Superficial CD34⁺ fibroblastic tumor with focal atypical presentation: A case report

Introduction

Superficial CD34-positive fibroblastic tumor (SCPFT) is a rare mesenchymal tumor of intermediate malignancy, with reported cases representing a wide range of ages from pediatrics to the elderly, although the highest incidence is among middle-aged people. The preoperative duration typically exceeds 1 year. SCPFT typically manifests as a long-standing, painless mass in the subcutaneous tissue, commonly occurring in the lower limbs, occasionally in the upper limbs and back, and rarely in the neck, chest, axilla, abdominal wall and breast (1). SCPFT was first reported by Carter et al (2) in 2014 and then described as a new entity in the 5th edition of the World Health Organization (WHO) classification of soft tissue tumors (3). Clinically, SCPFT presents as a slow-growing, painless, circumscribed mass that mainly occurs in adults, and it generally involves the deep dermis and subcutaneous tissues, showing a predilection for the lower extremities. Grossly, SCPFT sections are firm and yellow-to-tan in color. Microscopically, SCPFTs are composed of spindled to epithelioid cells arranged in a fascicular pattern. The tumor cells show apparent cellular heterogeneity and prominent nucleoli with apparent CD34 positivity (4).

Surgical resection is the optimal treatment for most patients, resulting in a good long-term prognosis. However, patients with positive surgical margins may experience local recurrence, while regional lymph node metastasis is uncommon (1,5). However, more cases are needed for long-term prognostic data to establish reliable indicators. The current report presents a case of a typical SCPFT with morphological, immunophenotypic and molecular characteristics, revealing internal morphological heterogeneity.

Case report

A 43-year-old male patient was admitted to the Shandong Provincial Qianfoshan Hospital (Jinan, China) in October 2022, following the discovery of a tumor in the left perineum. The lesion, initially presenting as a mildly tender mass at the thigh roots near the scrotum 6 months prior, exhibited slow growth without signs of irritation or inflammation. Upon physical examination, a contained solid mass measuring 5.5×4×3 cm was observed. Computed tomography revealed a hypoechoic enclosed mass, measuring 5×3.5×3 cm, at the base of the thigh (Fig. 1A). There were no abnormalities present in the hematological laboratory tests, including peripheral blood cell analysis, coagulation index and blood infection markers. The tumor was surgically excised under general anesthesia, involving complete capsule removal, and the excised tumor exhibited a white-colored, cartilage-like consistency on the cut surface (Fig. 1B).

Figure 1. (A) Gross appearance of the tumor specimen presenting with a grayish-white color in section and a tough texture. (B) Computed tomography image demonstrating a hypoechoic, well-demarcated mass situated at the base of the thigh. (C) Tumor cells situated beneath the dermis and exhibiting local infiltration into the dermis (HE, ×100 magnification). (D) Tumor cells arranged in bundles, accompanied by eosinophils and lymphocytes in the background (HE, ×200 magnification). (E) Tumor cells with distinct nucleoli, with visible intranuclear inclusion bodies (HE, ×400 magnification). (F) Tumor margins featuring cell-rich areas, as depicted in the inset (HE, ×400 magnification), showing tumor cells with abundant cytoplasm, atypical nuclei and a small size (HE, ×100 magnification). HE, hematoxylin and eosin.

The tumor was cut into flat tissue blocks ~2×1.5 cm in size and 0.3 cm in thickness. These tumor blocks were then fixed in a solution containing 10% formaldehyde in 0.01 M phosphate-buffered saline (PBS) for 2 h at room temperature. Following fixation, the tissue blocks were loaded into the Tissue-Tek VIP®6 AI Tissue Processor (Sakura Finetek USA, Inc.) and subsequently embedded in paraffin. Formalin-fixed paraffin-embedded sections were cut using a Leica RM 2155 Rotary Microtome (Leica Microsystems). The subsequent dewaxing process included sequential treatments with xylene, anhydrous ethanol, a decreasing concentration gradient of ethanol (95, 90, 80 and 70%), and water. Following this, the sections were immersed in Harris hematoxylin staining solution for 5 min and then differentiated with 0.3% acid alcohol, before being incubated with 0.6% ammonia. Eosin staining solution was applied for 1–3 min, followed by dehydration with ethanol and xylene. Finally, the samples were mounted with neutral gum to prepare the slides. Postoperative histopathological microscopy revealed that the lesion was confined within the subcutaneous tissue, and at low magnification, the dermis of the skin showed infiltration by tumor cells (Fig. 1C). The tumor displayed sparse-to-moderate cellularity, featuring a densely cellular, basophilic, nodular area at the periphery. Under high magnification, the tumor primarily consisted of spindle-shaped to epithelioid cells, arranged singly or in clusters within a collagenous stroma. The tumor cells exhibited marked nuclear pleomorphism with prominent nucleoli, abundant eosinophilic cytoplasm and occasional multilobed nuclei or intranuclear exudate inclusion bodies. Scattered lymphocytes and eosinophils were observed in the interstitium (Fig. 1D and E). Within the densely populated nodule, the tumor cells displayed the same morphological characteristics but were relatively smaller in size and more tightly arranged compared with other regions (Fig. 1F).

Paraffin sections of 5-µm were loaded into The Discovery ULTRA (Roche Tissue Diagnostics) for automated staining. The automated immunohistochemistry staining process utilized solutions and antibodies from Roche Tissue Diagnostics unless otherwise specified. Initially, slides underwent de-paraffinization with the EZ PREP solution (cat. no. 950-100), followed by antigen retrieval through Heat Induced Epitope Retrieval in Tris-EDTA buffer (cat. no. 950-124) at pH 7.8 and 95°C for 40 min. Subsequently, primary antibodies were manually applied and incubated at 37°C for 60 min. The targets were then linked using the OmniMap anti-rabbit (cat. no. 760-4311) and OmniMap anti-mouse (cat. no. 760-4310) HRP-conjugated secondary antibodies. Visualization of the different targets was achieved through DAB and H2O2 (cat. no. 760-159). Finally, tissue slides were counterstained with hematoxylin and mounted using a xylene-based mounting medium. The following ready-to-use primary antibodies were utilized: Cytokeratin [anion exchanger (AE)1/AE3; cat. no. Kit-0009], epithelial membrane antigen (EMA; cat. no. Kit-0011), ERG (cat. no. RMA-0748), STAT6 (cat. no. RMA-0845), CD34 (cat. no. MAB-1076), smooth muscle actin (SMA; cat. no. MAB-0890), Desmin (cat. no. MAB-0766), S100 protein (cat. no. MAB-0697), CD10 (cat. no. MAB-0668), CD117 (cat. no. Kit-0029), discovered on GIST-1 (DOG1; cat. no. MAB-0851), integrase interactor 1 (INI1)/SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily B member 1 (SMARCB1) (cat. no. MAB-0696), anaplastic lymphoma kinase (ALK; cat. no. MAB-0848) and Ki-67 (cat. no. MAB-0672). All antibodies were purchased from MaximBiotech, Inc. The tumor cells exhibited strong positive staining for CD34 in a diffuse pattern (Fig. 2A). Nuclear expression of INI1/SMARCB1 was evident in all cases (Fig. 2B). The Ki-67 labeling index was ~5% in most areas and ~20% in the focal cell-dense regions (Fig. 2C and D). Furthermore, the tumor cells displayed negative staining for AE1/AE3, CD117, STAT6, DOG1, SMA, ALK, Desmin, CD10, ERG, EMA and S-100 expression (Figs. S1 and S2).

Figure 2. (A) Immunohistochemical staining demonstrating tumor CD34 diffuse cytoplasmic positivity (×200 magnification). (B) Positive nuclear staining for integrase interactor 1 observed via immunohistochemical analysis (×200 magnification). (C) Cell-rich region displaying high Ki-67 expression (×200 magnification). (D) Tumor cells outside the cell-rich nodules with very low expression of Ki-67 (×200 magnification). (E) Dual-color break-apart probe analysis indicating a rearrangement in the PR/SET domain 10 gene (×1,000 magnification) and (F) transforming growth factor β receptor 3/MGEA5 dual-color fusion probe showing MGEA5 is negative for gene breaks (×1,000 magnification).

For fluorescence in situ hybridization (FISH), a commercial TGFBR3-MGEA5 Dual Fusion/Translocation FISH probe (cat. no. F.01228-01) and PRDM10 Break Apart FISH probe (cat. no. F.01377-01) were obtained from Guangzhou Anbiping Medical Laboratory Co., Ltd. The PRDM10 probe utilized two custom-labeled FISH DNA probes to label the PRDM10 gene flanks on chromosome 11. Specifically, the BAC clone D11S1083 (366 kb), positioned centromeric to PRDM10, was labeled with FITC dUTP, while the BAC clone RH104271 (189 kb), located telomeric to PRDM10, was labeled with 5(6)-TRITC dUTP. In the case of the TGFBR3-MGEA5 Dual Fusion/Translocation FISH Probe, the BAC clone BV444963-STSG24458 (872 kb) was labeled with FITC dUTP to identify the TGFBR3 gene, and the BAC clone SHGC153008-D10S2456 (765 kb) was labeled with 5(6)-TRITC dUTP for identifying the MGEAS gene.

Paraffin sections of 4 µm were dewaxed and hydrated. Chromosomal DNA was denatured on the slides in a 70% formamide-2XSSC solution at 68–70°C for 2 min. Subsequently, the slides were dehydrated and air-dried. The hybridization mixture, containing DNA probe at a concentration of 20–50 µg/ml, was added to the slide and covered with a cover slip. Subsequently, the slides were incubated in a moist plastic chamber at 37°C for 6–12 h. After incubation, the slides were washed and dried before being immersed in blocking buffer (1X PBS, 0.1% Triton-100) for 2 min. Following this, the slides were rinsed in PBS for 5 min at room temperature. The semi-dried slides were then treated with 100 µl of 1:100 rabbit antibiotin antibody and incubated in a humidity chamber at 37°C for 5 min. Subsequently, the slides were washed with PBS and immersed in 100 µl diluted antibody (FITC-conjugated goat anti-rabbit antibody at a ratio of 1:100 in dilution buffer). Slides were then incubated in the humidity chamber at 37°C for 30–60 min. After incubation, the slides were washed again, and 60 µl of an antifade solution (composed of p-phenylenediamine at a concentration of 10 mg/ml, 90% glycerol and propidium iodide at 1 µg/ml as a counterstain) was added to each slide. The slides were observed under fluorescence microscope (BM4000B, Leica Microsystems GmbH).

FISH analysis revealed a positive rearrangement of the PRDM10 gene (Fig. 2E), whilst TGFBR3/MGEA5 showed negative results (Fig. 2F). These findings aligned with a diagnosis of SCPFT. Post-surgery, the patient exhibited a rapid recovery. There is no clear academic consensus on the optimal follow-up time after SCPFT. The patient will be monitored every 6 months during the first postoperative year and annually thereafter. So far, the patient has not had recurrence or metastasis.

Discussion

SCPFT was initially identified and characterized in 2014 by Carter et al (2) in an assessment of 18 cases. The tumors were observed in the superficial subcutaneous layer, displaying notable pleomorphic cells, a low mitotic index, absence of necrosis and widespread strong positivity for CD34 in immunohistochemical staining. Carter et al designated this distinctive entity as SCPFT. In the 2020 revision of the WHO Classification of Soft Tissue and Bone Tumors, SCPFT was acknowledged as a novel tumor (3). SCPFT has garnered growing attention from pathologists, resulting in a rise in the number of published cases. Presently, ~150 cases of SCPFTs have been documented in the English-language literature (5–15).

Clinically, SCPFT predominantly affects young adults spanning an age range of 8–85 years, with a slight prevalence in males (5,7). The primary sites of SCPFT occurrence are commonly the lower limbs, particularly the thighs, along with other locations such as the calves, groin, foot, Achilles tendon, shoulder, vulva, neck, knee, buttocks and arms. The majority of cases are situated in the deep dermis to the superficial subcutis. Grossly, SCPFTs vary in size from 1.2-10 cm, typically exhibiting a circumscribed border but occasionally infiltrating surrounding soft tissues (16). Microscopically, SCPFTs consist of spindled to epithelioid cells arranged in fascicles or storiform formations. The cells exhibit prominent eosinophilic cytoplasm, notable nuclear pleomorphism with prominent macronucleoli and occasional pseudoinclusions. The interstitium displays branching capillaries and scattered lymphocytes, mast cells and foam cells. Mitotic figures are exceedingly rare (<1/50 high-power fields). Immunohistochemically, SCPFT cells consistently exhibit diffuse and strong positivity for CD34. Recently, synaptic cell adhesion molecule 3 has been identified as a sensitive marker expressed in 95% (56/59 cases) of SCPFT (17). The Ki-67 index is generally low in SCPFT, typically <5% (7).

In 2015, Hofvander et al identified recurrent PRDM10 gene fusions for the first time in 3/28 cases of undifferentiated pleomorphic sarcoma (UPS) with a morphological resemblance to SCPFT (18). It was hypothesized that these cases represented a subset of low-grade UPS. Subsequent studies have assessed the relationship between SCPFT and PRDM10-STT, highlighting their overlapping features and leaning toward considering them as part of the same spectrum (5,17,19). Due to the presence of marked cellular pleomorphism and nuclear atypia, an accurate diagnosis of SCPFT necessitates the exclusion of superficially located and high-grade sarcomas with morphological similarities (19).

The principal differential diagnoses include atypical fibrous histiocytoma (AFH), pleomorphic hyalinizing angiectatic tumor (PHAT), atypical fibroxanthoma (AFX), epithelioid sarcoma (EpS), UPS, pleomorphic dermal sarcoma (PDS) and myxoinflammatory fibroblastic sarcoma (MIFS). AFH typically exhibits a classic fibrous histiocytoma background interspersed with a population of atypical cells displaying nuclear hyperchromasia and prominent nucleoli, often with abundant cytoplasm. Dermal collagen at the tumor margins and pigmentation of the basal layer aid distinguishes AFH from SCPFT. Although AFH tumor cells may express CD34, it is usually focal and weak, and they generally do not express cytokeratins (20,21). PHAT shares many common features with SCPFT, including a superficial location, marked pleomorphism of tumor cells, inconspicuous mitotic activity, pseudo-inclusions in the nucleus and diffuse CD34+ staining. However, PHAT has distinctive features, such as thick-walled ectopic vessels with marked perivascular hyalinization (22). AFX typically occurs on sun-exposed skin in elderly patients, featuring highly pleomorphic cells with hyperchromatic nuclei, atypical mitoses and abundant cytoplasm, commonly arranged in a spindly architecture. Multinucleated giant cells and solar elastosis are often present in AFX specimens. AFX stains positive for several markers, including CD10 and p53, but is negative or focally positive for CD34 (23). EpS shares certain immunophenotypic similarities with SCPFT, exhibiting diffuse or patchy expression of cytokeratin and CD34 in ~50% of cases. However, EpS is characterized by epithelioid to spindled cells with central pseudogranulomatous architecture in the classic type, and predominant epithelioid and rhabdoid cells in the proximal type, distinguishing it from SCPFT. Loss of nuclear expression of SMARCB1 is a hallmark feature of EpS, present in the vast majority of cases (24). UPS typically occurs in middle-aged and older patients, originating in deeper tissues and morphologically showing marked pleomorphism, heterogeneity and aberrant mitosis. UPS may express CD34 focally, but not diffusely, in contrast to SCPFTs (25). PDS may infiltrate the subcutis, potentially causing confusion with SCPFT. PDS exhibits true pleomorphism and heterogeneity, with common appearances of mitotic figures and necrosis, similar to AFX. PDS expresses CD10 but not CD34 diffusely (26). Finally, MIFS typically exhibits a multinodular growth pattern with a variable combination of three morphologic zones: Myxoid, hyalinized and inflammatory. MIFS tumor cells can range from plump spindle cells to histiocytoid or epithelioid cells, featuring enlarged basophilic or eosinophilic macronucleoli reminiscent of virallyinfected nuclei. Variable expression of CD34 is not uncommon in MIFS and can be prominent in certain cases. Certain subset cases may involve molecular genetic alterations, such as MGEA5 or BRAF rearrangements (27).

SCPFT is considered to originate from CD34+ fibroblasts, which are a subset of fibroblasts and are assumed to be mesenchymal stem cells. A series of fibroblastic tumors that exhibit CD34+ activity, including leiomyosarcoma and primary fibrosarcoma, are believed to originate from CD34+ fibroblasts (28). CD34 is a transmembrane phosphoglycoprotein with a molecular weight of ~115 kDa, which was first discovered on hematopoietic stem and progenitor cells (29). CD34 has also been identified as a marker of several types of non-hematopoietic cells, including fibroblast progenitors and endothelial precursors (30). The exact functions of CD34 proteins remain complex and multifaceted. Researchers have proposed potential roles for CD34 in promoting cell proliferation and preventing cell differentiation (31). Thus, in SCPFT, CD34 maintains a high self-renewal capacity of fibroblasts, which means that they can continuously undergo cell division and generate more cells. On the other hand, CD34 is associated with endothelial progenitor cells and angiogenesis (32).

In the present study, a panel of antibodies was used for immunohistochemical staining to assist in the diagnosis and differential diagnosis. Concurrently, molecular analysis indicated the presence of a PRDM10 gene rearrangement, thereby confirming the diagnosis of SCPFT. While numerous SCPFT cases have been documented, it is important to note two key points in this specific case. Firstly, SCPFT usually develops in the thigh; however, in this instance, the tumor was located near the scrotum in the groin, which is a less common site for SCPFT. Secondly, various regions within the tumor exhibited distinct levels of proliferative activity. Notably, the cell-dense nodules displayed higher proliferative activity compared with other regions, as highlighted by an elevated Ki-67 proliferation index. This demonstrates that intra-tumor heterogeneity and the potential of local recurrence exists in SCPFT, indicating that clinicians should adjust their follow-up strategy to increase the frequency of follow-up visits when required.

In conclusion, the present study describes a case of SCPFT located in the groin, which was confirmed by using immunohistochemistry and FISH. Moreover, tumor heterogeneity was identified in this case. Despite an increasing number of reported SCPFT cases, further accumulation of cases is essential to unravel the intrinsic mechanisms and clinical characteristics of SCPFT.

Supplementary Material

Supporting Data

Acknowledgements

Not applicable.

Funding

Funding: No funding was received.

Availability of data and materials

The data generated in the present study may be requested from the corresponding author.

Authors' contributions

JS was the first-visit doctor who designed the study and the main writer of the manuscript. SH was the surgeon who acquired and analyzed the patient's clinical data. XY is a diagnostic pathologist who performed the pathological assessment of the patient and participated in the writing of the manuscript. All authors have read and approved the final version of the manuscript. JS and SH confirm the authenticity of all the raw data

Ethics approval and consent to participate

The present study was approved by the Medical Ethics Committee of the First Affiliated Hospital of Shandong First Medical University (Jinan, China; approval no. 2023-S434).

Patient consent for publication

Written informed consent was obtained from the patient for publication of this case report and any accompanying images.

Competing interests

The authors declare that they have no competing interests.

References