Apoptosis Inhibitor 5: A Multifaceted Regulator of Cell Fate.

Apoptosis, or programmed cell death, is a fundamental process that maintains tissue homeostasis, eliminates damaged or infected cells, and plays a crucial role in various biological phenomena. The deregulation of apoptosis is involved in many human diseases, including cancer. One of the emerging players in the intricate regulatory network of apoptosis is apoptosis inhibitor 5 (API5), also called AAC-11 (anti-apoptosis clone 11) or FIF (fibroblast growth factor-2 interacting factor). While it may not have yet the same level of notoriety as some other cancer-associated proteins, API5 has garnered increasing attention in the cancer field in recent years, as elevated API5 levels are often associated with aggressive tumor behavior, resistance to therapy, and poor patient prognosis. This review aims to shed light on the multifaceted functions and regulatory mechanisms of API5 in cell fate decisions as well as its interest as therapeutic target in cancer.

A phase 1 trial of the anti-KIR antibody IPH2101 in patients with relapsed/refractory multiple myeloma.

Natural killer (NK) cells elicit cytotoxicity against multiple myeloma (MM); however, MM cells express HLA class I molecules as ligands to NK cell inhibitory killer immunoglobulin-like receptors (KIRs) as a means of immunoevasion. KIR-ligand mismatch may improve outcomes in allogeneic transplantation for MM. Extrapolating on this concept, we conducted a phase 1 trial of IPH2101, an anti-KIR antibody, in patients with relapsed/refractory MM. IPH2101 was administered intravenously every 28 days in 7 dose-escalated cohorts (0.0003-3 mg/kg) for up to 4 cycles. Pharmacokinetic, pharmacodynamic, and correlative immunologic studies were completed. A total of 32 patients were enrolled. The biologic endpoint of full KIR2D occupancy across the dosing cycle was achieved without dose-limiting toxicity or maximally tolerated dose. One severe adverse event was noted. Pharmacokinetic and pharmacodynamic findings approximated preclinical predictions, and IPH2101 enhanced ex vivo patient-derived NK cell cytotoxicity against MM. No objective responses were seen. No evidence of autoimmunity was observed. These findings suggest that IPH2101 is safe and tolerable at doses that achieve full inhibitory KIR saturation, and this approach warrants further development in MM. This trial was registered at www.clinicaltrials.gov as #NCT00552396.

Phase I study of bromohydrin pyrophosphate (BrHPP, IPH 1101), a Vgamma9Vdelta2 T lymphocyte agonist in patients with solid tumors.

PURPOSE: Vgamma9Vdelta2 (gammadelta) T lymphocytes, a critical peripheral blood lymphocyte subset, are directly cytotoxic against many solid and hematologic tumor types. Vgamma9Vdelta2 T lymphocytes can be selectively expanded in vivo with BrHPP (IPH1101) and IL-2. The present phase I trial was conducted with the aim of determining the maximum-tolerated dose (MTD) and safety of IPH1101 combined with a low dose of IL-2 in patients with solid tumors. EXPERIMENTAL DESIGN: A 1-h intravenous infusion of IPH11 was administered alone at cycle 1, combined with a low dose of SC IL-2 (1 MIU/M(2) d1 to d7) in the subsequent cycles (day 1 every 3 weeks). The dose of IPH1101 was escalated from 200 to 1,800 mg/m(2). RESULTS: As much as 28 patients with solid tumors underwent a total of 109 treatment cycles. Pharmacodynamics data demonstrate that gammadelta T lymphocyte amplification in humans requires the co-administration of IL-2 and is dependent on IPH 1101 dose. Dose-limiting toxicity occurred in two patients at a dose of 1,800 mg/m(2): one grade 3 fever (1 patient) and one grade 3 hypotension (1 patient) suggesting cytokine release syndrome immediately following the first infusion. At lower doses the treatment was well tolerated; the most frequent adverse events were mild fever, chills and abdominal pain, without exacerbation in the IL-2 combined cycles. CONCLUSION: IPH1101 in combination with SC low-dose IL-2 is safe, well tolerated and induces a potent gammadelta T lymphocyte expansion in patients. Its clinical activity will be evaluated in phase II clinical trials.

Phase-I study of Innacell gammadelta, an autologous cell-therapy product highly enriched in gamma9delta2 T lymphocytes, in combination with IL-2, in patients with metastatic renal cell carcinoma.

PURPOSE: gamma9delta2 T lymphocytes have been shown to be directly cytotoxic against renal carcinoma cells. Lymphocytes T gammadelta can be selectively expanded in vivo with BrHPP (IPH1101, Phosphostim) and interleukin 2 (IL-2). A phase I Study was conducted in patients with metastatic renal cell carcinoma (mRCC) to determine the maximum-tolerated dose and safety of Innacell gammadelta, an autologous cell-therapy product based on gamma9delta2 T lymphocytes, in patients with mRCC. EXPERIMENTAL DESIGN: A 1-h intravenous infusion of gamma9delta2 T lymphocytes was administered alone during treatment cycle 1 and combined with a low dose of subcutaneous interleukin-2 (IL-2, 2 MIU/m2 from Day 1 to Day 7) in the two subsequent cycles (at 3-week intervals). The dose of gamma9delta2 T lymphocytes was escalated from 1 up to 8 x 10(9) cells. RESULTS: Ten patients underwent a total of 27 treatment cycles. Immunomonitoring data demonstrate that gamma9delta2 T lymphocytes are initially cleared from the blood to reappear at the end of IL-2 administration. Dose-limiting toxicity occurred in one patient at the dose of 8 x 10(9) cells (disseminated intravascular coagulation). Other treatment-related adverse events (AEs) included mainly gastrointestinal disorders and flu-like symptoms (fatigue, pyrexia, rigors). Hypotension and tachycardia also occurred, especially with co-administered IL-2. Six patients showed stabilized disease. Time to progression was 25.7 weeks. CONCLUSION: The data collected in ten patients with mRCC indicate that repeated infusions of Innacell gammadelta at different dose levels (up to 8 x 10(9) total cells), either alone or with IL-2 is well tolerated. These results are in favor of the therapeutic value of cell therapy with Innacell gammadelta for the treatment of cancers.

Large scale expansion of gamma 9 delta 2 T lymphocytes: Innacell gamma delta cell therapy product.

gamma9delta2 T lymphocytes are non-conventional lymphocytes presenting a direct cytotoxic effect against a broad range of tumour targets. These cells also secrete inflammatory cytokines that can boost the other components of the immune system. In contrast to conventional CD8(+) T cells, the cytotoxic effect of gamma9delta2 T lymphocytes does not depend on the expression of major histocompatibility complex molecules by target tumour cells. INNACELL gammadeltatrade mark is a cell therapy product obtained by ex vivo amplification of mononuclear cells. The stimulation is achieved by a specific synthetic agonist of gamma9delta2 T lymphocytes, bromohydrin pyrophosphate (BrHPP). After a single stimulation with BrHPP, gamma9delta2 T lymphocytes are expanded for 2 weeks in a closed system in culture medium with interleukin-2 (IL-2). On day 15, cells are washed and harvested in 4% human serum albumin. In this manufacturing process, the total cell population is expanded by approximately 10-fold and gamma9delta2 T lymphocytes undergo a specific 1000-fold expansion, corresponding to a gamma9delta2 T lymphocyte enrichment of more than 70% at the end of the culture. This manufacturing process is much simpler than most current cellular therapy approaches using conventional CD8(+) T-cell lines or clones: there is no final or initial separation, no purification step and no use of feeder cells; the specific T-cell receptor-mediated signal provided by BrHPP is sufficient to trigger the IL-2-dependent expansion of the gamma9delta2 subset, which then becomes predominant in the cell culture in large amounts.

In vivo immunomanipulation of V gamma 9V delta 2 T cells with a synthetic phosphoantigen in a preclinical nonhuman primate model.

Vgamma9Vdelta2(+) cells represent the major population of gammadelta T cells in primate blood and react in an MHC-unrestricted fashion to a set of low m.w. nonpeptide phosphoantigens. Two types of structurally related agonists have been discovered so far: the natural phosphoantigens (hydroxydimethyl allyl-pyrophosphate or isopentenyl-pyrophosphate (IPP)) acting directly on Vgamma9Vdelta2(+) TCR and aminobisphosphonates, which block the mevalonate pathway in target cells, leading to accumulation of natural phosphoantigens that in turn activate Vgamma9Vdelta2(+) cells. We demonstrate in the cynomolgus monkey that Vgamma9Vdelta2 can be manipulated in vivo with bromohydrin pyrophosphate (BrHPP)/Phosphostim, a potent synthetic agonist for which the mechanism of action is similar to natural phosphoantigens. Although of very short half-life, injection of BrHPP leads to strong activation of Vgamma9Vdelta2, inducing production of a high level of Th1 cytokines. Combination of BrHPP with low-dose rhIL-2 induces specific amplification of effector-memory peripheral Vgamma9Vdelta2 in blood in a dose-dependant manner. This transient response returns to baseline within 10-15 days. Successive infusions of BrHPP and rhIL-2 induce less vigorous expansions, suggesting a progressive exhaustion of the response. As no toxicity is detected with or without IL-2, this scheme represents a promising immunotherapeutic strategy for induction of systemic Th1 cytokines and massive expansion of gammadelta T cell subset with antitumor and anti-infectious properties.

Selective blockade of CD28 and not CTLA-4 with a single-chain Fv-alpha1-antitrypsin fusion antibody.

B7-1 and B7-2 are costimulatory molecules expressed on antigen-presenting cells. The CD28/B7 costimulation pathway is critical for T-cell activation, proliferation, and Th polarization. Blocking both cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) and CD28 interactions with a CTLA-4/Ig fusion protein inhibits various immune-mediated processes in vivo, such as allograft rejection and autoimmunity. However, selective blockade of CD28 may represent a better strategy for immunosuppression than B7 blockade, because CTLA-4/B7 interactions have been shown to participate in the extinction of the T-cell receptor-mediated activation signal and to be required for the induction of immunologic tolerance. In addition, selective CD28 inhibition specifically decreases the activation of alloreactive and autoreactive T cells, but not the activation of T cells stimulated by exogenous antigens presented in the context of self major histocompatibility complex (MHC) molecules. CD28 blockade cannot be obtained with anti-CD28 dimeric antibodies, which cluster their target and promote T-cell costimulation, whereas monovalent Fab fragments can block CD28 and reduce alloreactivity. In this study, we report the construction of a monovalent single-chain Fv antibody fragment from a high-affinity antihuman CD28 antibody (CD28.3) that blocked adhesion of T cells to cells expressing the CD28 receptor CD80. Genetic fusion with the long-lived serum protein alpha1-antitrypsin led to an extended half-life without altering its binding characteristics. The anti-CD28 fusion molecule showed biologic activity as an immuno-suppressant by inhibiting T-cell activation and proliferation in a mixed lymphocyte reaction.

Differential effect of CD28 versus B7 blockade on direct pathway of allorecognition and self-restricted responses.

Immunosuppression with B7 antagonists might have 2 opposite effects: reducing T-cell costimulation through CD28 but also preventing CTLA-4 from transmitting its negative regulatory signal. We therefore hypothesized that a selective blockade of CD28 might be qualitatively different from blocking B7. It was previously reported that CD28 modulation prolongs allograft survival in the rat and reverses induction of experimental autoimmune encephalomyelitis in mice. However, whether CD28 or B7 blockade results in similar immunosuppression on alloimmune and self-restricted responses to soluble antigens has not yet been investigated. Here, we addressed this issue in vitro with antagonist anti-CD28 Fab fragments and in vivo using the modulating anti-rat JJ319 monoclonal antibody. As in the inhibition of B7 with CTLA4 immunoglobulin, anti-CD28 Fab fragments inhibited allogenic T-cell proliferation in mixed cultures. In vivo modulation of CD28 blocked the expansion of alloreactive T cells and promoted their apoptosis. In contrast, selective blockade of CD28 did not modify T-cell proliferative responses and antibody production to soluble antigens, whereas blocking B7 with CTLA4 immunoglobulin did. Our data show that blocking CD28, while leaving CTLA4-B7 interactions undisturbed, inhibits alloreactive CD4+ T-cell expansion but does not modify the response to nominal antigens presented in the context of a self-major histocompatibility complex. That B7 engagement is needed for self-restricted responses whereas engagement of CD28 is not essential adds to the suggestion that another unidentified ligand of B7 might deliver a costimulatory signal in the absence of CD28.