Introduction

Anaplastic thyroid carcinoma(ATC) is an aggressive thyroid malignancy, accounting for 1-4% of thyroid carcinomas.(1) It typically affects older individuals in their eighth decade, with a higher incidence in females.(1) The clinical presentation often includes acute onset stridor and local pain, with patients exhibiting regional lymph node metastasis and recurrent laryngeal nerve involvement. ATC is known for its propensity for distant metastasis, particularly to the lungs and brain. Few patients have a history of long-standing goiter and an association with thyrotoxicosis.(2) The median survival of these patients is extremely bleak, ranging from 1-6 months.(1)

ATC originates from undifferentiated cells of thyroid follicles, resulting in aggressive and rapidly progressive tumors. Prior literature review suggests ATC arises from pre-existing differentiated thyroid carcinoma(DTC), constituting up to 23-55% of cases.(1) ATC has various morphological patterns, making it challenging to differentiate from metastatic tumors. The morphological spectrum includes spindle cell/sarcomatous, epithelioid, squamous, giant cell, pleomorphic and mixed patterns.(2) The molecular pathogenesis includes mutations in BRAF, RAS, CTNNB1, PIK3CA, and TP53 genes.(3) ATC typically exhibit tumor-associated macrophage(TAMs) infiltration, which accounts for 40 to 70% of the total tumor mass and acts as an immunosuppressive tumor stroma, contributing to treatment resistance and poor prognosis.(2)

Previously, ATC was by default classified as stage IV in the American Joint Committee on Cancer (AJCC) 7th edition. However, the 5th edition of World Health Organization (WHO) now stages it like other differentiated thyroid carcinomas according to AJCC 8th edition.(1) Multimodal therapy with a combination of surgery, external beam radiation therapy (EBRT), and chemotherapy along with targeted therapy has shown better overall survival in ATC.(2)

Material and Methods

A 6-year retrospective study (July 2018 to July 2024) of diagnosed cases of ATC in the Department of Pathology with institutional ethics committee (IEC) approval. Histopathologically confirmed cases of ATC were meticulously searched from the hospital database. Cases with viable tumour tissue and available slides and blocks were retrieved for detailed analysis.

The details on the age, sex, clinical, biochemical parameters like thyroid function tests, serum calcium, relevant past history, imaging details, type of excision, treatment details, metastatic sites involved, follow up data were collected from patient electronic medical records. Gross findings, pathological staging details according to TNM Classification, AJCC 8^th edition was documented.

Cases lacking histological confirmation of ATC, unavailability of data, slides, blocks, non-neoplastic thyroid lesions, inflammatory conditions, and benign tumors of thyroid were excluded from the study.

The available histopathology slides and stained or freshly cut from retrieved blocks were examined for histological features like ATC subtype, pattern, necrosis, mitosis, lymphatic/angioinvasion, perineural invasion and tumour-infiltrating lymphocytes (TILs). Other histological parameters like extrathyroidal extension, adjacent thyroid findings, other tissue or organ involvement, margin status, lymph node metastasis and extra-nodal extension were evaluated. DTC components were assessed wherever possible.

Representative tumor foci were marked for tissue microarray(TMA) excluding slides with necrotic or poorly preserved tissue. Among the 15 cases of ATC, 12 were suitable for TMA and underwent BRAFV600E immunohistochemistry(IHC) using clone IHC600. The IHC slide was interpreted as positive if >50% of tumour cells showed intense to moderate cytoplasmic granular stain and focal if <50% showed weak cytoplasmic granular staining. The already available IHC slides of each case were reviewed with a robust IHC panel of PAX8(MD-50), TTF1(SPT24), CK(AE1/AE3), Vimentin(SP20), p53(do-7), p40(Delta NPP), Synaptophysin(EP158), Chromogranin(EP38), CD56(123C3), CD34(QB-End/10), CMYC(Y69), Desmin(D33), HMB45(HMB-45), GFAP(GA5), CEA(CEAm), NapainA(BS10) and BRAF(IHC600). Based on the hospital records of the patients diagnosed with ATC, these patients were followed up till the last hospital visit or until death.

The collected data were entered in the Microsoft Excel 2016 and analysed with IBM SPSS Statistics for Windows, Version 29.0.(Armonk, NY: IBM Corp).To describe about the data descriptive statistics, T test was used to compare means, and Chi-square test was employed to study the association between the categorical variables. To find the Survival analysis the Kaplan Meier Curve with Log-rank method were used. In the above statistical tools, the probability value .05 is considered as significant level.

Results

This retrospective study spanning six years analysed fifteen histologically confirmed cases of ATC. The age ranged from 48-82 years, with a mean of 66.1 years and a male to female ratio of 1:2. The most prevalent clinical presentation was anterior neck swelling(n=12, 80%), followed by odynophagia(n=7, 46.6%), voice change(n=7, 46.6%), breathlessness(n=4, 26.6%) and stridor(n=3, 20%). Symptoms ranged from 2 days-8 months. 33.3% (n=5) had a long-standing goiter (20-25 years) with rapid progression. One patient reported biomass exposure for 20 years. 73.3%(n=11) were euthyroid, 20%(n=3) hypothyroid and 6.6%(n=1) hyperthyroid with toxic multinodular goiter. 73.3%(n=11) had leukocytosis (>10000 cells/cu mm). Serum calcium levels were within normal limits in all cases.

Radiologically, 60%(n=9) revealed distant metastasis at diagnosis, primarily affecting lung(n=8, 53.3%), followed by bone(vertebrae and rib)(n=4, 26.6%), pancreas(n=1, 6.6%) and 26.6% had multiple sites of distant metastasis.

80%(n=12) showed locally advanced lesions involving strap muscle, thyroid cartilage, prevertebral space, recurrent laryngeal nerve, trachea, larynx, and carotid vessel. Cervical lymph node metastasis was noted in 26.6%(n=4) cases. Tumor size ranged from 2-13cm with a mean of 7.1cm, presenting as either unifocal(n=9, 60%) or multifocal tumors(n=6, 40%). The clinicopathological features are summarized in Table 1.

Grossly, tumours appeared solid to cystic, exhibiting extensive necrosis, haemorrhage, and gross extrathyroidal invasion(Figure 1).

Figure 1: Anaplastic thyroid carcinoma showing large, fleshy, gray-white tumor replacing the entire lobe.

Frozen section was performed on 6 patients on an emergency basis in view of stridor and was diagnosed as ATC in 5 cases, while one was deemed negative for malignancy. Histopathology revealed varied morphological patterns like epithelioid(n=5, 33.3%)(Figure 2A), sarcomatoid(n=5, 33.3%)(Figure 2B), squamous(n=4, 26.6%)(Figure 2C) and pleomorphic/rhabdoid(n=1, 6.6%)(Figure 2D).

Figure 2: A. Anaplastic thyroid carcinoma, Epithelioid subtype(200x, H & E). B. Anaplastic thyroid carcinoma, Sarcomatoid subtype(200x, H & E). C. Squamous cell carcinoma(200x, H & E). D. Anaplastic thyroid carcinoma, Rhabdoid subtype(200x, H & E).

Mixed configurations of epithelioid and spindle pattern(n=3, 20%)(Figure 3A) and spindle with pleomorphic features (n=2, 13.3%) were identified. Other findings included osteoclast-like giant cells(n=1, 6.6%) (Figure 3B), myxoid stroma(n=1, 6.6%)(Figure 3C) and osteoid-like matrix, reminiscent of osteosarcoma(n=1, 6.6%)(Figure 3D).

Figure 3: A. Anaplastic thyroid carcinoma, mixed epithelioid and spindle subtype(200x, H & E). B. ATC with osteoclast like giant cell rich areas(200x, H & E). C. Sarcomatoid ATC with prominent myxoid stroma(200x, H & E). D. ATC with osteoid-like matrix mimicking osteosarcoma(200x, H & E).

Coexistence of ATC with differentiated thyroid carcinoma was observed in 46.6% (n=7) of cases. Of these, four cases (26.6%) were associated with papillary thyroid carcinoma (PTC) (Figure 4A), and three cases (20%) with follicular thyroid carcinoma (FTC) (Figure 4B).

Figure 4: A. Squamous cell carcinoma(lower left) with coexisting classic Papillary thyroid carcinoma(upper right) (200x, H & E) and inset showing p40 positivity. B. ATC with follicular thyroid carcinoma(200x, H & E).

Among the subtypes, 13.3% of squamous ATC were accompanying a PTC precursor. The epithelioid variant of ATC was associated with both PTC (n=2) and FTC (n=2) precursors. One case of pure sarcomatoid ATC showed an FTC precursor. The association of the clinicopathologic features with ATC subtypes are given in Table 2.

While DTC components were identifiable in larger excision specimens, their presence often went undetected in needle biopsies, underscoring the limitations of these approaches. Adjacent thyroid revealed lymphocytic thyroiditis(n=5, 33.3%) and multinodular goiter(n=5, 33.3%). A hallmark feature was the presence of neutrophilic inflammatory infiltrate. Extensive areas of necrosis, atypical mitosis, angioinvasion was observed in all cases.

High TILs(>10%) were noted in epithelioid(20%) and squamous(26.6%) ATC, while low TILs(<5%) in sarcomatoid(26.6%) and pleomorphic/rhabdoid(6.6%) ATC.

Immunohistochemistry(IHC) revealed strong, nuclear expression of PAX8 in 66.6%(n=10)(Figure 5A) and negative in sarcomatoid ATC. TTF1 showed focal weak nuclear positive(53.3%, n=8)(Figure 5A[Inset]), 73.3% expressed p53 mutant type diffuse nuclear positivity(Figure 5B) with one case showed null pattern. P40 was positive in all squamous ATC. Sarcomatoid ATC expressed vimentin, desmin and focal to negative CK. Napsin A was performed and all were negative. 53.3%(n=8) expressed BRAF V600E by IHC. Among these, four PTC-associated ATC and one pure pleomorphic/rhabdoid ATC showed diffuse intense BRAF positivity(Figure 5C). Focal BRAF staining was seen in two epithelioid ATC(Figure 5D).

Figure 5: A. Immunohistochemistry reveals diffuse nuclear expression of PAX8(200x), absence of TTF1 with positive internal control in thyroid follicles[Inset]. B. p53 mutant pattern(200x). C. Diffuse cytoplasmic BRAF expression(200x). D. Focal weak cytoplasmic staining of BRAF IHC(200x).

Immunohistochemistry findings are as highlighted in Table 3.

Only one patient received multimodal treatment with chemotherapy and radiotherapy post radical resection. Financial constraints precluded multimodal therapies, limiting options to palliative care in 14 patients. On follow-up, two patients were alive with disease(AWD), 12 patients died of disease(DOD) and one patient was lost on follow-up(LOF). The patients were followed up for a duration of 1 month-18 months. Treatment characteristics are illustrated in Table 4.

Two(13.3%) patients with squamous ATC and associated PTC precursor with BRAF mutation were AWD at 7 months and 18 months of last follow up respectively. Among these, one patient received multimodal treatment had local recurrence with lung metastasis and AWD at 18 months of follow up. The median overall survival of 15 cases was one month.

The survival analysis using the Kaplan-Meier curve and Log-rank method compared with age <60 and >60 years resulted in a p-value of 0.468, while between tumour size <5cm and >5cm yielded a p-value of 0.696 indicating no statistical significance(Figure 6A & 6B).

Figure 6: Survival curve of ATC patients for: (a) Age and (b) Tumour size.

Discussion

ATC is a highly aggressive, undifferentiated thyroid malignancy exhibits endpoint of progression from DTC.(4) In the current study 46.6% of cases had associated DTC. According to McIver et al., the progression from DTC to ATC spans 2-32 years, with a mean duration of 9.5 years.(5)

ATC accounts for 1-4% of all thyroid cancers worldwide and 1% in the USA and is responsible for 14-39% of thyroid cancer-related deaths.(1) ATC affects the geriatric group peaking in the eighth decade.(6,7) However, we observed a slightly younger cohort with a mean age of 66.1 years(7th decade). Literature review suggests a female preponderance consistent with our findings with male-to-female ratio of 1:2.(5,6,8)

ATC lacks a clearly defined clinical presentation, often manifesting with neck swelling and compressive symptoms involving the trachea, esophagus, and involvement of recurrent laryngeal nerve, similar to this study.(1) Long-standing goiter of 20-25 years, experience sudden progression within a month, were observed in 33.3% of cases in our cohort, mirroring findings by Pradhan et al.(9). Cervical lymph node metastasis has been reported in 29-64% in other studies(10), with the current study observing it in 26.6% of cases. Lung is the most common sites of distant metastasis, consistent with findings in literature.(9,11)

While previous studies reported associations with thyrotoxicosis, goiter, and paraneoplastic syndrome, our cohort was primarily euthyroid with leukocytosis(73.3%). Notably, 60% of patients were diagnosed with stage IVC disease, while 33.3% were in stage IVB, aligning closely with observations by Jannin and Zhang et al.(2,12)

ATC demonstrates diverse histology with heterologous elements like osteoid and cartilaginous components.(1,10) Similar to our findings, PTC forms the most associated DTC as shown in a study of 360 ATC cases.(13) It can also be seen with FTC and oxyphilic-Hürthle cell carcinoma.(14) Also, a significant association is seen between the tall cell variant of papillary carcinoma and squamous ATC as demonstrated in other studies.(13,15,16) In contrast, sarcomatoid ATC is more closely related to FTC and often harbors RAS mutations.(15) Sarcomatoid histological type is associated with the highest overall mortality, with all sarcomatoid cases in this study presenting at a higher stage with distant metastases. The WHO 5th Edition now recognizes squamous cell carcinoma (SCC) of the thyroid as a morphological subtype of ATC.(17) The molecular and clinical behavior reveal overlapping features between SCC and ATC, originating from follicular cells and expressing BRAFV600E, TTF1, and PAX8 with poor prognosis.(18)

According to Chen et al, BRAF-mutated ATC constitutes 36% and is associated with a high frequency of DTC, particularly PTC similar to the current study and shows a higher association with squamous patterns.(14) BRAF has therapeutic benefits in ATC. Therefore, BRAFV600E mutation testing is suggested in all cases. FDA has approved dabrafenib and trametinib (targeting BRAF and MEK1/2) for patients with BRAFV600E-mutant ATC(15,19)

We also found high TILs (>10-20%) in 53.3% of ATC, mainly in epithelioid(20%) and squamous(26.6%) subtype, in contrast to low TILs(<5%) in sarcomatoid ATC. The significance of TILs in ATC is still under investigation. A positive correlation between PD-L1 and BRAFV600E was seen along with association of epithelioid pattern exhibiting increased PD-L1 compared to sarcomatoid pattern of ATC, like the findings noticed by other researchers.(20–22) Immunotherapy, particularly spartalizumab (anti-PD-1 antibody), has shown promising results by Capdevila et al(23)

PAX8 is a sensitive marker compared to TTF1 and shows increased expression with epithelioid and squamous ATC, making it invaluable for distinguishing ATC from metastatic lung carcinoma. According to Nonaka et al, PAX8 expression was observed in 79% of ATC, compared to TTF-1 in 18%.(24) We observed PAX8 in 66.6% and TTF1 in 53.3%. In the present study, NapsinA was negative in all cases, however, Wu et al showed 11.1% of Napsin A expression in ATC aiding differentiation from metastatic lung carcinoma when combined with thyroid markers.(25) The diagnosis of ATC hinges on identifying DTC components, establishing thyroid as the primary tumour site, exclusion of metastasis or direct invasion from adjacent structures. Differentiating high-grade non-anaplastic thyroid carcinoma relies on thyroglobulin expression and lack of anaplasia.

According to prior literature, old age(>70 years), males, leukocytosis, extrathyroidal extension with distant metastasis at presentation are poor prognostic features.(6,11,12) Contrary to previous notions, age, tumour size and leukocytosis did not impact prognosis in the current study, similar to findings by Masui et al(26). Xu et al showed that the site of ATC, morphologic features, necrosis, mitoses and distant metastasis, was not associated with outcomes.(13) Our study revealed a better outcome with BRAF mutant squamous ATC, suggesting that this subset may derive particular benefit from targeted treatments. However, no statistically significant associations were observed between age, tumour size or morphological subtypes with survival.

One patient who received surgical intervention with multimodal treatment survived for 7 months, proving that incorporation of multimodal therapy including surgical intervention has shown promise in prolonging survival.(27) Patients undergoing total thyroidectomy demonstrate superior outcomes compared to partial resections, although lymph node dissection does not influence survival.(6) Despite surgical intervention, ATC prognosis remains grim, with only 20% of patients surviving beyond one year.(10) However, the limited survival time and poor statistical data on ATC underscore the need for more research and improved treatment strategies.

Conclusion

This six-year retrospective analysis highlights the aggressive nature of ATC, often diagnosed at an advanced stage with frequent metastasis. The patients presented in seventh decade with acute manifestations. ATC often arises from DTC, particularly papillary thyroid carcinoma in epithelioid and squamous subtypes with expression of BRAF. BRAF-mutated squamous ATC has prolonged survival in this cohort. Application of targeted therapy based on BRAFV600E status may aid in prognostication and improve outcomes in selected patients. Future research must focus on molecular risk stratification, integration of targeted therapy and immunotherapy into multimodal treatment strategies.

Limitations of the Study

The study faced several limitations, including a small sample size, retrospective nature, lack of molecular testing beyond IHC, inherent limitations of core needle biopsies, which fail to capture crucial elements such as DTC components and staging parameters. The statistical analysis was attempted; however, no significant values were obtained in view of small sample size.

Competing interests

The authors declare that they have no competing interests.

Ethics approval and consent to participate

The study was approved by Institutional ethical committee of Kasturba Medical College, Manipal (IEC1 – 404). Since it was retrospective study, consent from the participants was waived.

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