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Toxins Oct 2020Toxins, while harmful and potentially lethal, have been engineered to develop potent therapeutics including cytotoxins and immunotoxins (ITs), which are modalities with... (Review)
Review
Toxins, while harmful and potentially lethal, have been engineered to develop potent therapeutics including cytotoxins and immunotoxins (ITs), which are modalities with highly selective targeting capabilities. Currently, three cytotoxins and IT are FDA-approved for treatment of multiple forms of hematological cancer, and additional ITs are tested in the clinical trials or at the preclinical level. For next generation of ITs, as well as antibody-mediated drug delivery systems, specific targeting by monoclonal antibodies is critical to enhance efficacies and reduce side effects, and this methodological field remains open to discover potent therapeutic monoclonal antibodies. Here, we describe our application of engineered toxin termed a cell-based IT screening system. This unique screening strategy offers the following advantages: (1) identification of monoclonal antibodies that recognize cell-surface molecules, (2) selection of the antibodies that are internalized into the cells, (3) selection of the antibodies that induce cytotoxicity since they are linked with toxins, and (4) determination of state-specific activities of the antibodies by differential screening under multiple experimental conditions. Since the functional monoclonal antibodies with internalization capacities have been identified successfully, we have pursued their subsequent modifications beyond antibody drug conjugates, resulting in development of immunoliposomes. Collectively, this screening system by using engineered toxin is a versatile platform, which enables straight-forward and rapid selection for discovery of novel functional antibodies.
Topics: Animals; Antibodies, Monoclonal; Antibody Specificity; Bacterial Toxins; Biological Transport; Cell Membrane; Cytotoxicity, Immunologic; Diphtheria Toxin; Exotoxins; High-Throughput Screening Assays; Humans; Immunoconjugates; Immunotoxins; Interleukin-2; Liposomes; Recombinant Fusion Proteins
PubMed: 33076544
DOI: 10.3390/toxins12100658 -
Journal of Virology Sep 2020Regulatory T cells (Tregs) may be key contributors to the HIV/SIV latent reservoir, since they harbor high levels of HIV/SIV; reverse CD4 T cell immune activation...
Regulatory T cells (Tregs) may be key contributors to the HIV/SIV latent reservoir, since they harbor high levels of HIV/SIV; reverse CD4 T cell immune activation status, increasing the pool of resting CD4 T cells; and impair CD8 T cell function, favoring HIV persistence. We tested the hypothesis that Treg depletion is a valid intervention toward an HIV cure by depleted Tregs in 14 rhesus macaque (RM) controllers infected with SIVsab, the virus that naturally infects sabaeus monkeys, through different strategies: administration of an anti-CCR4 immunotoxin, two doses of an anti-CD25 immunotoxin (interleukin-2 with diphtheria toxin [IL-2-DT]), or two combinations of both. All of these treatments resulted in significant depletion of the circulating Tregs (>70%) and their partial depletion in the gut (25%) and lymph nodes (>50%). The fractions of CD4 T cells expressing -67 increased up to 80% in experiments containing IL-2-DT and only 30% in anti-CCR4-treated RMs, paralleled by increases in the inflammatory cytokines. In the absence of ART, plasma virus rebounded to 10 vRNA copies/ml by day 10 after IL-2-DT administration. A large but transient boost of the SIV-specific CD8 T cell responses occurred in IL-2-DT-treated RMs. Such increases were minimal in the RMs receiving anti-CCR4-based regimens. Five RMs received IL-2-DT on ART, but treatment was discontinued because of high toxicity and lymphopenia. As such, while all treatments depleted a significant proportion of Tregs, the side effects in the presence of ART prevent their clinical use and call for different Treg depletion approaches. Thus, based on our data, Treg targeting as a strategy for HIV cure cannot be discarded. Regulatory T cells (Tregs) can decisively contribute to the establishment and persistence of the HIV reservoir, since they harbor high levels of HIV/SIV, increase the pool of resting CD4 T cells by reversing their immune activation status, and impair CD8 T cell function, favoring HIV persistence. We tested multiple Treg depletion strategies and showed that all of them are at least partially successful in depleting Tregs. As such, Treg depletion appears to be a valid intervention toward an HIV cure, reducing the size of the reservoir, reactivating the virus, and boosting cell-mediated immune responses. Yet, when Treg depletion was attempted in ART-suppressed animals, the treatment had to be discontinued due to high toxicity and lymphopenia. Therefore, while Treg targeting as a strategy for HIV cure cannot be discarded, the methodology for Treg depletion has to be revisited.
Topics: Animals; Anti-Retroviral Agents; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cytokines; Diphtheria Toxin; Immunity, Cellular; Inflammation; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Lymphopenia; Macaca mulatta; Primates; Receptors, CCR4; Recombinant Fusion Proteins; Simian Immunodeficiency Virus; T-Lymphocytes, Regulatory; Virus Latency
PubMed: 32669326
DOI: 10.1128/JVI.00533-20 -
The Cochrane Database of Systematic... Jul 2020Mycosis fungoides (MF) is the most common type of cutaneous T-cell lymphoma, a malignant, chronic disease initially affecting the skin. Several therapies are available,... (Meta-Analysis)
Meta-Analysis
BACKGROUND
Mycosis fungoides (MF) is the most common type of cutaneous T-cell lymphoma, a malignant, chronic disease initially affecting the skin. Several therapies are available, which may induce clinical remission for a time. This is an update of a Cochrane Review first published in 2012: we wanted to assess new trials, some of which investigated new interventions.
OBJECTIVES
To assess the effects of interventions for MF in all stages of the disease.
SEARCH METHODS
We updated our searches of the following databases to May 2019: the Cochrane Skin Specialised Register, CENTRAL, MEDLINE, Embase, and LILACS. We searched 2 trials registries for additional references. For adverse event outcomes, we undertook separate searches in MEDLINE in April, July and November 2017.
SELECTION CRITERIA
Randomised controlled trials (RCTs) of local or systemic interventions for MF in adults with any stage of the disease compared with either another local or systemic intervention or with placebo.
DATA COLLECTION AND ANALYSIS
We used standard methodological procedures expected by Cochrane. The primary outcomes were improvement in health-related quality of life as defined by participants, and common adverse effects of the treatments. Key secondary outcomes were complete response (CR), defined as complete disappearance of all clinical evidence of disease, and objective response rate (ORR), defined as proportion of patients with a partial or complete response. We used GRADE to assess the certainty of evidence and considered comparisons of psoralen plus ultraviolet A (PUVA) light treatment as most important because this is first-line treatment for MF in most guidelines.
MAIN RESULTS
This review includes 20 RCTs (1369 participants) covering a wide range of interventions. The following were assessed as either treatments or comparators: imiquimod, peldesine, hypericin, mechlorethamine, nitrogen mustard and intralesional injections of interferon-α (IFN-α) (topical applications); PUVA, extracorporeal photopheresis (ECP: photochemotherapy), and visible light (light applications); acitretin, bexarotene, lenalidomide, methotrexate and vorinostat (oral agents); brentuximab vedotin; denileukin diftitox; mogamulizumab; chemotherapy with cyclophosphamide, doxorubicin, etoposide, and vincristine; a combination of chemotherapy with electron beam radiation; subcutaneous injection of IFN-α; and intramuscular injections of active transfer factor (parenteral systemics). Thirteen trials used an active comparator, five were placebo-controlled, and two compared an active operator to observation only. In 14 trials, participants had MF in clinical stages IA to IIB. All participants were treated in secondary and tertiary care settings, mainly in Europe, North America or Australia. Trials recruited both men and women, with more male participants overall. Trial duration varied from four weeks to 12 months, with one longer-term study lasting more than six years. We judged 16 trials as at high risk of bias in at least one domain, most commonly performance bias (blinding of participants and investigators), attrition bias and reporting bias. None of our key comparisons measured quality of life, and the two studies that did presented no usable data. Eighteen studies reported common adverse effects of the treatments. Adverse effects ranged from mild symptoms to lethal complications depending upon the treatment type. More aggressive treatments like systemic chemotherapy generally resulted in more severe adverse effects. In the included studies, CR rates ranged from 0% to 83% (median 31%), and ORR ranged from 0% to 88% (median 47%). Five trials assessed PUVA treatment, alone or combined, summarised below. There may be little to no difference between intralesional IFN-α and PUVA compared with PUVA alone for 24 to 52 weeks in CR (risk ratio (RR) 1.07, 95% confidence interval (CI) 0.87 to 1.31; 2 trials; 122 participants; low-certainty evidence). Common adverse events and ORR were not measured. One small cross-over trial found once-monthly ECP for six months may be less effective than twice-weekly PUVA for three months, reporting CR in two of eight participants and ORR in six of eight participants after PUVA, compared with no CR or ORR after ECP (very low-certainty evidence). Some participants reported mild nausea after PUVA but no numerical data were given. One participant in the ECP group withdrew due to hypotension. However, we are unsure of the results due to very low-certainty evidence. One trial comparing bexarotene plus PUVA versus PUVA alone for up to 16 weeks reported one case of photosensitivity in the bexarotene plus PUVA group compared to none in the PUVA-alone group (87 participants; low-certainty evidence). There may be little to no difference between bexarotene plus PUVA and PUVA alone in CR (RR 1.41, 95% CI 0.71 to 2.80) and ORR (RR 0.94, 95% CI 0.61 to 1.44) (93 participants; low-certainty evidence). One trial comparing subcutaneous IFN-α injections combined with either acitretin or PUVA for up to 48 weeks or until CR indicated there may be little to no difference in the common IFN-α adverse effect of flu-like symptoms (RR 1.32, 95% CI 0.92 to 1.88; 82 participants). There may be lower CR with IFN-α and acitretin compared with IFN-α and PUVA (RR 0.54, 95% CI 0.35 to 0.84; 82 participants) (both outcomes: low-certainty evidence). This trial did not measure ORR. One trial comparing PUVA maintenance treatment to no maintenance treatment, in participants who had already had CR, did report common adverse effects. However, the distribution was not evaluable. CR and OR were not assessable. The range of treatment options meant that rare adverse effects consequently occurred in a variety of organs.
AUTHORS' CONCLUSIONS
There is a lack of high-certainty evidence to support decision making in the treatment of MF. Because of substantial heterogeneity in design, missing data, small sample sizes, and low methodological quality, the comparative safety and efficacy of these interventions cannot be reliably established on the basis of the included RCTs. PUVA is commonly recommended as first-line treatment for MF, and we did not find evidence to challenge this recommendation. There was an absence of evidence to support the use of intralesional IFN-α or bexarotene in people receiving PUVA and an absence of evidence to support the use of acitretin or ECP for treating MF. Future trials should compare the safety and efficacy of treatments to PUVA, as the current standard of care, and should measure quality of life and common adverse effects.
Topics: Acitretin; Antineoplastic Agents; Bexarotene; Combined Modality Therapy; Humans; Immunologic Factors; Interferon-alpha; Mycosis Fungoides; Neoplasm Staging; PUVA Therapy; Photochemotherapy; Photopheresis; Randomized Controlled Trials as Topic; Skin Neoplasms
PubMed: 32632956
DOI: 10.1002/14651858.CD008946.pub3 -
The Journal of Clinical Investigation Jun 2020The critical role of suppressive myeloid cells in immune regulation has come to the forefront in cancer research, with myeloid-derived suppressor cells (MDSCs) as a main... (Review)
Review
The critical role of suppressive myeloid cells in immune regulation has come to the forefront in cancer research, with myeloid-derived suppressor cells (MDSCs) as a main oncology immunotherapeutic target. Recent improvement and standardization of criteria classifying tumor-induced MDSCs have led to unified descriptions and also promoted MDSC research in tuberculosis (TB) and AIDS. Despite convincing evidence on the induction of MDSCs by pathogen-derived molecules and inflammatory mediators in TB and AIDS, very little attention has been given to their therapeutic modulation or roles in vaccination in these diseases. Clinical manifestations in TB are consequences of complex host-pathogen interactions and are substantially affected by HIV infection. Here we summarize the current understanding and knowledge gaps regarding the role of MDSCs in HIV and Mycobacterium tuberculosis (co)infections. We discuss key scientific priorities to enable application of this knowledge to the development of novel strategies to improve vaccine efficacy and/or implementation of enhanced treatment approaches. Building on recent findings and potential for cross-fertilization between oncology and infection biology, we highlight current challenges and untapped opportunities for translating new advances in MDSC research into clinical applications for TB and AIDS.
Topics: Acquired Immunodeficiency Syndrome; HIV-1; Humans; Mycobacterium tuberculosis; Myeloid-Derived Suppressor Cells; Tuberculosis
PubMed: 32420917
DOI: 10.1172/JCI136288 -
Molecular Oncology May 2020The majority of clinically diagnosed cutaneous T-cell lymphomas (CTCL) highly express the cell-surface markers CC chemokine receptor 4 (CCR4) and/or CD25. Recently, we...
The majority of clinically diagnosed cutaneous T-cell lymphomas (CTCL) highly express the cell-surface markers CC chemokine receptor 4 (CCR4) and/or CD25. Recently, we have developed diphtheria toxin-based recombinant Ontak®-like human IL2 fusion toxin (IL2 fusion toxin) and anti-human CCR4 immunotoxin (CCR4 IT). In this study, we first compared the efficacy of the CCR4 IT vs IL2 fusion toxin for targeting human CD25 CCR4 CTCL. We demonstrated that CCR4 IT was more effective than IL2 fusion toxin. We further constructed an IL2-CCR4 bispecific IT. The bispecific IT was significantly more effective than either IL2 fusion toxin or CCR4 IT alone. The bispecific IT is a promising novel targeted therapeutic drug candidate for the treatment of refractory and recurrent human CD25 and/or CCR4 CTCL.
Topics: Animals; Diphtheria Toxin; Flow Cytometry; Humans; Immunotoxins; Inhibitory Concentration 50; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Lymphoma, T-Cell, Cutaneous; Mice; Receptors, CCR4; Recombinant Fusion Proteins; Recombinant Proteins; Skin Neoplasms; Xenograft Model Antitumor Assays
PubMed: 32107846
DOI: 10.1002/1878-0261.12653 -
Immunotherapy Sep 2019T regulatory cells (Tregs) are an important T cell population for immune tolerance, prevention of autoimmune diseases and inhibition of antitumor immunity. The... (Review)
Review
T regulatory cells (Tregs) are an important T cell population for immune tolerance, prevention of autoimmune diseases and inhibition of antitumor immunity. The tumor-promoting role played by Tregs in cancer has prompted numerous approaches to develop immunotherapeutics targeting Tregs. One approach to depletion of Treg cells is retargeting the highly potent cytotoxic activity of bacterial toxins. These agents capitalize on the well-characterized bacterial toxins, diphtheria toxin and exotoxin A-both of which harbor membrane translocation domains and enzymatic domains that catalytically halt protein synthesis within intoxicated eukaryotic cells and act at picomolar or subpicomolar concentrations. In this review, we summarize the preclinical and clinical development of several Treg-depleting cancer immunotherapies based on these two bacterial toxins.
Topics: ADP Ribose Transferases; Animals; Bacterial Toxins; Clinical Trials as Topic; Diphtheria Toxin; Drug Evaluation, Preclinical; Exotoxins; Humans; Immunity, Cellular; Immunotherapy; Lymphocyte Depletion; Neoplasms; T-Lymphocytes, Regulatory; Tumor Microenvironment; Virulence Factors; Pseudomonas aeruginosa Exotoxin A
PubMed: 31361167
DOI: 10.2217/imt-2019-0060 -
Frontiers in Medicine 2019Mycosis fungoides (MF) and Sézary syndrome (SS) are the most common subtypes of cutaneous T-cell lymphoma. The majority of MF cases present with only patches and... (Review)
Review
Mycosis fungoides (MF) and Sézary syndrome (SS) are the most common subtypes of cutaneous T-cell lymphoma. The majority of MF cases present with only patches and plaques and the lesions are usually limited to the skin. On the other hand, in some cases, patients show skin tumors or erythroderma followed by lymph node involvement and rarely visceral organ involvement. SS is a rare, aggressive cutaneous T-cell lymphoma marked by exfoliative erythroderma, lymphadenopathy, and leukemic blood involvement. Because patients with relapsed or refractory MF/SS display a poor prognosis and the current treatment options are characterized by high rates of relapse, there is unmet need for the efficient treatment. This review provides a discussion of the recent and future promising therapeutic approaches in the management of advanced MF/SS. These include mogamulizumab, brentuximab vedotin, alemtuzumab, immune checkpoint inhibitors, IPH4102 (anti-KIR3DL2 antibody), histone deacetylase inhibitors (vorinostat, romidepsin, panobinostat, belinostat, and resminostat), pralatrexate, forodesine, denileukin diftitox, duvelisib, lenalidomide, and everolimus.
PubMed: 31192214
DOI: 10.3389/fmed.2019.00116 -
Proceedings of the National Academy of... Feb 2019Denileukin diftitox (DAB-IL-2, Ontak) is a diphtheria-toxin-based fusion protein that depletes CD25-positive cells including regulatory T cells and has been approved for...
Denileukin diftitox (DAB-IL-2, Ontak) is a diphtheria-toxin-based fusion protein that depletes CD25-positive cells including regulatory T cells and has been approved for the treatment of persistent or recurrent cutaneous T cell lymphoma. However, the clinical use of denileukin diftitox was limited by vascular leak toxicity and production issues related to drug aggregation and purity. We found that a single amino acid substitution (V6A) in a motif associated with vascular leak induction yields a fully active, second-generation biologic, s-DAB-IL-2(V6A), which elicits 50-fold less human umbilical vein endothelial cell monolayer permeation and is 3.7-fold less lethal to mice by LD analysis than s-DAB-IL-2. Additionally, to overcome aggregation problems, we developed a production method for the fusion toxin using that secretes fully folded, biologically active, monomeric s-DAB-IL-2 into the culture medium. Using the poorly immunogenic mouse B16F10 melanoma model, we initiated treatment 7 days after tumor challenge and observed that, while both s-DAB-IL-2(V6A) and s-DAB-IL-2 are inhibitors of tumor growth, the capacity to treat with higher doses of s-DAB-IL-2(V6A) could provide a superior activity window. In a sequential dual-therapy study in tumors that have progressed for 10 days, both s-DAB-IL-2(V6A) and s-DAB-IL-2 given before checkpoint inhibition with anti-programmed cell death-1 (anti-PD-1) antibodies inhibited tumor growth, while either drug given as monotherapy had less effect. s-DAB-IL-2(V6A), a fully monomeric protein with reduced vascular leak, is a second-generation diphtheria-toxin-based fusion protein with promise as a cancer immunotherapeutic both alone and in conjunction with PD-1 blockade.
Topics: Amino Acid Substitution; Antibodies; Cell Proliferation; Corynebacterium diphtheriae; Diphtheria Toxin; Human Umbilical Vein Endothelial Cells; Humans; Immunosuppressive Agents; Immunotoxins; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Melanoma, Experimental; Programmed Cell Death 1 Receptor; Receptors, Interleukin-2; Recombinant Fusion Proteins; T-Lymphocytes, Regulatory
PubMed: 30718426
DOI: 10.1073/pnas.1815087116 -
Journal of Immunotherapy (Hagerstown,... 2019Cellular immunotherapy using allogeneic natural killer (NK) cells may overcome chemotherapy-refractory acute myeloid leukemia. Our goal was to document NK cell... (Clinical Trial)
Clinical Trial
Cellular immunotherapy using allogeneic natural killer (NK) cells may overcome chemotherapy-refractory acute myeloid leukemia. Our goal was to document NK cell homing/persistence in the bone marrow following adoptive immunotherapy. Our cohort included 109 patients who received NK cell therapy for refractory acute myeloid leukemia following lymphodepleting conditioning +/- denileukin diftitox, +/- low-dose total body irradiation. We evaluated the NK cell density in bone marrow core biopsies performed an average of 14 days after NK cell transfer using a CD56 immunohistochemical stain. The NK cell density in core biopsies showed only moderate correlation with NK cell percentage in bone marrow aspirates evaluated by flow cytometry (rs=0.48) suggesting that distribution of CD56 cells in the bone marrow niche offers unique insight into NK cell homing. Better leukemia control was associated with increased NK cell density, such that patients with <5% blasts had a higher NK cell density (P=0.01). As well, NK cell density above the median of reference group was significantly associated with morphologic remission of leukemia (P=0.01). Moreover, the NK cell density varied significantly between conditioning protocols. Our findings suggest that the use of low-dose irradiation or CD25-targeting immunocytokine (denileukin diftitox, IL2DT) as part of conditioning results in increased NK cell homing/persistence in the bone marrow. These novel results will help guide future immunotherapy with NK cells.
Topics: Adolescent; Adult; Aged; Bone Marrow; Child; Child, Preschool; Female; Humans; Immunotherapy, Adoptive; Killer Cells, Natural; Leukemia, Myeloid, Acute; Male; Middle Aged; Young Adult
PubMed: 30489431
DOI: 10.1097/CJI.0000000000000250 -
Cancer Science Mar 2018E7777, a recombinant cytotoxic fusion protein comprising diphtheria toxin fragments A and B and human interleukin-2, shares an amino acid sequence with denileukin...
E7777, a recombinant cytotoxic fusion protein comprising diphtheria toxin fragments A and B and human interleukin-2, shares an amino acid sequence with denileukin diftitox but has improved purity and an increased percentage of active protein monomer species. A phase I study was carried out to evaluate the tolerability, safety, pharmacokinetics, and antitumor activity of E7777 in Japanese patients with relapsed/refractory peripheral and cutaneous T-cell lymphoma. E7777 (6, 12, and expanded 9 μg/kg/day) was given to 13 patients by i.v. infusion on five consecutive days per 21-day cycle. Dose-limiting toxicities, including increased alanine aminotransferase, hyponatremia (n = 2), hypokalemia, lymphopenia, fatigue, hypoalbuminemia, rash, and increased lipase (n = 1), were observed in all three patients in the 12 μg/kg/day cohort, whereas two of six patients in the 9 μg/kg/day cohort showed decreased appetite or fatigue. The maximum tolerated and recommended dose of E7777 was 9 μg/kg/day for five consecutive days per 21-day cycle. The objective response rate was 38% (5/13) and did not appear to depend on tumor expression of CD25. E7777 was well tolerated, assuming careful management of adverse events during treatment, and preliminary but clinically meaningful antitumor activity was observed. Subsequent studies of E7777 for T-cell lymphomas are warranted. This study was registered with www.ClinicalTrials.gov (NCT1401530).
Topics: Administration, Oral; Adult; Aged; Antineoplastic Agents; Cohort Studies; Diphtheria Toxin; Drug Administration Schedule; Drug Resistance, Neoplasm; Female; Humans; Infusions, Intravenous; Interleukin-2; Lymphoma, T-Cell, Cutaneous; Male; Maximum Tolerated Dose; Middle Aged; Neoplasm Recurrence, Local; Recombinant Fusion Proteins; Skin Neoplasms; Treatment Outcome; Young Adult
PubMed: 29363235
DOI: 10.1111/cas.13513