Myeloid neoplasms with isolated deletion of the entire long arm del(16)(q11) are extremely rare, and only three adult cases have been reported (Rogers et al., 2017). Two patients were diagnosed as myelodysplastic syndrome (MDS) with del(16)(q11) and one patient gained del(16)(q11) during progression from MDS to AML (Rogers et al., 2017). We report an exceptionally rare pediatric case of de novo AML with isolated del(16)(q11) with basophilia and eosinophilia. There were marked dysplastic changes in neutrophils, basophils and erythroid precursors, most consistent with AML with myelodysplasia-related changes (AML-MRC). The striking dysplastic features include premature nuclear lobation in immature myeloid cells and basophils without basophilic granules. The agranular basophils were initially counted as atypical monocytes erroneously, which were only recognized after flow cytometric analysis. This case demonstrates the power of flow cytometry in the practice of hematology by identifying the dysplastic agranular basophils, which were not described previously.

A 15-year-old previously healthy male presented with progressive pallor and unintentional weight loss of 16 lb over 4 months. Complete blood count (CBC) revealed severe macrocytic anemia (hemoglobin: 6.1 g/dL, mean corpuscular volume: 106.4 fL), mild leukopenia (white blood cells [WBC]: 3.8 × 10⁹/L), and moderate thrombocytopenia (platelets: 75 × 10⁹/L). The initial WBC differential count on peripheral blood (PB) smear (aqueous Romanowsky stain) showed circulating blasts (15%), eosinophilia (15%, 570/μl), and relative monocytosis (22%). The majority of purported monocytes had atypical morphology.

Flow cytometric analysis of PB (Figure 1a) detected increased myeloblasts (red, 22.7%), eosinophilia (pink, 16.4%), basophilia (brown, 19.4%) with normal percentage of monocytes (dark green, 3.1%). Table 1 lists the detailed immunophenotype of different WBC subsets. The basophils were intermediate-sized with low side light scatter (SSC) and partially overlapped with the lymphocytes on the forward side scatter (FSC)/SSC plot. They had moderate CD45 (dimmer than lymphocytes and brighter than myeloblasts), and bright CD123 without HLA-DR (Han et al., 2008). Neutrophils (blue, 2.9%) had decreased SSC, consistent with hypo-granular. The flow cytometric finding was diagnostic for acute myeloid leukemia (AML) with eosinophilia and basophilia.

The findings of the flow cytometry prompted re-examination of the PB smear to identify the missed basophils. Typical basophils were not seen (Figure 1b, from a different patient). All basophils were counted as atypical monocytes initially due to lack of the typical basophilic cytoplasmic granules. Compared to a typical monocyte (Figure 1c), the basophils (Figure 1d–i) were smaller with a higher nuclear to cytoplasmic ratio, scant-to-moderate clear-to-pale cytoplasm with vacuoles, irregularly round-to oval, folded or multilobated nuclei with finely dispersed-to-condensed chromatin. Rare immature basophils contained inconspicuous nucleoli. Besides the lack of basophilic cytoplasmic granules, these basophils showed striking morphological similarity to normal basophils with respect to cell size, clear cytoplasm and lobulated nuclei. The varied nuclear shapes and heterogeneous levels of CD117 may reflect different maturation stages of basophils (Han et al., 2008).

Bone marrow (BM) aspirate smears were stained with Jenner-Giemsa method. Except rare immature and mature basophils contained chunky basophilic granules (Figure 1j–l), the majority basophils were agranular (Figure 1m). To further confirm the loss of granules in basophils, BM smears were stained with Diff-Quik stain (Figure 1n) and Romanowsky stain with Basofix. Both are alcohol-based stains to fix the basophilic granules, which may be washed away with an aqueous Romanowsky stain. Both methods revealed basophilic granules in control but not in most basophils of our patient. In conclusion, the lack of granules in basophils was verified by four different staining methods. Hypogranular or agranular basophils have not been described previously and are not recognized by most laboratory personnel.

Myeloblasts had large prominent nucleoli with varied morphological features, including prominent azurophilic granules with bilobed nuclei (Figure 1o), cytoplasmic blebs and/or Auer rods (Figure 1p), elongated nuclear grooves (Figure 1q). Marked dysplasia was observed in immature and mature neutrophilic granulocytes, eosinophils and erythroid precursors (Figure 1r–x). Ring sideroblasts were not seen. Megakaryocytes were decreased with unremarkable morphology on biopsy and were not seen on aspirate.

Karyotype was 46,XY,del(16)(q11.1)[16]/46,XY[4]. Interphase FISH showed one CBFB signal in 88.7% of nuclei on BM aspirate (Figure 1y. Break-apart probes, red: proximal; green: distal). A 50-gene next generation sequencing panel for myeloid neoplasms detected two novel missense mutations: EZH2 c.2084C>T p.S695L (VAF 19.0%), and ASXL1 c.4308C>G p.I1436M (VAF 47.2%). The EZH2 mutation occurred within the functional significant SET domain and is likely pathogenic. Mutations of ASXL1 and EZH2 are frequently detected in myeloid neoplasms, clonal hematopoiesis of indeterminate potential, and idiopathic hyper-eosinophilic syndrome.

This patient reached complete remission (CR) after induction therapy (AAML1031 without etoposide) and has remained in CR for 7 months. Adult patients with myeloid neoplasms with isolated del(16q) seemed to have worse progress but its prognostic value in children is not clear (Rogers et al., 2017).

AML with concurrent basophilia and eosinophilia is uncommon. Our case showed a novel combination of del(16q), ASXL1 and EZH2 mutations in a pediatric case of de novo AML with basophilia and eosinophilia. There was marked morphological dysplasia in the myeloid and erythroid lineages. Two prominent uncommon dysplastic features included premature nuclear lobation in immature neutrophils, and complete loss of the granules in most basophils. The lack of basophilic granules in basophils made them resemble atypical monocytes, causing erroneous WBC differential count initially. It is important to recognize this rarely described morphological dysplasia.

整个长臂 del(16)(q11) 孤立缺失的髓系肿瘤极为罕见，仅报告了 3 例成人病例（Rogers 等人，  2017 年）。两名患者被诊断为伴有 del(16)(q11) 的骨髓增生异常综合征 (MDS)，一名患者在从 MDS 进展为 AML 期间获得了 del(16)(q11)（Rogers 等人，  2017）。我们报告了一个异常罕见的新发 AML 儿科病例，伴有孤立的 del(16)(q11) 并伴有嗜碱性粒细胞增多和嗜酸性粒细胞增多。中性粒细胞、嗜碱性粒细胞和红系前体细胞有明显的异常增生变化，最符合伴有骨髓增生异常相关变化 (AML-MRC) 的 AML。显着的发育异常特征包括未成熟骨髓细胞中的过早核分叶和没有嗜碱性颗粒的嗜碱性粒细胞。无颗粒嗜碱性粒细胞最初被错误地计为非典型单核细胞，仅在流式细胞仪分析后才被识别。该病例通过识别发育不良的粒状嗜碱性粒细胞，证明了流式细胞术在血液学实践中的作用，这在以前没有描述过。

一名 15 岁以前健康的男性在 4 个月内出现渐进性苍白和 16 磅体重意外减轻。全血细胞计数 (CBC) 显示严重的大红细胞性贫血（血红蛋白：6.1 g/dL，平均红细胞体积：106.4 fL）、轻度白细胞减少症（白细胞 [WBC]：3.8 × 10 ⁹ /L）和中度血小板减少症（血小板： 75×10 ⁹ /L)。外周血 (PB) 涂片（水性 Romanowsky 染色）的初始 WBC 分类计数显示循环原始细胞 (15%)、嗜酸性粒细胞增多 (15%, 570/μl) 和相对单核细胞增多症 (22%)。大多数声称的单核细胞具有非典型形态。

PB 的流式细胞术分析（图 1a）检测到成髓细胞增加（红色，22.7%）、嗜酸性粒细胞增多（粉色，16.4%）、嗜碱性粒细胞增多（棕色，19.4%），单核细胞百分比正常（深绿色，3.1%）。表 1 列出了不同 WBC 亚群的详细免疫表型。嗜碱性粒细胞中等大小，具有低侧光散射 (SSC)，在前向散射 (FSC)/SSC 图上与淋巴细胞部分重叠。它们具有中等的 CD45（比淋巴细胞更暗，比成髓细胞更亮），以及没有 HLA-DR 的亮 CD123（Han 等人，  2008 年）。中性粒细胞（蓝色，2.9%）的 SSC 降低，与低颗粒一致。流式细胞仪检测结果可诊断急性髓性白血病 (AML) 伴嗜酸性粒细胞增多和嗜碱性粒细胞增多。

流式细胞术的发现促使重新检查 PB 涂片以识别漏诊的嗜碱性粒细胞。没有看到典型的嗜碱性粒细胞（图 1b，来自不同的患者）。由于缺乏典型的嗜碱性细胞质颗粒，所有嗜碱性粒细胞最初都被视为非典型单核细胞。与典型的单核细胞（图 1c）相比，嗜碱性粒细胞（图 1d-i）更小，核质比更高，细胞质稀少至中等，透明至苍白，有空泡，不规则圆形至椭圆形，折叠或多叶核，染色质分散至凝聚。罕见的未成熟嗜碱性粒细胞含有不显眼的核仁。除了缺乏嗜碱性细胞质颗粒外，这些嗜碱性粒细胞在细胞大小、清晰的细胞质和分叶状细胞核方面表现出与正常嗜碱性粒细胞惊人的形态相似性。 2008 年）。

用 Jenner-Giemsa 方法对骨髓 (BM) 抽吸涂片进行染色。除了罕见的未成熟和成熟嗜碱性粒细胞含有厚实的嗜碱性颗粒（图 1j-l）外，大多数嗜碱性粒细胞是无颗粒的（图 1m）。为了进一步确认嗜碱性粒细胞中颗粒的损失，BM涂片用Diff-Quik染色（图1n）和Basofix染色的Romanowsky染色。两者都是用于固定嗜碱性颗粒的酒精类染色剂，可以用水性 Romanowsky 染色剂将其洗掉。两种方法都显示对照中有嗜碱性颗粒，但我们患者的大多数嗜碱性粒细胞中没有。总之，通过四种不同的染色方法证实了嗜碱性粒细胞中缺乏颗粒。以前没有描述过颗粒不足或无颗粒的嗜碱性粒细胞，并且大多数实验室人员不认识。

成髓细胞具有大而突出的核仁，具有不同的形态特征，包括具有双叶核的突出嗜天青颗粒（图 1o）、细胞质泡和/或 Auer 棒（图 1p）、细长的核凹槽（图 1q）。在未成熟和成熟的中性粒细胞、嗜酸性粒细胞和红系前体细胞中观察到明显的发育异常（图 1r-x）。未见环状铁粒幼细胞。巨核细胞减少，活检形态不明显，抽吸物未见。

核型为 46,XY,del(16)(q11.1)[16]/46,XY[4]。间期 FISH 在 BM 抽吸物中 88.7% 的细胞核中显示一个CBFB信号（图 1y。分离探针，红色：近端；绿色：远端）。用于骨髓肿瘤的 50 基因下一代测序面板检测到两个新的错义突变：EZH2 c.2084C>T p.S695L (VAF 19.0%) 和 ASXL1 c.4308C>G p.I1436M (VAF 47.2%)。EZH2突变发生在功能重要的 SET 域内，并且可能是致病的。ASXL1和EZH2的突变经常在骨髓肿瘤、不确定的克隆性造血和特发性高嗜酸性粒细胞综合征中检测到。

该患者在诱导治疗（不含依托泊苷的 AAML1031）后达到完全缓解 (CR)，并在 CR 中保持了 7 个月。患有孤立性 del(16q) 的髓系肿瘤成年患者的进展似乎更糟，但其在儿童中的预后价值尚不清楚 (Rogers et al.,  2017 )。

并发嗜碱性粒细胞增多症和嗜酸性粒细胞增多症的 AML 并不常见。我们的案例显示了一种新的 del(16q)、ASXL1和EZH2突变组合，在一个伴有嗜碱性粒细胞和嗜酸性粒细胞增多的新发 AML 儿科病例中。在髓系和红系谱系中存在明显的形态发育异常。两个突出的不常见的发育异常特征包括未成熟中性粒细胞的过早核分叶和大多数嗜碱性粒细胞的颗粒完全丧失。嗜碱性粒细胞中缺乏嗜碱性颗粒使它们类似于非典型单核细胞，最初导致错误的 WBC 分类计数。重要的是要认识到这种很少描述的形态发育异常。