To the Editor,

In the 2016 iteration of the World Health Organisation (WHO) classification of myeloid neoplasms, myelodysplastic syndrome/myeloproliferative neoplasm with ring sideroblasts and thrombocytosis (MDS/MPN‐RS‐T) has been included as a unique entity, defined by the presence of erythroid lineage dysplasia in the presence or absence of multilineage dysplasia and persistent thrombocytosis (platelet count ≥450 × 10⁹/l), without other disease‐defining genetic abnormalities (Arber et al., 2016). Frequent somatic alterations in patients with MDS/MPN‐RS‐T include SF3B1 (85%), JAK2V617F (50%), TET2 (25%), ASXL1 (20%), DNMT3A (15%) and SETBP1 (10%) (Patnaik et al., 2016a). With an estimated median overall survival of approximately 76 months, outcomes are better than in patients with MDS‐RS and single lineage dysplasia, but inferior when compared to patients with essential thrombocythemia (Broseus et al., 2012).

Due to ineffective erythropoiesis related to locking of intracellular iron in mitochondria and accelerated erythroid precursor apoptosis, anaemia is a major source of morbidity, not only due to complications associated with impaired oxygen transport but also due to red blood cell (RBC) transfusion‐associated iron overload (Patnaik & Tefferi, 2019). Erythropoiesis‐stimulating agents (ESAs) are commonly used to reverse this erythroid maturation arrest and decrease the RBC transfusion frequency, but literature on specific response rates in patients with MDS/MPN‐RS‐T is limited (Patnaik & Tefferi, 2019). This is especially relevant as patients with MDS/MPN‐RS‐T are already at an increased risk for thrombosis, which may be compounded by the use of concomitant ESA (Broseus et al., 2012). We conducted a single institution retrospective study to assess ESA responses among WHO‐defined patients with MDS/MPN‐RS‐T.

After Institutional Review Board approval, adult patients with WHO‐defined MDS/MPN‐RS‐T, diagnosed from years 2002 to 2014 at our institution, were included in the study. Diagnostic bone marrow (BM) aspirate and trephine biopsy morphology and cytogenetics were reviewed to ensure compliance with the latest WHO criteria (Arber et al., 2016). Twenty‐four (60%) study patients underwent next generation sequencing (NGS) for the following genes: IKZF1, PTPN11, SH2B3, SUZ12, ZRSR2, ASXL1, CALR, CBL (exons 8, 9), CEBPA (exon 1), CSF3R (exons 14, 17), DNMT3A (exons 4, 8, 13, 15, 16, 18, 19, 20, 22 and 23), EZH2 (exons 8, 17 and 18), FLT3 (exons 14, 20), IDH1 (exon 4), IDH2 (exon 4), JAK2 (exons 12, 14), KIT, MPL (exon 10), NPM1 (exon 11), NRAS (exons 2, 3), RUNX1 (exons 3, 4 and 8), SETBP1 (exon 3), SF3B1 (exons 13, 15 and 17), SRSF2 (exon 1), TET2 (exons 3, 9, 10, 11), TP53 (exons 5, 6, 7, and 8) and U2AF1 (exons 2, 6) obtained on BM mononuclear cells, at diagnosis, or at first referral, by previously described methods (Patnaik et al., 2016a). Treatment details, including the type of ESAs used, the dose administered, side effects and the reason for discontinuation were retrospectively abstracted. Erythroid responses were assessed using the 2015 IWG (International Working Group) MDS/MPN response criteria (Savona et al., 2015).

Forty patients with WHO‐defined MDS/MPN‐RS‐T, who received ESA treatment at any time during their disease course, were included in the study; median age 72·1 years (range, 52–93·3), 45% male. Table 1 describes the clinical and laboratory characteristics of these patients. The median haemoglobin (Hb) at diagnosis was 9·2 gm/dl (range, 6·6–12·1); median baseline Hb level (prior to ESA treatment) was 9 gm/dl (range, 6·6–12·1 ‒ Hb levels were spuriously high in six patients due to recent RBC transfusions and levels prior to referral were not available) and median baseline erythropoietin (EPO) level (available in 15 patients ‒ nine responders, six non‐responders) 44 iu/l (range, 8–500). Seventeen (43%) patients had a bone marrow reticulin stain evaluation at diagnosis, with seven (41·2%) patients showing evidence of bone marrow fibrosis (six grade 1, one grade 2). Median LDH level at diagnosis was 152·5 (range: 145–270) U/l; however, this information was only available for four patients. Abnormal karyotype was seen only in three (7·5%) patients, with abnormalities including der(13;14)(q10;q10), ‐Y and inv(3)(q21;q26). Eleven (31%) patients were RBC transfusion‐dependent (TD) prior to start of ESA therapy. None of the 40 ESAs‐treated patients were on concomitant cytoreductive therapy for thrombocytosis. Twenty‐one (53%) patients were treated with epoietin‐α (EA), 15 (38%) with darbepoietin‐α (DA), and four (10%) received both drugs sequentially. Median administered doses were 40,000 iu weekly (range, 20 000–80 000) for EA and 200 mcg every two weeks (range, 100–500), for DA (Figure S1A). Dynamic dosing alterations due to toxicity or response could not be reliably ascertained due to the retrospective nature of this study. Median treatment duration was 14 months (range, 1–172) and causes for discontinuation included ESAs failure/loss of response in 21 (53%), sustained Hb rise in one (3%), and dose‐limiting (≥grade 3) adverse events (AEs) in four (10%) patients. These dose‐limiting AEs included treatment emergent hypertension in two [5%, one at Hb 10·8 (increase from 10·2 at baseline) gm/dl and platelet count 1193 (increase from 481 at baseline) × 10⁹/l and other at Hb of 7·6 (decrease from 9 at baseline) gm/dl with platelet count unknown (472 × 10⁹/l at baseline)], venous thromboembolism [at Hb 9·1 (increase from 8·6 at baseline) gm/dl, platelet count 318 (decrease from 537 at baseline) × 10⁹/l], ischemic stroke [at Hb 8·5 (increase from 7·7 at baseline) gm/dl, platelet count 569 (increase from 492 at baseline) × 10⁹/l] and splenic infarction (Hb and platelet count unknown) in one (3%) patient each. Overall, aspirin use was documented in 17 (43%) patients [14 (35%) at 81 mg dose and three (8%) at 325 mg dose] and all patients with thrombotic/ischemic events (n = 3) had been on low‐dose aspirin prophylaxis. The start and indication of aspirin therapy could not be reliably assessed in our cohort, which prevents us from recommending routine aspirin prophylaxis for all MDS/MPN‐RS‐T patients. Two (5%) patients transformed to acute myeloid leukemia (AML) two and five years after the last dose of ESA (NGS testing results not available).

Erythroid responses, as adjudicated by the 2015 IWG MDS/MPN overlap syndrome response criteria, were assessable in 38 (95%) patients; 18 (45%) met the criteria for an erythroid response, with the median duration of response being 20 months (range, 2–172). There was a trend towards a lower baseline EPO level in patients who responded to EPO, in comparison to non‐responders (median, 29 iu/l vs. 112 iu/l, P = 0·08). After a receiver‐operating characteristic (ROC) analysis, patients with an EPO level of ≤44 iu/l were most likely to achieve erythroid responses to ESA (P = 0·03*, Figure S1B), while pretreatment RBC transfusion independence was not predictive of response (P = 0·3, Figure S1C). Age (P = 0·33), gender (P = 0·2), baseline white blood cell count (P = 0·67), haemoglobin (P = 0·96), and platelet counts (P = 0·45), BM RS% (P = 0·84), leukemic transformation rates (P = 0·9) and mortality rates (P = 0·62) did not differ between ESA responders and non‐responders. The distribution of molecular aberrations was also not different between responders and non‐responders (details in Table 1). While there was a trend towards a better Kaplan–Meier estimate of overall survival (OS) in ESA responders versus non‐responders, this did not reach statistical significance (median OS, 76 months vs. 46 months; P = 0·7, Figure S1D). After ESA failure, seven (18%) patients received hypomethylating agents (HMAs ‒ prior to therapy, a bone marrow biopsy was available for four patients and none had evidence for disease progression/excess blasts), while five (13%) received lenalidomide [none with del(5q) abnormalities]. There were no erythroid responses to HMA, while one out of three (33%) assessable lenalidomide treated patients achieved a morphological complete response with transfusion independence. Due to the limited numbers in our cohort, we separately assessed a cohort of 23 ESA‐treated MDS/MPN‐RS‐T patients from the H. Lee Moffitt Cancer Center for validation purposes, and found a comparable erythroid response rate in 13 (57%) patients (details in Table SI). Due to limited clinical annotation (absence of important clinical parameters such as baseline EPO level), this cohort was not combined with the Mayo Clinic cohort.

Several clinical trials studying ESA therapy among MDS patients have reported 18–45% erythroid response rates among patients with MDS‐RS (Ferrini et al., 1998; Greenberg et al., 2009; Platzbecker et al., 2017b; Fenaux et al., 2018). However, data specifically for patients with MDS/MPN‐RS‐T are limited (Patnaik & Tefferi, 2019). While ESA therapy has long been used to manage anaemia in MDS/MPN‐RS‐T, ours is the first study to meticulously document ESA response rates in these patients by using the IWG MDS/MPN overlap syndrome response criteria (Savona et al., 2015). Encouragingly, MDS/MPN‐RS‐T patients treated with ESA had an erythroid response rate of 45%, with low endogenous EPO level (≤44 iu/l) being the best and only predictive factor of response. Unlike in MDS, acknowledging limitations related to smaller patient numbers, a lower pretreatment RBC transfusion need (≤2 units per month) did not predict response to ESA therapy (Hellstrom‐Lindberg et al., 1998; Hellstrom‐Lindberg et al., 2003).

There are distinct clinical and biological differences among patients with MDS‐RS and MDS/MPN‐RS‐T, such as the presence of thrombocytosis, a higher degree of BM fibrosis, and a higher risk of developing arterial and venous thrombosis in the latter group (Broseus et al., 2012; Patnaik et al., 2016b). Although controversial, there is evidence supporting an increased risk of vascular events and thrombosis with ESA use among patients with MDS (Steurer et al., 2003; Niazy et al., 2008). This raises a concern whether ESA use can compound an already elevated thrombosis risk in patients with MDS/MPN‐RS‐T. In our cohort, three patients developed vascular events (venous thrombosis, splenic infarction and ischemic stroke) despite being on low dose aspirin therapy. Given the retrospective nature of this study, further inferences on causality and association with ESA use are limited. However, future prospective randomised studies are necessary to assess for safety and efficacy. The advent of novel therapeutic agents, such as luspatercept which acts on late stages of erythropoiesis through the TGF‐beta pathway also bring further hope to patients with MDS/MPN‐RS‐T (Platzbecker et al., 2017a). Future studies, both in ESA refractory/ineligible patients and in the front line setting are needed to see if these drugs could offer safer and more effective alternatives.

In summary, ESA are effective first line therapies for the management of anaemia in patients with MDS/MPN‐RS‐T, with 45% achieving an erythroid response (median duration of response of 20 months). Low baseline endogenous EPO level (≤44 iu/l) was found to be the best and only predictor of response. Erythroid responses did not translate into clear survival benefits in affected patients. Future prospective studies are needed to document the safety of these agents, especially related to thrombosis/vascular adverse events.

致编辑

在2016年世界卫生组织（WHO）的髓类肿瘤分类中，骨髓增生异常综合征/骨髓增生性肿瘤伴有环铁粒母细胞和血小板增多症（MDS / MPN-RS-T）被列为独特的实体，由红系的存在定义存在或不存在多谱系发育异常和持续性血小板增多症（血小板计数≥450×10 ⁹ / l）的谱系发育不良，而没有其他定义疾病的遗传异常（Arber et al。，2016）。MDS / MPN‐RS‐T患者的频繁体细胞改变包括SF3B1（85％），JAK2V617F（50％），TET2（25％），ASXL1（20％），DNMT3A（15％）和SETBP1（10％）（Patnaik等，2016a）。估计中位总生存期约为76个月，其结局优于MDS-RS和单谱系发育异常的患者，但与原发性血小板增多症患者相比则差（Broseus等人，2012年）。

由于与线粒体中细胞内铁的锁定相关的无效的红细胞生成和加速的类红血球前体细胞凋亡，贫血是发病的主要来源，不仅是由于与氧气运输受损相关的并发症，而且是由于红细胞（RBC）输血相关的铁超载（Patnaik＆Tefferi，2019）。促红细胞生成促进剂（ESA）通常用于逆转这种红系成熟停滞并降低RBC输注频率，但有关MDS / MPN-RS-T患者特定反应率的文献有限（Patnaik＆Tefferi，2019）。这尤其重要，因为MDS / MPN-RS-T患者已经增加了血栓形成的风险，可能会伴随使用ESA（Broseus等人，2012）。我们进行了一项单一机构回顾性研究，以评估WHO定义的MDS / MPN-RS-T患者的ESA反应。

经机构审查委员会批准后，从2002年至2014年在我们机构诊断出的具有WHO定义的MDS / MPN-RS-T的成年患者被纳入研究。审查了诊断性骨髓（BM）抽吸和曲华因活检的形态和细胞遗传学，以确保符合最新的WHO标准（Arber等，2016）。二十四（60％）的研究患者接受下一代测序（NGS），用于下列基因：IKZF1，PTPN11，SH2B3，SUZ12，ZRSR2，ASXL1，CALR，CBL（外显子8，9），CEBPA（外显子1），CSF3R（第14、17外显子），DNMT3A（第4、8、13、15、16、18、19、20、22和23外显子），EZH2（第8、17和18外显子），FLT3（第14外显子），IDH1（第4外显子），IDH2（第4外显子），JAK2（第12外显子），KIT，MPL（第10外显子），NPM1（第11外显子），NRAS（第2外显子3），RUNX1（外显子3、4和8），SETBP1（外显子3），SF3B1（外显子13、15和17），SRSF2（外显子1），TET2（外显子3、9、10、11），TP53（外显子5、6 ，7和8）和BM单核细胞上诊断或首次转诊时通过先前描述的方法获得的U2AF1（外显子2、6）（Patnaik等，2016a）。回顾性分析治疗细节，包括使用的ESA类型，给药剂量，副作用和停药原因。使用2015 IWG（国际工作组）MDS / MPN响应标准（Savona等，2015）评估类红细胞响应。

研究纳入了四十名世卫组织定义的MDS / MPN-RS-T患者，他们在病程中随时接受了ESA治疗。中位年龄72·1岁（范围52–93·3），男性45％。表1描述了这些患者的临床和实验室特征。诊断时的中值血红蛋白（Hb）为9·2 gm / dl（范围：6·6-12·1）；中位基线Hb水平（ESA治疗之前）为9 gm / dl（范围为6·6-12·1 six由于最近的RBC输血，六名患者的Hb水平虚高，转诊前尚无水平）和中位基线促红细胞生成素（EPO）水平（15例患者中有respond 9位反应者，6位无反应者）44 iu / l（范围8–500）。17名（43％）患者在诊断时进行了骨髓网蛋白染色评估，其中7名（41·2％）患者显示出骨髓纤维化的证据（6级，1级）1年级2）。诊断时LDH中位数为152·5（范围：145–270）U / l；但是，此信息仅适用于四名患者。仅在三名（7·5％）患者中观察到异常核型，其异常包括der（13; 14）（q10; q10），-Y和inv（3）（q21; q26）。在开始ESA治疗之前，有11名（31％）患者为RBC输血依赖性（TD）。40例接受ESAs治疗的患者中，没有一个接受过细胞减灭疗法治疗血小板增多症。21例（53％）患者接受了epoietin-α（EA）治疗，15例（38％）接受了darbepoietin-α（DA）治疗，4例（10％）依次接受了这两种药物。EA的中位给药剂量为每周40,000 iu（范围为20 000至80 000），DA的中位给药剂量为每两周200 mcg（范围为100至500）（图S1A）。由于这项研究的回顾性，不能可靠地确定由于毒性或反应引起的动态剂量变化。中位治疗时间为14个月（范围1至172），中止治疗的原因包括ESA失败/应答丧失21（53％），持续血红蛋白升高1（3％）和剂量限制（≥3级）四（10％）位患者的不良事件（AEs）。这些限制剂量的AE包括两种治疗突发性高血压[5％，一种为Hb 10·8（基线时从10·2增加），gm / dl和血小板计数1193（基线时为481增加）×10⁹ / l，其他在Hb为7·6（从基线的9降低）到gm / dl，血小板计数未知（基线为472×10 ⁹ / l）]，静脉血栓栓塞[在Hb 9·1，从8·基线为6）gm / dl，血小板计数318（基线为537减少）×10 ⁹ / l]，缺血性卒中[Hb 8·5（基线为7·7升高），克/分升，血小板计数569（每位患者（3％）的脾脏梗塞（血红蛋白和血小板计数未知）从基线时的492×10 ⁹ / l]增加）。总体而言，有17例（43％）患者使用阿司匹林的记录[81 mg剂量时为14（35％），325 mg剂量时为3（8％）]和所有有血栓/缺血事件的患者（n = 3）曾接受小剂量阿司匹林预防。在我们的队列中无法可靠地评估阿司匹林治疗的开始和适应症，这使我们无法建议对所有MDS / MPN-RS-T患者进行常规阿司匹林预防。在最后一剂ESA注射后两年和五年，有两名（5％）患者转变为急性髓细胞性白血病（AML）（尚无NGS检测结果）。

根据2015年IWG MDS / MPN重叠综合征反应标准判定，类红细胞反应在38例（95％）患者中得到评估；18名（45％）达到了类红细胞反应的标准，中位反应时间为20个月（范围2–172）。与无反应者相比，对EPO有反应的患者的基线EPO水平有降低的趋势（中位数为29 iu / l对112 iu / l，P  = 0·08）。经过接受者操作特征（ROC）分析后，EPO水平≤44iu / l的患者最有可能实现对ESA的类红血球反应（P  = 0·03 *，图S1B），而治疗前RBC的输血独立性则没有预测响应（P  = 0·3，图S1C）。年龄（P  = 0·33），性别（P = 0·2），基线白细胞计数（P  = 0·67），血红蛋白（P  = 0·96）和血小板计数（P  = 0·45），BM RS％（P  = 0·84）， ESA应答者和非应答者之间的白血病转化率（P  = 0.9）和死亡率（P = 0.62）没有差异。响应者和非响应者之间的分子像差分布也没有差异（详情见表1）。尽管有一种趋势是更好地估计ESA应答者相对于非应答者的总体生存期（OS），但Kaplan–Meier估计值没有统计学意义（中位OS分别为76个月和46个月；P = 0·7，图S1D）。ESA失败后，有七名（18％）患者接受了低甲基化药物治疗（HMAs治疗前，有四名患者进行了骨髓活检，没有证据表明疾病进展/过度爆炸），而五名（13％）接受来那度胺[没有del（5q）异常]。没有对HMA的类红血球反应，而三分之二（33％）可评估来那度胺治疗的患者达到了形态完全反应，且具有输血独立性。由于我们队列中的人数有限，我们分别对H.Lee Moffitt癌症中心的23名经ESA治疗的MDS / MPN-RS-T患者进行了评估，以进行验证，并在13例中发现了类似的类红细胞反应率（57 （％）患者（表S1中的细节）。

一些在MDS患者中研究ESA治疗的临床试验报告说，MDS-RS患者中的类红素反应率为18–45％（Ferrini等，1998； Greenberg等，2009； Platzbecker等，2017b； Fenaux等。 ，2018）。但是，专门针对MDS / MPN‐RS‐T患者的数据有限（Patnaik＆Tefferi，2019）。虽然ESA治疗长期以来一直用于治疗MDS / MPN-RS-T中的贫血，但我们的研究是第一项通过使用IWG MDS / MPN重叠综合征反应标准来精确记录这些患者的ESA反应率的研究（Savona等，2015年）。令人鼓舞的是，接受ESA治疗的MDS / MPN-RS-T患者的类红细胞反应率为45％，低内源性EPO水平（≤44iu / l）是最好且唯一的预测反应因素。与MDS不同的是，由于认识到局限性在于患者人数较少，因此较低的RBC输血预处理需求（每月≤2个单位）不能预测对ESA治疗的反应（Hellstrom-Lindberg等，1998； Hellstrom-Lindberg等，2003）。）。

MDS-RS和MDS / MPN-RS-T患者之间存在明显的临床和生物学差异，例如存在血小板增多症，BM纤维化程度较高，以及后者发生动脉和静脉血栓形成的风险较高（Broseus等人，2012； Patnaik等人，2016b）。尽管存在争议，但有证据支持在MDS患者中使用ESA会增加血管事件和血栓形成的风险（Steurer等，2003； Niazy等，2008）。）。这引起了人们的担忧，即在MDS / MPN-RS-T患者中使用ESA是否会增加已经升高的血栓形成风险。在我们的队列中，尽管接受低剂量阿司匹林治疗，但三名患者仍发生了血管事件（静脉血栓形成，脾梗塞和缺血性中风）。考虑到这项研究的回顾性，对因果关系以及与ESA使用的关联的进一步推论是有限的。但是，未来的前瞻性随机研究对于评估安全性和有效性是必要的。新型治疗药物的出现，例如通过TGF-β途径作用于红细胞生成后期的luspatercept，也为MDS / MPN-RS-T患者带来了更多希望（Platzbecker等，2017a）。需要在ESA难治性/不合格患者以及前线患者中进行进一步的研究，以了解这些药物是否可以提供更安全，更有效的替代药物。

总之，ESA是治疗MDS / MPN-RS-T患者贫血的有效一线疗法，其中有45％的患者达到红系反应（中位反应时间为20个月）。基线内源性EPO水平低（≤44iu / l）被认为是最佳且唯一的反应预测因子。在受影响的患者中，类红血球反应并未转化为明显的生存获益。需要进一步的前瞻性研究来证明这些药物的安全性，尤其是与血栓形成/血管不良事件有关的安全性。