|
  
- 积分
- 723
- 威望
- 723
- 包包
- 1773
|
1 f* l; Q) ?( l1 M5 P: ~
粗略翻译了一下，不准确的地方还请各位网友指正！- C7 o! w: M! _* w1 h7 |+ U8 m
( v1 I: I1 K1 h8 A. R
Cytokine-Induced NK-Like T Cells: From Bench to Bedside
# a Z% | t. p9 x& D9 [( d" x细胞因子诱导NK样T淋巴细胞（CIK细胞）：从实验室到临床
, }" ^9 w: s+ \6 E
% W! d/ _$ S9 }. M TCytokine-induced killer (CIK) cells are polyclonal T effector cells generated when cultured under cytokine stimulation. CIK cells exhibit potent, non-MHC-restricted cytolytic activities against susceptible tumor cells of both autologous and allogeneic origins. Over the past 20 years, CIK cells have evolved from experimental observations into early clinical studies with encouraging preliminary efficacy towards susceptible autologous and allogeneic tumor cells in both therapeutic and adjuvant settings. This paper is our attempt to summarize the available published literature related to CIK cells. Looking into the future, we anticipate that the continuous therapeutic application of CIK cells will likely be developed along two major directions: overcoming the challenge to organize large prospective randomized clinical trials to define the roles of CIK cells in cancer immunotherapy and expanding its spectrum of cytotoxicity towards resistant tumor cells through experimental manipulations.
/ B3 J2 j# p7 U* k5 S细胞因子诱导杀伤性细胞（CIK细胞）是多克隆效应T淋巴细胞在细胞因子诱导下产生的一群异质细胞。CIK细胞有强大的抗瘤活性，非MHC限制性杀瘤优点，对易感的自体和异体肿瘤细胞都有很强的杀伤活性。在过去20年中，无论是从实验观察还是到早期的临床研究，都显示了CIK细胞作为辅助治疗方法可以有效地杀伤易感的自体和异体肿瘤细胞。这份报告的目的意在探讨关于CIK细胞的有效的公开性文献。放眼未来，我们可以预想到以后关于CIK细胞免疫治疗研究会沿着以下两个大方向进行：克服组织数量大随机的临床试验这个难题，说明CIK细胞在肿瘤免疫治疗中的作用；通过实验扩大CIK细胞对肿瘤细胞的细胞毒活性范围。+ n* I( n1 u9 e& U. k! @
1. Background2 ]3 k$ _5 E+ W" r) `) ]
One of the first prototypes of cytokine-induced immune-effector cells is the Lymphokine-Activated Killer (LAK) cells. Firstly described in the early 1980s, LAK cells are cytotoxic effector lymphocytes whose cytolytic activities are not restricted by major histocompatibility complex (MHC) and have the ability to kill fresh tumor cells and NK-resistant tumor cell lines [1]. LAK cells are generated following expansion in the presence of IL-2 for a relatively short culture period of approximately 5 days. After culture, the heterogeneous LAK cell population consists of CD3−Leu19+NK cells, CD3+Leu19+ cells, and CD3+Leu19− T cells. Leu19 was subsequently redesignated as CD56 and these CD3+CD56+ cells are also termed non-MHC-restricted cyto- toxic T cells. The two cell subsets the CD3+Leu19+ T cells and CD3−Leu19+NK cells contribute to the cytolytic property of LAK cells [2]. - R- ?$ Y8 ]" u) l8 W$ R
1. 目的
0 c( g! r. l1 u; v. g% W: e0 i; N第一个细胞因子诱导的免疫效应细胞是淋巴因子激活的杀伤细胞（LAK细胞），首先最后的是在1980年有对LAK细胞的描述，LAK细胞是有细胞毒性的效应淋巴细胞，其溶瘤活性不受主要组织相容性复合物限制（即非MHC限制性），LAK细胞可以有效杀伤新生肿瘤细胞以及NK不敏感肿瘤细胞。加入IL-2诱导激发，培养约5天时间，即可以大量扩增LAK细胞。然后加入一群异质的LAK细胞群，其中包括CD3−Leu19+NK细胞，CD3+Leu19+细胞以及CD3+Leu19−T细胞。Leu19细胞随后如CD56和那些CD3+CD56+一样被称为非MHC限制性细胞毒T细胞。CD3+Leu19+T淋巴细胞和CD3−Leu19+NK细胞这两个细胞亚群为LAK细胞的主要效应细胞。- z8 `& u `7 \- w1 L+ I& k
Over the years, various improvements in the methods to culture LAK cells have been developed. These included the addition of OKT3 at the initiation of culture, prolongation of culture duration, and the addition of various different types of cytokines at the end of culture. These improved methodologies to culture LAK cells resulted in better expansion over the originally described method [3]. LAK cells demonstrated potent in vitro cytotoxicity against susceptible tumor cells and led to the regression of established tumors in animal models [4, 5]. In clinical studies, LAK cells had demonstrated modest efficacy against metastatic cancer such as renal cell carcinoma and melanoma[6]. In a randomized controlled trial in the 1990s, adoptive immunotherapy using ex vivo activated T cells showed clinical efficacy in terms of prolongation of relapse-free survival for patients with hepatocellular carcinoma following resection of the primary tumor [7]. 9 d& c! I1 F% R
经过几年时间，人们对LAK细胞培养方法进行了改良并得到了各种各样的培养方法，这些培养方法包括培养开始时添加OKT3，延长培养时间，培养最后添加各种不同类型的细胞激活因子。LAK细胞培养方法经过改良后，LAK细胞的扩增有效率比以往更高了[3]。动物实验中，体外条件下LAK细胞有效杀伤肿瘤细胞并使人工建立的荷瘤小鼠模型内的肿瘤消退[4, 5]。在临床研究中，LAK细胞也表现出了对肾细胞癌和黑色素瘤的肿瘤细胞杀伤活性。1990年的随机对照试验中，肿瘤过继免疫治疗的作用机制是通过激活人体内T淋巴细胞抗肿瘤效应来延长肝细胞癌患者手术后无复发生存时间。$ d# H3 V4 r1 F% @- P
The discovery, generation, and therapeutic use of immune-active host effector cells that can kill cancer cells are continuously being developed. The pioneering work that accelerated the field of cellular immunotherapy with CIK cells was performed in Stanford. The authors described CIK cells as non-MHC-restricted T cells with marked ability to proliferate and demonstrated superiority over LAK cell in cytolytic activities against B cell lymphoma [8]. Further- more, CIK cells exhibit potent in vivo cytolytic activities without the need for coadministration of IL-2. CIK cells are generated by the timed addition of IFN-γ 1000 u/ml on day 1 of culture, followed 24 hours later by the addition of anti- CD3 at 50 ng/ml and IL-2 at 300 IU/ml. Together with the periodic addition of IL-2, the culture medium is regularly replenished throughout the culture period of 21–28 days [8].
: T" d; U: U# H2 Z8 E7 C8 S研究发现，增殖方式，以及对肿瘤有杀伤效应的宿主免疫效应细胞医疗用途都在不断的进展。关于促进CIK细胞免疫治疗范围的最早研究工作是在斯坦福完成的。这个创始人提出了CIK细胞如非MHC限制性T淋巴细胞一样增殖能力强，并且CIK细胞在治疗B细胞淋巴瘤[8]上与LAK相比，有着更强的抗瘤活性。此外，在没有添加IL-2的情况下CIK细胞体外仍然有抗瘤活性。CIK的培养方法是：培养第1天添加浓度为1000 u/ml的IFN-γ，24小时后添加浓度50 ng/ml的抗CD3单抗以及浓度300 IU/ml的IL-2，之后定期添加IL-2，并且在21-28天的培养期内是由始至终都保持足够的培养基[8]。
( Z2 q1 p8 i2 _ FAt the end of the culture, the CD3+CD56+ cells, derived from CD3+CD56− cells, could expand by up to 1000-fold and gave the greatest cytotoxicity against various tumor cell targets, including K562 and B cell lymphoma cell lines, as compared to CD3+CD56− cells [9]. The expression of CD56 on these non-MHC-restricted effector T cells was found to be the result of IFN-γ-priming that induced the production of IL-12 by monocytes and the upregulation of CD58 (LFA-3), both of which were demonstrated to be crucial for the expansion of CD56+ T cells [10]. Subsequently, this unique subset of non-MHC-restricted CD3+CD56+ T cells was referred to as NK-like T cells since, similar to the NK cells, they do not require prior specific sensitization to induce the recognition of target cells.
/ B$ j( [( {" v7 a, @! l0 T5 @在培养的最后，从CD3+CD56−细胞衍生出的CD3+CD56+细胞浓度高达1000个国际单位，与CD3+CD56−细胞相比，CD3+CD56+细胞对不同的肿瘤靶细胞如K562和B淋巴瘤细胞株有着超强的杀伤活性[9]。研究发现IFN-γ可诱导表达CD56的非限制性T淋巴细胞，CD56非限制性T淋巴细胞可通过单核细胞及增加CD58（LFA-3）诱导产生大量IL-2，因此IFN-γ和IL-2是CD56+T淋巴细胞[10]有效扩增的两个不可或缺的重要因素。后来，因为非MHC限制性CD3+CD56+T淋巴细胞与NK细胞相似，不需特殊抗原致敏便可识别靶细胞，因此又被称为NK细胞样T淋巴细胞。) Y9 U7 `4 B) f" \2 n
( s, H. p! X- A+ z
$ Y& x; [3 z; c( c6 m2. Functional, Phenotypic, and Genotypic Characterization of CIK Cells
' v( `4 H3 C* W: e, y4 d# UFollowing bulk culture in vitro, the effector cell population obtained is heterogeneous and comprises of a small fraction (∼2%) of CD3−CD56+ NK cells and >90% of CD3+ cells of which ∼35% expresses CD56 while the remaining cells are CD3+CD56− [11]. The sorted NK cell fraction (CD3−CD56+) behaves like classical NK cells and the killing of autologous acute myeloid leukemia (AML) target cells mediated by these cells can be enhanced by blocking the HLA class I molecules on the target cells [12]. The CD3+CD56+ subset, termed non-MHC-restricted T cells, is able to kill both autologous and allogeneic susceptible tumor targets such as primary AML cells of disparate HLA types [11].
# i+ p5 K6 _6 j& S5 ]3 p9 y* A) G3. CIK细胞功能、表型及基因型的鉴定% X3 d6 T8 v! z3 x
经过体外的批量培养后，获得一群异质的效应细胞，这群异质的效应细胞里面有小部分（约2％）CD3−CD56+NK细胞，90％以上的CD3+ CD56+细胞以及少量的CD3+CD56−细胞[11]。CD3−CD56+NK细胞的杀伤机制和典型NK细胞很相似，通过阻断AML靶细胞上人类白细胞抗原（HLA）Ⅰ类分子来增强介导自体髓细胞性白血病（AML）靶细胞凋亡[12]。CD3+CD56+细胞是非MHC限制性T淋巴细胞，可以有效杀伤自体和异体易感肿瘤靶细胞，如不同HLA类型的原发性AML细胞。
/ U( D+ s! j% B# Y/ T% }/ h: |0 r# C However, we have earlier shown that this subset of cells recognizes target cells through the T cell receptor (TCR) and requires the presence of MHC molecules on the target cells, a phenomenon similar to that observed for classical cytotoxic T lymphocytes. Furthermore, blockade of either TCR on the effector cells or the MHC class I molecules on the target cells abrogates the killing of target cells [12]. This discrepant observation of being not restricted by the MHC specificity in its cytolytic activities but with the effector function dependent on the presence of MHC is inconsistent with the current T cell paradigm, which dictates that cytotoxic effector T lymphocytes recognize target peptides in the context of self-MHC molecules. Besides our report, similar observations had also been independently made by others [13–15]. & J. N" {: `# P! Y; Z5 C
然而，早期研究报道说此种表型细胞识别靶细胞需要通过T细胞受体（TCR）和靶细胞上的MHC分子，就像细胞毒T淋巴细胞一样。而且，无论是阻断效应细胞上TCR还是靶细胞上的MHCⅠ类分子，都不能对靶细胞发挥杀伤效应[12]。此种表型T细胞抗瘤活性为非MHC限制性，但效应功能却是MHC限制性的这种特殊现象，使细胞毒效应T淋巴细胞有效识别表达自体MHC分子的靶细胞。此种情况除了我们报道之外，还有其他的一些类似的相关研究报道。: r2 Q% a2 g, I1 n
A hypothetical working explanation is that T cells in an “activated” state induced by cytokines could express elevated levels of adhesion molecules which result in the recognition of target cells more readily and efficiently, overriding the requirement for stringent matching of the MHC molecules. While these T cells are labeled as “NK-like”, we demonstrated that the well-characterized conventional NK receptors, including the KIR, NKG2C/E, NKG2D, and DNAM-1, are not involved in the recognition and killing of AML targets [12]. The exact molecules responsible for the recognition of AML targets await further molecular characterization. While CIK cells have been shown to kill the myeloma cell line U266 via NKG2D-mediated recognition of its cognate ligands MICA and MICB on the target cells [16], the same mechanism is unlikely to be operative for AML targets as these target cells do not express MICA and MICB [17].
$ N* T' u% M4 {/ \+ I一种假设性说明就是T细胞经过细胞激活因子诱导激活后表达高水平的粘附分子，更容易更有效的识别靶细胞，但前提是要在跟靶细胞MHC分子严格匹配的条件下。然而通过观察发现，NK细胞样T细胞的特征性NK细胞受体，其中包括KIR，NKG2C/E，NKG2D以及DNAM-1，与识别和杀伤AML靶细胞没有必然联系[12]。这种识别AML靶细胞的确切分子还有待进一步进行分子鉴定。而CIK细胞已经被证实是通过NKG2D介导识别靶细胞上的同源配体MICA和MICB而对骨髓瘤细胞株U266进行杀伤作用[16]。但相同的机制不能说明AML靶细胞和不表达MICA和MICB的靶细胞是一样的。
+ H( r% o+ p3 V, `( RInvestigation into the possible explanations for the better cytolytic activities against tumor cells demonstrated for the CD3+CD56+ cells over its CD3+CD56− counterpart revealed that the CD3+CD56+ cells consist of a higher proportion of CD8+ cells compared to the CD3+CD56− cell subset. Furthermore, the CD3+CD56+ subset is a more terminally differentiated late effector T cell population bearing the CD27+CD28− or CD27−CD28− phenotypes. In contrast, the CD3+CD56− cells are early effector T cells expressing mainly the CD27+CD28+ and CD62L+ phenotypes. The granzyme content is also higher in the CD3+CD56+ T cells, consistent with the report that late effector T cells possess more potent cytotoxicity than early effector T cells [12]. Table 1 summarizes the characteristics of the three subsets of cells in the bulk CIK culture.
W D/ }9 k( r$ b7 S4 f/ YCD3+CD56+细胞与CD3+CD56−细胞相比具有更高的抗瘤活性，可能解释就是CD3+CD56+细胞比的CD3+CD56−细胞含有比率更高的CD8+细胞。除此之外，CD3+CD56+细胞亚群是在分化的同时表达CD27+CD28−或CD27−CD28−表型的晚期效应T细胞群。相反，CD3+CD56−细胞是主要表达CD27+CD28+和CD62L+表型的早期效应T细胞。并且CD3+CD56+细胞内分泌颗粒酶内容物含量也比CD3+CD56−细胞高。这与报告中报道的晚期效应T细胞的细胞毒性比早期效应T细胞的细胞毒性强的结论是一致的[12]。表1简要概述了在CIK细胞批量培养中的三种表型细胞的特征。 f4 ~9 }3 K) b
Our research focus is on the cellular immunotherapy of hematological malignancies and we have observed that CIK cells kill AML blasts efficiently but not acute lymphoblastic leukemia (ALL) blasts [11]. The resistance of ALL blasts to killing by immune effector cells has also been well reported by others [18, 19]. We therefore also studied the possible molecular event that might explain this discrepancy. We observed that the CD3+CD56+ fraction of CIK cells cultured from acute leukemic marrow at diagnosis expressed high levels of immune function genes including IFN-γ, TNFα , CXCR3, CCR1, granzyme B, CCL3 (MIP-1α), CCL4 (MIP-1β), CCL5 (RANTES), IL-7R, IL-12Rβ2, and caspase-1, consistent with a Th1 and Tc1 polarization [20]. Upon stimula-' [/ H$ Z! c7 J
tion by the corresponding leukemic targets, that is, CIK cells derived from AML patients coincubating with autologous or allogeneic AML blasts and similarly, CIK cells derived from ALL patients coincubating with autologous or allogeneic ALL blasts, immune response-related genes like IFN-γ, GM-CSF, IL-4, IL-8, cytokine receptor genes IL-2Rβ,IL-2Rα, IL-4R, IL-12Rβ, IL-15Rα, chemokine and chemokine receptor genes, and genes belonging to the TNF and TNF
$ I+ w% G# \# _ k* Creceptor superfamily were upregulated.6 A5 p# f: U! e1 H
我们研究目的是探讨关于细胞免疫治疗血液恶性肿瘤以及CIK细胞可以有效杀伤AML原始细胞但无法杀伤急性淋巴细胞性白血病（ALL）原始细胞[11]。ALL原始细胞抵制免疫效应细胞对其的杀伤作用在其他的文献中也有报道[18, 19]。所以我们也对这个可能分子理论进行研究希望可以通过这个研究来解释这个偏差。通过鉴定，我们观察到在CIK细胞与急性髓性白血病细胞共培养时部分CD3+CD56+细胞表达了高水平的有免疫功能的基因，包括IFN-γ，TNFα，CXCR3，CCR1，颗粒酶B，CCL3 (MIP-1α)，CCL4 (MIP-1β)，CCL5 (RANTES)，IL-7R，IL-12Rβ2以及半胱天冬氨酶-1，这与Th1和Tc1亚群[20]是一样的。在通过相应的白血病肿瘤细胞抗原刺激基础上，这样，CIK细胞从ALL患者自体或异体ALL原始细胞中获得抗原特异性，使免疫应答相关基因如IFN-γ，GM-CSF，IL-4，IL-8，细胞因子受体基因IL-2Rβ，IL-2Rα，IL-4R，IL-12Rβ，IL-15Rα，趋化因子和趋化因子受体基因，TNF和TNF受体超级家族的基因数量增加。
@4 \' s7 ?" R Many proapoptotic genes were downregulated whilst antiapoptotic genes were upregulated. One observation of great interest was that genes encoding the NK receptors NKG2C, NKG2E, and CD94 were exclusively upregulated in CIK cells stimulated with myeloid blasts but not lymphoid blasts. In contrast, TGFβ-1 gene transcript was upregulated in CIK stimulated with lymphoid blasts. The differential regulation of these immune-related genes corroborated with our observed findings of good CIK killing against myeloid but poor killing against lymphoid blasts [20]. However we did not detect surface expression of NKG2C and NKG2E molecules on the CIK cells both before and after stimulation with myeloid target cells [12]. Therefore, the molecular and cellular significance of the upregulation of these immune-related genes needs to be further investigated.$ ]! C, b& Z0 M% F
很多诱导凋亡基因下调的同时抗凋亡基因上调。其中一个有趣的发现是用急变髓细胞而非淋巴细胞刺激CIK细胞能够上调CIK细胞表面NK细胞样受体如NKG2C，NKG2E，和CD94。相反，在淋巴细胞刺激CIK细胞后可上调β型转化生长因子TGFβ-1基因转录。这种免疫相关基因的调节差异证实了我们研究所发现的CIK细胞可以有效抵抗骨髓样细胞但对淋巴细胞只有微弱的抵抗作用[20]。但是在髓样靶细胞刺激前后我们都无法检测出CIK细胞表面表达有NKG2C和NKG2E分子[12]。所以，这些免疫相关基因的分子和细胞有效性上调机制还需要进一步的调查研究。8 M0 K/ T1 N ?0 g W: C
; |+ M$ _' N- Y/ h
Figure 1:Marrow cells obtained from newly diagnosed AML samples analyzed by side scatter versus CD45 staining show a large proportion of leukemic blasts that are CD45dim and a much smaller fraction of normal lymphocytes that are CD45bright(a). Culture of the bulk marrow cells comprising of majority of leukemic blasts with a small fraction of lymphocytes under CIK condition (b) is able to generate an end product comprising of a majority of CD3+ cells with a variable CD3+CD56+ fraction (c).) p7 y; Q. R' q, N: |# P
图1：+ P7 I) j, q5 W) L) g0 u2 e, u
E) \+ Z( k/ F: T
. |4 H9 s0 l2 d3 r Q* f" ?3. In Vitro Antitumor Activity of CIK Cells- g! h( G/ s1 {* p$ M+ K" o
Over the years, CIK cells have been tested for its antitumor activity against a variety of tumor targets. It was first shown to be able to kill t(14;18)-positive lymphoma cell lines but not against normal human hemopoietic precursors[8]. Doxorubicin- and vinblastine-resistant tumor cell lines& B ]0 O' B3 B, H, K, |4 {1 f) P
expressing high level of Pgp (P-glycoprotein) were also susceptible to CIK-mediated lysis [21]. Furthermore, chronic myeloid leukemia (CML) colony growth was reported to be suppressed by CIK cells and after 28 days of coincubation, the remaining colonies in culture were exclusively composed of Philadelphia- (Ph-) negative cells [22, 23]. Previously, we have shown that CIK cells could be generated from the marrow or peripheral blood samples from acute leukemia patients collected at diagnosis. Figure 1 shows the change in the composition of cells cultured under CIK condition from a starting cell population consisting of a majority of leukemic blasts with typically less than 10% T cells, to a majority of T cells at maturity. These cells were lytic against both autologous and allogeneic AML targets [11]. Additionally, CIK cells could also be generated from untreated chronic lymphocytic leukemia (CLL) patients, where cytotoxicity against autologous CLL targets could be induced by addition of anti-CD3 MoAb [24, 25].
/ d: F$ k7 o7 V* S/ v+ V8 o3. CIK细胞体外抗癌活性
2 \* g( q+ s6 f# z1 N* o经过几年时间，试验证实CIK细胞对很多种肿瘤靶细胞都有抗瘤活性。第一次试验CIK细胞可以杀伤t(14;18)-阳性淋巴瘤细胞株但对人体自身造血细胞没有毒副作用[8]。多柔比星和长春碱对表达高水平P糖蛋白Pgp的肿瘤细胞株有抑制作用，这种肿瘤细胞株对于CIK细胞介导凋亡作用同样敏感[21]。此外，有报告称CIK细胞可以有效抑制慢性髓细胞样白血病CML细胞生长，培养28天后剩余的细胞株组成费城阴性细胞[22, 23]。以前，已经知道CIK细胞可以从急性白血病患者骨髓或外周血样中产生。图1也展示了CIK细胞与包含着大量白血病细胞和不到10％T细胞的细胞群共培养后白血病细胞含量的改变以及诱导扩增出了大量的成熟T细胞的过程。这些成熟T细胞对自体和异体AML靶细胞都有抑制作用[11]。另外，慢性淋巴细胞性白血病CLL患者也可产生CIK细胞，并且这种CIK细胞在抗CD3单抗的诱导下可以有效杀伤自体CLL靶细胞[24, 25]。
+ H* c: ~$ O6 T1 K. n7 |1 H- vImmunological manipulations aimed to potentiate the antitumor activity of CIK cells have been explored. Hence, dendritic cells (DC) were cultured and engineered to present tumor antigens to CIK cells hoping that this might enhance the recognition of tumor cells and its subsequent killing.0 A% Z% T& N2 K, l2 M) f4 \
This was shown to be feasible for multiple myeloma (MM) when the DC was pulsed with target-derived idiotype before coculture with the CIK cells [26]. CIK-resistant pancreatic carcinoma target cells also became susceptible to CIK cells that were cocultured with DC loaded with tumor-restricted RNA and the CA19-9 peptide [26, 27]. Furthermore, trans-fection of IL-2 genes into CIK cells to enhance its IL-2 production potentiated its cytotoxicity against a pancreatic% ? x; A: G' ?" K
cancer cell line following coculture with DC when compared to non-IL-2-transfected CIK cells [28].
" F0 z% Q, |- ^2 F1 p' X以免疫治疗为目的对CIK细胞抗瘤活性的研究已经开展了。因此，树突状细胞（DC细胞）递呈肿瘤抗原给共培养的CIK细胞，希望可以增强CIK细胞识别肿瘤的能力以及提高其肿瘤杀伤活性。这种假设在用肿瘤抗原衍生的个体基因型脉冲致敏DC细胞，然后与CIK共培养后可以有效杀伤多发性骨髓瘤MM细胞的试验中得到了证实[26]。之前对CIK细胞不敏感的胰腺癌靶细胞也对当CIK细胞与负载肿瘤抑制RNA和胃癌相关抗原肽（CA19-9肽）的DC细胞共培养后的CIK细胞敏感了。此外， 和没有经过IL-2基因转染的CIK细胞相比，IL-2基因转染CIK细胞与DC细胞共培养诱导分泌IL-2的能力更强，数量更多，对胰腺癌细胞的杀伤效应更强[28]。6 U( u1 U( P% `; o+ L7 b
4. Mice Studies3 f* g, C; |, b8 |; o
Earlier work had shown the antitumor activity of CIK cells in mice. Human B cell lymphoma cell lines harboring the t(14;18) chromosomal translocation were injected into SCID mice to evaluate the efficacy of CIK cells. When allogeneic CIK cells were injected 1 day after inoculation of tumor cells, the SCID mice receiving the CIK cells had prolonged survival compared to control mice and LAK cell-treated mice, with long-term survival of 30% and no molecular evidence of lymphoma [9]. In SCID mice engrafted with human CML, autologous CIK cells transfer 4 weeks after inoculation of tumor cells resulted in eradication of bcr-abl which remained detectable in control mice [22]. This study showed that CIK cells, which are Ph chromosome-negative, could be expanded from patients with CML and had potent in vitro and in vivo efficacy against autologous tumor cells. In the same study, an additional series of SCID mice was not engrafted with CML but they instead unexpectedly developed large human Epstein-Barr virus-associated lymphomas. Within this series, the CIK cells-treated group developed no or small tumor as compared to large tumor developed in the untreated group[22]. These experimental data served as convincing early evidence of in vivo efficacy of CIK cells.
A" B+ ]& X" s( n6 k9 D( X) G% v4. 小鼠试验) A/ J9 s; P: G) u' h# ^( V. ?
早期的小鼠试验显示了CIK细胞的抗瘤活性。将t(14;18)染色体易位的人B淋巴细胞株输注到SCID小鼠中来证实CIK细胞的抗瘤活性。在小鼠体内构建肿瘤1天后，对小鼠输注同源CIK细胞，发现与输注LAK细胞治疗相比，输注CIK细胞可以使SCID小鼠的生存时间更长，其中分子水平上无淋巴瘤复发的长期存活率达30%[9]。输注人CML细胞到SCID小鼠体内并且产生肿瘤细胞后，输注自体CIK细胞，4周后，发现小鼠模型内剩余的bcr-abl融合基因被清除干净 [22]。这就证明了ph 染色体阴性CIK细胞可以在CML患者体内扩增，并且在体内外都对自体肿瘤细胞有强的抗瘤活性。在相同的研究中，在一组SCID小鼠内输注CML肿瘤细胞，意外的发现CML肿瘤细胞在小鼠体内诱发了人EB病毒感染淋巴瘤。在这组小鼠中，CIK细胞治疗组，小鼠体内的肿瘤有了明显的消退甚至消失，而没有进行治疗的对照组小鼠体内的肿瘤没有消退迹象反而不断生长[22]。这对于早期CIK细胞治疗肿瘤提供了可信的实验数据。
2 h" w$ N7 _ b: ^3 IWith the availability of the bioluminescence imaging (BLI) technology [29], the in vivo functional activities of CIK cells could be visualized in a real-time fashion in mice inoculated with bioluminescent gene-transfected tumor cells and treated with CIK cells for cellular immunotherapy. To monitor tumor regression by BLI, mice were implanted intraperitoneal with HeLa-luc (a luciferase gene-transfected human cervical carcinoma cell line) and subsequently treated with CIK cells. Mice treated with CIK cells had significant tumor regression or complete eradication compared to saline-treated mice [30]. Similar tumor response was visualized for murine lymphoma cell lines [31]. Using the same strategy, CIK cells were transfected with the gfp/luc genes to visualize its trafficking by BLI [31]. Following injection, it was observed that luc+ CIK cells first reached the lungs within 30minutes followed by a general distribution to other sites of the body. By the 7th hour, a population of the labeled CIK cells migrated to the tumor sites and remained detectable at these sites for an additional 9 days with resultant tumor regression [31]. Importantly, this antitumor effect of CIK cells occurred without the need for exogenous IL-2, a clinically relevant observation.! D/ g1 E" t% S! }& f; L
随着活体成像（BIL）技术的发展[29]，我们可以对生物荧光标记基因转染肿瘤细胞移植小鼠进行CIK细胞免疫治疗的全过程进行实时直观观察。在小鼠腹膜处移植HeLa-luc细胞（一种荧光标记基因转染的人宫颈癌细胞株）后运用CIK细胞免疫治疗，通过活体成像技术观察肿瘤的消退情况。结果发现与saline治疗相比，经过CIK细胞免疫治疗的小鼠体内肿瘤有了明显消退甚至是完全消失[30]。同样的肿瘤免疫反应也发生在CIK细胞杀伤鼠淋巴瘤细胞株[31]。相同的原理，通过gfp（绿色荧光蛋白）/luc（荧光素酶）双标基因转染CIK细胞，我们可以利用活体成像系统观察CIK细胞治疗情况 [31]。输注luc+标记CIK细胞后，观察发现不到30分钟CIK细胞已到达肺部，紧接着到达体内的各个部位。7小时后标记CIK细胞移行到肿瘤部位，并停留在肿瘤部位，9天后发现肿瘤有了明显消退迹象[31]。重要的是，CIK细胞在没有外源性IL-2的辅助作用下也能发挥抗瘤活性，这是又一个临床上的重要发现。
$ ^9 L" q+ a2 f0 k& r. H* P5 @# M5. CIK Cells Across MHC Barrier
3 ?) O9 l( M8 F C' hDonor lymphocyte infusion (DLI) is used to increase the graft-versus-tumor (GVT) effect after allogeneic hematopoietic cell transplant. The role of DLI in the management of malignancies remains restricted mainly due to the limited spectrum of activity and high risk of graft-versus-host! b) g% k7 l1 G# w* x9 m) E2 S
disease (GVHD). The finding of new cell populations for adoptive immunotherapy with the ability to separate GVT from GVHD would be useful. Being non-MHC-restricted and active against autologous and allogeneic tumor cells with comparable efficacy [11], CIK cells have therefore been exploited in studies related to both its efficacy and possible toxicity due to GVHD across MHC barrier. In an allogeneic model where transplant with purified hemopoietic stem cell was done across major MHC barrier from H-2b donors into murine lymphoma-inoculated H-2d recipient mice, transplanted mice died from persistent lymphoma [32].4 N# x* j6 c8 ]+ i J
5. CIK细胞非MHC限制性* `- A7 ~( s% g- E' }
异基因造血细胞移植后，供者淋巴细胞输注（DLI）可用于提高移植物抗肿瘤效应(GVT)。但DLI治疗恶性肿瘤的应用仍然存在着限制，主要是因为有限的抗瘤活性以及高危险性的移植物抗宿主病(GVHD)。因此急需发现一种用于过继免疫治疗的新型细胞群，这种新型细胞群不仅可有效将GVT从GVHD中分离出来，而且还可以非MHC限制性有效杀伤自体和异体肿瘤细胞，有强的抗瘤活性[11]。而研究发现CIK细胞是GVHD非MHC限制性且具有强的抗瘤活性以及细胞毒性。通过提纯造血干细胞移植诱导H-2b供体非MHC限制性的异基因模型，然后将异基因模型移植到鼠淋巴瘤H-2d移植小鼠体内，移植小鼠死于持久性淋巴瘤[32]。
) U9 W. E0 g6 `' o8 w! i! h9 QHowever, recipient mice infused with CIK cells expanded from donor mice had reduction of lymphoma and 50% survived long term, with no or minimal GVHD. In contrast, recipient mice that were treated with unmanipulated donor splenocytes died from acute GVHD [32]. This observation suggests that expanded CIK cells mediate significant graft-versus-tumor effiect without significant GVHD even when transplanted across MHC barrier. The mechanism of reduced GVHD induction was postulated to be related to IFN-γ production [33].
2 A/ B& U& W% j6 A- v然而，给受体小鼠输注由供体小鼠扩增的CIK细胞，可使受体小鼠体内的淋巴瘤消退，并且50%小鼠无或最低限度移植物抗宿主病（GVHD）长期生存。相反，受体小鼠进行未经处理供者脾细胞治疗后死于移植物抗宿主病GVHD[32]。提示了扩增后的CIK细胞无诱导显著移植物抗宿主病GVHD的间接非MHC限制性抗肿瘤效应。而CIK细胞降低GVHD发病率的作用机制被认为是与IFN-γ的产生有关[33]。
- r2 v& s, u$ R$ uIn an MHC-mismatched model when mice bearing the A20 leukemia/lymphoma B cells were treated with allogeneic CIK cells, the donor CIK cells were observed to infiltrate GVHD target tissues to a lesser extent and for a shorter duration than unmanipulated splenocytes, resulting in milder histological changes in the gut and liver [34]. Instead, CIK cells were found to accumulate and persist at tumor sites resulting in eradication of established tumor [34]. These encouraging experimental data may be important information for application in human allogeneic transplantation where donor CIK cells may prove to be superior and hopefully safer than unmanipulated DLI in the prevention or treatment of relapse.
9 r+ _& P$ n/ g0 W9 e用异基因CIK细胞治疗构建A20白血病/B细胞淋巴瘤MHC不合小鼠，供体CIK细胞可有效降低GVHD靶组织程度并且所用时间比应用未经过处理的脾细胞更短，还可诱导缓和肠道和肝脏组织学变化 [34]。另外，CIK细胞可在肿瘤部位累积并持续存在，诱导肿瘤凋亡直至肿瘤消失[34]。这些激动人心的实验数据为人异基因移植的应用提供重要根据，与未经过处理的供者淋巴细胞输注DLI方法相比，供体CIK细胞免疫治疗有望成为防治肿瘤复发抗瘤活性更强更安全的治疗方法。
" c, d' f2 m: x, V' W( C: ~! f- x r7 F! h* J. p
6. Novel Development
) F0 z8 F; x7 {, m. _6 Z! o5 UExciting development that promised to broaden the application of CIK cell was reported over recent years. Primary ovarian carcinoma cells are resistant to CIK-mediated lysis. However, it was demonstrated recently that the addition of bispecific antibodies (BsAb) CD3xCA125 or CD3xHer2 (heteroconjugates of anti-CD3 with anti-CA125 or Her2, resp.) could efficiently target the CD3+ CIK cells to ovarian carcinoma bearing the specific ovarian tumor antigens and overcome the resistance. This strategy was shown to be effective against autologous primary ovarian tumor cells in vitro and in ovarian tumor-bearing mice. The survival of mice with ovarian carcinoma treated with CIK cells redirected by the BsAb was prolonged compared to control mice treated with CIK cells alone [35]. In an attempt to treat the Ewing’s family tumors (EFTs), the low levels of cell surface Her2/neu expression were employed to redirect ex vivo activated CIK cells to tumor targets using the BsAb CD3xHer2/neu [36]. It was demonstrated that CD3xHer2/neu could be used to redirect CIK cells to mediate cytotoxicity against EFTs.- z+ p. Q Q0 t0 t
6. 新进展/ U% ^* z6 c- e8 e
近年来关于CIK细胞应用的研究取得了令人欣喜的进展。上皮性卵巢癌细胞耐受CIK细胞介导凋亡。但是，最近有研究发现添加双特异性抗体(BsAb) CD3xCA125或CD3xHer2（分别是有抗CA125的抗CD3单抗或有Her2的抗CD3单抗）可以诱导CD3+CIK细胞抗原特异性并且有效识别杀伤卵巢癌细胞。这种杀伤机制可以用于CIK细胞体外和在负载卵巢癌小鼠中杀伤自体上皮性卵巢癌细胞。和CIK细胞单独治疗相比，运用添加双特异性抗体的CIK细胞治疗负载卵巢癌小鼠可使小鼠的存活时间更长[35]。有人尝试在表面Her2/neu低表达的CIK细胞中加入双特异性抗体CD3xHer2/neu ，治疗Ewing家族肿瘤（EFTs）[36]。结果表明添加双特异性抗体CD3xHer2/neu可使CIK细胞对EFTs间接发挥杀伤效应。, h$ \& o( Q$ D, Y; C5 e: {
Besides using BsAb, other methods of harnessing antigen-antibody affinity to redirect CIK cells to target tumor cells involved the transfection of CIK cells to express tumor-specific receptors. CIK cells are known to be inactive against B cell ALL targets in vitro, but CIK cells engineered to express the chimeric receptor specific for the CD19 antigen could redirect CIK cells and become cytotoxic towards CD19-expressing B cell ALL targets [37].# E7 I& P4 G: I7 ]& h$ g
基于双特异性抗体，固定抗原/抗体亲和让CIK细胞有效识别肿瘤靶细胞的各种方法包括转染技术使CIK细胞表达抗原特异性受体。已知CIK细胞体外对B细胞急性淋巴细胞白血病靶细胞无杀伤活性，但可以利用CD19抗原诱导CIK细胞表达特异性嵌合受体，使CIK细胞有效识别表达CD19的B细胞急性淋巴瘤白血病靶细胞并且发挥杀伤效应[37]。$ H, M. m: ^- S/ ^6 t/ P
' d' p- s* o5 ]3 [7 Y8 [# M5 M
Another innovative genetic modification of CIK cells was the transfection of CIK cells with oncolytic viruses. These are viruses with the ability to infect only transformed (tumor) cells. A modified double-deleted vaccinia virus (vvDD) was able to infect CIK cells without affecting its activity. CIK cells were employed as a carrier vehicle bringing the oncolytic virus to tumor cells through the NKG2D receptor which is highly expressed by CIK cells to its ligands MICA and MICB on the tumor cells. VvDD-carrying CIK cells migrated to the tumor sites where the oncolytic viruses were then released to specifically lyse tumor cells. This approach has been successfully shown to work using mouse models for human ovarian tumor and murine breast carcinoma, resulting in reduction of tumor burden and prolongation of survival [38].# ?8 c3 C; w0 a& O3 V
另外一种遗传修饰创新技术就是通过溶瘤病毒转染CIK细胞。这种病毒能够感染病变(肿瘤)细胞。经过修饰去除双基因痘苗病毒（VVDD）能够感染CIK细胞而不会影响其活性。CIK细胞通过其高表达的NKG2D受体和肿瘤细胞表面与受体相匹配的配体MICA和MICB将负载的溶瘤病毒运输给肿瘤细胞。负载去除双基因痘苗病毒的CIK细胞运行到肿瘤所在部位，然后释放溶瘤病毒有效诱导肿瘤细胞凋亡。此种诱导肿瘤细胞凋亡作用途径已成功用在治疗负载卵巢癌和乳腺癌的小鼠模型上，并且有效诱导降低小鼠模型肿瘤负荷和延长小鼠生存时间。
& ]4 G) K% w6 B7. Clinical Trials
; P0 y" u1 `( E7 Z. VCIK cells can be generated successfully from healthy donors as well as patients treated with chemotherapy for various malignancies and undergoing peripheral blood progenitor cell (PBSC) leukapheresis [39]. Feasibility of large-scale expansion was also reported for cord blood and even from washout of leftover mononuclear cells from cord blood unit bags [40]. Several phase I trials to test the clinical efficacy of CIK cells on a small number of patients have been reported. The first clinical trial using CIK cells was reported in 1999 by Schmidt-Wolf et al. in Germany using autologous CIK cells electroporated with IL-2 genes for infusion into 10 patients with metastatic renal carcinoma, colorectal cancer, and lymphoma. One patient with follicular lymphoma showed# h2 P, n X) N, L
complete response. No major side effect was observed, except for 3 patients who developed fever that spontaneously resolved [41]. % L" r) Y1 R, P4 i2 P! s) S
7. 临床试验
, t4 {% c4 g( B. N& _4 r通过白细胞分离技术，从健康者和经过化疗的各种恶性肿瘤患者中收集外周血造血干细胞(PBSC)，体外条件下成功诱导扩增培养CIK细胞[39]。有报道称脐带血或清除脐带血后剩余的单核细胞都可以用来进行大规模培养CIK细胞[40]。只有若干个少数病人的I期临床试验报道了CIK细胞的临床效应。第一个运用CIK细胞治疗的临床试验是由Schmidt-Wolf et al.在德国于1999年完成。当时Schmidt-Wolf et al.将 IL-2电穿孔转染CIK细胞，然后输注到包括肾细胞转移癌患者、结肠直肠癌患者以及淋巴瘤患者总共10例患者体内。其中一例滤泡性淋巴瘤患者通过CIK细胞治疗后病情得到了完全缓解。试验过程中，没有发现患者出现严重毒副反应，只有3例患者出现了发热现象，但随后发热自行消退。
2 I' N3 o: G& x" O6 O# `5 V0 UNegrin’s group at Stanford reported giving 9 patients autologous CIK cells to treat relapsed Hodgkin disease (HD) and non-Hodgkin lymphoma (NHL) postautologous transplantation. Besides demonstrating the feasibility of large-scale expansion of CIK cells and the absence of adverse reaction, this trial achieved 2 partial responses and 2 stabilization of disease in the recipients [42]. In another phase I/II study, autologous CIK cells were given as an adjuvant immunotherapy postautologous transplantation for 9 high risk HD and 4 MM patients, again demonstrating safety as well as the maintenance of disease response in these high risk patients in the first year post-transplantation [43]. In the adjuvant therapy of acute leukemia, a study from China reported that 73.4% of the 19 patients who received 1–4 courses of autologous CIK cell infusions in combination with consolidation chemotherapy remained in continuous remission for a follow-up period of 4 years, compared to 27.3% in the chemotherapy-only group [44].0 a2 T2 v7 x% M! m- l& W
斯坦福内格林试验组给予9例霍奇金病（HD）和非霍奇金淋巴瘤(NHL)患者自体CIK细胞免疫治疗，有2例患者病情部分缓解，2例患者病情稳定[42]。本试验不仅证明了CIK细胞大规模扩增培养的可行性，还证明了CIK细胞治疗无明显的毒副作用。在另一个Ⅰ/Ⅱ期临床试验研究中，自体CIK细胞治疗作为一种辅助免疫治疗，治疗9例高危霍奇金病HD患者和4例恶性黑色素瘤MM患者，再一次证明了CIK细胞过继免疫治疗一年内，CIK细胞治疗的安全性和有效的抗肿瘤免疫应答效应[43]。在中国进行的一个CIK细胞免疫治疗作为一种辅助疗法的临床试验研究中，接受1-4疗程自体CIK细胞治疗联合强力化疗的19例患者中有73.4％在4年随访期中病情得到持续性完全缓解，而单独运用化疗组只有27.3％病情得到持续性缓解[44]。
' E; s8 I9 s! u" Y! p( JCIK cells have also been explored in clinical trials to treat solid tumors by investigators in China. In one report, patients with stage IV stomach cancer who were treated with chemotherapy and received additional CIK cell infusions had a higher overall short-term remission rate of 56.3% as6 F7 O3 @" T' y0 S, [
compared to 48% in the chemotherapy-only group [45]. In another study, patients with hepatocellular carcinoma who achieved complete remission with transcatheter arterial chemoembolization and radiofrequency ablation were divided into two groups, where intrahepatic arterial CIK cell infusion was given to a study group of 45 patients. Relapse at 1.5 years occurred in 31.1% of patients in the CIK group as compared to 85% in the control group, with the difference mainly accounted for by the significantly lower relapses seen at local sites (15.56% versus 65%, resp.) [46]. The same group also reported their experience in 10 patients with renal
" W7 @. ]3 e' x/ R# C% Rcell carcinoma postnephrectomy who were given intradermal autologous dendritic cells pulsed with autologous tumor lysate and followed by autologous CIK cell infusions. In 4 patients with evaluable disease, one case of partial response in terms of resolution and reduction in the size of metastatic lung nodules was observed [47].4 m8 U; N6 ~9 l3 f
在中国也开始进行了CIK细胞免疫治疗实体瘤的临床试验。有一报道，IV期胃癌患者在经过化疗后，再进行CIK细胞免疫治疗，患者短期缓解率高达56.3%，而单独化疗组短期缓解率只有48%[45]。在另一临床试验中，将经导管动脉化学栓塞和射频消融使病情达到完全缓解的肝细胞癌患者分为研究组和对照组两组，给予研究组共45例患者肝动脉输注CIK细胞。CIK细胞治疗组1.5年后复发率是31.1%，对照组复发率为85%，而这两组的差异最主要体现在研究组局部复发率比对照组更低 (分别是，研究组15.56%、对照组65%)[46]。给予相同试验组中的10例肾细胞癌患者输注负载自体肿瘤裂解物的皮内树突状细胞（DC细胞），再输注自体CIK细胞。观察发现，4例患者病情可评估，其中一种部分反应方式是以肿瘤消退和减少转移性肺结节的免疫反应方式。, j2 x g+ @. y6 ?* |) L
In the allogeneic setting, an early abstract from Stanford in 2006 reported 10 patients with various hematological malignancies relapsing after allogeneic transplant-received cytoreductive treatment followed by donor CIK cell infusions in escalating doses up to 1 × 108 CD3/kg. Grade I acute
5 f% L7 @9 ]0 ~) x! F8 a& e/ tGVHD occurred in 1 patient and limited chronic GVHD in 2 patients [48]. Another report from Italy studied 11 patients in similar clinical settings and with variable doses administered for each patient. As often occurs in such settings where other salvage therapies are being used concurrently, it was difficult to assess the efficacy. Nevertheless, 3 patients (1 each with MDS, HD, and AML) achieved measurable response, in terms of improvement in donor chimerism or clearance of disease , that could be solely attributable to donor CIK cell infusions. Similarly, GVHD rate was not higher than that of unmanipulated DLI, with grade I and II acute GVHD in 4 patients, 2 of whom progressed to extensive chronic GVHD [49]. In our own experience using allogeneic donor CIK cells to treat patients in relapse who are refractory to chemotherapy and DLI, response attributable solely to CIK infusions was also seen in a few patients. GVHD involving mainly skin and liver was sometimes observed but was easily controlled (Linn and Hui, unpublished data).
6 d- s( l9 H( k$ y& G v关于异基因移植，2006年斯坦福的早期摘要报道10例各种血液恶性肿瘤患者异基因移植后复发，再用剂量增大到1 × 108 CD3/kg的供体CIK细胞进行免疫治疗。1例患者出现I度急性GVHD，2例患者出现有限的慢性GVHD [48]。意大利一临床试验研究，11例患者在相同的临床环境条件下，给予每个患者不同药物剂量。但是这种试验方案会经常出现突发状况同时需要采用挽救治疗，这样就很难去评估CIK细胞治疗的疗效了。然而，由于进行了供体CIK细胞输注，通过检测供体嵌合现象的改善或肿瘤清除率，3例患者（分别患有骨髓增生异常综合征MDS、霍奇金病HD、急性骨髓系白血病AML）均达到可测量反应。同时，与输注未经处理过的供体淋巴细胞相比，输注供体CIK细胞诱导GVHD发生率更低，4例病例出现I～ II度急性GVHD，其中2例病例发展为广泛慢性GVHD [49]。在我们以往试验中，用异体CIK细胞治疗对化疗和DLI不敏感的肿瘤复发患者，只有少数患者对CIK细胞治疗有效。有时在皮肤和肝脏可以观察到GVHD，但很容易控制。（Linn 和 Hui，未注明日期）。: k* q* l5 l5 m! L" k$ y
8. Future Directions8 h% s" D5 v# b8 F& w0 |; }
Over the years, investigators have learnt to generate various types of cytotoxic effector cells and have used them for the cellular immunotherapy of human cancer. CIK cells, either in autologous or allogeneic context, have demonstrated encouraging results both in vitro and in clinical studies
7 W3 [% i# `* W3 U* ^% d4 L: \$ h) r2 iby various groups, as summarized in Table 2.Advancesin genetic engineering technologies could further broaden the potential clinical applications of CIK cells if the challenge to conform these genetic engineering technologies to GMP compliance could be overcome. However, clinical studies with CIK cells are still in their infancy and only involved a relatively small number of patients in most of these studies. The relatively robust and simple cell culture procedures to expand CIK cells have enabled this approach of adoptive cellular immunotherapy to be increasingly studied across the world. As an immunotherapeutic modality, infusion of CIK cells is most likely to show efficacy in a relatively low tumor burden stage or in an adjuvant setting, rather8 E& O% [# Z, |( e6 e; ^: y
than in high tumor burden diseases. With the amount of encouraging experimental and clinical evidence currently available, randomized clinical trials are justifiable and have to be done under stringent compliance with the CONSORT principles. This will likely involve a substantial number of patients in order to demonstrate statistical significance for a modest degree of outcome superiority. Such studies are urgently needed in order to provide unequivocal evidence of the clinical usefulness of CIK cells, so as to enable its integration into cancer treatment protocols to improve overall cure rate.
6 \! f* F$ i) a j+ d% M* _8. 展望
+ z/ Q" x. u j/ A/ v经过几年时间，研究者发现了多种细胞毒效应细胞，并且将其运用到细胞免疫治疗人类肿瘤中。无论是自体还是异体CIK细胞，在体外实验还是临床试验中都有很强的杀伤效应，在表2中有概括。随着基因工程技术不断发展，在遵循基因工程技术GMP情况下，我们可克服种种困难探索CIK细胞在临床上应用并使其得以进一步扩展。然而，CIK细胞的临床研究仍然处于不成熟阶段，大多数相关的临床试验也是只有少量病例数的临床试验。相对粗糙和简单的CIK细胞扩增培养方法使得过继性免疫治疗道路还需进行不断的研究探索。如免疫治疗模式，在肿瘤负荷较低阶段可以通过输注CIK细胞有效杀伤甚至清除肿瘤细胞，在肿瘤负荷较高的情况下免疫细胞治疗只能作为一种辅助疗法进行治疗。以当前实验和临床试验取得的一系列有效数据为根据，可以在严格遵守CONSORT声明条件下开展随机临床试验。为了试验结果有统计学意义，这就可能涉及到一定量的临床试验病例数。目前急迫需要开展此类临床试验，为CIK细胞临床治疗的有效性提供明确数据根据，同时也是为了改善CIK细胞免疫治疗肿瘤方案，提高CIK细胞治疗肿瘤的全治愈率。
! J. x, ?) ]* r3 I, S' [& ~! a |
-
总评分: 威望 + 50
包包 + 100
查看全部评分
|
发表于 2011-5-5 09:40
发表于 2011-5-13 16:41
发表于 2011-7-3 23:48
请更正