Cord blood-derived cytokine-induced killer cells biotherapy combined with second-line chemotherapy in the treatment of advanced solid malignancies

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Abstract

This study investigated the efficacy of cord blood-derived cytokine-induced killer (CB-CIK) biotherapy combined with second-line chemotherapy in treating advanced solid malignancies after first-line chemotherapy failure. Forty patients with advanced solid malignancies after first-line chemotherapy failure were divided into two groups: CB-CIK cells transfusion plus second-line chemotherapy (CB-CIK + Chemotherapy) group and second-line chemotherapy alone (Chemotherapy) group. The ORR and DCR were 30% and 80% in CB-CIK + Chemotherapy group compared with 15% and 70% in Chemotherapy group (P = 0.451 for ORR and P = 0.716 for DCR) respectively. The time to progression and the median survival time were 3.45 months (95% CI 2.30–4.60 months) and 11.17 months (95% CI 9.05–13.28 months) in CB-CIK + Chemotherapy group compared with 2.03 months (95% CI 1.23–2.82 months) and 7.52 months (95% CI 5.97–9.06 months) in Chemotherapy group respectively. Compared with patients in Chemotherapy group, the patients in CB-CIK + Chemotherapy group had significantly longer PFS (P = 0.031) and overall survival (P = 0.048). In vitro studies further revealed that CB-CIK cells could overcome drug resistance in cisplatin-resistant lung adenocarcinoma cell line A549/CDDP through downregulating ABCG-2 and P-gp and induce cytotoxicity through the high level expression of CD3, CD56, FasL, and CD69. This could explain why CB-CIK could have synergistic effects with second-line chemotherapy shown in this clinical study. We concluded CB-CIK cells combined with second-line chemotherapy can significantly improve PFS and median survival compared with second-line chemotherapy alone in patients with advanced solid malignancies after first-line chemotherapy failure.

Introduction

For advanced solid malignancies, the efficiency of second-line chemotherapy is often low. A main cause of the failure of chemotherapy is the resistance of tumor cells against anticancer compounds, and a major form of such a resistance is called multidrug resistance (MDR), caused by the increased expression of ABC (ATP binding cassette) multidrug transporter proteins, which are active in an ATP-dependent manner extruding a large variety of drugs or drug conjugates from the cells. ABC proteins comprise a large superfamily of transmembrane proteins [1], [2], [3]. By far the best known major drug transporters, i.e. ABCB1 (P-glycoprotein, p-gp, or MDR1), ABCG2 (BCRP), ABCC1 (MRP1), and ABCC2 (MRP2, cMOAT) have been characterized in detail with respect to their structure and function [4], [5], [6]. P-glycoprotein (p-gp ) encoded by the mdr-1 gene is a 170 kDa plasma membrane protein that functions as an ATP-driven drug export pump. Cytotoxic drugs of natural origin with different chemical structures and mechanisms of action, such as vinca alkaloids, anthracyclines, epipodophyllotoxins and taxanes, can be extruded by p-gp through the cell membranes of resistant cells and cross-resistance occurs [7]. ATP-binding cassette protein G2 (ABCG2) is a 655-amino-acid polypeptide transporter that forms a homodimer and has been reported as a tetramer in plasma membranes. ABCG2 can extrude the antineoplastic drugs such as anthracyclines and camptothecins, which plays an important role in chemotherapy drug resistance [8].

New anticancer approaches to overcome chemotherapy drug resistance and increase survival for advanced solid malignancies who failed to first-line chemotherapy are needed. Immunotherapy stimulating the immune system and enhancing the patient's own anti-tumor ability is an alternative and promising way to treat cancer and overcome chemotherapy drug resistance [9], [10]. In the 1980s, the lymphocytes activated by IL-2, were used for treating malignancies at advanced stage [11], [12], [13]. Since then, several kinds of immunological effector cells such as lymphokine-activated killer (LAK) cells, tumor infiltrating lymphocytes (TIL), anti-CD3 induced activated killer cells (CD3AK) and cytokine-induced-killer (CIK) cells have been used as immunotherapy, and a protocol by which CIK cells were considerably induced in vitro was developed [12], [14], [15], [16], [17], [18], [19], [20]. CIK cells are MHC-unrestricted cytotoxic lymphocytes generated by incubation of peripheral blood lymphocytes with anti-CD3 monoclonal antibody, IL-2, IL-1 and IFN-γ. CIK cells represent the lymphocytes with increased anti-tumor cytotoxic activity in vitro and in vivo [18], and high proliferation ability as compared to LAK cells [21], [22]. The increase of lysis activity to tumor cells is mainly due to the high proliferation potential of CD3 and CD56 double positive cells in CIK cells [19], [21], [23], [24]. CIK cells are therefore suitable for the immunotherapy against advanced solid malignancies. Cord blood (CB) is progressively becoming an extensively used treatment for patients with both malignant and nonmalignant disorders. The CIK cells obtained from CB have an overlapping phenotype with the traditional CIK cells obtained from adult peripheral blood. Both CD3 + CD56− and CD3 + CD56+ cells contribute to their cytotoxicity. 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, as compared to CD3 + CD56− cells [25]. The activation of T lymphocytes, both in vivo and in vitro, inducing expression of CD69 is involved in lymphocyte proliferation and functions as a signal-transmitting receptor in lymphocytes, such as CIK cells [26]. Fas ligand (FasL or CD178), a type-II transmembrane protein, binding with its receptor induces target cell apoptosis. Fas ligand/receptor interactions play an important role in the regulation of the immune system, the progression of cancer and the apoptosis of carcinoma cells inducted by T lymphocytes [27]. HLA-DR is a major histocompatibility complex, MHC class II, cell surface receptor encoded by the human leukocyte antigen complex. The primary function of HLA-DR is to present peptide antigens, potentially foreign in origin, to the immune system for the purpose of eliciting or suppressing T-(helper)-cell responses that eventually lead to the production of antibodies against the same peptide antigen [28]. CD25 is the alpha chain of the IL-2 receptor. The IL-2/IL-2R interaction then stimulates the growth, differentiation and survival of antigen-selected cytotoxic T cells via the activation of the expression of specific genes [29]. In addition, they express NKG2D and perforin, the two proposed molecules which have been demonstrated to play an important role in CIK-mediated cytotoxicity so far [30], [31]. Studies by Nishimura et al. showed that adoptive transfer of allogeneic CIK cells caused minimal graft-versus-host disease (GVHD) because CIK cells infiltrated GVHD target tissues much less and transiently [32]. CIK cells produce large amounts of IFN-γ, which has protective effects against GVHD [33]. Moreover, the cord blood-derived cytokine-induced killer (CB-CIK) cells have been shown to be able to kill variety of human solid and hematological malignancies both in vitro and in vivo [34], [35]. Clinical studies with CB-CIK cells to treat malignancies showed promising efficacy with negligible side effects such as transient fever and Myalgia which mostly disappeared even without needs of anti-inflammatory drug administration [36]. All these suggest that CB-CIK cells have great potentials to be clinically used in treating human malignancies as a safe and effective approach especially for advanced cancer patients who cannot tolerate chemotherapy or radiotherapy. Recently, Deng et al. reported that CB-CIK cells alone have been used in treating advanced human lung cancer with minimal side effects and some efficiency [36].

Currently, CIK cells transfusion has evolved to become a promising way for overcoming chemotherapy resistance [9], [10]. Liu et al. reported that CIK cells possessed a higher antitumor cytotoxic activity against a drug-resistant lung adenocarcinoma cell line in vitro and they found CIK cells plused with docetaxel demonstrated a prominent augmentation of antitumor activity against multidrug resistance lung adenocarcinoma cell lines both in vitro and in vivo [10]. Since most of the clinical studies so far showed that the efficiencies of CIK cells especially autologous ones to treat human malignancies were low [37], [38] and studies have showed that CB-CIK cells are more potent in cytotoxic activity against various tumor cells than autologous ones with similar phenotype both in vitro and in vivo, we hypothesized CB-CIK cells could act synergistically with chemotherapeutic agents on malignant cells after first-line chemotherapy failure and CB-CIK cells might help second-line chemotherapeutic agents to overcome cancer cells' drug resistance, and thus improve PFS and median survival of patients with advanced solid malignancies. To test this hypothesis, we performed a randomized block design study to evaluate the potential effect of CB-CIK combined with second-line chemotherapy on survival, response evaluation and tolerability in advanced solid malignancies after first-line chemotherapy failure. Also, we performed in vitro studies to reveal the potential role of CB-CIK in this combination therapy.

Section snippets

CB-CIK induction

The mononuclear cells were prepared from cord blood with Ficoll-Hypaque centrifugation method, and then induced with the recombinant cytokines IFN-γ at 1000 U/ml (Boehringer Mannheim, Germany), IL-2 at 300 U/ml (Boehringer Mannheim, Germany), IL-1 at 100 U/ml (Boehringer Mannheim, Germany) and anti-CD3 antibody at 50 ng/ml (Boehringer Mannheim, Germany) for 15 days. CB-CIK cell growth was observed under the microscope and CB-CIK cells phenotypes were determined by flow cytometry [31]. Unless the

Population

Between May 2007 and May 2009, a total of 40 patients with advanced solid malignancies after first-line chemotherapy failure were enrolled in this study. Of these, 20 patients received second-line chemotherapy plus CB-CIK treatment (CB-CIK + Chemotherapy group), 20 patients received standard second-line chemotherapy alone (Chemotherapy group). Most of the baseline characteristics of the patients were well balanced between the two groups (Table 1, Table 2). In this study, there were 8 lung cancer

Discussions

CIK cells are a novel population of immune effect cells with high proliferation rate and potent antitumor activity against a variety of tumor cells [40]. After 18 days of incubation, our CB-CIK cells expressed high level of CD3, CD56, CD69, and FasL, which implied that mature CB-CIK cells had strong proliferation and cytotoxicity against tumor cells. The CD3+CD56+ cells gave the greatest cytotoxicity against various tumor cell targets [25]. CD69 appears to be the earliest inducible cell surface

Acknowledgements

This project is sponsored by the Scientific Research Foundation for the Returned Overseas Chinese Scholars, China State Education Ministry (No. 2007-1108) and we thank Dr. Longbang Chen for his kind suggestions.

References (49)

  • M. Dean et al.

    The human ATP-binding cassette (ABC) transporter superfamily

    Genome Res

    (2001)
  • H. Green et al.

    Spontaneous reversal of P-Glycoprotein expression in multidrug resistant cell lines

    Pharmacol Toxicol

    (2003)
  • A.G. Niethammer et al.

    Multidrug resistance-1 (MDR-1): a new target for T-cell-based immunotherapy

    FASEB J

    (2005)
  • P.Y. Liu et al.

    The antitumor effects of CIK cells combined with docetaxel against drug-resistant lung adenocarcinoma cell line SPC-A1/DTX in vitro and in vivo

    Cancer Biother Radiopharm

    (2009)
  • S.A. Rosenberg

    Adoptive immunotherapy of cancer using lymphokine activated killer cells and recombinant interlukine-2

    Important Adv Oncol

    (1986)
  • S.A. Rosenberg et al.

    A new approach to the adoptive immunotherapy of cancer with tumor infiltrating lymphocytes

    Science

    (1986)
  • S.A. Rosenberg et al.

    Gene transfer into humans' immunotherapy of patients with advanced melanoma using tumor infiltrating lymphocytes modified by retroviral gene transduction

    N Engl J Med

    (1990)
  • E.A. Grimm et al.

    Lymphokine activated killer cell phenomenon: lysis of natural killer resistant fresh solid tumor cells by IL-2 activated autologous human peripheral blood lymphocytes

    J Exp Med

    (1982)
  • S.A. Rosenberg

    Lymphokine-activated killer cells. A new approach to immunotherapy of cancer

    J Natl Cancer Inst

    (1985)
  • D.H. Lynch et al.

    Induction of murine lymphokine-activated killer cells by recombinant IL-7

    J Immunol

    (1990)
  • Y.S. Yun et al.

    In vivo antitumor activity of anti-CD3 induced activated killer cells

    Cancer Res

    (1989)
  • I.G. Schmidt-Wolf et al.

    Use of a SCID mouse/human lymphoma model to evaluate cytokine-induced killer cells with potent antitumor cell activity

    J Exp Med

    (1991)
  • I.G. Schmidt-Wolf et al.

    Propagation of large numbers of T cells with natural killer cell markers

    Br J Haematol

    (1994)
  • P.H. Lu et al.

    A novel population of expanded human CD3 + CD56+ cells with potent in vivo antitumor activity in mice with severe combined immunodeficiency

    J Immunol

    (1994)
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