Extra copies of ALK gene locus is a recurrent genetic aberration and favorable prognostic factor in both ALK-positive and ALK-negative anaplastic large cell lymphomas

Systemic anaplastic large cell lymphoma (ALCL) is subtyped into ALK-positive ALCL and ALK-negative ALCL based on the presence or absence of ALK protein expression. ALK-positive ALCL is characterized by t(2;5)(p23;q35)/NPM-ALK or variant ALK-involved translocations, while little is known about the genetic changes in ALK-negative ALCL. We investigated the structural and numerical aberrations of the ALK gene using interphase fluorescence in situ hybridization (FISH) in 81 cases with ALCL and analyzed their association with clinical outcome of the patients. ALK gene rearrangement was found in 47 of 50 (94%) ALK-positive ALCLs but in none of 31 ALK-negative ALCLs. Extra copies of the ALK gene locus, representing mainly extra copies of chromosome 2, were seen in 19 ALK-positive (38%) and 15 ALK-negative (48%) cases (P=0.357). In 55 cases with follow-up information, the mean survival time of the 38 ALK positive cases (58 months) was significantly longer than that of 17 ALK-negative cases (22.5 months) (P=0.038). Interestingly, the cases with extra copies of ALK had a significantly longer mean survival time than those without (64.4 months vs 35.3 months) (P=0.023) and this difference was observed in both ALK-positive (72.3 vs 45.9 months) and ALK-negative (34.7 vs 9.9 months) cases. Multivariate analysis showed that both ALK protein expression and extra copies of ALK gene were independent predictors for better survival (P=0.008). Our results suggest that the extra copies of ALK gene locus are a frequent genetic aberration in both ALK-positive and ALK-negative ALCL and is a favorable prognostic marker for the patients.     

Prognostic significance of MUC-1 expression in systemic anaplastic large cell lymphoma.

Systemic anaplastic large cell lymphoma (ALCL) frequently carries the t(2;5)(p23;q35) and overexpresses anaplastic lymphoma kinase (ALK). MUC-1, a highly glycosylated transmembrane protein, is detected in normal and malignant epithelial cells and has been associated with a poorer patient survival in various human malignancies. We have shown previously that MUC-1 is expressed as a consequence of t(1;14)(q21;32) in a subset of diffuse large B-cell lymphomas. ALCLs are known to express MUC-1, but its clinical significance is undefined. For this study, eligible patients with ALCL were HIV negative, received anthracycline-containing regimens, and had pretreatment archival tissue. Expression of MUC-1 and ALK was determined immunohistochemically after heat-induced antigen retrieval. A 10% cutoff for MUC-1 positivity was used. We identified 63 patients with systemic ALCL (22 ALK+, 41 ALK-) with a median age of 47 years, and 41 were male. MUC-1 was detected in 16 of 22 (73%) ALK-positive and 20 of 41 (49%) ALK-negative ALCL (P = 0.06, chi(2) test). MUC-1 expression was not associated with apoptotic rate as detected by terminal deoxynucleotidyl transferase-mediated nick end labeling assay or proliferation index as evaluated by MIB-1 antibody. For 48 patients with ALCL (16 ALK+, 32 ALK-) and complete clinical follow-up, 5-year progression-free survival (PFS) was 39.7% for patients with MUC-1-positive tumors versus 75.2% (P = 0.027 by Log-rank) for patients with MUC-1-negative tumors. For the ALK-negative ALCL group of 32 patients, the 5-year PFS was 26 versus 70.8% for patients with MUC-1-positive versus MUC-1-negative tumors (P = 0.0096 by Log-rank). For the ALK-positive ALCL group of 16 patients, the 5-year PFS was 52 versus 100% for patients with MUC-1-positive versus MUC-1-negative tumors (P, not significant). In summary, MUC-1 is frequently expressed in systemic ALCL, and its expression is associated with significantly inferior outcome in patients untreated previously with ALK-negative tumors. Future studies should explore the underlying molecular mechanisms of MUC-1 expression in these tumors and its role as a target for novel therapeutic strategies.     

B and CTL responses to the ALK protein in patients with ALK-positive ALCL

Anaplastic lymphoma kinase (ALK)-positive anaplastic large cell lymphoma (ALCL) has a good prognosis compared to ALK-negative ALCL, possibly as a result of the immune recognition of the ALK proteins. The aim of our study was to investigate the presence of both a B and cytotoxic T cell (CTL) response to ALK in ALK-positive ALCL. We confirmed the presence of an antibody response to ALK in all 9 ALK-positive ALCL patients investigated. An ELISpot assay was used to detect a gamma-interferon (IFN) T cell response after short term culture of mononuclear blood cells with 2 ALK-derived HLA-A*0201 restricted peptides: ALKa and ALKb. A significant gamma-IFN response was identified in all 7 HLA-A*0201-positive ALK-positive ALCL patients but not in ALK-negative ALCL patients (n = 2) or normal subjects (n = 6). CTL lines (>95% CD8-positive) raised from 2 ALK-positive ALCL patients lysed ALK-positive ALCL derived cell lines in a MHC-Class I restricted manner. This is the first report of both a B cell and CTL response to ALK in patients with ALK-positive ALCL. This response persisted during long-term remission. The use of modified vaccinia virus Ankara (MVA) to express ALK is also described. Our findings are of potential prognostic value and open up therapeutic options for those ALK-positive patients who do not respond to conventional treatment.     

CD4 T-helper responses to the anaplastic lymphoma kinase (ALK) protein in patients with ALK-positive anaplastic large-cell lymphoma

We have previously shown both humoral and CTL responses to anaplastic lymphoma kinase (ALK) in patients with ALK-positive anaplastic large-cell lymphoma (ALCL). However, because CD4(+) T-helper (Th) cells also play a vital role in developing and maintaining tumor immunity, we investigated the presence of a CD4(+) Th response in ALK-positive ALCL. Using an IFN-gamma ELISPOT assay, we identified two ALK-derived DRB1-restricted 24-mer promiscuous peptides, ALK1(278-301) and ALK2(233-256), as being immunogenic in six ALK-positive ALCL patients but not in two ALK-negative ALCL patients or five normal subjects. A significant interleukin-4 response to the ALK peptides was detected in only one ALK-positive patient. CD4(+) Th cell lines lysed ALK-positive ALCL cell lines in a MHC class II-restricted manner. This first report of a CD4(+) Th response to ALK provides valuable information for developing future immunotherapeutic options for ALK-positive ALCL patients who fail to respond well to conventional therapies.     

Cytomorphologic examination of anaplastic large cell lymphoma by fine-needle aspiration cytology

BACKGROUND: The cytomorphology of anaplastic large cell lymphoma (ALCL) is distinctive yet variable. To the authors' knowledge, to date only small case series have described the cytologic findings noted in patients with ALCL. The current series is the largest case series presented to date to retrospectively review the cytomorpholgic findings noted in patients with ALCL, with specific attention paid to those with anaplastic lymphoma kinase (ALK)-negative ALCL. METHODS: Over a 13-year period, the available Diff-Quik cytology smears and surgical excision specimens taken from patients with ALCL were evaluated. Different clinical and morphologic parameters were evaluated, including ALK status. RESULTS: A total of 37 cases were retrieved and evaluated, 19 of which had both cytology and surgical pathology specimens available for review. ALK-negative ALCL cytology smears were found to have a high number of anaplastic cells compared with ALK-positive cases. The hallmark cells in the ALK-negative cases were not classic. CONCLUSIONS: ALCL can be diagnosed accurately by fine-needle aspiration cytology (FNAC) alone when aided by immunocytochemistry in ALK-positive cases. Ancillary studies should be anticipated such that material for cell block preparation and molecular studies is taken at the time of FNAC. The results of the current study demonstrate the varied FNAC morphology of ALCL. The presence of severe pleomorphism and anaplasia was found to correlate with ALK-negative status.     

ALK-negative anaplastic large cell lymphoma with extensive peripheral blood and bone marrow involvements manifested as “leukemic phase”

CD30-positive anaplastic large cell lymphoma (ALCL) is a distinctive malignant large cell lymphoma of T-cell lineage, often presenting in lymph node or extranodal sites. ALCL cases with extensive bone marrow and peripheral blood involvement manifested as “leukemic phase” are extremely rare and the most of those cases reported are anaplastic large cell lymphoma kinase (ALK) positive ALCL in childhood population. Here we report four adult cases of ALK-negative ALCL with extensive bone marrow and peripheral blood involvement manifested as “leukemic phase”. Circulating large lymphoma cells varied from 20 to 80% in peripheral blood and bone marrow biopsy showed various nodular or interstitial infiltrates. By reviewing the clinicopathologic data of previously reported ALCL cases with extensive bone marrow and peripheral blood involvement, there appears to be of large variations in regard to the patient's age, morphologic variants, immunophenotypic or genotypic characteristics of the disease. While most cases of ALCL with peripheral blood and bone marrow involvement were ALK-positive or carrying t(2;5) translocation, rare ALK-negative cases were also present. Leukemic ALCL patients usually have unfavourable prognosis, regardless of ALK expression.     

原发系统性间变性大细胞淋巴瘤临床病理特征及免疫表型分析

 目的　分析原发系统性间变性大细胞淋巴瘤（ALCL）的临床病理特征和免疫组织化学特点,提高诊治水平。方法　选取22例ALCL患者,均进行分期、国际预后指数（IPI）、乳酸脱氢酶（LDH）检测,应用免疫组织化学SP法检测间变性淋巴瘤激酶（ALK）、Ki-67、Caspase-3、CD30、EMA、Granzyme B等,回顾性分析患者临床、病理形态学资料、免疫表型及生物学特性,并进行预后分析。结果　22例均为原发系统性ALCL,ALK+ 15例（68.2 %）,ALK- 7例（31.8 %）;ALK+患者发病年龄、Ki-67增殖指数较ALK-患者低,Caspase-3表达率高,差异有统计学意义（χ2＝4.618,P＝0.032）;15例ALK+ALCL均表达CD30和EMA。ALCL中ALK的表达与Ki-67、Caspase-3的表达呈负相关（r＝-0.581,P＝0.006;r＝0.458,P＝0.032）。ALK+病例较ALK-病例Granzyme B（χ2＝0.11,P＝0.74）、 TIA-1（χ2＝0.01,P＝0.92）的表达率高,但差异无统计学意义（P＞0.05）。有效率为 54.5 ％（12／22）,其中完全缓解率为18.2 %（4／22）;全组中位生存期12个月,1年生存率为59.1 %（13／22）,2年生存率为50.0 ％（11／22）。Ann Arbor分期、LDH及IPI与疾病预后相关。结论　ALK+较ALK-ALCL患者核增殖低,恶性程度低,临床特征和免疫表型具有一定的特征性;ALK、Ki-67、Caspase-3、分期、血清LDH及IPI对预测ALCL患者的生存和指导治疗有帮助。     

间变性淋巴瘤激酶阳性和阴性间变性大细胞淋巴瘤分子遗传学研究

目的：研究间变性大细胞淋巴瘤（ALCL）的遗传学特征,探讨其发病机制,寻找有助于ALCL诊断、分类及预后评估的新分子靶标。方法收集ALCL病例石蜡包埋组织10例,其中4例间变性淋巴瘤激酶（ALK）阳性,6例ALK阴性。采用免疫组织化学染色及荧光原位杂交技术分别检测表型及2p23重排,利用OncoScan芯片在全基因组水平上扫描分析10例ALCL的拷贝数变异。结果10例ALCL均存在拷贝数变异,拷贝数获得者多于拷贝数缺失者。拷贝数获得主要累及17q11.2、Xp22.3、Xq28,拷贝数缺失主要累及3q26.1、14q11.2、22q11.23。 ALK阴性者拷贝数变异比ALK阳性者更为复杂：ALK阴性者拷贝数获得主要累及9q24.3-24.1、14q32.33,拷贝数缺失主要累及2p11.2、16p13.3,而ALK阳性者并无此变异。结论 ALCL存在复杂的遗传学不平衡,染色体片段获得多于缺失,ALK阴性ALCL遗传学不平衡更为复杂。 ALCL是异质性明显的一类肿瘤。     

p53 and β-Catenin Expression Predict Poorer Prognosis in Patients With Anaplastic Large-Cell Lymphoma

BackgroundThe Wnt/β-catenin signaling pathway is a major target of p53. β-Catenin/p53 coexpression predicts poorer survival in carcinoma patients. Conversely, CD99 inhibits tumor metastasis through the Wnt/β-catenin pathway. We therefore assessed p53, β-catenin, and CD99 by immunohistochemistry.Patients and MethodsWe studied 45 patients with systemic anaplastic large-cell lymphoma (ALCL), including 20 anaplastic lymphoma kinase (ALK)-positive and 25 ALK-negative ALCL. β-Catenin expression was analyzed using phospho-β-catenin-S552 antibody because its nuclear localization indicates Wnt signaling.ResultsIn this cohort, p53 expression was associated with ALK-negative ALCL. Furthermore, p53 or β-catenin expression alone or β-catenin/p53 double expression showed poorer overall survival and disease-free survival in patients with ALCL overall and in patients with ALK-negative ALCL. CD99 expression was more frequent in ALK-positive ALCL but had no prognostic significance.ConclusionThis is the first study to evaluate phospho-β-catenin-S552 expression in ALCL. The results of this study, although limited by small patient size, suggest that β-catenin and p53 may play a role in pathogenesis and may be helpful in risk stratification of ALCL patients.     

Anaplastic large cell lymphoma--a rare disorder in southern Taiwan

Anaplastic large cell lymphoma (ALCL) is a subgroup of non-Hodgkin's lymphomas with large lymphoma cells expressing CD30 antigen. This entity has rarely been reported in Taiwan. We performed a retrospective clinicopathologic study in a medical center in southern Taiwan during a 13-year period and identified 13 cases. There were 10 males and 3 females with a median age of 49 years old. Seven presented with pure nodal disease and 5 had bony involvement. The staging results were stage I (5 patients), II (1), III (1), and IV (4). The pathologic subtypes were common variant (10), lymphohistiocytic variant (2), and small cell variant (1). Eleven tumors were of T-cell lineage; 2, null-cell. Immunohistochemically, 5 tumors (38.5%) expressed cytotoxic markers, T-cell intracellular antigen-1 and/or granzyme B. Two tumors (15.4%) expressed anaplastic lymphoma kinase (ALK). Long-term follow-up information was available in 8 patients. The 2 patients with ALK-expressing tumors (37 and 49 years old) were free of disease for 61 and 54 months, respectively. The other 6 patients were either died of disease (5 patients) or experienced relapse with progressive disease (1). In conclusion, we reported the largest series of ALCL in Taiwan. We confirmed ALK-expressing ALCL carries favorable prognosis and ALK-negative ALCL has similar poor prognosis as non-anaplastic T-cell lymphoma. As compared to the previous reports from the West, our ALK positive rate was lower and the age of our ALK-positive patients was older. A larger national or multi-institutional study is needed for further characterization of ALCL in Taiwan.     

Histology impacts the outcome of peripheral T-cell lymphomas after high dose chemotherapy and stem cell transplant

The role of high dose chemotherapy (HDC) and stem cell transplant (SCT) in peripheral T-cell lymphoma (PTCL) was studied in 28 patients, from 1988 to 2002. The aim was to determine if subsets recognized by the REAL/WHO classification have different prognoses. Outcome was compared to 86 patients with diffuse large B-cell lymphoma (DLBCL) transplanted during 1986-2000. The 3-year overall survival (OS) and event free survival (EFS) were 69% and 50%. Patients with anaplastic large cell lymphoma (ALCL) had a better 3-year OS compared to those with non-ALCL histology (86% vs. 47%, P=0.0122). Anaplastic lymphoma kinase (ALK)- positive ALCL patients had a superior EFS compared to ALK-negative ALCL (100% vs. 0; P=0.0228). Patients with cutaneous ALCL (ALK-negative) relapsed, but had an indolent course after SCT. Low International Prognostic Index score at relapse predicted for a better 3-year OS (85% vs. 34%, P=0.0238). When compared to DLBCL, patients with ALCL had a superior OS (86% vs. 36%, P=0.0034) and patients with non-ALCL had a comparable OS. ALCL histology confers better survival compared to non-ALCL and DLBCL histologies. ALK-positive ALCL is associated with the best EFS after relapse with HDC and SCT. The timing of SCT for non-ALCL histology remains to be determined.     

57例原发系统型间变大细胞性淋巴瘤临床分析

背景与目的：原发系统型间变大细胞性淋巴瘤（anaplastic large cell lymphoma,ALCL）的临床特征和疾病转归国内外报道的结果存在一定的差异。间变性淋巴瘤激酶蛋白（anaplastic lymphoma kinase,ALK）被认为是原发系统型ALCL较为特异的标志,表达与否和患者的临床特征及预后相关。本研究目的是探讨原发系统型ALCL患者的临床特点和预后,检测ALK的表达并探讨其临床意义。方法：回顾性分析1997年1月至2006年6月中山大学肿瘤防治中心收治的57例原发系统型ALCL患者的临床资料。采用免疫组化SP法检测肿瘤标本中ALK的表达。结果：57例患者的中位年龄为30岁,男女比例为1．9：1,有B症状、Ⅲ～Ⅳ期和结外侵犯患者分别占57．9％、40．4％和40．4％。所有患者接受化疗,56例可评价疗效,40例（71．4％）取得完全缓解,客观有效率为96．4％;中位随访49．1个月,中位无疾病进展生存期为35．2个月,预计5年生存率为65．2％：国际预后指数评分低／低中危患者和中高／高危患者的预计5年生存率分别为68．9％和33．3％（P=0．010）。46例中ALK阳性29例（63％）,发病年龄小于阴性者（P=0．001）,预计5年生存率优于阴性者（尸〈0．01）。多因素分析显示,ALK表达状况、原发部位和乳酸脱氢酶水平是ALCL患者的独立预后因素。结论：原发系统型ALCL发病年龄较年轻,化疗有效率高,预后良好,但ALK阴性、IPI中高／高危、尤其合并结外侵犯和乳酸脱氢酶水平升高者预后较差。     

p53 gene mutations are uncommon but p53 is commonly expressed in anaplastic large-cell lymphoma.

Anaplastic large-cell lymphoma (ALCL), as defined in the World Health Organization, is a heterogeneous category in which a subset of cases is associated with the t(2;5)(p23;q35) or variant translocations resulting in overexpression of anaplastic lymphoma kinase (ALK). p53 has not been assessed in currently defined subsets of ALCL tumors. In this study, we assessed ALK+ and ALK- ALCL tumors for p53 gene alterations using PCR, single-strand conformation polymorphism and direct sequencing methods. We also immunohistochemically assessed ALCL tumors for p53 expression. Three of 36 (8%) ALCL tumors (1/14 ALK+, 2/22 ALK-) with adequate DNA showed p53 gene mutations. By contrast, p53 was overexpressed in 36 of 55 (65%) ALCL tumors (16 ALK+, 20 ALK-). p21, a target of p53, was expressed in 15 of 31 (48%) ALCL tumors including seven of 15 (47%) p53-positive tumors. p21 expression in a subset of ALCL suggests the presence of functional p53 protein. Apoptotic rate was significantly higher in p53-positive than p53-negative tumors (mean 2.78 vs 0.91%, P = 0.0003). We conclude that the p53 gene is rarely mutated in ALK+ and ALK- ALCL tumors. Nevertheless, wild-type p53 gene product is commonly overexpressed in ALCL and may be functional in a subset of these tumors.     

儿童原发中枢神经系统间变性淋巴瘤激酶阳性间变性大细胞淋巴瘤一例并文献复习

目的:探讨儿童原发中枢神经系统（CNS）间变性大细胞淋巴瘤（ALCL）的临床特征及预后。方法:回顾性分析福建医科大学附属协和医院收治的1例原发CNS间变性淋巴瘤激酶（ALK）阳性ALCL患儿的临床资料,并复习相关文献。结果:患儿以头痛、发热为主要症状就诊外院,颅脑磁共振成像提示右侧小脑肿块,术前无CNS以外淋巴瘤浸润证据,行小脑肿瘤切除术,术后病理明确诊断为ALK阳性ALCL,未及时化疗。术后第27天转入福建医科大学附属协和医院,化疗前评估肿瘤已扩散至骨髓、睾丸、椎骨等部位,外周血NPM-ALK融合基因阳性。2个疗程化疗后达完全缓解,但最终死于化疗相关并发症。结论:原发CNS的ALK阳性ALCL病例罕见,易误诊,病情进展快,总体预后不良;及时进行活组织病理检查以确诊,早期行以化疗为主的综合治疗可能改善患者预后。