Up-regulation of inhibitory natural killer receptors CD94/NKG2A with suppressed intracellular perforin expression of tumor-infiltrating CD8+ T lymphocytes in human cervical carcinoma

Inhibitory signals that govern the cytolytic functions of CD8(+) T lymphocytes have been linked to the expression of natural killer cell receptors (NKRs) on CTLs. There is limited knowledge about the induction of inhibitory NKR (iNKR) expression in vivo. Up-regulation of iNKRs has been linked to the modulation of the virus- and/or tumor-specific immune responses in animal models. In the present study, we directly examined the expression of various NKRs on tumor-infiltrating lymphocytes (TILs) derived from human cervical cancer. We found that in human cervical cancer, the percentage expression of immunoglobulin-like NKR(+)CD8(+) T lymphocytes were similar in gated CD8(+)-autologous TILs and peripheral blood mononuclear cells. On the contrary, cervical cancer-infiltrating CD8(+) T lymphocytes expressed up-regulated C-type lectin NKRs CD94/NKG2A compared with either peripheral blood CD8(+) T cells or normal cervix-infiltrating CD8(+) T lymphocytes. Dual NKR coexpression analyses showed that CD94 and NKG2A were mainly expressed on CD56(-)CD161(-)CD8(+) TILs within the cancer milieu. Immunohistochemical study showed that cervical cancer cells expressed abundant interleukin 15 (IL-15) and transforming growth factor-beta (TGF-beta). In kinetic coculture assay, cervical cancer cells can promote the expression of CD94/NKG2A on CD8(+) T lymphocytes. The cancer-derived effects can be reversed by addition of rIL-15Ralpha/Fc and anti-TGF-beta antibody. Functional analyses illustrated that intracellular perforin expression of CD8(+) T cells was minimal upon up-regulation of CD94/NKG2A. Kinetic cytotoxicity assays showed that up-regulated expressions of CD94/NKG2A restrain CD8(+) T lymphocyte cytotoxicity. Our study strongly indicated that cervical cancer cells could promote the expression of iNKRs via an IL-15- and possibly TGF-beta-mediated mechanism and abrogate the antitumor cytotoxicity of TILs.     

Lysis of fresh human tumor cells by autologous large granular lymphocytes from peripheral blood and pleural effusions

Human lymphocytes and their subpopulations from the peripheral blood and pleural effusions of cancer patients were tested for cytotoxicity against fresh tumor cells isolated from carcinomatous pleural effusions of the same patients. Fresh tumor cells were relatively resistant to lysis by autologous unseparated lymphocytes in a 4 h Cr-release assay. Positive reactions were recorded in 10 of 38 blood samples and 10 of 37 effusion specimens. Purification of large granular lymphocytes (LGL) by discontinuous Percoll gradient centrifugation resulted in enhancement of cytotoxic activity against autologous tumor cells and K562 cells, with no reactivity in LGL-depleted small T-lymphocyte populations. Significant lysis of effusion tumor cells by autologous LGL was observed in 15 of 22 blood specimens and 15 of 21 effusion samples. Further depletion of high-affinity sheep erythrocyte rosetting cells from Percoll-purified LGL populations gave an increase in autologous tumor-killing activity. Depletion of LGL/K562 conjugates from LGL populations decreased lysis of autologous tumor cells and K562 cells. Effusion tumor cells that were susceptible to lysis by allogeneic normal LGL were also killed by autologous LGL, and effusion tumor cells resistant to lysis by allogeneic NK cells were not lysed by autologous LGL. In a single-cell cytotoxicity assay in agarose, 4-26% LGL bound autologous tumor cells and 0.2-5% LGL killed these target cells, while 12-45% LGL bound K562 cells and 2-20% LGL lysed them. These results indicate that cytotoxic potential for autologous effusion tumor cells is present in the peripheral blood and pleural effusions of cancer patients; it is strongly associated with a minor proportion of LGL and restricted to the cell population that can lyse NK-sensitive K562 cells.     

Lymphokine-activated Killer Induction and Its Regulation by Macrophages in Malignant Pleural Effusions

Mononuclear cells (MNC) from pleural effusions and peripheral blood of 18 patients with primary lung cancer with malignant pleural effusion were studied. Pleural and blood MNC generated lymphokine-activated killer (LAK) activity similarly when cultured for 4 days with an optimal concentration of interleukin 2 (IL-2). Highly purified lymphocytes (>98%) and monocyte-macrophages (>90%) were isolated by discontinuous Percoll gradient centrifugation from pleural and blood MNC. Pleural macrophages, as well as blood monocytes, showed significant augmenting effects on in vitro LAK cell induction from pleural and blood lymphocytes by IL-2. During daily intrapleural administration of IL-2, significant induction of LAK activity in vivo was observed after 3 days, but then this LAK activity in pleural MNC decreased almost to zero by day 15. Daily injections of IL-2 resulted in reduction in the up-regulation of LAK induction by pleural macrophages and also in increase in the levels of soluble IL-2 receptors in pleural effusions. These findings indicate that in vivo LAK induction of lymphocytes in malignant effusions by IL-2 may be regulated by macrophages in the effusions.     

Natural killer and NK-Like T-cell activation in colorectal carcinoma patients treated with autologous tumor-derived heat shock protein 96

Heat shock proteins (HSPs) are involved in the activation of both adaptive and innate immune systems. Here, we report that vaccination with autologous tumor-derived HSP96 of colorectal cancer patients, radically resected for liver metastases, induced a significant boost of natural killer (NK) activity detected as cytokine secretion and cytotoxicity in the presence of NK-sensitive targets. Increased NK activity was associated with a raise in CD3-CD56+ NK and/or CD3+CD56+ NK-like T cells, displaying enhanced expression of NKG2D and/or NKp46 receptors. Up-regulated expression of CD83 and CD40 and increased interleukin-12 release on stimulation were observed in CD14+ cells from post-HSP96 peripheral blood mononuclear cells, suggesting an indirect pathway of NK stimulation by HSP96-activated monocytes. Additionally, CD3-CD56+ and CD3+CD56+ lymphocytes were found to undergo functional and phenotypic activation on in vitro exposure to HSP96 even in the absence of monocytes, supporting a potential direct activity of HSP96 on these cell subsets. This evidence was confirmed by the specific binding of FITC-conjugated HSP96 to a subset of both CD3-CD56+ and CD3+CD56+ cells in peripheral blood mononuclear cells from colorectal cancer patients. Altogether, these findings identify the activation of the NK compartment as an additional immunologic effect of autologous tumor-derived HSP96 administration in cancer patients.     

Lymphokine-activated killer induction and its regulation by macrophages in malignant pleural effusions

Mononuclear cells (MNC) from pleural effusions and peripheral blood of 18 patients with primary lung cancer with malignant pleural effusion were studied. Pleural and blood MNC generated lymphokine-activated killer (LAK) activity similarly when cultured for 4 days with an optimal concentration of interleukin 2 (IL-2). Highly purified lymphocytes (greater than 98%) and monocyte-macrophages (greater than 90%) were isolated by discontinuous Percoll gradient centrifugation from pleural and blood MNC. Pleural macrophages, as well as blood monocytes, showed significant augmenting effects on in vitro LAK cell induction from pleural and blood lymphocytes by IL-2. During daily intrapleural administration of IL-2, significant induction of LAK activity in vivo was observed after 3 days, but then this LAK activity in pleural MNC decreased almost to zero by day 15. Daily injections of IL-2 resulted in reduction in the up-regulation of LAK induction by pleural macrophages and also in increases in the levels of soluble IL-2 receptors in pleural effusions. These findings indicate that in vivo LAK induction of lymphocytes in malignant effusions by IL-2 may be regulated by macrophages in the effusions.     

抑制性杀伤细胞受体在NK细胞和T细胞上的表达及其与急性移植物抗宿主病的关系

 目的 研究异基因造血干细胞移植（allo-HSCT）患者移植前后外周血NK及T细胞上4种抑制性杀伤细胞受体（CD158a、CD158b、NKB1和CD94／NKG2A）的表达及其与急性移植物抗宿主病（aGVHD）的关系。方法 采用流式细胞术检测NK及T细胞上抑制性杀伤细胞受体的表达。结果 NK细胞上CD158a和CD158b的表达于移植后3～4个月、NKB1和CD94／NKG2A的表达于移植后2个月恢复移植前水平。移植前后CD158a和NKB1在CD+3 T细胞上持续低水平表达;移植前CD158b和CD94／NKG2A在CD+3 T细胞上的表达水平较高,且主要表达于CD+8 T细胞上,移植后其在CD+8 T细胞上的表达增加。CD+8 T细胞上CD158b的表达在发生Ⅰ度aGVHD时显著增高,与无aGVHD组及Ⅱ~Ⅳ度aGVHD组相比,差异均有统计学意义（P均＜0.05）;在发生Ⅱ~Ⅳ度aGVHD时增高不明显,与无aGVHD组相比,差异无统计学意义（P＞0.05）。结论 CD158b在CD+8 T细胞上高表达可能有助于降低T细胞同种反应性,减轻aGVHD的程度。     

食管鳞癌患者外周血及癌组织中浸润性NK细胞表面受体的表达

目的 分析食管鳞癌患者外周血及组织中NK细胞的比例及其表面受体的表达。方法 流式细胞仪检测食管鳞癌患者外周血及组织中NK细胞的比例及其表面受体的表达。结果 食管鳞癌组织中NK细胞比例明显高于癌旁5 cm以远的非肿瘤正常组织。CD16在食管鳞癌患者外周血中的表达与健康者外周血并无明显差异，但在鳞癌组织中的表达则明显低于正常组织；癌组织中NKp30、NKG2D的表达均明显低于正常组织；NKp44的表达明显高于正常组织。在同一患者体内，与外周血相比，癌组织中的CD16、NKp30、NKG2D的表达更低；而NKp44、NKG2A的表达更高。外周血与癌组织中浸润的NK细胞数量与肿瘤进展呈负相关。结论 食管鳞癌组织中NK细胞表面受体表达失衡可介导食管鳞癌细胞发生免疫逃逸。     

Lysis of fresh human tumor cells by autologous large granular lymphocytes and T-lymphocytes: two distinct killing activities induced by coculture with autologous tumor

The specific and natural killer (NK)-restricted nature of autologous tumor killing by blood lymphocytes was studied in patients with carcinomatous pleural effusions. Large granular lymphocytes (LGL) and small T-lymphocytes were isolated by centrifugation on discontinuous Percoll density gradients. Tumor cells freshly isolated from pleural effusions of cancer patients were classified according to their susceptibility to purified LGL from normal donors in a 4-hour 51Cr release assay. Of 15 NK-sensitive tumors, 14 were lysed by fresh autologous LGL, whereas only 2 were killed by T-cells. Neither LGL nor T-cells were cytotoxic to NK-resistant autologous tumor. LGL and T-cells were then cultured in vitro with autologous tumor cells for 6 days. In 13 of 15 autologous mixed lymphocyte-tumor cultures (MLTC) NK-sensitive tumor-cultured LGL maintained their autotumor killing activity, whereas LGL cultured alone lost the activity. Depletion of high-affinity sheep erythrocyte-rosetting cells from Percoll-purified LGL resulted in an enrichment of effector cells. LGL from autologous MLTC were able to kill NK-susceptible allogeneic effusion tumor and K562 as were fresh LGL. No lysis of NK-resistant autologous tumor was observed with cultured LGL. In contrast, activation of T-cells in autologous MLTC resulted in the generation of autotumor killer cells in 10 of 15 NK-sensitive and 3 of 6 NK-resistant tumor samples. However, cultured T-cells were incapable of killing allogeneic tumor and K562. In autologous MLTC T-cells proliferated in response to autologous tumor, whereas no proliferation was observed in the culture of LGL. The enrichment of blasts from cultured T-cells on discontinuous Percoll gradients induced an augmentation of autotumor cytotoxicity, with no reactivity in blast-depleted, small, resting T-lymphocytes. These results indicated that 2 distinct types of autotumor-recognizing lymphocytes, LGL and T-cells, are present in the peripheral blood of cancer patients.     

非小细胞肺癌EGFR基因突变异质性研究进展

 表皮生长因子受体（EGFR）酪氨酸激酶抑制剂（TKIs）在EGFR基因活化突变阳性晚期非小细胞肺癌（NSCLC）患者中疗效显著,然而,同样是EGFR活化突变阳性的NSCLC患者接受EGFR-TKIs治疗后,疗效仍有明显差异;同一个患者的不同瘤灶也会对EGFR-TKIs出现不同的反应,这些现象可能与EGFR基因突变异质性有关,同一瘤灶的不同部分、同一患者的不同瘤灶、以及同一瘤灶治疗前后EGFR突变状态都有可能不一致。本文将从这三个方面对EGFR突变异质性研究进展作一综述。     

The clonogenic potential of selected CD34+ cells from patients with MDS appear preserved when tested ex vivo

Our aim was to examine in 17 patients with MDS the effects of PMA activated and non-activated autologous lymphocytes on selected bone marrow CD34+ progenitors, in dose response studies. We used a double layer culture technique. Compared with controls, there was no difference in the colony growth promoting capacity of autologous PMA stimulated or unstimulated blood lymphocytes from MDS patients. In addition, similar to control studies, increasing numbers of lymphocytes, (0, 1 × 10⁵, 1 × 10⁶) led to a corresponding increase in the number of CFU-GM (p = 0.04). We conclude that MDS blood mononuclear cells have the ability to stimulate colony growth of autologous CD34+ cells while these selected progenitors show a proliferative capacity that is similar to normal when they are isolated from the bone marrow accessory cells.     

Apolipoprotein E expression promotes lung adenocarcinoma proliferation and migration and as a potential survival marker in lung cancer

Many human lung cancer cell lines express apolipoprotein E (ApoE), especially cells derived from malignant pleural effusions (MPE) in patients with lung adenocarcinoma. This study aimed to investigate the influence of ApoE expression on lung cancer. In lung cancer tissues, ApoE expression was more frequently found in malignant pleural effusions (MPE)-associated lung adenocarcinoma than in lung adenocarcinoma or squamous cell carcinoma without MPE (P < 0.05), indicating that ApoE is associated with the pathogenesis of MPE in patients with lung adenocarcinoma. Next, we examined the roles of ApoE in an MPE-derived lung adenocarcinoma cell line that endogenously over-expresses ApoE, PC14PE6/AS2 (AS2). In that experiment we inhibited ApoE expression by transfection of a plasmid carrying ApoE siRNAs into AS2 cells to generate AS-S2 and AS-S3 cells. Compared to vector-control cells and parental AS2 cells, AS2-S2 and AS2-S3 cells grew slower (P < 0.05), were more sensitive to cisplatin, and had significantly impaired cellular migration (P < 0.05). Furthermore, over-expression of ApoE was independently associated with poor survival in lung adenocarcinoma patients who had MPE at the time of diagnosis (P < 0.001). Conclusively, ApoE over-expression promotes cancer proliferation and migration and contributes to an aggressive clinical course in patients with lung adenocarcinoma and MPE.     

肺癌患者调节性T细胞与NK细胞活化受体NKG2D的相关性及其临床意义

目的 分析肺癌患者外周血中CD+4 CDHi25 CDLo127调节性T细胞及自然杀伤细胞(NK细胞)活化受体NKG2D的表达水平之间的关系,探讨其在肿瘤免疫逃逸机制中的作用及临床意义.方法 选择70例肺癌患者,均经病理确诊.采用流式细胞术(FCM)检测患者外周血中CD+4 CDHi25 CDLo127调节性T细胞、NK细胞及NKG2D表达水平,并以50名健康人为对照.结果 肺癌患者外周血中CD+4 CDHi25 CDLo127调节性T细胞比例较健康对照组明显升高[(8.4±4.1)％与(6.7±1.7)％],差异有统计学意义(t=3.09,P＜0.05);肺癌组NK细胞比例与健康对照组比较[(15.6±8.3)％与(17.2±4.2)％],差异无统计学意义(t=-1.33,P＞0.05);肺癌组NKG2D较健康对照组明显降低[(83.3±4.9)％与(87.4±2.9)％],差异有统计学意义(t=3.16,P＜ 0.05).CD+4 CDHi25 CDLo127调节性T细胞与NKG2D呈负相关性(r=-0.302,P＜0.05).结论 在肺癌患者外周血中调节性T细胞可能通过下调NKG2D,参与肿瘤免疫逃逸机制,二者可作为评估肺癌患者免疫功能状态及预后的参考指标.     

乳腺癌患者可溶性MICA对自然杀伤细胞受体的影响

目的观察乳腺癌患者外周血自然杀伤（NK）细胞杀伤活性及受体的变化,探讨可溶性MICA（sMICA）对NK细胞受体及杀伤活性的影响。方法ELISA法检测外周血血清sMICA的含量。流式细胞术（FCM）检测NK细胞百分比、NK细胞活化性受体NKG2D、抑制性受体KIR（CD158b）表达。MTT法检测NK细胞对乳腺癌细胞株MCF-7的杀伤活性。结果与健康人比较,乳腺癌患者中81．6％表达sMICA,含量为（205．36±71．27）ng／L,且sMICA含量与TNM分期呈正相关。乳腺癌患者外周血NK细胞所占百分比无明显差异,但血清sMICA阳性的乳腺癌患者中NK细胞杀伤活性明显降低,NKG2D表达下降,CD158b表达增高。当NK细胞培养体系中加入sMICA阳性的乳腺癌血清时,其杀瘤活性明显降低【（76．2±6．7）％与（48．4±4．1）％】,NKG2D的表达明显下调[（92．5±7．1）％与（62．5±6．4）％],而CD158b的表达明显上升【（10．6±3．2）％与（43．6±3．4）％】。sMICA阳性的乳腺癌患者NK细胞与细胞因子IL-15共培养,NK细胞的杀瘤活性、NKG2D的表达明显升高,KIR（CD158b）的表达明显下降。结论乳腺癌外周血血清中sMICA可通过下调NK细胞NKG2D表达以及上调KIR表达,降低NK细胞杀瘤活性。IL-15可逆转sMICA对NK细胞的免疫下调作用。     

Lysis of fresh human tumor cells by autologous peripheral blood lymphocytes and pleural effusion lymphocytes activated by OK432

Lymphocytes from peripheral blood (PBL) and from pleural effusions (PEL) of cancer patients were tested for cytotoxicity against tumor cells freshly isolated from carcinomatous pleural effusion of the same patient. Significant lysis of autologous tumor cells was recorded for 4 of 28 PBL samples and for 5 of 28 PEL cases when investigated in a 4-hour 51Cr release assay. In vitro treatment of lymphocytes for 20 hours with the streptococcal preparation OK432 resulted in an induction or augmentation of cytotoxicity against autologous tumor cells in 21 of 28 PBL and PEL specimens. OK432-induced cytotoxicity required active cell metabolism, RNA and protein syntheses, but not DNA synthesis of lymphocytes. Supernatants of OK432-stimulated lymphocytes, and interferon and interleukin 2 failed to induce autologous tumor killing. Nylon wool-nonadherent lymphocytes were involved in both spontaneous and OK432-induced lysis of fresh autologous tumor cells. OK432-activated lymphocytes from normal donors and cancer patients caused lysis of fresh allogeneic tumor cells and also K562 cells.     

晚期肺癌患者外周血Treg细胞与NK细胞及其活化受体检测临床意义的探讨

目的:观察CD4+CD25highCD127low调节性T细胞(Treg细胞)、NK细胞及其活化受体(NKG2D)在初诊晚期肺癌患者外周血中的变化,探讨它们在肺癌发病中的作用及临床意义。方法:流式细胞术检测90例初诊晚期肺癌患者及45位健康人外周静脉血CD4+CD25highCD127lowTreg细胞、NK细胞及NKG2D的水平,并就其中27例患者化疗前后数据对比。结果:与对照组比较,肺癌组CD4+CD25highCD127lowTreg细胞比例明显升高,t=3.500,P=0.001;NK细胞比例及NKG2D的表达明显降低,t值分别为-3.534和-5.228,P值均为0.001。Ⅳ期患者CD4+CD25highCD127lowTreg细胞比例较ⅢA期患者明显升高,F=3.657,P=0.030。化疗后患者NK细胞及NKG2D较化疗前明显降低,t值分别为3.176和2.771,P值分别为0.004和0.01。化疗前后患者CD4+CD25highCD127lowTreg细胞水平差异无统计学意义,t=1.419,P=0.168。肺癌患者外周血CD4+CD25highCD127lowTreg细胞比例与NK细胞比...     

The Putative Role of Natural Killer Cells in Patients with Hepatitis C Virus-Related Hepatocellular Carcinoma

Background: Natural Killer (NK) cells have crucial roles in immune responses against malignant transformation including hepatocellular carcinoma (HCC). The NKG2D receptor has a critical role in the NK recognition of target cells. Aim: We assessed NKG2D receptor expression as a diagnostic biomarker for HCC detection and progression in Egyptian patients with hepatitis C virus (HCV)-related HCC. Methods: We classified 81 patients into three groups: chronic hepatitis (21), cirrhotic (30) and HCC (30) patients, with 36 individuals enrolled to the control group. We analyzed NK levels in peripheral blood and NKG2D receptor expression in NK cells using flow cytometry. Results: We observed a significant decrease in NKG2D (CD314) expression on circulating NK cells and frequency of NK cells expressing NKG2D (CD314) in HCC patients. Also, in patients, larger foci lesions significantly correlated with decreased NK cell numbers. Multiple foci numbers and patients with a Child score C significantly correlated with decreased circulating NK cells expressing NKG2D and decreased NKG2D expression. Conclusion: The percentage of NK cells in peripheral blood and NKG2D receptor expression could function as potential biomarkers for HCC detection and progression.     

Clinical application of adoptive immunotherapy by cytotoxic T lymphocytes induced from tumor-infiltrating lymphocytes

The tumor-infiltrating lymphocytes (TIL) were cultured with interleukin 2 (IL-2) to induce the cytotoxic T lymphocytes possessing autologous tumor-killing activity from 21 cancer patients (11 with solid tumor and 10 with malignant peritoneal or pleural effusions), and transferred into 7 patients as IL-2-activated TIL adoptively. The clinical application of activated TIL by adoptive transfer could result the complete regression of malignant pleural effusions in a patient with pancreatic cancer, and the nearly complete regression of malignant ascites in a patient with gastric cancer. The autologous tumor cells were isolated at the purity of more than 90% by Ficoll-Hypaque and Percoll discontinuous gradients, and then the TIL were cultured with IL-2 until 4 weeks. The optimal concentration of IL-2 was 1,500 IU/ml to obtain maximum proliferation and autologous tumor killing activity. The cytotoxic activities of activated TIL at 3 weeks-incubation was 72 +/- 15, 42 +/- 26, 27 +/- 21 and 22 +/- 15% against K562, Daudi, KATO-III and autologous tumor, respectively. By negative selection method, it was clarified that the killer cells recognizing autologous tumor consisted of CD4 or CD8 positive T lymphocyte in 43% of patients. The CD8 positive cells and CD56 positive cells increased, the CD4 positive cells and CD16 positive cells decreased by flow cytometry. The activated TIL could lyse not only cultured tumor cell lines, also other autologous tumor cells. The CD56+ cells were isolated by the Panning method, these cells could not lyse autologous tumor cells. Thus, it was indicated that the cytotoxic T lymphocytes recognizing autologous tumor could be generated from TIL and the adoptive immunotherapy of activated TIL was effective in cancer therapy.     

The increase of CD57+ T cells in the peripheral blood and their impaired immune functions in patients with advanced gastric cancer

In addition to natural killer (NK) cells, T cells expressing natural killer cell markers, CD56 or CD57 (NK type T cells), have been considered to play an important role in antitumor immunity. We examined the proportion of NK cell and NK type T cell subsets in the peripheral blood from patients with gastric cancer. The IFN-gamma production capacity and population of cytoplasmic perforin positive cells in peripheral blood mononuclear cells (PBMC) were evaluated. Peripheral blood samples were obtained from 56 patients with gastric cancer and 21 healthy volunteers. The proportion of CD56- CD57+ T cells (CD57+ T cells) was significantly higher in advanced gastric cancer patients than those in healthy volunteers and patients with early stage gastric cancer, whereas no correlation was observed between the proportion of CD56+ T cells or NK cells and tumor progression. Furthermore, a significant decrease of CD8+ CD57+ T cells was found in patients with advanced gastric cancer. The proportion of CD57+ T cells did not correlate with interferon-gamma (IFN-gamma) production from PBMC in gastric cancer patients, although a significant correlation was found between them in healthy volunteers. The proportion of perforin positive CD57+ T cells, especially CD8+ CD57+ T cells, in patients with gastric cancer was markedly lower than that in healthy volunteers. Collectively, although the proportion of CD57+ T cells in PBMC was found to increase with tumor progression, their function in antitumor immunity is impaired in patients with gastric cancer.     

Up-regulation of activating and inhibitory NKG2 receptors in allogeneic and autologous hematopoietic stem cell grafts

Background

Hematopoietic Stem Cell Transplantation (HSCT) is known to induce the inhibitory immune receptor NKG2A on NK cells of donor origin. This occurs in allogeneic recipients, in both the haploidentical and HLA-matched settings.

Methods

To gain further insight, not only NKG2A, but also the activating receptors NKG2C and NKG2D were assessed by flow cytometry. Immunophenotyping was carried out not only on CD56⁺ but also on CD8⁺ lymphocytes from leukemia and lymphoma patients, receiving both HLA-matched (n = 7) and autologous (n = 5) HSCT grafts. Moreover, cognate NKG2 ligands (HLA-E, MICA, ULBP-1, ULBP-2 and ULBP-3) were assessed by immunohistochemistry in diagnostic biopsies from three autotransplanted patients, and at relapse in one case.

Results

All the NKG2 receptors were simultaneously up-regulated in all the allotransplanted patients on CD8⁺ and/or CD56⁺ cells between 30 and 90 days post-transplant, coinciding with, or following, allogeneic engraftment. Up-regulation was of lesser entity and restricted to CD8⁺ cells in the autotransplantation setting. The phenotypic expression ratio between activating and inhibitory NKG2 receptors was remarkably similar in all the patients, except two outliers (a long survivor and a short survivor) who surprisingly displayed a similar NKG2 activation immunophenotype. Tumor expression of 2 to 3 out of the 5 tested NKG2 ligands was observed in 3/3 diagnostic biopsies, and 3 ligands were up-regulated post-transplant in a patient.

Conclusions

Altogether, these results are consistent with a dual (activation-inhibition) NK cell re-education mode, an innate-like T cell re-tuning, and a ligand:receptor interplay between the tumor and the immune system following HSCT including, most interestingly, the up-regulation of several activating NKG2 ligands. Turning the immune receptor balance toward activation on both T and NK cells of donor origin may complement ex vivo NK cell expansion/activation strategies in unmanipulated patients.

Electronic supplementary material

The online version of this article (doi:10.1186/s13046-015-0213-y) contains supplementary material, which is available to authorized users.