YKL-40蛋白在犬隻造血系腫瘤中作為預後生物標記的角色

Abstract

生物標記的開發是一個從發現、驗證到臨床應用的複雜過程。根據美國食品藥物管理局與國家衛生院生物標記工作小組的定義，生物標記是可用於評估生理過程、疾病狀態或治療反應的指標物。其中，預後生物標記與監測生物標記在臨床腫瘤治療策略中扮演重要角色，探索新的生物標記具有重要意義。YKL-40是一種分泌型醣蛋白，在人類癌症中參與腫瘤細胞增殖、轉移及血管新生。研究顯示，YKL-40的高表現量與較高的臨床分期及不良預後相關。在獸醫學領域，罹患癌症的犬血中YKL-40水平也有升高的現象，但其作為生物標記的潛力尚未被深入探討。由於犬類血液腫瘤的生物標記研究仍在進發展階段，開發新的生物標記具有迫切需求。本研究目的為探討YKL-40在犬造血腫瘤疾病中的生物標記潛力，分析YKL-40的表現量與腫瘤的臨床特徵、病理特徵、治療與生存時間的相關性。 在組織表現量的研究中，我們以皮膚型肥大細胞瘤作為目標，收集40例罹患皮膚型肥大細胞瘤犬的組織，包含20例低病理分級與20例高病理分級的樣本，透過免疫反應性評分進行YKL-40的半定量分析。結果顯示，低分級MCT組織的YKL-40表現顯著高於高分級腫瘤（p < .01）。而YKL-40表現水平與腫瘤直徑、分裂指數及血管密度呈負相關。在患犬生存分析中，YKL-40弱表現的犬（n = 15）中位生存時間為219天，而中等表現（n = 19）及強表現（n = 6）的犬中位生存時間顯著較長（p < .01）。此外，在低分級患犬組別中，YKL-40弱表現與較差的預後相關，而中至強表現則與較長的生存時間相關（p < .01）。 在血液表現量的研究中，我們以多中心淋巴瘤為目標，收集來自30隻犬的血液檢體，分析時間點包含治療前、治療過程中、治療後與疾病復發時的樣本，並使用酶聯免疫吸附測定定量血清YKL-40的濃度。結果顯示，治療前多中心淋巴瘤犬血清的YKL-40水平（394.0 pg/mL, n = 30）顯著高於健康犬對照組（218.6 pg/mL, n = 11; p = 0.012），其中臨床分期第五期的犬YKL-40水平最高（p = 0.027）。治療完成後，YKL-40水平顯著下降（p = 0.030），然而治療過程中，YKL-40的變化與治療反應未呈顯著相關性。在患犬生存分析中，治療前YKL-40水平與無疾病進展生存期（p = 0.830）及總生存期（p = 0.267）之間無顯著相關性。 總結來說，YKL-40在不同犬腫瘤類型中的表現量的意義具有差異性。在皮膚型肥大細胞瘤中，中等至強YKL-40表現與良好預後相關，而弱表現則與較差的預後相關，顯示YKL-40具有作為肥大細胞瘤患犬的預後生物標記的潛力。在犬多中心淋巴瘤中，血清YKL-40水平可能與疾病嚴重程度相關，並隨著疾病進展而變化，然而，其作為預後與監控生物標記的角色仍需進一步驗證。未來需要進行大樣本數的前瞻性研究，以確認YKL-40在犬造血腫瘤中作為生物標記的臨床應用價值，這些資訊將有助於提升犬臨床腫瘤學中的治療決策。

The development of biomarkers is a complex process involving biomarker discovery, validation, and clinical application. According to the FDA-NIH Biomarker Working Group, biomarkers serve as indicators of physiological processes, disease states, or treatment responses. Prognostic and monitoring biomarkers play crucial roles in clinical oncology, emphasizing the importance of exploring novel biomarkers. YKL-40, a secretory glycoprotein, is involved in tumor cell proliferation, metastasis, and angiogenesis in human cancers. Studies have demonstrated that elevated YKL-40 expression is associated with advanced clinical stages and poor prognosis. In veterinary medicine, elevated blood YKL-40 levels have been observed in dogs with cancers; however, its potential as a biomarker remains underexplored. As the development of biomarkers for canine hematopoietic tumors is still in the early stages, identifying novel biomarkers is critically important. This study aims to investigate the potential of YKL-40 as a biomarker for canine hematopoietic tumors by evaluating its association with clinical characteristics, pathological features, treatment response, and survival. For tissue expression analysis, cutaneous mast cell tumors (MCTs) were studied. Tissue samples from 40 dogs, including 20 low-grade and 20 high-grade cMCTs, were collected, and YKL-40 expression was semi-quantified using the immunoreactivity score. The results showed that YKL-40 expression was significantly higher in low-grade tumors than in high-grade tumors (p < .01). YKL-40 expression levels were inversely correlated with tumor diameter, mitotic count, and vessel density. Survival analysis demonstrated that dogs with mild YKL-40 expression (n = 15) had a median survival time (MST) of 219 days, whereas those with moderate (n = 19) or strong expression (n = 6) exhibited significantly longer MSTs (p < .01). In low-grade cMCTs, mild YKL-40 expression was associated with poorer prognosis (MST: 319 days), while moderate to strong expression was correlated with longer survival time (p < .01). For blood level analysis, serum samples were collected from 30 dogs with multicentric lymphoma at different time points, including pre-treatment, during-treatment, post-treatment, and disease relapse. Serum YKL-40 levels were measured using an enzyme-linked immunosorbent assay. Pre-treatment serum YKL-40 levels in dogs with lymphoma (394.0 pg/mL, n = 30) were significantly higher than in those in healthy controls (218.6 pg/mL, n = 11; p = 0.012), with the highest levels observed in dogs at clinical stage V (p = 0.027). Post-treatment YKL-40 levels significantly decreased (p = 0.030). However, the change in YKL-40 levels during treatment was not significantly associated with treatment responses. Survival analysis revealed no significant association between pre-treatment YKL-40 levels and progression-free survival (PFS, p = 0.830) or overall survival (OS, p = 0.267). In conclusion, the significance of YKL-40 expression could vary across canine tumor types. In cMCTs, moderate to strong YKL-40 expression could be associated with a favorable prognosis, while mild expression could be correlated with poorer outcomes, supporting its potential as a prognostic biomarker. In canine multicentric lymphoma, serum YKL-40 levels may be correlated with disease severity and could change as the disease progresses. However, its role as a prognostic and monitoring biomarker requires further validation. Future prospective studies with larger sample sizes are necessary to confirm the clinical utility of YKL-40 as a biomarker in canine hematopoietic tumors. This information could aid decision-making in canine clinical oncology.