RESUMO
Tertiary lymphoid structures (TLS) resemble follicles of secondary lymphoid organs and develop in nonlymphoid tissues during inflammation and cancer. Which cell types and signals drive the development of TLS is largely unknown. To investigate early events of TLS development in the lungs, we repeatedly instilled p(I:C) plus ovalbumin (Ova) intranasally. This induced TLS ranging from lymphocytic aggregates to organized and functional structures containing germinal centers. We found that TLS development is independent of FAP+ fibroblasts, alveolar macrophages, or CCL19 but crucially depends on type I interferon (IFN-I). Mechanistically, IFN-I initiates two synergistic pathways that culminate in the development of TLS. On the one hand, IFN-I induces lymphotoxin (LT)α in lymphoid cells, which stimulate stromal cells to produce the B-cell-attracting chemokine CXCL13 through LTßR-signaling. On the other hand, IFN-I is sensed by stromal cells that produce the T-cell-attracting chemokines CXCL9, CXCL10 as well as CCL19 and CCL21 independently of LTßR. Consequently, B-cell aggregates develop within a week, whereas follicular dendritic cells and germinal centers appear after 3 weeks. Thus, sustained production of IFN-I together with an antigen is essential for the induction of functional TLS in the lungs.
RESUMO
Radiotherapy induces DNA damage and cell death, but recent data suggest that concomitant immune stimulation is an integral part of the therapeutic action of ionizing radiation. It is poorly understood how radiotherapy supports tumor-specific immunity. Here we report that radiotherapy induced tumor cell death and transiently activated complement both in murine and human tumors. The local production of pro-inflammatory anaphylatoxins C3a and C5a was crucial to the tumor response to radiotherapy and concomitant stimulation of tumor-specific immunity. Dexamethasone, a drug frequently given during radiotherapy, limited complement activation and the anti-tumor effects of the immune system. Overall, our findings indicate that anaphylatoxins are key players in radiotherapy-induced tumor-specific immunity and the ensuing clinical responses.
Assuntos
Complemento C3a/imunologia , Complemento C5a/imunologia , Regulação Neoplásica da Expressão Gênica/imunologia , Imunidade Inata/efeitos da radiação , Melanoma Experimental/imunologia , Neoplasias Cutâneas/imunologia , Animais , Antineoplásicos Hormonais/farmacologia , Ativação do Complemento , Complemento C3a/genética , Complemento C5a/genética , Dexametasona/farmacologia , Raios gama , Humanos , Imunidade Inata/efeitos dos fármacos , Melanoma Experimental/genética , Melanoma Experimental/patologia , Melanoma Experimental/radioterapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Complemento/genética , Receptores de Complemento/imunologia , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/radioterapia , Carga Tumoral/efeitos da radiaçãoRESUMO
Mononuclear phagocytes consisting of monocytes, macrophages, and DCs play a complex role in tumor development by either promoting or restricting tumor growth. Cutaneous squamous cell carcinoma (cSCC) is the second most common nonmelanoma skin cancer arising from transformed epidermal keratinocytes. While present at high numbers, the role of tumor-infiltrating and resident myeloid cells in the formation of cSCC is largely unknown. Using transgenic mice and depleting antibodies to eliminate specific myeloid cell types in the skin, we investigated the involvement of mononuclear phagocytes in the development of UV-induced cSCC in K14-HPV8-E6 transgenic mice. Although resident Langerhans cells were enriched in the tumor, their contribution to tumor formation was negligible. Equally, dermal macrophages were dispensable for the development of cSCC. In contrast, mice lacking circulating monocytes were completely resistant to UV-induced cSCC, indicating that monocytes promote tumor development. Collectively, these results demonstrate a critical role for classical monocytes in the initiation of skin cancer.
Assuntos
Carcinogênese/patologia , Epiderme/patologia , Monócitos/patologia , Neoplasias Cutâneas/patologia , Raios Ultravioleta/efeitos adversos , Animais , Carcinogênese/efeitos da radiação , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Epiderme/efeitos da radiação , Feminino , Queratinócitos/patologia , Queratinócitos/efeitos da radiação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Monócitos/efeitos da radiação , Pele/patologia , Pele/efeitos da radiaçãoRESUMO
Detection of CD4(+) T cells specific for tumor-associated antigens is critical to investigate the spontaneous tumor immunosurveillance and to monitor immunotherapy protocols in patients. We investigated the ability of HLA-DR 1101 multimers to detect CD4(+) T cells specific for three highly promiscuous MAGE-A3 derived peptides: MAGE-A3(191-205) (p39), MAGE-A3(281-295) (p57) and MAGE-A3(286-300) (p58). Tetramers stained specific CD4(+) T cells only when loaded with p39, although all peptides activated the specific T cells when presented by plastic-bound HLA-DR 1101 monomers. This suggested that tetramer staining ability was determined by the mode rather than the affinity of peptide binding to HLA-DR 1101. We hypothesized that peptides should bear a single P1 anchor residue to bind all arms of the multimer in a homogeneous register to generate peptide-HLA-DR conformers with maximal avidity. Bioinformatics analysis indicated that p39 contained one putative P1 anchor residue, whereas the other two peptides contained multiple ones. Designing p57 and p58 analogues containing a single anchor residue generated HLA-DR 1101 tetramers that stained specific CD4(+) T cells. Producing HLA-DR 1101 monomers linked with the optimized MAGE-A3 analogues, but not with the original epitopes, further improved tetramer efficiency. Optimization of CD4(+) T-cell epitope-binding registers is thus critical to generate functional HLA-DR tetramers.
Assuntos
Antígenos de Neoplasias/imunologia , Linfócitos T CD4-Positivos/imunologia , Epitopos de Linfócito T/imunologia , Antígenos HLA-DR/imunologia , Proteínas de Neoplasias/imunologia , Fragmentos de Peptídeos/imunologia , Linhagem Celular , Separação Celular , Citometria de Fluxo , Humanos , Reação em Cadeia da PolimeraseRESUMO
Some breast tumors metastasize aggressively whereas others remain dormant for years. The mechanism governing metastatic dormancy remains largely unknown. Through high-parametric single-cell mapping in mice, we identify a discrete population of CD39+PD-1+CD8+ T cells in primary tumors and in dormant metastasis, which is hardly found in aggressively metastasizing tumors. Using blocking antibodies, we find that dormancy depends on TNFα and IFNγ. Immunotherapy reduces the number of dormant cancer cells in the lungs. Adoptive transfer of purified CD39+PD-1+CD8+ T cells prevents metastatic outgrowth. In human breast cancer, the frequency of CD39+PD-1+CD8+ but not total CD8+ T cells correlates with delayed metastatic relapse after resection (disease-free survival), thus underlining the biological relevance of CD39+PD-1+CD8+ T cells for controlling experimental and human breast cancer. Thus, we suggest that a primary breast tumor could prime a systemic, CD39+PD-1+CD8+ T cell response that favors metastatic dormancy in the lungs.
Assuntos
Antígenos CD/imunologia , Apirase/imunologia , Neoplasias da Mama/imunologia , Linfócitos T CD8-Positivos/imunologia , Neoplasias Mamárias Experimentais/imunologia , Receptor de Morte Celular Programada 1/imunologia , Animais , Antígenos CD/metabolismo , Apirase/metabolismo , Neoplasias da Mama/patologia , Neoplasias da Mama/terapia , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , Humanos , Imunoterapia , Pulmão/imunologia , Pulmão/patologia , Neoplasias Mamárias Experimentais/patologia , Neoplasias Mamárias Experimentais/terapia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos Knockout , Metástase Neoplásica , Receptor de Morte Celular Programada 1/metabolismoRESUMO
Sensing of cytoplasmic DNA by cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) synthase (cGAS) results in production of the dinucleotide cGAMP and consecutive activation of stimulator of interferon genes (STING) followed by production of type I interferon (IFN). Although cancer cells contain supra-normal concentrations of cytoplasmic DNA, they rarely produce type I IFN spontaneously. This suggests that defects in the DNA-sensing pathway may serve as an immune escape mechanism. We find that cancer cells produce cGAMP that is transferred via gap junctions to tumor-associated dendritic cells (DCs) and macrophages, which respond by producing type I IFN in situ. Cancer-cell-intrinsic expression of cGAS, but not STING, promotes infiltration by effector CD8+ T cells and consequently results in prolonged survival. Furthermore, cGAS-expressing cancers respond better to genotoxic treatments and immunotherapy. Thus, cancer-cell-derived cGAMP is crucial to protective anti-tumor CD8+ T cell immunity. Consequently, cancer-cell-intrinsic expression of cGAS determines tumor immunogenicity and makes tumors hot. These findings are relevant for genotoxic and immune therapies for cancer.
Assuntos
Neoplasias/imunologia , Nucleotidiltransferases/metabolismo , Animais , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Dano ao DNA , Células Dendríticas/metabolismo , Progressão da Doença , Humanos , Imunoterapia , Interferon Tipo I/metabolismo , Proteínas de Membrana , Camundongos Endogâmicos C57BL , Repetições de Microssatélites/genética , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Nucleotídeos Cíclicos/metabolismoRESUMO
MHC-class I tetramers technology enabled the characterization of peptide-specific T cells at the single cell level in a variety of studies. Several laboratories have also developed MHC-class II multimers to characterize Ag-specific CD4+ T cells. However, the generation and use of MHC-class II multimers seems more problematic than that of MHC-I multimers. We have generated HLA-DR*1101 tetramers in a versatile empty form, which can be loaded after purification with peptides of interest. We discuss the impact of critical biological and structural parameters for the optimal staining of Ag-specific CD4+ T cells using HLA-DR*1101 tetramers, such as: (i) activation state of CD4+ T cells; (ii) membrane trafficking in the target CD4+ T cells; (iii) binding characteristics of the loaded CD4 epitope. Our data indicate that reorganization of TCR on the plasma membrane upon CD4+ T cell activation, as well as an homogenous binding frame of the CD4 epitopes to the soluble HLA-DR monomer, are critical for a stable TCR/MHC-class II tetramer interaction. These factors, together with the low frequencies and affinities of specific CD4+ T cells, explain the need for in vitro expansion or ex vivo enrichment of specific T cells for the optimal visualization with MHC-class II tetramers.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/imunologia , Animais , Antígenos/imunologia , Drosophila , Mapeamento de Epitopos , Antígenos HLA-DR/imunologia , Cadeias HLA-DRB1 , Humanos , Ativação Linfocitária , Microdomínios da Membrana , Peptídeos/imunologia , Multimerização Proteica , Reprodutibilidade dos Testes , TemperaturaRESUMO
Human papillomavirus (HPV)-driven cutaneous squamous cell carcinoma (cSCC) is the most common cancer in immunosuppressed patients. Despite indications suggesting that HPV promotes genomic instability during cSCC development, the molecular pathways underpinning HPV-driven cSCC development remain unknown. We compared the transcriptome of HPV-driven mouse cSCC with normal skin and observed higher amounts of transcripts for Porcupine and WNT ligands in cSCC, suggesting a role for WNT signaling in cSCC progression. We confirmed increased Porcupine expression in human cSCC samples. Blocking the secretion of WNT ligands by the Porcupine inhibitor LGK974 significantly diminished initiation and progression of HPV-driven cSCC. Administration of LGK974 to mice with established cSCC resulted in differentiation of cancer cells and significant reduction of the cancer stem cell compartment. Thus, WNT/ß-catenin signaling is essential for HPV-driven cSCC initiation and progression as well as for maintaining the cancer stem cell niche. Interference with WNT secretion may thus represent a promising approach for therapeutic intervention.
Assuntos
Aciltransferases/metabolismo , Carcinoma de Células Escamosas/patologia , Proteínas de Membrana/metabolismo , Papillomaviridae/patogenicidade , Infecções por Papillomavirus/patologia , Neoplasias Cutâneas/patologia , Proteínas Wnt/metabolismo , Aciltransferases/antagonistas & inibidores , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/virologia , Inibidores Enzimáticos/farmacologia , Perfilação da Expressão Gênica , Humanos , Proteínas de Membrana/antagonistas & inibidores , Camundongos , Células-Tronco Neoplásicas/patologia , Papillomaviridae/genética , Pirazinas/farmacologia , Piridinas/farmacologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/virologia , Nicho de Células-Tronco/fisiologia , Via de Sinalização Wnt/genéticaRESUMO
Myeloid leukocytes are essentially involved in both tumor progression and control. We show that neo-adjuvant treatment of mice with an inhibitor of CSF1 receptor (CSF1R), a drug that is used to deplete tumor-associated macrophages, unexpectedly promoted metastasis. CSF1R blockade indirectly diminished the number of NK cells due to a paucity of myeloid cells that provide the survival factor IL-15 to NK cells. Reduction of the number of NK cells resulted in increased seeding of metastatic tumor cells to the lungs but did not impact on progression of established metastases. Supplementation of mice treated with CSF1R-inhibitor with IL-15 restored numbers of NK cells and diminished metastasis. Our data suggest that CSF1R blockade should be combined with administration of IL-15 to reduce the risk of metastasis.
Assuntos
Células Matadoras Naturais/metabolismo , Células Mieloides/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Animais , Linhagem Celular Tumoral , CamundongosRESUMO
PURPOSE: Combination of immune checkpoint inhibitors with chemotherapy is under investigation for cancer treatment. EXPERIMENTAL DESIGN: We studied the rationale of such a combination for treating mesothelioma, a disease with limited treatment options. RESULTS: The combination of gemcitabine and immune checkpoint inhibitors outperformed immunotherapy alone with regard to tumor control and survival in a preclinical mesothelioma model; however, the addition of dexamethasone to gemcitabine and immune checkpoint inhibitors nullified the synergistic clinical response. Furthermore, treatment with gemcitabine plus anti-PD-1 resulted in an objective clinical response in two patients with mesothelioma, who were resistant to gemcitabine or anti-PD-1 as monotherapy. CONCLUSIONS: Thus, treatment of mesothelioma with a combination of gemcitabine with immune checkpoint inhibitors is feasible and results in synergistic clinical response compared with single treatment in the absence of steroids.
Assuntos
Antineoplásicos Imunológicos/farmacologia , Biomarcadores Tumorais , Desoxicitidina/análogos & derivados , Resistencia a Medicamentos Antineoplásicos , Imunomodulação/efeitos dos fármacos , Neoplasias Pulmonares/imunologia , Mesotelioma/imunologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Biópsia , Linhagem Celular Tumoral , Desoxicitidina/farmacologia , Modelos Animais de Doenças , Sinergismo Farmacológico , Expressão Gênica , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Mesotelioma/diagnóstico , Mesotelioma/tratamento farmacológico , Mesotelioma/metabolismo , Mesotelioma Maligno , Camundongos , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Prognóstico , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto , GencitabinaRESUMO
BACKGROUND: MHC class I-peptide tetramers are currently utilised to characterize CD8+ T cell responses at single cell level. The generation and use of MHC class II tetramers to study antigen-specific CD4+ T cells appears less straightforward. Most MHC class II tetramers are produced with a homogeneously built-in peptide, reducing greatly their flexibility of use. We attempted the generation of "empty" functional HLA-DR*1101 tetramers, receptive for loading with synthetic peptides by incubation. No such reagent is in fact available for this HLA-DR allele, one of the most frequent in the Caucasian population. RESULTS: We compared soluble MHC class II-immunoglobulin fusion proteins (HLA-DR*1101-Ig) with soluble MHC class II protein fused with an optimised Bir site for enzymatic biotynilation (HLA-DR*1101-Bir), both produced in insect cells. The molecules were multimerised by binding fluorochrome-protein A or fluorochrome-streptavidin, respectively. We find that HLA-DR*1101-Bir molecules are superior to the HLA-DR*1101-Ig ones both in biochemical and functional terms. HLA-DR*1101-Bir molecules can be pulsed with at least three different promiscuous peptide epitopes, derived from Tetanus Toxoid, influenza HA and the tumour associated antigen MAGE-3 respectively, to stain specific CD4+ T cells. Both staining temperature and activation state of CD4+ T cells are critical for the binding of peptide-pulsed HLA-DR*1101-Bir to the cognate TCR. CONCLUSION: It is therefore possible to generate a soluble recombinant HLA-DR*1101 backbone that is receptive for loading with different peptides to stain specific CD4+ T cells. As shown for other HLA-DR alleles, we confirm that not all the strategies to produce soluble HLA-DR*1101 multimers are equivalent.
Assuntos
Apresentação de Antígeno , Antígenos de Neoplasias/imunologia , Antígenos HLA-DR/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Proteínas de Neoplasias/imunologia , Fragmentos de Peptídeos/imunologia , Toxoide Tetânico/imunologia , Animais , Antígenos de Neoplasias/química , Biotinilação , Linfócitos T CD4-Positivos/imunologia , Linhagem Celular , DNA Complementar/genética , Drosophila melanogaster/citologia , Epitopos/imunologia , Genes de Imunoglobulinas , Genes MHC da Classe II , Genes Sintéticos , Antígenos HLA-DR/química , Antígenos HLA-DR/genética , Cadeias HLA-DRB1 , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Humanos , Fragmentos Fc das Imunoglobulinas/genética , Imunoglobulina G/genética , Zíper de Leucina , Proteínas de Neoplasias/química , Fragmentos de Peptídeos/química , Conformação Proteica , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/imunologia , TransfecçãoRESUMO
Aldara is a cream used for topical treatment of non-melanoma skin cancer, and is thought to act through stimulation of anti-tumour immunity. The active ingredient, imiquimod, has been shown to stimulate toll-like receptor 7. Aldara also induces psoriasis-like lesions when applied to naive murine skin, and as such is used as a mouse model for psoriasis. Here we find that in naive murine skin, Aldara induces inflammation largely independently of toll-like receptor 7. Surprisingly, inflammasome activation, keratinocyte death and interleukin 1 release also occur in response to the vehicle cream in the absence of imiquimod. We show that isostearic acid, a major component of the vehicle, promotes inflammasome activation in cultured keratinocytes, and so may contribute to the observed effects of Aldara on murine skin. Aldara therefore stimulates at least two immune pathways independently, and both imiquimod and vehicle are required for a full inflammatory response. Although it remains to be tested, it is possible that imiquimod-independent effects also contribute to the therapeutic efficacy of Aldara.
Assuntos
Aminoquinolinas/farmacologia , Imunidade/efeitos dos fármacos , Imunidade/imunologia , Receptor 7 Toll-Like/imunologia , Aminoquinolinas/efeitos adversos , Aminoquinolinas/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Carcinoma Basocelular/tratamento farmacológico , Carcinoma Basocelular/imunologia , Carcinoma Basocelular/patologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Epiderme/efeitos dos fármacos , Epiderme/imunologia , Epiderme/patologia , Epiderme/ultraestrutura , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Imiquimode , Inflamassomos/metabolismo , Interferon Tipo I/imunologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/imunologia , Queratinócitos/patologia , Ceratose Actínica/tratamento farmacológico , Ceratose Actínica/imunologia , Ceratose Actínica/patologia , Camundongos , Modelos Imunológicos , Infiltração de Neutrófilos/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Regulação para Cima/efeitos dos fármacosRESUMO
Although malignant cells can be recognized and controlled by the immune system, in patients with clinically apparent cancer immunosurveillance has failed. To better understand local immunoregulatory processes that impact on cancer progression, we correlated intratumoral immunological profiles with the survival of patients affected by primary clear cell renal cell carcinoma (ccRCC). A retrospective analysis of 54 primary ccRCC samples for 31 different immune response-related transcripts, revealed a negative correlation of CD68 (a marker of tumor-associated macrophages, TAMs) and FOXP3 (a marker of regulatory T cells, Tregs) with survival. The subsequent analysis of 12 TAM-related transcripts revealed an association between the genes coding for CD163, interferon regulatory factor 4 (IRF4) and fibronectin 1 (FN1), all of which have been linked to the M2 TAM phenotype, with reduced survival and increased tumor stage, whereas the opposite was the case for the M1-associated gene coding for inducible nitric oxide synthetase (iNOS). The M2 signature of (CD68+) TAMs was found to correlate with CD163 expression, as determined in prospectively collected fresh ccRCC tissue samples. Upon co-culture with autologous tumor cells, CD11b+ cells isolated from paired blood samples expressed CD163 and other M2-associated proteins, suggesting that the malignant cells promote the accumulation of M2 TAMs. Furthermore, the tumor-associated milieu as well as isolated TAMs induced the skewing of autologous, blood-derived CD4+ T cells toward a more immunosuppressive phenotype, as shown by decreased production of effector cytokines, increased production of interleukin-10 (IL-10) and enhanced expression of the co-inhibitory molecules programmed death 1 (PD-1) and T-cell immunoglobulin mucin 3 (TIM-3). Taken together, our data suggest that ccRCC progressively attracts macrophages and induces their skewing into M2 TAMs, in turn subverting tumor-infiltrating T cells such that immunoregulatory functions are increased at the expense of effector functions.
RESUMO
Inhibition of phosphodiesterase IV by N-(3,5-dichloropyrid-4-yl)-3-cyclopentyloxy-4-methoxybenzamide (piclamilast) enhances the myeloid differentiation induced by all-trans-retinoic acid (ATRA), retinoic acid receptor alpha (RARalpha), or retinoic acid receptor X agonists in NB4 and other retinoid-sensitive myeloid leukemia cell types. ATRA-resistant NB4.R2 cells are also partially responsive to the action of piclamilast and retinoic acid receptor X agonists. Treatment of NB4 cells with piclamilast or ATRA results in activation of the cAMP signaling pathway and nuclear translocation of cAMP-dependent protein kinase. This causes a transitory increase in cAMP-responsive element-binding protein phosphorylation, which is followed by down-modulation of the system. ATRA + piclamilast have no additive effects on the modulation of the cAMP pathway, and the combination has complex effects on cAMP-regulated genes. Piclamilast potentiates the ligand-dependent transactivation and degradation of RARalpha through a cAMP-dependent protein kinase-dependent phosphorylation. Enhanced transactivation is also observed in the case of PML-RARalpha. In NB4 cells, increased transactivation is likely to be at the basis of enhanced myeloid maturation and enhanced expression of many retinoid-dependent genes. Piclamilast and/or ATRA exert major effects on the expression of cEBP and STAT1, two types of transcription factors involved in myeloid maturation. Induction and activation of STAT1 correlates directly with enhanced cytodifferentiation. Finally, ERK and the cAMP target protein, Epac, do not participate in the maturation program activated by ATRA + piclamilast. Initial in vivo studies conducted in severe combined immunodeficiency mice transplanted with NB4 leukemia cells indicate that the enhancing effect of piclamilast on ATRA-induced myeloid maturation translates into a therapeutic benefit.