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Am J Clin Exp Immunol. 2017; 6(3): 27–42.
Published online 2017 May 15.
PMCID: PMC5545683

Combination of celecoxib (Celebrex®) and CD19 CAR-redirected CTL immunotherapy for the treatment of B-cell non-Hodgkin’s lymphomas


The nonsteroidal anti-inflammatory drug (NSAID) Celecoxib (Celebrex®) received Food and Drug Administration (FDA) approval in 1998 for treatment of osteoarthritis and rheumatoid arthritis, and in recent years, its use has been extended to various types of malignancies, such as breast, colon, and urinary cancers. To maintain the survival of malignant B cells, non-Hodgkin’s Lymphoma (NHL) is highly dependent on inflammatory microenvironment, and is inhibited by celecoxib. Celecoxib hinders tumor growth interacting with various apoptotic genes, such as cyclooxygenase-2 (Cox-2), B-cell lymphoma 2 (Bcl-2) family, phosphor-inositide-3 kinase/serine-threonine-specific protein kinase (PI3K/Akt), and inhibitors of apoptosis proteins (IAP) family. CD19-redirected chimeric antigen-receptor (CD19 CAR) T cell therapy has shown promise in the treatment of B cell malignancies. Considering its regulatory effect on apoptotic gene products in various tumor types, Celecoxib is a promising drug to be used in combination with CD19 CAR T cell therapy to optimize immunotherapy of NHL.

Keywords: Non-Hodgkin lymphoma, immunotherapy, CD19, chimeric antigen receptor, apoptosis, signal transduction, celecoxib, Bcl-2 family, apoptosome, resistance, mitochondria, rituximab, CHOP, adoptive cell transfer

Inflammation and cancer

Inflammation is a defense mechanism by which immune cells, such as neutrophils, monocytes, and macrophages, are mobilized to the areas that have foreign bodies due to infection or injuries. These immune cells release pro-inflammatory cytokines, a necessary physiological step in fighting infection and healing wounds. Inflammation is self-limiting in normal cells, while it remains chronic in tumor cells [1]. Several studies have shown that there is a close relationship between tumor growth and inflammation. The nuclear factor for Kappa B cells (NF-κB), a group of transcription factors related to v-Rel oncogene, is an important link between cancer and inflammation [2]. Underlying inflammation or the formation of inflammatory microenvironment caused by malignant progression, activates NF-κB. Once activated, NF-κB upregulates tumor promoting cytokines like interleukin-6 (IL-6) and tumor necrosis factors-α (TNF-α), and pro-survival proteins like Bcl-xL, an antiapoptotic member of Bcl-2 family [2]. Additionally, it is proven that cell proliferation alone does not cause cancer, but an environment rich in DNA mutations, growth factors, activated stroma, and inflammatory cells is required [1]. Inflammatory signals trigger the release of several soluble factors, one of which is prostaglandin E2 (PGE2), an enzymatic product of two Cox isoenzymes, Cox-1 and Cox-2 [3].

Physiological roles of cyclooxygenase (Cox)

Cyclooxygenase-1 (Cox-1) and Cox-2 have different physiological functions due to their differences in tissue expression and regulation [4]. Cox-1 is a house-keeping gene, constitutively expressed in almost all tissues. It produces prostaglandins (PGs) that are involved in homeostatic functions. Cox-2, encoded by the gene Ptgs2, is tightly regulated and is highly inducible during inflammation. It is significantly upregulated in cells with inflammatory arthritis, proinflammatory cytokines, and tumorigenic potential [4]. With its inflammatory property, Cox-2 can promote tumor growth. Using ApcΔ716 knockout mice, a mouse model of human familial adenomatous polyposis (FAP), Oshima and colleagues showed that Cox-2 overexpression could induce tumorigenesis. In regular ApcΔ716 mice, significant amount of Cox-2 was expressed at very early stage of polyp formation. In Apc mice with Ptgs2 knockout and Apc mice treated with Cox-2 inhibitor, MF Tricyclic, substantial decrease in polyps was noted [5]. Cox-2 catalyzes the conversion of arachidonic acid (AA) to prostaglandin endoperoxide H2, and the reaction results in the formation of several mutagenic metabolites, such as malondialdehyde [6]. The peroxidase activity of Cox-2 can also convert xenobiotics into mutagens [6].

As mentioned above, Cox-2 produces PGE2, a protein that facilitates tumor growth. PGE2 induces the expression of IL-6 and haptoglobin, both of which are important regulators of angiogenesis; moreover, PGE2 also creates an immunosuppressive environment and a tumor microenvironment that support angiogenesis [3]. Cox-2 and PGE2 are implicated in the development of colorectal cancer [7]. Overexpression of PGE2 can also increase the protein levels of myeloid cell leukemia-1 (Mcl-1) through a PI3K/Akt-dependent pathway in human adenocarcinoma cells [8]. Mcl-1, a member of the anti-apoptotic Bcl-2 family, is involved in the intrinsic apoptotic signaling pathway. Moreover, Cox-2 mRNA stability is regulated by p38 mitogen-activated protein kinase (MAPK), a signal transduction pathway involved in extrinsic apoptotic signaling pathway [9]. Inflammatory stimuli, such as lipopolysaccharides (LPs), IL-1, and TNF-α, activate p38 MAPK, which in turn activates Cox-2 transcription. More research is needed to determine the exact mechanisms by which Cox-2 promotes tumor growth, but from the studies conducted so far, Cox-2 most likely induces tumorigenicity not simply by its activation of carcinogens, but by interacting with other factors, mainly the apoptotic machinery.

Major apoptotic pathways

There are two main pathways the cell uses to initiate apoptosis: the extrinsic pathway and the intrinsic pathway. They are tightly regulated by antiapoptotic signal transduction pathways such as NF-κB, PI3K/Akt, and MAPK, all of which are frequently dysregulated in tumors [10]. The extrinsic pathway is activated by death ligands, such as TNF-α, Fas ligand (FasL), lymphotoxin, and Apo2L/TNF-related apoptosis-inducing ligand (Apo2L/TRAIL). When these ligands bind to their cognate receptors on the cell surface, Fas-associated protein with death domain (FADD) and TNFRSF1A-Associated via death domain (TRADD) are recruited to activate the initiator caspases, including pro-caspases-8, -2, and -10. The initiator caspases then activate the caspase cascade, resulting in the activation of executioner caspases. The intrinsic pathway is activated by cell stress such as chemotherapy. The cell stress triggers the cytoplasmic release of cytochrome c and second mitochondria-derived activator of caspases/direct IAP binding protein with low PI (SMAC/DIABLO) from the mitochondria, both of which are important pro-apoptotic effector proteins. Cytochrome c, together with dATP, cytoplasmic factors like Apoptotic protease activating factor-1 (Apaf-1) and pro-caspase-9, forms the large multiprotein complex called apoptosome, which then triggers the autocatalytic process and activation of caspase-9, which ultimately activates executioner caspases-3, -6, -7. Executioner caspases cleave several intracellular substrates, such as death substrate poly ADP-ribose polymerase (PARP), and trigger apoptosis. Many cases in which tumors become resistance to apoptosis are caused by downregulation or shedding of death receptors. Death receptors, such as Fas, TRAIL-R1 (DR4) and TRAIL-R2 (DR5) will not be expressed on the cell surface, so the death ligands cannot activate the extrinsic signaling pathway [10].

Besides the mentioned above factors, the intrinsic apoptotic signaling pathway is also regulated by Bcl-2 protein family. The pro-survival subfamily of Bcl-2 (Bcl-xL, Bcl-w, Mcl-1, Bfl-1/A1 and Bcl-B) promotes cell survival upon exposure to cytotoxic stimuli, while the Bax-like pro-apoptotic subfamily (Bax, Bak, and Bok) and BH3-only proteins subfamily (Bik, Bad, Bid, Bim, Bmf, Hrk, Noxa and Puma) promote cell death [11]. Whether the cell undergoes apoptosis depends on the balance between the pro-survival and pro-death signals from these three subfamilies. BH3-only proteins subfamily serves as cellular stress censor. Upon receiving cellular stress, BH3-only proteins inactivate Bcl-2 like proteins, resulting in the activation of Bax-like proteins. Activated Bax-like proteins permeabilize the outer mitochondrial membrane, triggering cytochrome c release and initiating the intrinsic pathway leading to apoptosis. Loss of Bax function and reduced levels of Bax are linked to resistance to chemotherapy and poor prognosis in pancreatic and ovarian cancer cells [12].

There are other proteins involve in the development of apoptosis-resistant tumor cells. Hypoxia inducible factor-1 (HIF-1) is a key regulator of hypoxia, a process that creates environmental stress and induces apoptosis. However, tumor cells, after several periods of hypoxia, adapt to the environmental stress and become resistant to apoptosis. HIF-1 can trigger hypoxia-mediated apoptosis by increasing the level of BCL2/Adenovirus E1B 19 kDa Interacting Protein 3 (BNIP3) and its homologue NIX, which in turn inhibit the pro-survival effect of Bcl-2; however, HIF-1 can also prevent apoptosis by inducing the expression of IAP-2 [13]. HIF-1 is an important apoptosis mediator, and cancer cells seem to selectively use HIF-1 to avoid undergoing hypoxia-mediated apoptosis.

Besides IAP-2, there are other proteins within the IAPs family that are important in intrinsic apoptotic signaling pathway, namely cellular inhibitors of apoptosis (cIAP)-1, cIAP-2, X-linked inhibitors of apoptosis protein (XIAP), and survivin. Under normal conditions, these proteins inhibit the activation of executioner caspases. SMAC/DIABLO, upon activation, physically associates with these proteins and removes the inhibitors of caspase activation.

Celecoxib as a selective Cox-2 inhibitor

As mentioned above, Cox-2 expression increases during inflammation. Therefore, Celecoxib, with its anti-inflammatory property, is theoretically a novel drug for cancer treatment. Celecoxib inhibits Cox-2 by interfering with prostaglandin-mediated upregulation of anti-apoptotic proteins such as Mcl-1 [14]. In Cox-2 overexpressed cells (Cox-2/cl.4), treatment of cytotoxic dose of 10 μM celecoxib and 25 μM NS-398, another Cox-2 inhibitor, significantly reduces the level of Mcl-1 [8]. Moreover, under the same experimental conditions, both inhibitors can inhibit PGE2 by 70-80% [8]. Celecoxib hinders Cox-2 activities in several cell lines. It exerts antiproliferative effects on Raji and Ramos Burkitt lymphomas in vitro [15] and in nude mice having intracranial lymphomas, which mimic human central nervous system (CNS) lymphomas [16]. Celecoxib is also an effective apoptotic inducer of B cells lymphoma, but not necessarily of T cells lymphoma [17].

In a study using cells of hemapoietic origin, treatment with high doses of celecoxib was very effective in patients with multiple myeloma (MM) [18]. More than 30% of malignant cells in MM had overexpression of Cox-2. Patients with MM tend to develop resistance to chemotherapy, so celecoxib is a good alternative therapeutic drug. In a phase II clinical trial, patients with relapsed and refractory MM were given thalidomide with celecoxib at doses ranging from 200 to 800 mg/day [19]. The results were promising: those who took doses greater than 400 mg/day had greater progression-free survival than those who took doses equal to or less than 400 mg/day (12.7 months compared to 4.6 months). Patients who took higher doses also had a better overall survival rate (OSR) than those who took the lesser dose (29.6 months compared to 18.9 months). However, adverse effects (AEs), such as peripheral edema and renal complication, were observed in some patients [19].

Celecoxib is also an effective drug to use in treatment of patients with NHL. In a phase II study, 35 patients with relapsed or refractory NHL were treated with high doses of celecoxib (400 mg [20]. The median progression-free rate was 4.7 months and median overall survival rate was 14.4 months with 8.4 months median follow-up. Even though celecoxib was used in high doses, the AEs observed were minimal. Gastrointestinal toxicity was observed with no interference with compliance. Most AEs were grade 1 and 2, including nausea, hypertension, and fatigue. Pharmacokinetics data showed that celecoxib was stable for a prolonged period. Per a preclinical model of Kerbel and colleagues, a plasma concentration having more than 500 μg/L was antiangiogenic [21]. The plasma concentrations were taken during the 12-hour period after the administration of the first dose of celecoxib. After a single dose of 400 mg, the peak concentration (Cmax) was 2,369 ± 1,586 μg/L at a median time of 3.2 hours, while Cmin after a single dose was 539 ± 335 μg/L. Additionally, celecoxib has an apparent clearance (Cl/F) of t 0.6 ± 0.4 L/h/kg and an elimination half-life (t 1/2) of 4.1 ± 0.9 hours [22]. Celebrex in being clinically used in various tumor models (summarized in Table 1).

Table 1
Summary of clinical data using Celebrex in various tumors

The pro-apoptotic effect of celecoxib does not depend entirely on Cox-2 inhibition. Several studies have shown that celecoxib can also induce apoptosis in Cox-2 negative cells. Celecoxib showed substantial antiproliferative effects on epithelial cancer cell lines, which had no detectable levels of Cox-2 expression [23]. In a study by Song and colleagues, Cox-2 depletion did not induce cell death and some of celecoxib derivatives that did not have Cox-2 inhibitory activity could facilitate apoptosis [24]. Interestingly, there are reports on the inhibition of cell proliferation in in vitro and in vivo models of Burkitt’s lymphoma due to downregulation of cyclins A and B and the loss of cyclin-dependent kinase (CDK) activity upon treatment with dimethyl-celecoxib (DMC), a celecoxib analog that lacks Cox-2 inhibitory function [15]. Therefore, Cox-2 presence in the cell is not required for celecoxib pro-apoptotic effect.

Modulation of apoptotic machinery by celecoxib

Celecoxib induces apoptosis via the intrinsic signaling pathway. The apoptotic effect is Bcl-2-independent and apoptosome-dependent. In Jurkat cells, Apaf-1 and pro-caspase-9 were required for celecoxib-induced apoptosis, while the presence or absence of Bcl-2 did not interfere with celecoxib-induced apoptosis [25]. Overexpression of Bcl-2 did not affect the effectiveness of celecoxib in Jurkat cells and Bcl-2 expression levels were not modified by celecoxib, as seen by the unaltered size and abundance of nonphosphorylated Bcl-2 protein levels. Additionally, Bcl-xL lacks significant inhibitory effects on celecoxib-induced apoptosis.

In lymphomas, arachidonic acid is converted to prostaglandins by Cox-2, leading to the upregulation of several anti-apoptotic proteins such as Bcl-2, PI3K/Akt, and Mcl-1 [26]. Surprisingly, only Mcl-1 and Bcl-xL, not Bcl-2 or PI3K/Akt, form a high affinity complex with Bak, thus blocking apoptosis [27]. Cytotoxic signals activate BH3-only proteins, which interact with Mcl-1 and Bcl-xL, thus displacing Bzx and apoptosis ensues. Both Mcl-1 and Bcl-xL are required to inhibit Bak pro-apoptotic activity; when Mcl-1 and Bcl-xL do not bind Bak, apoptosis is induced. Celecoxib interferes with pro-survival signals by downregulating Mcl-1. In Jurkatt T lymphoma cells treated with celecoxib, there was a sharp decline of Mcl-1, allowing Bak to trigger apoptosis. An abundance of Bcl-xL compensates for the function of Mcl-1 and blocked apoptosis in Mcl-1-negative cells [28]. The presence of Bak is more crucial than the presence of Bcl-2 for celecoxib-induced apoptosis to proceed. In fact, Jurkat cells that were Bak/Bax-negative showed nearly complete resistance to celecoxib-induced apoptosis, while overexpression of Bcl-2 only had limited anti-apoptotic effects [29]. Overall, the mechanism through which celecoxib induces apoptosis in intrinsic signaling pathway requires the presence of functional Bak. In human T-cell leukemia virus type I (HTLV-I), celecoxib blocks Akt/GSK3β survival signaling pathway to induce apoptosis via the intrinsic pathway that is accomplished by activating Bax and inhibiting the PI3K/Akt signaling pathway. This is associated with malignant transformation [30]. Possible mechanisms of Celebrex-mediated apoptosis are depicted in Figure 1.

Figure 1
Potential mechanisms of celebrex-mediated apoptosis. A. Either through direct cox-2 inhibition, or via induction of cyclins A, B and inhibition of cyclin-dependent kinase (CDK) activity, Celebrex inhibits tumor growth and induces cell cycle arrest. B. ...

Celecoxib interacts with other factors to induce apoptosis

Kaposi’s sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) are the etiological agents involved in aggressive NHL. PGE2 affects the eicosonoid (EP) receptors, epspecially EP1 and EP4, blocking apoptosis. By using Cox-2 inhibitor in combination with EP receptors antagonists, significant apoptosis was induced in EBV and KSHV-positive cells [31].

Cytotoxic signals that cause cells to undergo apoptosis are also activated by endorecticulum (ER)-stress. In Raji cells, the apoptotic effects of celecoxib could be improved in combination with bortezomib, the proteosome inhibitor that is also capable of inducing ER-stress [32].

In acute myeloid leukemia (AML), celecoxib and doxorubixin drastically reduced cell proliferation and increased apoptosis [33]. Downregulation of cyclin E and CDK-2, both of which are key regulators of cell cycle progression, was observed. The pro-apoptotic effect was also linked to G0/G1 phase cell arrest and survivin downregulation. Survivin, a multifunctional member of IAP family, interferes with caspases activation to suppress apoptosis [14]. In the presence of celecoxib and DMC, reduced survivin level prevents tumor growth more effectively. Survivin is potentially a Cox-2 independent target of celecoxib (Figure 1).

Immunotherapy for the treatment of non-Hodgkin’s lymphoma (NHL)

Several patients with NHL have undergone a novel immunotherapy modality that utilizes chimeric antigen receptors (CARs). CARs are fusion proteins that have both T-cell activation domain and antigen recognition moieties [34]. By genetically modifying T cells to express CARs, these T cells can specifically recognize specific surface markers such as CD19, a protein that is only expressed in B-cell lineages and not on hematopoeitic stem cells, thus effectively targeting CD19+ NHL. In one clinical trial, 4 of the 8 patients who had an infusion of anti-CD19-CAR-transduced T cells and a course of IL-2 in combination with chemotherapy exhibited long-term depletion of normal polyclonal CD19+ B-lineage cells, CD19 CAR T cells were detected in the blood of all patients [34]. However, like chemotherapy, tumors develop resistance through inherent or acquired anti-apoptotic mechanisms. New approaches are necessary to overcome this issue, and the use of celecoxib in treatment is a promising one.

Adoptive T cell therapy as an alternative approach to immunotherapy

Among all recent immune-based therapeutic strategies, adoptive T cell (ATC) therapy is a powerful tool that has promising potential in eradicating apoptosis-resistant tumor cells. Tumor-reactive lymphocytes are selected ex vivo and then adoptively transferred into patients. These lymphocytes are often administered with growth factors to enhance their survival, expansion, and cytotoxic potential in vivo [35]. In ATC using tumor-infiltrating lymphocytes (TILs), T cells are isolated from fresh biopsy specimens obtained from patients and tumor-specific T cells are selected and expanded using high levels of interleukin-2 (IL-2) [36]. Besides TIL-based ATC, in recent years, genetically modified T cells, such as T cell receptor (TCR) modified and chimeric antigen receptor (CAR) T cells [37], have gained great interest. These modified T cells have enhanced anti-tumor effects and higher specificity for tumor cells compared to regular T cells, thus improving the efficacy of the immune system of immunosuppressed patients.

T cell receptor (TCR) transgenic T cells’ mode of action and limitation

For TCR-engineered T cells, the desired TCR that is specific to a particular tumor associated antigen (TAA) is transferred to T cells via genetic means. When genes encoding TCR α and β chains are transferred into peripheral blood T lymphocytes, the antigen-specific recognition property of these lymphocytes is enhanced significantly [38]. T cells with highly expressed TCRs for MART-1 and gp100 antigens, both of which are expressed on melanomas and melanocytes, are more reactive to metastatic melanoma than regular T cells [39]. Inducing T cell immunity using TCR transgenic T cells is an appealing immunotherapeutic approach, but it has its own limitations. After prolonged exposure to transduced T cells, tumor cells undergo a change in which they express less or no antigens that these T cells can bind to, rendering the treatment ineffective. Administration of transduced T cells is limited to vaccination, which decreases the optimal response in patients with immunodeficiency [40]. Moreover, tumor recognition by TCR-engineered T cells is human leukocyte antigen (HLA)-dependent, making the induction of T cells immunity difficult. HLA is the human version of the major histocompability complex (MHC), a group of genes that encodes cell surface proteins to help lymphocytes distinguish host cells from foreign substances (self-recognition and non-self-recognition). Each individual has a specific MHC haplotype. If the transduced T cells derived from an individual that has an incompatible MHC haplotype with the recipient (MHC mismatch), then induction of immunity might not be successful [40].

Chimeric antigen receptor (CAR) T cells: structures and development

Unlike TCR-engineered T cells, CAR-engineered T cells recognize tumor cells in an HLA independent fashion. CAR T cells allow the use of a variety of different combination of signaling and costimulatory domains for optimal T cell recognition and activation. A CAR typically has a ligand-binding domain, such as a single-chain variable fragment (scFv) derived from a monoclonal antibody or an antigen-binding fragment (Fab), and a signaling domain, which usually has CD3ζ, a component of the TCR complex [41,42]. Antigen recognition mediated by scFv allows CAR-T cells to recognize their target independently of MHC [43]. CD3ζ serves as an activation domain. T cells activation is promoted by the phosphorylation of the tyrosines in immunoreceptor tyrosine-based activation motifs (ITAMs) of CD3ζ; the first and third ITAMs are linked to apoptosis [44].

Three generations of CAR-engineered T cells have been created so far. The first generation consists of only a ligand-binding domain and signaling domain without any co-stimulation [45]. The second and third generation have different co-stimulatory domains that help enhance the specificity of T cells as well as other effector functions such as proliferation and cytokine production [46]. In second generation CAR-T cells, the activation domain is fused with the co-stimulatory domain, which can be CD28, 4-1BB, OX40 or DAP10 [41]. Dual-signaling CAR-T cells help enhance the strength of signaling and the persistence of transduced T cells in the body [47]. Third generation CAR T cells have a second costimulatory domain added to the primary costimulatory domain that is used in the second generation. The additional costimulatory domain enhances the cytotoxic potential and effector functions of T cells, including proliferation, expansion and cytokine production, against tumors [48]. A common example of a third-generation CAR T cell would be CD28/4-1BB/CD3ζ [49].

Chimeric antigen receptor (CAR)-transduced T cell therapy for NHL

Lymphoma is a cancer caused by malignant lymphocytes. It has two subtypes: Hodgkin lymphoma (HL) and NHL. They differ in gene expression profiles. HL is commonly found as nodular sclerosis with high level of Hodgkin Reed-Sternberg (HRS) cells [50]. It is linked to primary mediastinal B cell lymphoma (PMBL), since both have amplified JAK2 gene expression [51]. Compared to HL, NHL is much more prevalent, accounting for 90% of lymphoma cases [52]. NHL has several subtypes, 85% of which arises from malignant B cells and the others arise from malignant T cells and natural killer (NK) lymphoma [52]. The subtypes of NHL are classified based on the stage of B cell differentiation that they are derived from and the type of protooncogenes that they expressed. There have been several revisions of NHL classification, but the generally accepted subtypes are: lymphoplasmacytic lymphoma, follicular lymphoma, mantle-cell lymphoma, marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT) lymphoma, diffuse large B-cell lymphoma, Burkitt’s lymphoma, and anaplastic large T-cell lymphoma [53-55]. Most of these lymphomas are of B cell origin, making them ideal targets for treatment using CAR-transduced T cells.

Anti-CD19 CAR-engineered T cell therapy for NHL

Among several CAR-transduced T cells designed so far, anti-CD19 CAR T cells are especially effective against B cells malignancies [42]. Anti-CD19 CAR-transduced T cells therapy is an appealing alternative to rituximab and CHOP in the treatment of NHL. CHOP is a first generation combination chemotherapy comprised of cyclophosphamide, doxorubicin, vincristine, and prednisone used for diffuse large-B-cell lymphoma [56]. It only induces complete response in 40% of elderly patients and the overall survival rate is only 35% [57]. CHOP is very toxic for elderly patients, but if given reduced CHOP regimens, the treatment would not be as effective [58]. Several attempts to increase the efficacy of CHOP have been made, and one of the most effective strategies is combining CHOP with rituximab, a chimeric anti-CD20 IgG1 monoclonal antibody. Rituximab binds specifically to CD20, an antigen expressed in 90% of B cell lymphomas [59]. It is quite effective when given as a single-agent to treat patients with indolent lymphoma [60]. In one study, a complete response of 76% was achieved in patients treated with CHOP and rituximab compared to a complete response of 63% in patients treated with CHOP alone [61]. CHOP plus rituximab (R-CHOP) treatment is also very effective in younger patients. Two groups of 18 to 60 years old were given either CHOP or CHOP plus rituximab [62]. The group given both CHOP and rituximab had a 79% of 3-year event-free survival compared to 59% of 3-year event-free survival in group given only CHOP [62]. Although R-CHOP has better efficacy than treatment with CHOP alone, tumor cells eventually develop resistance to this treatment in a similar fashion to other types of chemotherapy. To overcome this challenge, anti-CD19 CAR-engineered T cells were developed to make use of the immune system, reducing the severity of AEs due to toxic chemicals and increasing TAA specificity.

CD19 is an ideal antigen for immunotherapy, since it is expressed only on B-cell leukemia, lymphomas, and normal B cells, but not on other types of cells [63,64]. Anti-CD19 CAR T cells can target B-cell leukemia and lymphomas specifically without inducing apoptosis in other cell types, giving fewer side effects compared to other types of CAR-T cells that target more ubiquitous antigens. Gene that codes for anti-CD19 is put into scFv and the activation domain can be either CD28/CD3ζ or 4-1BB/CD3ζ. The costimulatory molecule CD28 is required for T cell activation and survival. It binds to B7.1 (CD80) and B7.2 (CD86) on tumor cells to trigger apoptosis [65]. In one trial, 5 patients with B-cell acute lymphoid leukemia (ALL) were treated with CD28-containing CD19-CAR T cells [66]. After lymphodepletion and CAR-T cell infusion, all patients achieved complete remission.

4-1BB, a member of TNF receptor superfamily, has a high affinity for 4-1BBL, a ligand that is expressed on activated macrophages and B cells [67]. A clinical trial using antiCD19scFv/4-1BB/CD3ζ to treat children with relapsed or refractory CD19+ ALL [68]. The overall survival (OS) was 78% and the persistence of CTL019 (CAR targeting CD19) cells continued for 1 to 26 months after infusion. In another clinical trial, out of the 8 patients with B cell lymphomas, 4 patients receiving an infusion of anti-CD19 CAR-T cells (antiCD19scFv/4-1BB/CD3ζ) and a course of IL-2 in combination with chemotherapy had long-term depletion of normal polyclonal CD19+ B-lineage cells [34]. Different levels of anti-CD19 CAR gene could be detected in the blood of all patients. For patient 1 to patient 6, the gene was detected within 20 days of initial infusion, while it was detected after 14 weeks for patient 7 and 8 weeks for patient 8 [34].

Mechanisms by which tumor cells avoid recognition by CAR T cells

Downregulation of CD19 expression on tumor cell surface

Treatment using CAR T cells has great potential, but there are some limitations that needed to be overcome. Since anti-CD19 CAR T cells mode of action relies heavily on the antigen recognition provided by the binding of anti-CD19 scFv on T cells to CD19 receptors on tumor cells, tumor cells can avoid anti-CD19 CAR T cells by downregulating or inhibiting CD19 expression. For example, the expression of C/EBPα and C/EBPβ in differentiated B cells will efficiently reprogram the cells into macrophages [69]. C/EBPs inhibits Pax5, a transcription factor of B cells, causing downregulation of CD19. Pax5 gene codes for several B-cell activator proteins (BSAPs), which are expressed exclusively in B-lymphoid lineage cells [70]. Therefore, downregulation of Pax5 will ultimately result in CD19 downregulation. In a timed course analysis, IL-7 enhanced the expression of CD19 on the surface of progenitor B lineage cells originated from human bone marrow [71]. In one study, patients with antibody-deficiency syndrome were found to have low level of CD19 expression due to a homozygous mutation in the CD19 gene despite having normal level of B cells [72]. In another study, immunodeficient patients who had normal amount of CD19 alleles but defective CD81 gene also had low level of CD19 expression [73]. In these cases, CD19 gene sequence remains intact, yet the cells lack the CD19 antigenic epitope, possibly caused by mutations in mRNA splicing [74].

Alternative B cell differentiation pathways

One patient with CLL undergoing anti-CD19 CAR-transduced T cells was reported to have mature lymphoma that could avoid T cells recognition by differentiating in a pathway different from that of its normal counterpart [75]. Prior to receiving anti-CD19 CAR-engineered T cells therapy, the patient was shown to already have partial loss of CD19 and other B cell markers by flow cytometry. This suggested that B cell differentiated abnormally before anti-CD19 CAR T cells were administered. During the course of treatment, CLL transformed to plasmablastic lymphoma (PBL) and CD19- leukemia. PBL had a mutation in complementarity-determining region 1 (CDR1), rendering the immunoglobulin heavy chain (IGH) reading frame unproductive. TP52 sequencing revealed a p.Gly245Ser in PBL [75].

Normal mature B cell repertoire requires the engagement of B cell activating factor receptor (BAFF-R) by BAFF [76]. CD19+ cells also have BAFF expression, but low or no mBAFF, which is expressed at a higher level in NHL macrophages than in healthy macrophages. The elevated level of BAFF might be linked to tumor B cell differentiation pathway that results in apoptosis triggered by CD19 antigen recognition.

Aberrant B cell antigen receptor (BCR) signaling pathways

B-cell antigen receptor (BCR) signaling pathways are highly regulated in normal B cells, but in NHL, they are aberrantly activated [77]. Many proteins involved in these pathways are activated by CD19. BCR signaling requires activation of protein tyrosine kinase (PTK). CD19 and Scr family PTKs undergo several phosphorylations, creating an amplification loop that greatly enhances B cell activation upon CD19 engagement [78]. Lyn, a member of Src family PTK and initiator of BCR signaling, is recruited by CD19 [79,80]. When BCR is activated, Lyn phosphorylates Vav and some residues of CD19 [80]. The activated Vav in turn initiates mitogen-activated protein kinase (MAPK) pathways. In malignant B cells, MAPKs, namely extracellular signal-regulated kinase (ERK) and p38, and Lyn are constitutively activated [81].

CD19 also initiates phosphoinositide-3-kinase (PI3K)/Akt signaling pathway. It activates PI3K by binding to the regulatory subunit p85 of PI3K [82]. PI3K in turn activates Akt downstream via phosphorylation, leading to activation of mTOR, the mammalian target of rampamycin [83]. In primary effusion lymphoma (PEL), a subtype of NHL, PI3K/Akt/mTOR pathway is constitutively active and dual inhibition of both PI3K and mTOR effectively inhibits tumor proliferation [84].

Bcl-2, a family of regulatory proteins, is commonly overexpressed in NHL, leading to resistance to apoptosis and promotion of tumorigenesis [85]. Over 40% of patients with diffuse large B cell lymphoma were reported to have high level of Bcl-2 expression [86]. Bcl-2 family consists of both pro-apoptotic and anti-apoptotic proteins. In most tumor types, the anti-apoptotic members of Bcl-2 tend to be overexpressed. Out of eight leukemia or lymphoma cell lines, seven were shown to have high levels of anti-apoptotic Bcl-2 proteins, especially Bfl-1, Mcl-1, and Bcl-xL [87]. Many different signaling pathways were potentially responsible for promoting Bcl-2 expression. Elevated levels of insulin-like growth factor-1 (IGF-1) were observed in malignant effusions, making it a possible marker for solid tumors [88]. When its receptor, IGF-1R, a receptor tyrosine kinase (RTK), is inhibited, tumor progression in chronic lymphocytic leukemia (CLL) is limited [89]. IGF-1 induces Bcl-2 promoter containing cAMP-response element (cAMP) site via cAMP-binding protein (CREB) signaling pathway [90]. Akt signaling was also found to play a role in this pathway. Cell lines expressing Akt showed increased CREB activity, which resulted in higher levels of Bcl-2 expression [91]. It is possible that by inhibiting Akt, NHL would become responsive again to apoptotic signals from the immune system or chemotherapy, as this method worked in a study using pancreatic tumor cell lines [92].

Aberrant expression of Bcl-2 family proteins is linked with drug resistance in various types of cancer [93]. In MCF-7 human breast cancer cells, estrogen induces the expression of Bcl-2 proto-oncogene transcripts significantly, leading to resistance to Adriamycin, a chemotherapy drug [94]. Lymphomas that have prolonged exposure to rituximab, chimeric anti-CD20 monoclonal antibody, become unresponsive to both rituximab and the chemotherapy drugs that rituximab is used with. Rituximab was not effective in chemosensitizing rituximab-resistant (RR) clones, developed from lymphoma lines, potentially due to hyperactivation of NF-κB and ERK1/2 pathways, resulting in over-expression of Bcl-2 and Mcl-1 in these clones [95]. Bcl-2 proteins are capable of enhancing drug resistance in cancer cells mainly due to their anti-apoptotic properties. By blocking Bax and Bak, which are pro-apoptotic proteins of Bcl-2 family, overexpressed anti-apoptotic Bcl-2 proteins (Bcl-2, Mcl-2, and Bcl-xL) inhibit the intrinsic apoptotic machinery of the cell [96]. Since anti-CD19 CAR therapy works by inducing both extrinsic and intrinsic apoptotic pathways, overexpression of anti-apoptotic Bcl-2 proteins might render the treatment ineffective. Using Bcl-2 family inhibitors, such as ABT-737, in combination with anti-CD19 CAR therapy can overcome this problem, increasing the efficacy of CAR T cells and managing tumor growth [97].

A study using CD19-/- mice demonstrated that CD19 propagated BCR-induced survival signals [82]. In clinical studies, mutated CD19 is linked to autoimmune diseases. A patient with a mutation in the splice acceptor site of intron 5 of maternal allele of CD19 had hypogammaglobinemia and no detectable antibodies against measles, rubella, tetanus and pertussis toxin, even though he was vaccinated [98]. In addition to BCR-dependent signaling pathways, CD19 is also capable of initiating MYC-driven lymphomagenesis [99]. Malignant B cell lymphomas have elevated levels of c-MYC, which in turn is greatly enhanced by CD19; therefore, by inhibiting CD19, the oncogenic capabilities of c-MYC is limited [100].

With CD19 as the upstream activator of several BCR-dependent signaling pathways, using anti-CD19 CAR therapy seems to be a promising therapeutic approach, as it will prevent the transient association of CD19 to BCR, thus inhibiting subsequent BCR-dependent signaling pathways that promote proliferation in NHL. Moreover, since CD19 is also involved in BCR-independent pathways, anti-CD19 CAR therapy potentially has more applications than previously reported.

Immunosuppressive microenvironment

The immunosuppressive microenvironment in which these cells grow is also an obstacle in using CAR T cells for cancer treatment. Tumor microenvironment has a high number of cytokines and immunosuppressive growth factors, including vascular endothelial growth factor (VEGF), interleukin (IL)-10, and transforming growth factor (TGF)-β [101]. These cytokines impede the anti-tumor activity of T cells, thus lowering the efficacy of CAR T cells. Before CAR T cells are administered, lymphodepletion is conducted. However, lymphodepletion might exacerbate the immunodeficiency that exists in patients, resulting in severe AEs caused by opportunistic diseases.

Anti-CD19 CAR T cells therapy has a larger range of cell depletion than anti-CD20 therapy, since it also affects pro-B cells and plasma cells (PCs) [102]. Anti-CD19 therapy works in a manner similar to anti-CD20 therapy, so it is possible that anti-CD19 therapy has the same drawbacks as anti-CD20 therapy. When treated with rituximab, a majority of patients had significantly impaired response to serological antibodies [103,104]. A possible way to mediate this problem is to supply patients with Ig antibodies via intravenous infusion [34], although the effectiveness of this approach remains unclear.

Another problem with CAR T cells therapy is that engineered T cells can persist in the body longer than the time they are designed to be. CAR T cells can cause several AEs such as fevers, hypotension, hypoxia, and neurologic changes [34,66,68], so the timing and doses of CAR T cells infusion need to be optimized to avoid tumor resistance to CAR T cells, high grade AEs, and exacerbated immunodeficiency.

Conclusions and future directions

The development and selective outgrowth of apoptosis-resistant tumor cells is a major hurdle in successful cancer therapy. Aberrant apoptotic machinery culminates in tumor cells that develop cross-resistance to a wide array of structurally and functionally distinct anti-cancer agents. Therefore, it is empirical to design novel approaches to modulate apoptotic machinery in order to bypass tumor resistance. Among new therapeutic drugs developed in recent years, celecoxib is a promising alternative. Its mechanism of action is flexible: it nduces apoptosis in the presence or absence of Cox-2 via intrinsic signaling pathway in a Bcl-2 independent and apoptosome-dependent manner. Additionally, several studies have shown that celecoxib further enhances apoptosis of tumor cells with minimal AEs when used in combination with other drugs, such as bortezomib, doxorubixin, and thalidomide.

Recent modern developments in utilization of the immune system to harness NHL suggest a promising role of CD19 CAR T cell therapy in NHL. However, a subset of tumor cells either inherently resistant or develop resistance to CAR-mediated immunotherapy. Based on the apoptotic gene regulatory effects of celecoxib, we propose that combination of CD19 CAR T cell therapy and celecoxib can potentially improve the treatment outcome of NHL patients (Figure 2).

Figure 2
CD19-redirected CAR T cell therapy of NHL. A. CD19-redirected CAR T cells can induce apoptotic cell death in CD19+ sensitive NHL cells, while those tumor cells with distorted apoptotic machinery exhibit resistance despite adequate surface CD19 expression. ...

Future research is warranted to understand the details of induction of apoptosis by celecoxib in various tumor models. This insight will allow more generalized, optimized, and effective treatments for patients. It will also help researchers expand the possible usage of celecoxib in combination with other anti-cancer modalities, such as histone deacetylase inhibitors (HDACi) in immunotherapy.

Disclosure of conflict of interest



1. Coussens LM, Werb Z. Inflammation and cancer. Nature. 2002;420:860–867. [PMC free article] [PubMed]
2. Karin M. NF-κB as a critical link between inflammation and cancer. Cold Spring Harb Perspect Biol. 2009;1:a000141. [PMC free article] [PubMed]
3. Gallouet AS, Travert M, Bresson-Bepoldin L, Guilloton F, Pangault C, Caulet-Maugendre S, Lamy T, Tarte K, Guillaudeux T. COX-2-independent effects of celecoxib sensitize lymphoma B cells to TRAIL-mediated apoptosis. Clin Cancer Res. 2014;20:2663–2673. [PubMed]
4. Crofford LJ. COX-1 and COX-2 tissue expression: implications and predictions. J Rheumatol Suppl. 1997;49:15–19. [PubMed]
5. Oshima M, Dinchuk JE, Kargman SL, Oshima H, Hancock B, Kwong E, Trzaskos JM, Evans JF, Taketo MM. Suppression of intestinal polyposis in ApcΔ716 knockout mice by inhibition of cyclooxygenase 2 (COX-2) Cell. 1996;87:803–809. [PubMed]
6. Ghosh N, Chaki R, Mandal V, Mandal SC. COX-2 as a target for cancer chemotherapy. Pharmacol Rep. 2010;62:233–244. [PubMed]
7. Greenhough A, Smartt HJ, Moore AE, Roberts HR, Williams AC, Paraskeva C, Kaidi A. The COX-2/PGE2 pathway: key roles in the hallmarks of cancer and adaptation to the tumour microenvironment. Carcinogenesis. 2009;30:377–386. [PubMed]
8. Lin MT, Lee RC, Yang PC, Ho FM, Kuo ML. Cyclooxygenase-2 inducing Mcl 1-dependent survival mechanism in human lung adenocarcinoma cl1. 0 cells involvement of phosphatidylinositol 3-KINASE/Akt pathway. J Biol Chem. 2001;276:48997–49002. [PubMed]
9. Dean JL, Brook M, Clark AR, Saklatvala J. p38 mitogen-activated protein kinase regulates cyclooxygenase-2 mRNA stability and transcription in lipopolysaccharide-treated human monocytes. J Biol Chem. 1999;274:264–269. [PubMed]
10. Jazirehi AR, Wenn PB, Damavand M. Therapeutic implications of targeting the PI3Kinase/AKT/mTOR signaling module in melanoma therapy. Am J Cancer Res. 2012;2:178–191. [PMC free article] [PubMed]
11. Adams JM, Cory S. The Bcl-2 apoptotic switch in cancer development and therapy. Oncogene. 2007;26:1324–1337. [PMC free article] [PubMed]
12. Bentires-Alj M, Dejardin E, Viatour P, Van Lint C, Froesch B, Reed JC, Merville MP, Bours V. Inhibition of the NF-κB transcription factor increases Bax expression in cancer cell lines. Oncogene. 2001;20:2805–2813. [PubMed]
13. Greijer AE, Van der Wall E. The role of hypoxia inducible factor 1 (HIF-1) in hypoxia induced apoptosis. J Clin Pathol. 2004;57:1009–1014. [PMC free article] [PubMed]
14. Jendrossek V. Targeting apoptosis pathways by Celecoxib in cancer. Cancer Lett. 2013;332:313–324. [PubMed]
15. Kardosh A, Wang W, Uddin J, Petasis NA, Hofman FM, Chen TC, Schonthal AH. Dimethyl-celecoxib (DMC), a derivative of celecoxib that lacks cyclooxygenase-2-inhibitory function, potently mimics the anti-tumor effects of celecoxib on Burkitt’s lymphoma in vitro and in vivo. Cancer Biol Ther. 2005;4:571–582. [PubMed]
16. Wang W, Kardosh A, Su YS, Schonthal AH, Chen TC. Efficacy of celecoxib in the treatment of CNS lymphomas: an in vivo model. Neurosurg Focus. 2006;21:E14. [PubMed]
17. Johansson AS, Pawelzik SC, Larefalk Å, Jakobsson PJ, Holmberg D, Lindskog M. Lymphoblastic T-cell lymphoma in mice is unaffected by celecoxib as single agent or in combination with cyclophosphamide. Leuk Lymphoma. 2009;50:1198–1203. [PubMed]
18. Bernard MP, Bancos S, Sime PJ, Phipps RP. Targeting cyclooxygenase-2 hematological malignancies: rationale and promise. Curr Pharm Des. 2008;14:2051. [PMC free article] [PubMed]
19. Prince HM, Mileshkin L, Roberts A, Ganju V, Underhill C, Catalano J, Bell R, Seymour JF, Westerman D, Simmons PJ, Lillie K. A multicenter phase II trial of thalidomide and celecoxib for patients with relapsed and refractory multiple myeloma. Clin Cancer Res. 2005;11:5504–5514. [PubMed]
20. Buckstein R, Kerbel RS, Shaked Y, Nayar R, Foden C, Turner R, Lee CR, Taylor D, Zhang L, Man S, Baruchel S. High-dose celecoxib and metronomic “low-dose” cyclophosphamide is an effective and safe therapy in patients with relapsed and refractory aggressive histology non-hodgkin’s lymphoma. Clin Cancer Res. 2006;12:5190–5198. [PubMed]
21. Kerbel RS, Klement G, Pritchard KI, Kamen B. Continuous low-dose anti-angiogenic/metronomic chemotherapy: from the research laboratory into the oncology clinic. Ann Oncol. 2002;13:12–15. [PubMed]
22. Buckstein R, Kerbel RS, Shaked Y, Nayar R, Foden C, Turner R, Lee CR, Taylor D, Zhang L, Man S, Baruchel S, Stempak D, Bertolini F, Crump M. High-dose celecoxib and metronomic “low-dose” cyclophosphamide is an effective and safe therapy in patients with relapsed and refractory aggressive histology non-Hodgkin’s lymphoma. Clin Cancer Res. 2006;12:5190–5198. [PubMed]
23. Waskewich C, Blumenthal RD, Li H, Stein R, Goldenberg DM, Burton J. Celecoxib exhibits the greatest potency amongst cyclooxygenase (COX) inhibitors for growth inhibition of COX-2-negative hematopoietic and epithelial cell lines. Cancer Res. 2002;62:2029–2033. [PubMed]
24. Song X, Lin HP, Johnson AJ, Tseng PH, Yang YT, Kulp SK, Chen CS. Cyclooxygenase-2, player or spectator in cyclooxygenase-2 inhibitor-induced apoptosis in prostate cancer cells. J Natl Cancer Inst. 2002;94:585–591. [PubMed]
25. Jendrossek V, Handrick R, Belka C. Celecoxib activates a novel mitochondrial apoptosis signaling pathway. FASEB J. 2003;17:1547–1549. [PubMed]
26. Vogel CF, Li W, Sciullo E, Newman J, Hammock B, Reader JR, Tuscano J, Matsumura F. Pathogenesis of aryl hydrocarbon receptor-mediated development of lymphoma is associated with increased cyclooxygenase-2 expression. Am J Pathol. 2007;171:1538–1548. [PubMed]
27. Willis SN, Chen L, Dewson G, Wei A, Naik E, Fletcher JI, Adams JM, Huang DC. Proapoptotic Bak is sequestered by Mcl-1 and Bcl-xL, but not Bcl-2, until displaced by BH3 only proteins. Genes Dev. 2005;19:1294–1305. [PubMed]
28. Rudner J, Elsaesser SJ, Jendrossek V, Huber SM. Anti-apoptotic Bcl-2 fails to form efficient complexes with pro-apoptotic Bak to protect from celecoxib-induced apoptosis. Biochem Pharmacol. 2011;81:32–42. [PubMed]
29. Müller AC, Handrick R, Elsaesser SJ, Rudner J, Henke G, Ganswindt U, Belka C, Jendrossek V. Importance of Bak for celecoxib-induced apoptosis. Biochem Pharmacol. 2008;76:1082–1096. [PubMed]
30. Sinha-Datta U, Taylor JM, Brown M, Nicot C. Celecoxib disrupts the canonical apoptotic network in HTLV-I cells through activation of Bax and inhibition of PKB/Akt. Apoptosis. 2008;13:33–40. [PubMed]
31. Paul AG, Chandran B, Sharma-Walia N. Concurrent targeting of EP1/EP4 receptors and COX-2 induces synergistic apoptosis in KSHV and EBV associated non Hodgkin lymphoma cell lines. Transl Res. 2013;161:447. [PMC free article] [PubMed]
32. Chen ST, Thomas S, Gaffney KJ, Louie SG, Petasis NA, Schönthal AH. Cytotoxic effects of celecoxib on Raji lymphoma cells correlate with aggravated endoplasmic reticulum stress but not with inhibition of cyclooxygenase-2. Leuk Res. 2010;34:250–253. [PubMed]
33. Chen C, Xu W, Wang CM. Combination of celecoxib and doxorubicin increases growth inhibition and apoptosis in acute myeloid leukemia cells. Leuk Lymphoma. 2013;54:2517–2522. [PubMed]
34. Kochenderfer JN, Dudley ME, Feldman SA, Wilson WH, Spaner DE, Maric I, Stetler-Stevenson M, Phan GQ, Hughes MS, Sherry RM, Yang JC, Kammula US, Devillier L, Carpenter R, Nathan DA, Morgan RA, Laurencot C, Rosenberg SA. B-cell depletion and remissions of malignancy along with cytokine-associated toxicity in a clinical trial of anti-CD19 chimeric-antigen-receptor-transduced T cells. Blood. 2012;119:2709–2720. [PubMed]
35. Rosenberg SA, Restifo NP, Yang JC, Morgan RA, Dudley ME. Adoptive cell transfer: a clinical path to effective cancer immunotherapy. Nat Rev Cancer. 2008;8:299–308. [PMC free article] [PubMed]
36. June CH. Adoptive T cell therapy for cancer in the clinic. J Clin Invest. 2007;117:1466–1476. [PMC free article] [PubMed]
37. Wang M, Yin B, Wang HY, Wang RF. Current advances in T-cell-based cancer immunotherapy. Immunotherapy. 2014;6:1265–1278. [PMC free article] [PubMed]
38. Kessels HW, Wolkers MC, van den Boom MD, van den Valk MA, Schumacher TN. Immunotherapy through TCR gene transfer. Nat Immunol. 2001;2:957–961. [PubMed]
39. Johnson LA, Morgan RA, Dudley ME, Cassard L, Yang JC, Hughes MS, Kammula US, Royal RE, Sherry RM, Wunderlich JR, Lee CC. Gene therapy with human and mouse T-cell receptors mediates cancer regression and targets normal tissues expressing cognate antigen. Blood. 2009;114:535–546. [PubMed]
40. Schumacher TN. T-cell-receptor gene therapy. Nat Rev Immunol. 2002;2:512–519. [PubMed]
41. Davila ML, Brentjens R, Wang X, Rivière I, Sadelain M. How do CARs work? Early insights from recent clinical studies targeting CD19. Oncoimmunology. 2012;1:1577–1583. [PMC free article] [PubMed]
42. Kenderian SS, Ruella M, Gill S, Kalos M. Chimeric antigen receptor T-cell therapy to target hematologic malignancies. Cancer Res. 2014;74:6383–6389. [PubMed]
43. Eshhar Z. From the mouse cage to human therapy: a personal perspective of the emergence of T-bodies/chimeric antigen receptor T cells. Hum Gene Ther. 2014;25:773–778. [PMC free article] [PubMed]
44. Combadiere B, Freedman M, Chen L, Shores EW, Love P, Lenardo MJ. Qualitative and quantitative contributions of the T cell receptor zeta chain to mature T cell apoptosis. J Exp Med. 1996;183:2109–2117. [PMC free article] [PubMed]
45. Till BG, Jensen MC, Wang J, Chen EY, Wood BL, Greisman HA, Qian X, James SE, Raubitschek A, Forman SJ, Gopal AK. Adoptive immunotherapy for indolent non-Hodgkin lymphoma and mantle cell lymphoma using genetically modified autologous CD20-specific T cells. Blood. 2008;112:2261–2271. [PubMed]
46. Milone MC, Fish JD, Carpenito C, Carroll RG, Binder GK, Teachey D, Samanta M, Lakhal M, Gloss B, Danet-Desnoyers G, Campana D. Chimeric receptors containing CD137 signal transduction domains mediate enhanced survival of T cells and increased antileukemic efficacy in vivo. Mol Ther. 2009;17:1453–1464. [PMC free article] [PubMed]
47. Sadelain M, Brentjens R, Rivière I. The basic principles of chimeric antigen receptor design. Cancer Discov. 2013;3:388–398. [PMC free article] [PubMed]
48. Ritchie DS, Neeson PJ, Khot A, Peinert S, Tai T, Tainton K, Chen K, Shin M, Wall DM, Hönemann D, Gambell P. Persistence and efficacy of second generation CAR T cell against the LeY antigen in acute myeloid leukemia. Mol Ther. 2013;21:2122–2129. [PubMed]
49. Gilham DE, Debets R, Pule M, Hawkins RE, Abken H. CAR-T cells and solid tumors: tuning T cells to challenge an inveterate foe. Trends Mol Med. 2012;18:377–384. [PubMed]
50. Rosenwald A, Wright G, Leroy K, Yu X, Gaulard P, Gascoyne RD, Chan WC, Zhao T, Haioun C, Greiner TC, Weisenburger DD. Molecular diagnosis of primary mediastinal B cell lymphoma identifies a clinically favorable subgroup of diffuse large B cell lymphoma related to Hodgkin lymphoma. J Exp Med. 2003;198:851–862. [PMC free article] [PubMed]
51. Joos S, Küpper M, Ohl S, von Bonin F, Mechtersheimer G, Bentz M, Marynen P, Möller P, Pfreundschuh M, Trümper L, Lichter P. Genomic imbalances including amplification of the tyrosine kinase gene JAK2 in CD30+ Hodgkin cells. Cancer Res. 2000;60:549–552. [PubMed]
52. Shankland KR, Armitage JO, Hancock BW. Non-Hodgkin lymphoma. Lancet. 2012;380:848–857. [PubMed]
53. Evans LS, Hancock BW. Non-hodgkin lymphoma. Lancet. 2003;362:139–146. [PubMed]
54. Bennett M, Farrer-Brown G, Henry K, Jelliffe AM, Gerard-Marchant R, Hamlin I, Lennert K, Rilke F, Stansfeld AG, Van Unnik JA. Classification of non-Hodgkin’s lymphomas. Lancet. 1974;304:405–408.
55. Armitage JO, Weisenburger DD. New approach to classifying non-Hodgkin’s lymphomas: clinical features of the major histologic subtypes. Non-Hodgkin’s lymphoma classification project. J. Clin. Oncol. 1998;16:2780–2795. [PubMed]
56. Fisher RI, Gaynor ER, Dahlberg S, Oken MM, Grogan TM, Mize EM, Glick JH, Coltman CA Jr, Miller TP. Comparison of a standard regimen (CHOP) with three intensive chemotherapy regimens for advanced non-Hodgkin’s lymphoma. New Engl J Med. 1993;328:1002–1006. [PubMed]
57. Sonneveld P, de Ridder M, van der Lelie H, Nieuwenhuis K, Schouten H, Mulder A, van Reijswoud I, Hop W, Lowenberg B. Comparison of doxorubicin and mitoxantrone in the treatment of elderly patients with advanced diffuse non-Hodgkin’s lymphoma using CHOP versus CNOP chemotherapy. J. Clin. Oncol. 1995;13:2530–2539. [PubMed]
58. Meyer RM, Browman GP, Samosh ML, Benger AM, Bryant-Lukosius D, Wilson WE, Frank GL, Leber BF, Sternbach MS, Foster GA. Randomized phase II comparison of standard CHOP with weekly CHOP in elderly patients with non-Hodgkin’s lymphoma. J. Clin. Oncol. 1995;13:2386–2393. [PubMed]
59. McLaughlin P, Grillo-López AJ, Link BK, Levy R, Czuczman MS, Williams ME, Heyman MR, Bence-Bruckler I, White CA, Cabanillas F, Jain V. Rituximab chimeric anti-CD20 monoclonal antibody therapy for relapsed indolent lymphoma: half of patients respond to a four-dose treatment program. J. Clin. Oncol. 1998;16:2825–2833. [PubMed]
60. McLaughlin P, Hagemeister FB, Grillo-López AJ. Rituximab in indolent lymphoma: the single-agent pivotal trial. Semin Oncol. 1999;26:79–87. [PubMed]
61. Coiffier B, Lepage E, Briere J, Herbrecht R, Tilly H, Bouabdallah R, Morel P, Van Den Neste E, Salles G, Gaulard P, Reyes F. CHOP chemotherapy plus rituximab compared with CHOP alone in elderly patients with diffuse large-B-cell lymphoma. New Engl J Med. 2002;346:235–242. [PubMed]
62. Pfreundschuh M, Trümper L, Österborg A, Pettengell R, Trneny M, Imrie K, Ma D, Gill D, Walewski J, Zinzani PL, Stahel R. CHOP-like chemotherapy plus rituximab versus CHOP-like chemotherapy alone in young patients with good-prognosis diffuse large-B-cell lymphoma: a randomised controlled trial by the Mabthera international trial (MInT) group. Lancet Oncol. 2006;7:379–391. [PubMed]
63. Li YS, Wasserman R, Hayakawa K, Hardy RR. Identification of the earliest B lineage stage in mouse bone marrow. Immunity. 1996;5:527–535. [PubMed]
64. Wang K, Wei G, Liu D. CD19: a biomarker for B cell development, lymphoma diagnosis and therapy. Exp Hematol Oncol. 2012;1:1. [PMC free article] [PubMed]
65. Linsley PS, Greene JL, Brady W, Bajorath J, Ledbetter JA, Peach R. Human B7-1 (CD80) and B7-2 (CD86) bind with similar avidities but distinct kinetics to CD28 and CTLA-4 receptors. Immunity. 1994;1:793–801. [PubMed]
66. Brentjens RJ, Davila ML, Riviere I, Park J, Wang X, Cowell LG, Bartido S, Stefanski J, Taylor C, Olszewska M, Borquez-Ojeda O, Qu J, Wasielewska T, He Q, Bernal Y, Rijo IV, Hedvat C, Kobos R, Curran K, Steinherz P, Jurcic J, Rosenblat T, Maslak P, Frattini M, Sadelain M. CD19-targeted T cells rapidly induce molecular remissions in adults with chemotherapy-refractory acute lymphoblastic leukemia. Sci Transl Med. 2013;5 177ra38. [PMC free article] [PubMed]
67. Melero I, Shuford WW, Newby SA, Aruffo A, Ledbetter JA, Hellström KE, Mittler RS, Chen L. Monoclonal antibodies against the 4-1BB T-cell activation molecule eradicate established tumors. Nat Med. 1997;3:682–685. [PubMed]
68. Grupp SA, Kalos M, Barrett D, Aplenc R, Porter DL, Rheingold SR, Teachey DT, Chew A, Hauck B, Wright JF, Milone MC. Chimeric antigen receptor-modified T cells for acute lymphoid leukemia. New Engl J Med. 2013;368:1509–1518. [PMC free article] [PubMed]
69. Xie H, Ye M, Feng R, Graf T. Stepwise reprogramming of B cells into macrophages. Cell. 2004;117:663–676. [PubMed]
70. Nutt SL, Heavey B, Rolink AG, Busslinger M. Commitment to the B-lymphoid lineage depends on the transcription factor Pax5. Nature. 1999;401:556–562. [PubMed]
71. Billips LG, Nuñez CA, Bertrand FE, Stankovic AK, Gartland GL, Burrows PD, Cooper MD. Immunoglobulin recombinase gene activity is modulated reciprocally by interleukin 7 and CD19 in B cell progenitors. J Exp Med. 1995;182:973–982. [PMC free article] [PubMed]
72. van Zelm MC, Reisli I, van der Burg M, Castaño D, van Noesel CJ, van Tol MJ, Woellner C, Grimbacher B, Patiño PJ, van Dongen JJ, Franco JL. An antibody-deficiency syndrome due to mutations in the CD19 gene. New Engl J Med. 2006;354:1901–1912. [PubMed]
73. van Zelm MC, Smet J, Adams B, Mascart F, Schandené L, Janssen F, Ferster A, Kuo CC, Levy S, van Dongen JJ, van der Burg M. CD81 gene defect in humans disrupts CD19 complex formation and leads to antibody deficiency. J Clin Invest. 2010;120:1265–1274. [PMC free article] [PubMed]
74. Onea AS, Jazirehi AR. CD19 chimeric antigen receptor (CD19 CAR)-redirected adoptive T-cell immunotherapy for the treatment of relapsed or refractory B-cell non-Hodgkin’s lymphomas. Am J Cancer Res. 2016;6:403. [PMC free article] [PubMed]
75. Evans AG, Rothberg PG, Burack WR, Huntington SF, Porter DL, Friedberg JW, Liesveld JL. Evolution to plasmablastic lymphoma evades CD19-directed chimeric antigen receptor T cells. Br J Haematol. 2015;171:205–209. [PubMed]
76. He B, Chadburn A, Jou E, Schattner EJ, Knowles DM, Cerutti A. Lymphoma B cells evade apoptosis through the TNF family members BAFF/BLyS and APRIL. J Immunol. 2004;172:3268–3279. [PubMed]
77. Woyach JA, Johnson AJ, Byrd JC. The B-cell receptor signaling pathway as a therapeutic target in CLL. Blood. 2012;120:1175–1184. [PubMed]
78. Fujimoto M, Fujimoto Y, Poe JC, Jansen PJ, Lowell CA, DeFranco AL, Tedder TF. CD19 regulates Src family protein tyrosine kinase activation in B lymphocytes through processive amplification. Immunity. 2000;13:47–57. [PubMed]
79. van Noesel CJ, Lankester AC, van Schijndel GM, van Lier RA. The CR2/CD19 complex on human B cells contains the src-family kinase Lyn. Int Immunol. 1993;5:699–705. [PubMed]
80. Roifman CM, Ke S. CD19 is a substrate of the antigen receptor-associated protein tyrosine kinase in human B cells. Biochem Biophys Res Commun. 1993;194:222–225. [PubMed]
81. Ogasawara T, Yasuyama M, Kawauchi K. Constitutive activation of extracellular signal-regulated kinase and p38 mitogen-activated protein kinase in B-cell lymphoproliferative disorders. Int J Hematol. 2003;77:364–370. [PubMed]
82. Otero DC, Anzelon AN, Rickert RC. CD19 function in early and late B cell development: I. Maintenance of follicular and marginal zone B cells requires CD19 dependent survival signals. J Immunol. 2003;170:73–83. [PubMed]
83. Hahn-Windgassen A, Nogueira V, Chen CC, Skeen JE, Sonenberg N, Hay N. Akt activates the mammalian target of rapamycin by regulating cellular ATP level and AMPK activity. J Biol Chem. 2005;280:32081–32089. [PubMed]
84. Bhatt AP, Bhende PM, Sin SH, Roy D, Dittmer DP, Damania B. Dual inhibition of PI3K and mTOR inhibits autocrine and paracrine proliferative loops in PI3K/Akt/mTOR-addicted lymphomas. Blood. 2010;115:4455–4463. [PubMed]
85. Webb A, Cunningham D, Cotter F, Clarke PA, Di Stefano F, Ross P, Corbo M, Dziewanowska Z. BCL-2 antisense therapy in patients with non-Hodgkin lymphoma. Lancet. 1997;349:1137–1141. [PubMed]
86. Hermine O, Haioun C, Lepage E, d’Agay MF, Briere J, Lavignac C, Fillet G, Salles G, Marolleau JP, Diebold J, Reyas F, Gaulard P. Prognostic significance of bcl-2 protein expression in aggressive non-Hodgkin’s lymphoma. Groupe d’Etude des Lymphomes de l’Adulte (GELA) Blood. 1996;87:265–272. [PubMed]
87. Placzek WJ, Wei J, Kitada S, Zhai D, Reed JC, Pellecchia M. A survey of the anti-apoptotic Bcl-2 subfamily expression in cancer types provides a platform to predict the efficacy of Bcl-2 antagonists in cancer therapy. Cell Death Dis. 2010;1:e40. [PMC free article] [PubMed]
88. Olchovsky D, Shimon I, Goldberg I, Shulimzon T, Lubetsky A, Yellin A, Pariente C, Karasik A, Kanety H. Elevated insulin-like growth factor-1 and insulin-like growth factor binding protein-2 in malignant pleural effusion. Acta Oncol. 2002;41:182–187. [PubMed]
89. Yaktapour N, Übelhart R, Schüler J, Aumann K, Dierks C, Burger M, Pfeifer D, Jumaa H, Veelken H, Brummer T, Zirlik K. Insulin-like growth factor-1 receptor (IGF1R) as a novel target in chronic lymphocytic leukemia. Blood. 2013;122:1621–1633. [PubMed]
90. Pugazhenthi S, Miller E, Sable C, Young P, Heidenreich KA, Boxer LM, Reusch JE. Insulin-like growth factor-I induces bcl-2 promoter through the transcription factor cAMP-response element-binding protein. J Biol Chem. 1999;274:27529–35. [PubMed]
91. Pugazhenthi S, Nesterova A, Sable C, Heidenreich KA, Boxer LM, Heasley LE, Reusch JE. Akt/protein kinase B up-regulates Bcl-2 expression through cAMP-response element-binding protein. J Biol Chem. 2000;275:10761–10766. [PubMed]
92. Fahy BN, Schlieman M, Virudachalam S, Bold RJ. AKT inhibition is associated with chemosensitisation in the pancreatic cancer cell line MIA-PaCa-2. Br J Cancer. 2003;89:391–397. [PMC free article] [PubMed]
93. Kang MH, Reynolds CP. Bcl-2 inhibitors: targeting mitochondrial apoptotic pathways in cancer therapy. Clin Cancer Res. 2009;15:1126–1132. [PMC free article] [PubMed]
94. Teixeira C, Reed JC, Pratt MC. Estrogen promotes chemotherapeutic drug resistance by a mechanism involving Bcl-2 proto-oncogene expression in human breast cancer cells. Cancer Res. 1995;55:3902–3907. [PubMed]
95. Jazirehi AR, Vega MI, Bonavida B. Development of rituximab-resistant lymphoma clones with altered cell signaling and cross-resistance to chemotherapy. Cancer Res. 2007;67:1270–1281. [PubMed]
96. Emily HY, Wei MC, Weiler S, Flavell RA, Mak TW, Lindsten T, Korsmeyer SJ. BCL-2, BCL-XL sequester BH3 domain-only molecules preventing BAX-and BAK-mediated mitochondrial apoptosis. Mol Cell. 2001;8:705–711. [PubMed]
97. Karlsson SC, Lindqvist AC, Fransson M, Paul-Wetterberg G, Nilsson B, Essand M, Nilsson K, Frisk P, Jernberg-Wiklund H, Loskog SI. Combining CAR T cells and the Bcl-2 family apoptosis inhibitor ABT-737 for treating B-cell malignancy. Cancer Gene Ther. 2013;20:386–393. [PubMed]
98. Kanegane H, Agematsu K, Futatani T, Sira MM, Suga K, Sekiguchi T, van Zelm MC, Miyawaki T. Novel mutations in a Japanese patient with CD19 deficiency. Genes Immun. 2007;8:663–670. [PubMed]
99. Chung EY, Psathas JN, Yu D, Li Y, Weiss MJ, Thomas-Tikhonenko A. CD19 is a major B cell receptor-independent activator of MYC-driven B-lymphomagenesis. J Clin Invest. 2012;122:2257–2266. [PMC free article] [PubMed]
100. Poe JC, Minard-Colin V, Kountikov EI, Haas KM, Tedder TF. A c-Myc and surface CD19 signaling amplification loop promotes B cell lymphoma development and progression in mice. J Immunol. 2012;189:2318–2325. [PMC free article] [PubMed]
101. De Souza AP, Bonorino C. Tumor immunosuppressive environment: effects on tumor-specific and nontumor antigen immune responses. Expert Rev Anticancer Ther. 2009;9:1317–1332. [PubMed]
102. Mei HE, Schmidt S, Dorner T. Rationale of anti-CD19 immunotherapy: an option to target autoreactive plasma cells in autoimmunity. Arthritis Res Ther. 2012;14:S1. [PMC free article] [PubMed]
103. van der Kolk LE, Baars JW, Prins MH, van Oers MH. Rituximab treatment results in impaired secondary humoral immune responsiveness. Blood. 2002;100:2257–2259. [PubMed]
104. Bingham CO, Looney RJ, Deodhar A, Halsey N, Greenwald M, Codding C, Trzaskoma B, Martin F, Agarwal S, Kelman A. Immunization responses in rheumatoid arthritis patients treated with rituximab: results from a controlled clinical trial. Arthritis Rheum. 2010;62:64–74. [PubMed]

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