle, without evidence of extraosseous involvement (Figure 1C). The patient underwent radical resection of the distal femur. On gross examination, a 5.0 × 4.5 × 4.0 cm ill-defined white–gray mass was identified in the medial epiphysis. The mass was partially admixed with bone cement from previous surgery and demonstrated focal hemorrhage and necrosis (Figure 2A). Histologic examination confirmed recurrent CCC (Figure 2B) with areas showing features overlapping with conventional chondrosarcoma (Figure 2C). The tumor was confined to bone and completely excised, with 5.0 cm of clearance from the proximal resection margin.

FIGURE 1 |.

(A) AP radiograph of the knee obtained 5 years after resection and stabilization demonstrates a large, expansile lucent lesion at the prior resection site, with internal areas of hyperattenuation suggestive of chondroid matrix. (B) Coronal T1-weighted image demonstrates a lobulated lesion in the medial femoral condyle, extending laterally into the intercondylar notch, distally to the articular surface, and proximally toward the physeal line, with associated cortical expansion and medial cortical thinning. (C) Axial T2-weighted fat-suppressed images demonstrate a lobulated T2-hyperintense lesion with internal low-signal foci in a ring-and-arc configuration (red arrow), consistent with chondroid matrix. Additional posterior satellite lesions are present (green arrow).

FIGURE 2 |.

(A) Gross photograph (coronal section of the distal femur) showing an ill-defined white–gray mass centered in the medial epiphysis. (B) H&E stain demonstrating sheets of uniform round cells with clear to pale cytoplasm and distinct cell borders, interspersed between trabeculae of woven bone. (C) Foci of hyaline cartilage differentiation resembling low-grade conventional chondrosarcoma.

Based on the findings of locally recurrent disease and a mixed histologic appearance, molecular profiling studies were pursued. Next-generation sequencing of the tumor was performed at a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory (Caris Life Sciences, Phoenix, AZ) using whole exome sequencing and whole transcriptome sequencing, as previously described by Smith et al. [10] Two pathogenic loss-of-function alterations were identified, a VHL splice site mutation (exon 2; c.341–2A>C; variant allelic frequency [VAF], 45%) and a complex COL2A1 splicing mutation (exon 20; c.1262_1266+2delinsTCC; VAF, 34%). In addition, an FH p.K477dup variant was identified. The tumor was microsatellite stable, with low tumor mutational burden (4 mut/Mb) and low genomic loss of heterozygosity (3%). Copy number analysis revealed multiple chromosomal abnormalities, including monosomy of chromosome 3, consistent with loss of the VHL locus. Transcriptomic analysis demonstrated markedly reduced VHL mRNA expression (10th percentile) and markedly elevated VEGFA expression (99th percentile), reported relative to an internal reference cohort. Molecular classification using Caris GPSai [11] failed to assign the tumor to a recognized cancer lineage.

On the basis of the identified VHL alteration, the patient was referred for genetic counseling. Germline testing of peripheral blood was performed using a 76-gene hereditary cancer DNA/RNA panel (Invitae) at a CLIA-certified constitutional genetics laboratory (Labcorp Genetics, San Francisco, California, USA), as described by Lincoln et al. [12] and in the Invitae panel technical specifications. The assay identified an FH c.1431_1433dupAAA (p.Lys477dup) variant, which had also been detected in the tumor analysis and represents a recessive fumarate hydratase deficiency allele not associated with increased cancer risk in the heterozygous state [13]. No germline VHL alteration was detected, supporting biallelic somatic VHL inactivation.

3 |. Discussion

Clear cell chondrosarcoma (CCC) is a diagnostically challenging entity due to morphologic overlap with other bone tumors showing chondroid differentiation, including conventional chondrosarcoma, chondroblastoma, and chondroblastic osteosarcoma. In contrast to chondroblastoma for which H3K36M immunohistochemistry is a useful diagnostic tool [14], there is no specific immunohistochemical or molecular marker for CCC. IDH1/2 mutations are identified in ~50% of conventional central chondrosarcomas, whereas CCC is IDH-wildtype [2, 5]. Due to the lack of a pathognomonic marker, distinction between CCC and conventional chondrosarcoma can be challenging, particularly in small biopsies with clear cells and focal hyaline cartilage. This distinction is clinically important because CCC has clinically meaningful metastatic potential, whereas Grade 1 chondrosarcoma (atypical cartilaginous tumor) only rarely, if ever, metastasizes [2, 15, 16].

Although CCC is not a recognized tumor type associated with VHL syndrome, two cases with clinical or germline evidence of VHL mutation have been reported [8, 9]. Dreijerink et al. described a CCC arising in the ulna of a patient with a germline VHL mutation (c.500G>A; p.Arg167Gln), in which the tumor demonstrated loss of heterozygosity at the VHL locus with monosomy of chromosome 3, consistent with biallelic inactivation [8]. Similarly, Song et al. reported a CCC of the talus in a patient with clinically established VHL syndrome. In that case, immunohistochemistry demonstrated HIF-1α and VEGF expression, suggesting activation of hypoxia-related signaling pathways associated with VHL dysfunction, although somatic mutational analysis was not performed [9]. These reports raise the possibility that VHL pathway dysregulation contributes to CCC pathogenesis in the setting of hereditary VHL syndrome. In contrast, the present tumor arose in a patient without a germline VHL alteration and represents, to our knowledge, the first molecularly confirmed example of biallelic somatic VHL inactivation in sporadic CCC (Table 1).

TABLE 1 |.

Reported cases of clear cell chondrosarcoma associated with VHL pathway alteration.

Feature	Dreijerink et al. [8]	Song et al. [9]	Present case
Age/sex	48-year-old female	32-year-old male	35-year-old male
Clinical context	VHL disease	VHL disease	Sporadic
Germline VHL status	VHL c.500G>A (p.Arg167Gln)	Germline VHL mutation reported, type not specified	Negative
Prior VHL-associated tumors	Hemangioblastoma, retinal hemangioma	Hemangioblastoma, clear cell renal cell carcinoma, pheochromocytoma	None
Tumor location	Distal ulna	Talus	Distal femur
VHL-related evidence in tumor	LOH at VHL locus with chromosome 3 monosomy; reduced VHL expression	HIF-1α and VEGF positive by IHC; no somatic VHL testing reported	Somatic VHL c.341-2A>C splice site mutation; chromosome 3 monosomy; reduced VHL mRNA expression; markedly elevated VEGFA
Proposed mechanism	Germline VHL mutation with second hit somatic inactivation	Possible VHL/HIF pathway activation in hereditary VHL setting	Biallelic somatic VHL inactivation in sporadic CCC
Outcome/follow-up	Not specified	No recurrence at 2 years	Local recurrence at 5 years; radical resection performed

VHL, a tumor suppressor gene on 3p25, encodes a component of an E3 ubiquitin ligase complex that targets HIF-α for degradation, thereby suppressing hypoxia signaling including angiogenesis mediated in part by VEGFA. Inactivation of VHL leads to HIF protein stabilization and activation of hypoxia-responsive genes promoting angiogenesis, metabolic reprogramming, and tumorigenesis [17]. Transcriptomic analysis in our case revealed underexpression of VHL, suggesting functional impairment of the pathway.

In addition to VHL inactivation, this tumor carried a pathogenic COL2A1 alteration. COL2A1, the gene encoding the α1 chain of type II collagen, is one of the most commonly mutated genes in conventional chondrosarcoma, with truncating, splice site, and missense mutations identified in approximately one third of cases [18–20]. These mutations are relatively specific to chondrosarcoma among chondroid and osseous neoplasms and are thought to disrupt the extracellular matrix and chondrocyte differentiation. Focal areas of the recurrent tumor that underwent genomic profiling in this case resembled conventional chondrosarcoma. As suitable tissue from the primary neoplasm was not available for molecular analysis, it remains unclear whether the COL2A1 mutation was an early event or a secondary alteration acquired at recurrence.

Molecular classification using Caris GPSai did not assign the tumor to a recognized cancer lineage, including chondrosarcoma. While this may reflect limited representation of CCC in the training sets for the molecular-based classifiers, it also suggests that CCC may have a distinct transcriptional signature that is not identical to that of conventional chondrosarcoma. These findings support the concept that CCC is not simply a clear cell variant of chondrosarcoma but rather a distinct entity.

In summary, this case provides molecular evidence of somatic VHL pathway inactivation in CCC, expanding the emerging genomic spectrum of this rare tumor beyond its established IDH-wildtype background. The integrated genomic and transcriptomic profiling used in this case illustrates how comprehensive molecular characterization can inform tumor biology and refine classification as additional cases are studied. Although the prevalence of VHL alterations in CCC remains unknown, these findings raise the possibility that VHL pathway dysregulation may contribute to tumorigenesis in a subset of cases, a hypothesis that warrants evaluation in larger, comprehensively profiled cohorts.

Data Availability Statement

The data presented in this report were generated as part of routine CLIA-certified diagnostic testing and are available from the corresponding author in de-identified form upon reasonable request. Access to the underlying raw data is restricted by the testing laboratories and patient privacy regulations and therefore cannot be shared by the authors.