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1.
Cancer Res ; 83(23): 3901-3919, 2023 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-37702657

RESUMO

Multiple myeloma remains an incurable malignancy due to acquisition of intrinsic programs that drive therapy resistance. Here we report that casein kinase-1δ (CK1δ) and CK1ε are therapeutic targets in multiple myeloma that are necessary to sustain mitochondrial metabolism. Specifically, the dual CK1δ/CK1ε inhibitor SR-3029 had potent in vivo and ex vivo anti-multiple myeloma activity, including against primary multiple myeloma patient specimens. RNA sequencing (RNA-seq) and metabolic analyses revealed inhibiting CK1δ/CK1ε disables multiple myeloma metabolism by suppressing genes involved in oxidative phosphorylation (OxPhos), reducing citric acid cycle intermediates, and suppressing complexes I and IV of the electron transport chain. Finally, sensitivity of multiple myeloma patient specimens to SR-3029 correlated with elevated expression of mitochondrial genes, and RNA-seq from 687 multiple myeloma patient samples revealed that increased CSNK1D, CSNK1E, and OxPhos genes correlate with disease progression and inferior outcomes. Thus, increases in mitochondrial metabolism are a hallmark of multiple myeloma progression that can be disabled by targeting CK1δ/CK1ε. SIGNIFICANCE: CK1δ and CK1ε are attractive therapeutic targets in multiple myeloma whose expression increases with disease progression and connote poor outcomes, and that are necessary to sustain expression of genes directing OxPhos.


Assuntos
Caseína Quinase Idelta , Mieloma Múltiplo , Humanos , Caseína Quinase Idelta/genética , Caseína Quinase Idelta/metabolismo , Mieloma Múltiplo/genética , Sobrevivência Celular , Fosforilação , Progressão da Doença
2.
J Proteome Res ; 20(6): 3134-3149, 2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-34014671

RESUMO

Multiple myeloma is an incurable hematological malignancy that impacts tens of thousands of people every year in the United States. Treatment for eligible patients involves induction, consolidation with stem cell rescue, and maintenance. High-dose therapy with a DNA alkylating agent, melphalan, remains the primary drug for consolidation therapy in conjunction with autologous stem-cell transplantation; as such, melphalan resistance remains a relevant clinical challenge. Here, we describe a proteometabolomic approach to examine mechanisms of acquired melphalan resistance in two cell line models. Drug metabolism, steady-state metabolomics, activity-based protein profiling (ABPP, data available at PRIDE: PXD019725), acute-treatment metabolomics, and western blot analyses have allowed us to further elucidate metabolic processes associated with melphalan resistance. Proteometabolomic data indicate that drug-resistant cells have higher levels of pentose phosphate pathway metabolites. Purine, pyrimidine, and glutathione metabolisms were commonly altered, and cell-line-specific changes in metabolite levels were observed, which could be linked to the differences in steady-state metabolism of naïve cells. Inhibition of selected enzymes in purine synthesis and pentose phosphate pathways was evaluated to determine their potential to improve melphalan's efficacy. The clinical relevance of these proteometabolomic leads was confirmed by comparison of tumor cell transcriptomes from newly diagnosed MM patients and patients with relapsed disease after treatment with high-dose melphalan and autologous stem-cell transplantation. The observation of common and cell-line-specific changes in metabolite levels suggests that omic approaches will be needed to fully examine melphalan resistance in patient specimens and define personalized strategies to optimize the use of high-dose melphalan.


Assuntos
Transplante de Células-Tronco Hematopoéticas , Mieloma Múltiplo , Humanos , Melfalan/farmacologia , Metabolômica , Mieloma Múltiplo/tratamento farmacológico , Transplante Autólogo
3.
Hum Immunol ; 70(1): 1-5, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19027044

RESUMO

Dendritic cell-associated lectin-1 (DCAL-1), also known as C-type lectin-like-1 (CLECL1), is a novel C-type lectin-like molecule expressed by antigen presenting cells including dendritic cells (DCs). Here we report that incubation of immature DCs (iDCs) with an anti-DCAL-1 monoclonal antibody (mAb) induced downstream signaling, including phosphorylation of c-Jun N-terminal kinase (JNK) and p44/42 MAP kinase. Furthermore, ligation of DCAL-1 expressed by iDCs specifically enhanced HLA-DR expression, whereas the expression of other co-stimulatory molecules remained unchanged and minimal cytokine secretion was detected. DCs that express high levels of major histocompatibility complex (MHC) class II in the absence of high levels of other co-stimulatory molecules and inflammatory cytokine secretion may play an important role in the maintenance of immune tolerance. Therefore, our data suggests an important role for DCAL-1 in the regulation of the immune response.


Assuntos
Células Dendríticas/imunologia , Lectinas Tipo C/imunologia , Proteínas de Membrana/imunologia , Monócitos/imunologia , Anticorpos Monoclonais/imunologia , Células Cultivadas , Citocinas/metabolismo , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Antígenos HLA-DR/metabolismo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Ativação Linfocitária , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Monócitos/citologia , Monócitos/metabolismo , Fosforilação
4.
Blood ; 107(4): 1459-67, 2006 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-16239426

RESUMO

Dendritic-cell (DC)-associated C-type lectin receptors (CLRs) take up antigens to present to T cells and regulate DC functions. DCAL-2 is a CLR with a cytosolic immunoreceptor tyrosine-based inhibitory motif (ITIM), which is restricted to immature DCs (iDCs), monocytes, and CD1a+ DCs. Cross-linking DCAL-2 on iDCs induced protein tyrosine phosphorylation and MAPK activation as well as receptor internalization. To test if DCAL-2 is involved in DC maturation and cytokine expression, we stimulated iDCs with anti-DCAL-2 mAb with or without LPS, zymosan, or CD40L. While anti-DCAL-2 did not induce iDCs to mature, it did up-regulate CCR7 expression and IL-6 and IL-10 production. DCAL-2 signals augmented DC maturation induced by LPS or zymosan, increasing both CCR7 and DC-LAMP expression. Of interest, DCAL-2 ligation had the opposite effects on TLR versus CD40L signaling: anti-DCAL-2 suppressed TLR-induced IL-12 expression, but significantly enhanced CD40L-induced IL-12 production. DCAL-2 ligation also suppressed the ability of TLR-matured DCs to induce IFN-gamma-secreting Th1 cells but augmented the capacity of CD40L-matured DCs to polarize naive T cells into Th1 cells. Thus, DCAL-2 may program DCs differently depending on whether DCs are signaled via TLRs or by T cells. DCAL-2 may be a potential immunotherapeutic target for modulating autoimmune diseases or for developing vaccines.


Assuntos
Citocinas/biossíntese , Células Dendríticas/imunologia , Lectinas Tipo C/imunologia , Receptores Mitogênicos/genética , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Antígenos CD/sangue , Sequência de Bases , Divisão Celular , Humanos , Lectinas Tipo C/genética , Ativação Linfocitária , Dados de Sequência Molecular , Monócitos/imunologia , Filogenia , Receptores Mitogênicos/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de Aminoácidos
5.
Blood ; 107(4): 1537-45, 2006 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-16249377

RESUMO

Dendritic-cell (DC) migration to secondary lymphoid organs is crucial for the initiation of adaptive immune responses. Although LPS up-regulates CCR7 on DCs, a second signal is required to enable them to migrate toward the chemokine CCL19 (MIP-3beta). We found that the nitric oxide (NO) donor NOR4 provides a signal allowing LPS-stimulated DCs to migrate toward CCL19. NO affects DC migration through both the initial activation of the cGMP/cGMP kinase (cGMP/cGK) pathway and a long-term effect that reduced cGK activity via negative feedback. Indeed, migration of DCs toward CCL19, unlike migration toward CXCL12 (SDF-1alpha), required inhibition of cGK. LPS increased both cGK expression and cGK activity as measured by phosphorylation of the key cGK target vasodilator-stimulated phosphoprotein (VASP). Because cGK phosphorylation of VASP can disrupt focal adhesions and inhibit cell migration, LPS-induced VASP phosphorylation may prevent DCs from migrating without a second signal. Long-term NOR4 treatment inhibited the increase in cGK-dependent VASP phosphorylation, releasing this brake so that DCs can migrate. NO has been implicated in the regulation of autoimmunity through its effect on T cells. Our results suggest that NO regulation of DC migration and cytokine production may contribute to the protective effects of NO in autoimmune disorders.


Assuntos
Quimiocinas CC/fisiologia , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , Células Dendríticas/imunologia , Linfonodos/imunologia , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico/fisiologia , Piridinas/farmacologia , Movimento Celular/efeitos dos fármacos , Quimiocina CCL19 , GMP Cíclico/metabolismo , Citocinas/análise , Citocinas/sangue , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/fisiologia , Citometria de Fluxo , Humanos , Lipopolissacarídeos/farmacologia , Reação em Cadeia da Polimerase
6.
Blood ; 104(5): 1404-10, 2004 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-15142882

RESUMO

The effects of estrogen on the immune system are still largely unknown. We have investigated the effect of 17beta-estradiol (E(2)) on human monocyte-derived immature dendritic cells (iDCs). Short-term culture in E(2) had no effect on iDC survival or the expression of cell surface markers. However, E(2) treatment significantly increased the secretion of interleukin 6 (IL-6) in iDCs and also increased secretion of osteoprotegerin (OPG) by DCs. Furthermore, E(2) significantly increased secretion of the inflammatory chemokines IL-8 and monocyte chemoattractant protein 1 (MCP-1) by iDCs, but not the production of the constitutive chemokines thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC). However, after E(2) pretreatment the lipopolysaccharide (LPS)-induced production of MCP-1, TARC, and MDC by DCs was clearly enhanced. Moreover, mature DCs pretreated with E(2) stimulated T cells better than control cells. Finally, we found that E(2) provides an essential signal for migration of mature DCs toward CCL19/macrophage inflammatory protein 3beta (MIP3beta). In summary, E(2) may affect DC regulation of T-cell and B-cell responses, as well as help to sustain inflammatory responses. This may explain, in part, the reason serum levels of estrogen correlate with the severity of certain autoimmune diseases.


Assuntos
Citocinas/metabolismo , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Estradiol/imunologia , Estradiol/farmacologia , Antígenos CD40/metabolismo , Movimento Celular/efeitos dos fármacos , Movimento Celular/imunologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Células Cultivadas , Quimiocina CCL17 , Quimiocina CCL19 , Quimiocina CCL2/metabolismo , Quimiocina CCL22 , Quimiocinas CC/metabolismo , Quimiocinas CC/farmacologia , Células Dendríticas/citologia , Glicoproteínas/metabolismo , Humanos , Imunofenotipagem , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Lipopolissacarídeos/farmacologia , Monócitos/citologia , Osteoprotegerina , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores do Fator de Necrose Tumoral
7.
J Immunol ; 171(12): 6421-30, 2003 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-14662841

RESUMO

Recruitment of monocytes into tissues and their differentiation into macrophages or dendritic cells (DCs) depend on the microenvironment of the inflammatory site. Although many factors affecting this process have been identified, the intracellular signaling pathways implicated are poorly understood. We found that cyclic nucleotides regulate certain steps of monocyte differentiation into DCs. Increased levels of the cyclic nucleotides, cAMP or cGMP, inhibit differentiation of CD14(+)/CD1a(low) monocytes into CD14(-)/CD1a(high) DCs. However, DC-specific ICAM-3-grabbing nonintegrin (CD209) up-regulation was not affected by cyclic nucleotides, indicating that DC development was not blocked at the monocyte stage. Interestingly, Ag-presenting function was increased by cyclic nucleotides, as measured by the higher expression of MHC class II, CD86, and an increased ability to stimulate CD4(+) T cell proliferation in allogeneic MLRs. Although cyclic nucleotides do not completely block DC differentiation, they do block the ability of DCs to be induced to mature by LPS. Treatment during DC differentiation with either cAMP or cGMP analogues hampered LPS-induced expression of CD83, DC-LAMP, and CCR7 and the ability of DCs to migrate toward CCL19/macrophage-inflammatory protein 3beta. Interestingly, the induction of a CD16(+) subpopulation of cells was also observed. Thus, signals causing an increase in either cAMP or cGMP levels during monocyte recruitment to inflammatory sites may restrain the activation of acquired immunity by blocking DC development and migration to lymph nodes. At the same time, these signals promote development of an active intermediate cell type having properties between those of macrophages and DCs, which might contribute to the innate immune response in the periphery.


Assuntos
Moléculas de Adesão Celular/biossíntese , AMP Cíclico/fisiologia , GMP Cíclico/fisiologia , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Inibidores do Crescimento/fisiologia , Lectinas Tipo C/biossíntese , Monócitos/citologia , Monócitos/metabolismo , Receptores de Superfície Celular/biossíntese , 8-Bromo Monofosfato de Adenosina Cíclica/farmacologia , Adjuvantes Imunológicos/biossíntese , Adjuvantes Imunológicos/fisiologia , Apresentação de Antígeno/fisiologia , Antígenos CD/biossíntese , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Divisão Celular/imunologia , Células Cultivadas , Quimiocina CCL19 , Quimiocinas CC/fisiologia , Quimiotaxia de Leucócito/imunologia , AMP Cíclico/biossíntese , GMP Cíclico/biossíntese , Citocinas/antagonistas & inibidores , Citocinas/biossíntese , Citocinas/metabolismo , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/fisiologia , Inibidores do Crescimento/biossíntese , Humanos , Imunoglobulinas/biossíntese , Líquido Intracelular/imunologia , Líquido Intracelular/metabolismo , Lipopolissacarídeos/farmacologia , Ativação Linfocitária/imunologia , Proteínas de Membrana Lisossomal , Glicoproteínas de Membrana/biossíntese , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Receptores CCR7 , Receptores de Quimiocinas/biossíntese , Linfócitos T/citologia , Linfócitos T/imunologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/imunologia , Antígeno CD83
8.
Blood ; 101(11): 4464-71, 2003 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-12531790

RESUMO

Macrophages and dendritic cells play an important role in regulating B-cell responses, including proliferation to antigens such as trinitrophenyl (TNP)-Ficoll and TNP-Brucella abortus. However, the mechanisms and molecule(s) that regulate these processes are relatively undefined. In this report, we show that human macrophages generated in vitro strongly costimulate proliferation of dense human tonsillar B cells ligated via their B-cell antigen receptor (BCR) but not proliferation via CD40. Similarly, dendritic cells also markedly enhance BCR-activated B-cell proliferation. Soluble molecule(s) are required for human macrophages to costimulate proliferation of B cells triggered via their BCR. Importantly, a TACI (trans-membrane activator and CAML interactor)-Fc fusion protein inhibits both macrophage- and dendritic cell (DC)-dependent BCR-activated B-cell proliferation, indicating a requirement for at least one of the known TACI ligands, BAFF and/or APRIL. Consistent with a major role for BAFF, macrophages release BAFF at levels sufficient to potently costimulate BCR-induced B-cell proliferation. In addition, BAFF is more than 100-fold more potent than APRIL in enhancing BCR-mediated human B-cell proliferation. Furthermore, immunodepletion of APRIL under conditions that prevent APRIL-mediated B-cell costimulation does not block macrophage enhancement of B-cell proliferation. Finally, there is no correlation between the high levels of a proliferation-inducing ligand (APRIL) expressed by macrophages compared with DCs and the similar abilities of macrophages and DCs to enhance BCR-stimulated B-cell proliferation. In summary, our results suggest that macrophage- and DC-derived B-cell-activating factor belonging to the TNF family (BAFF) represents a key molecule by which macrophages and DCs directly regulate human B-cell proliferative responses to T-cell-independent stimuli.


Assuntos
Linfócitos B/imunologia , Células Dendríticas/imunologia , Ativação Linfocitária , Macrófagos/imunologia , Proteínas de Membrana/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Adolescente , Adulto , Fator Ativador de Células B , Comunicação Celular/imunologia , Células Dendríticas/citologia , Feminino , Humanos , Ativação Linfocitária/imunologia , Macrófagos/citologia , Masculino , Proteínas de Membrana/imunologia , Pessoa de Meia-Idade , Neuropeptídeos/imunologia , Proteínas Nucleares/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Receptores do Fator de Necrose Tumoral/imunologia , Proteína Transmembrana Ativadora e Interagente do CAML , Fator de Necrose Tumoral alfa/imunologia
9.
J Immunol ; 169(10): 5638-48, 2002 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-12421943

RESUMO

We have characterized dendritic cell (DC)-associated lectin-1 (DCAL-1), a novel, type II, transmembrane, C-type lectin-like protein. DCAL-1 has restricted expression in hemopoietic cells, in particular, DCs and B cells, but T cells and monocytes do not express it. The DCAL-1 locus is within a cluster of C-type lectin-like loci on human chromosome 12p12-13 just 3' to the CD69 locus. The consensus sequence of the DCAL-1 gene was confirmed by RACE-PCR; however, based on sequence alignment with genomic DNA and with various human expressed sequence tags, we predict that DCAL-1 has two splice variants. C-type lectins share a common sequence motif of 14 invariable and 18 highly conserved aa residues known as the carbohydrate recognition domain. DCAL-1, however, is missing three of the cysteine residues required to form the standard carbohydrate recognition domain. DCAL-1 mRNA and protein expression are increased upon the differentiation of monocytes to CD1a(+) DCs. B cells also express high levels of DCAL-1 on their cell surface. Using a DCAL-1 fusion protein we identified a population of CD4(+) CD45RA(+) T cells that express DCAL-1 ligand. Coincubation with soluble DCAL-1 enhanced the proliferation of CD4(+) T cells in response to CD3 ligation and significantly increased IL-4 secretion. In contrast, coincubation with soluble DC-specific ICAM-3-grabbing nonintegrin (CD209) fusion protein as a control had no effect on CD4(+) T cell proliferation or IL-4 and IFN-gamma secretion. Therefore, the function of DCAL-1 on DCs and B cells may act as a T cell costimulatory molecule, which skews CD4(+) T cells toward a Th2 response by enhancing their secretion of IL-4.


Assuntos
Adjuvantes Imunológicos/fisiologia , Células Dendríticas/imunologia , Interleucina-4/metabolismo , Lectinas Tipo C/fisiologia , Proteínas de Membrana/fisiologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Adjuvantes Imunológicos/genética , Adjuvantes Imunológicos/isolamento & purificação , Adjuvantes Imunológicos/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Células Sanguíneas/imunologia , Células Sanguíneas/metabolismo , Células Cultivadas , Mapeamento Cromossômico , Células Dendríticas/metabolismo , Regulação da Expressão Gênica/imunologia , Humanos , Lectinas Tipo C/genética , Lectinas Tipo C/isolamento & purificação , Lectinas Tipo C/metabolismo , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Ligantes , Ativação Linfocitária/imunologia , Tecido Linfoide/imunologia , Tecido Linfoide/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/isolamento & purificação , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Tonsila Palatina/imunologia , Tonsila Palatina/metabolismo , Ligação Proteica/imunologia , Estrutura Terciária de Proteína/genética , Células Th2/imunologia
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