Revistas
Revista:
NATURE MEDICINE
ISSN:
1078-8956
The historical lack of preclinical models reflecting the genetic heterogeneity of multiple myeloma (MM) hampers the advance of therapeutic discoveries. To circumvent this limitation, we screened mice engineered to carry eight MM lesions (NF-kappa B, KRAS, MYC, TP53, BCL2, cyclin D1, MMSET/NSD2 and c-MAF) combinatorially activated in B lymphocytes following T cell-driven immunization. Fifteen genetically diverse models developed bone marrow (BM) tumors fulfilling MM pathogenesis. Integrative analyses of similar to 500 mice and similar to 1,000 patients revealed a common MAPK-MYC genetic pathway that accelerated time to progression from precursor states across genetically heterogeneous MM. MYC-dependent time to progression conditioned immune evasion mechanisms that remodeled the BM microenvironment differently. Rapid MYC-driven progressors exhibited a high number of activated/exhausted CD8(+) T cells with reduced immunosuppressive regulatory T (T-reg) cells, while late MYC acquisition in slow progressors was associated with lower CD8(+) T cell infiltration and more abundant T-reg cells. Single-cell transcriptomics and functional assays defined a high ratio of CD8(+) T cells versus T-reg cells as a predictor of response to immune checkpoint blockade (ICB). In clinical series, high CD8(+) T/T-reg cell ratios underlie early progression in untreated smoldering MM, and correlated with early relapse in newly diagnosed patients with MM under Len/Dex therapy. In ICB-refractory MM models, increasing CD8(+) T cell cytotoxicity or depleting T-reg cells reversed immunotherapy resistance and yielded prolonged MM control. Our experimental models enable the correlation of MM genetic and immunological traits with preclinical therapy responses, which may inform the next-generation immunotherapy trials.
New experimental models provide much-needed tools for understanding how genetically diverse multiple myeloma progresses and evolves in response to therapy.
Revista:
SCIENCE ADVANCES
ISSN:
2375-2548
Año:
2022
Vol.:
8
N°:
3
Págs.:
eabl4644
Normal cell counterparts of solid and myeloid tumors accumulate mutations years before disease onset; whether this occurs in B lymphocytes before lymphoma remains uncertain. We sequenced multiple stages of the B lineage in elderly individuals and patients with lymphoplasmacytic lymphoma, a singular disease for studying lymphomagenesis because of the high prevalence of mutated MYD88. We observed similar accumulation of random mutations in B lineages from both cohorts and unexpectedly found MYD88(L265P) in normal precursor and mature B lymphocytes from patients with lymphoma. We uncovered genetic and transcriptional pathways driving malignant transformation and leveraged these to model lymphoplasmacytic lymphoma in mice, based on mutated MYD88 in B cell precursors and BCL2 overexpression. Thus, MYD88(L265P) is a preneoplastic event, which challenges the current understanding of lymphomagenesis and may have implications for early detection of B cell lymphomas.
Revista:
DISEASE MODELS & MECHANISMS
ISSN:
1754-8411
Año:
2022
Vol.:
15
N°:
1
Págs.:
dmm049137
There is a paucity of adequate mouse models and cell lines available to study lung squamous cell carcinoma (LUSC). We have generated and characterized two models of phenotypically different transplantable LUSC cell lines, i.e. UN-SCC679 and UN-SCC680, derived from A/J mice that had been chemically induced with N-nitroso-tris-chloroethylurea (NTCU). Furthermore, we genetically characterized and compared both LUSC cell lines by performing whole-exome and RNA sequencing. These experiments revealed similar genetic and transcriptomic patterns that may correspond to the classic LUSC human subtype. In addition, we compared the immune landscape generated by both tumor cells lines in vivo and assessed their response to immune checkpoint inhibition. The differences between the two cell lines are a good model for the remarkable heterogeneity of human squamous cell carcinoma. Study of the metastatic potential of these models revealed that both cell lines represent the organotropism of LUSC in humans, i.e. affinity to the brain, bones, liver and adrenal glands. In summary, we have generated valuable cell line tools for LUSC research, which recapitulates the complexity of the human disease.
Revista:
CANCER DISCOVERY
ISSN:
2159-8274
Año:
2021
Vol.:
11
N°:
5
Págs.:
1268 - 1285
For millions of years, endogenous retroelements have remained transcriptionally silent within mammalian genomes by epigenetic mechanisms. Modern anticancer therapies targeting the epigenetic machinery awaken retroelement expression, inducing antiviral responses that eliminate tumors through mechanisms not completely understood. Here, we find that massive binding of epigenetically activated retroelements by RIG-I and MDA5 viral sensors promotes ATP hydrolysis and depletes intracellular energy, driving tumor killing independently of immune signaling. Energy depletion boosts compensatory ATP production by switching glycolysis to mitochondrial oxidative phosphorylation, thereby reversing the Warburg effect. However, hyperfunctional succinate dehydrogenase in mitochondrial electron transport chain generates excessive oxidative stress that unleashes RIP1-mediated necroptosis. To maintain ATP generation, hyperactive mitochondrial membrane blocks intrinsic apoptosis by increasing BCL2 dependency. Accordingly, drugs targeting BCL2 family proteins and epigenetic inhibitors yield synergistic responses in multiple cancer types. Thus, epigenetic therapy kills cancer cells by rewiring mitochondrial metabolism upon retroelement activation, which primes mitochondria to apoptosis by BH3-mimetics. SIGNIFICANCE: The state of viral mimicry induced by epigenetic therapies in cancer cells remodels mitochondrial metabolism and drives caspase-independent tumor cell death, which sensitizes to BCL2...
Autores:
Wojdacz, T. K. (Autor de correspondencia); Amarasinghe, H. E.; Kadalayil, L.; et al.
Revista:
BLOOD ADVANCES
ISSN:
2473-9529
Año:
2019
Vol.:
3
N°:
16
Págs.:
2474 - 2481
Chronic lymphocytic leukemia patients with mutated immunoglobulin heavy-chain genes (IGHV-M), particularly those lacking poor-risk genomic lesions, often respond well to chemo immunotherapy (CIT). DNA methylation profiling can subdivide early-stage patients into naive B-cell-like CLL (n-CLL), memory B-cell-like CLL (m-CLL), and intermediate CLL (i-CLL), with differing times to first treatment and overall survival. However, whether DNA methylation can identify patients destined to respond favorably to CIT has not been ascertained. We classified treatment-naive patients (n = 605) from 3 UK chemo and CIT clinical trials into the 3 epigenetic subgroups, using pyrosequencing and microarray analysis, and performed expansive survival analysis. The n-CLL, i-CLL, and m-CLL signatures were found in 80% (n = 245/305), 17% (53/305), and 2% (7/305) of IGHV-unmutated (IGHV-U) cases, respectively, and in 9%, (19/216), 50% (108/216), and 41% (89/216) of IGHV-M cases, respectively. Multivariate Cox proportional analysis identified m-CLL as an independent prognostic factor for overall survival (hazard ratio [HR], 0.46; 95% confidence interval [CI], 0.24-0.87; P = .018) in CLL4, and for progression-free survival (HR, 0.25; 95% CI, 0.10-0.57; P = .002) in ARCTIC and ADMIRE patients. The analysis of epigenetic subgroups in patients entered into 3 first-line UK CLL trials identifies m-CLL as an independent marker of prolonged survival and may aid in the identification of patients destined to demonstrate prolonged survival after CIT.
Revista:
JOURNAL OF PATHOLOGY
ISSN:
0022-3417
Año:
2018
Vol.:
245
N°:
1
Págs.:
61 - 73
The increased risk of Richter transformation (RT) in patients with chronic lymphocytic leukaemia (CLL) due to Epstein-Barr virus (EBV) reactivation during immunosuppressive therapy with fludarabine other targeted agents remains controversial. Among 31 RT cases classified as diffuse large B-cell lymphoma (DLBCL), seven (23%) showed EBV expression. In contrast to EBV- tumours, EBV+ DLBCLs derived predominantly from IGVH-hypermutated CLL, and they also showed CLL-unrelated IGVH sequences more frequently. Intriguingly, despite having different cellular origins, clonally related and unrelated EBV+ DLBCLs shared a previous history of immunosuppressive chemo-immunotherapy, a non-germinal centre DLBCL phenotype, EBV latency programme type II or III, and very short survival. These data suggested that EBV reactivation during therapy-related immunosuppression can transform either CLL cells or non-tumoural B lymphocytes into EBV+ DLBCL. To investigate this hypothesis, xenogeneic transplantation of blood cells from 31 patients with CLL and monoclonal B-cell lymphocytosis (MBL) was performed in Rag2(-/-) IL2c(-/-) mice. Remarkably, the recipients' impaired immunosurveillance favoured the spontaneous outgrowth of EBV+ B-cell clones from 95% of CLL and 64% of MBL patients samples, but not from healthy donors. Eventually, these cells generated monoclonal tumours (mostly CLL-unrelated but also CLL-related), recapitulating the principal features of EBV+ DLBCL in patients. Accordingly, clonally related and unrelated EBV+ DLBCL xenografts showed indistinguishable cellular, virological and molecular features, and synergistically responded to combined inhibition of EBV replication with ganciclovir and B-cell receptor signalling with ibrutinib in vivo. Our study underscores the risk of RT driven by EBV in CLL patients receiving immunosuppressive therapies, and provides the scientific rationale for testing ganciclovir and ibrutinib in EBV+ DLBCL. Copyright (c) 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Revista:
JOURNAL OF THORACIC ONCOLOGY
ISSN:
1556-0864
Año:
2016
Vol.:
11
N°:
4
Págs.:
524 - 536
INTRODUCTION:
Anti-programmed cell death 1 (anti-PD-1) and anti-programmed cell death ligand 1 (PD-L1) antagonist monoclonal antibodies (mAbs) against metastatic non-small cell lung cancer with special efficacy in patients with squamous cell lung cancer are being developed in the clinic. However, robust and reliable experimental models to test immunotherapeutic combinations in squamous lung tumors are still lacking.
METHODS:
We generated a transplantable squamous cell carcinoma cell line (UN-SCC680AJ) from a lung tumor induced by chronic N-nitroso-tris-chloroethylurea mutagenesis in A/J mice. Tumor cells expressed cytokeratins, overexpressed p40, and lacked thyroid transcription factor 1, confirming the squamous lineage reported by histological analysis. More than 200 mutations found in its exome suggested potential for antigenicity. Immunocompetent mice subcutaneously implanted with this syngeneic cell line were treated with anti-CD137 and/or anti-PD-1 mAbs and monitored for tumor growth/progression or assessed for intratumoral leukocyte infiltration using immunohistochemical analysis and flow cytometry.
RESULTS:
In syngeneic mice, large 12-day-established tumors derived from the transplantable cell line variant UN-SCC680AJ were amenable to curative treatment with anti-PD-1, anti-PD-L1, or anti-CD137 immunostimulatory mAbs. Single-agent therapies lost curative efficacy when treatment was started beyond day +17, whereas a combination of anti-PD-1 plus anti-CD137 achieved complete rejections. Tumor cells expressed weak baseline PD-L1 on the plasma membrane, but this could be readily induced by interferon-¿. Combined treatment efficacy required CD8 T cells and induced a leukocyte infiltrate in which T lymphocytes co-expressing CD137 and PD-1 were prominent.
CONCLUSIONS:
These promising results advocate the use of combined anti-PD-1/PD-L1 plus anti-CD137 mAb immunotherapy for the treatment of squamous non-small cell lung cancer in the clinical setting.
Revista:
PLOS ONE
ISSN:
1932-6203
Año:
2015
Vol.:
10
N°:
4
Págs.:
e0124670
Sirtuin 1 (SIRT1) and sirtuin 2 (SIRT2) are NAD+-dependent protein deacetylases involved in the regulation of key cancer-associated genes. In this study we evaluated the relevance of these deacetylases in lung cancer biology.
Material and Methods
Protein levels of SIRT1 and SIRT2 were determined in non-small cell lung cancer (NSCLC) cell lines and primary tumors from 105 patients. Changes in proliferation were assessed after SIRT1 and SIRT2 downregulation in lung cancer cell lines using siRNA-mediated technology or tenovin-1, a SIRT1 and SIRT2 inhibitor.
Results
High SIRT1 and SIRT2 protein levels were found in NSCLC cell lines compared with nontumor lung epithelial cells. The expression of SIRT1 and SIRT2 proteins was also significantly higher in lung primary tumors than in normal tissue (P<0.001 for both sirtuins). Stronger nuclear SIRT1 staining was observed in adenocarcinomas than in squamous cell carcinomas (P=0.033). Interestingly, in NSCLC patients, high SIRT1 and SIRT2 expression levels were associated with shorter recurrence-free survival (P=0.04 and P=0.007, respectively). Moreover, the combination of high SIRT1 and SIRT2 expression was an independent prognostic factor for shorter recurrence-free survival (P=0.002) and overall survival (P=0.022). In vitro studies showed that SIRT1 and/or SIRT2 downregulation significantly decreased proliferation of NSCLC.
Conclusions
Our results support the hypothesis that SIRT1 and SIRT2 have a protumorigenic role in lu
Revista:
EMBO MOLECULAR MEDICINE
ISSN:
1757-4684
Año:
2014
Vol.:
6
N°:
4
Págs.:
539 - 550
The vascular endothelial growth factor (VEGF) pathway is a clinically validated antiangiogenic target for non-small cell lung cancer (NSCLC). However, some contradictory results have been reported on the biological effects of antiangiogenic drugs. In order to evaluate the efficacy of these drugs in NSCLC histological subtypes, we analyzed the anticancer effect of two anti-VEGFR2 therapies (sunitinib and DC101) in chemically induced mouse models and tumorgrafts of lung adenocarcinoma (ADC) and squamous cell carcinoma (SCC). Antiangiogenic treatments induced vascular trimming in both histological subtypes. In ADC tumors, vascular trimming was accompanied by tumor stabilization. In contrast, in SCC tumors, antiangiogenic therapy was associated with disease progression and induction of tumor proliferation. Moreover, in SCC, anti-VEGFR2 therapies increased the expression of stem cell markers such as aldehyde dehydrogenase 1A1, CD133, and CD15, independently of intratumoral hypoxia. In vitro studies with ADC cell lines revealed that antiangiogenic treatments reduced pAKT and pERK signaling and inhibited proliferation, while in SCC-derived cell lines the same treatments increased pAKT and pERK, and induced survival. In conclusion, this study evaluates for the first time the effect of antiangiogenic drugs in lung SCC murine models in vivo and sheds light on the contradictory results of antiangiogenic therapies in NSCLC.
Revista:
MOLECULAR CANCER RESEARCH
ISSN:
1541-7786
Año:
2014
Vol.:
12
N°:
5
Págs.:
660 - 669
The TNF receptor-associated protein 1 (TRAP1) is a mitochondrial HSP that has been related to drug resistance and protection from apoptosis in colorectal and prostate cancer. Here, the effect of TRAP1 ablation on cell proliferation, survival, apoptosis, and mitochondrial function was determined in non-small cell lung cancer (NSCLC). In addition, the prognostic value of TRAP1 was evaluated in patients with NSCLC. These results demonstrate that TRAP1 knockdown reduces cell growth and clonogenic cell survival. Moreover, TRAP1 downregulation impairs mitochondrial functions such as ATP production and mitochondrial membrane potential as measured by TMRM (tetramethylrhodamine methylester) uptake, but it does not affect mitochondrial density or mitochondrial morphology. The effect of TRAP1 silencing on apoptosis, analyzed by flow cytometry and immunoblot expression (cleaved PARP, caspase-9, and caspase-3) was cell line and context dependent. Finally, the prognostic potential of TRAP1 expression in NSCLC was ascertained via immunohistochemical analysis which revealed that high TRAP1 expression was associated with increased risk of disease recurrence (univariate analysis, P = 0.008; multivariate analysis, HR: 2.554; 95% confidence interval, 1.085-6.012; P = 0.03). In conclusion, these results demonstrate that TRAP1 impacts the viability of NSCLC cells, and that its expression is prognostic in NSCLC.
Revista:
Journal of Clinical Oncology
ISSN:
0732-183X
Año:
2012
Vol.:
30
N°:
10
Págs.:
1129 - 1136
Purpose: Antiangiogenic therapies targeting the vascular endothelial growth factor (VEGF) pathway have yielded more modest clinical benefit to patients with non-small-cell lung cancer (NSCLC) than initially expected. Clinical data suggest a distinct biologic role of the VEGF pathway in the different histologic subtypes of lung cancer. To clarify the influence of histologic differentiation in the prognostic relevance of VEGF-mediated signaling in NSCLC, we performed a concomitant analysis of the expression of three key elements of the VEGF pathway in the earliest stages of the following two principal histologic subtypes: squamous cell carcinoma (SCC) and adenocarcinoma (ADC).
Patients and Methods: We evaluated tumor cell expression of VEGF, VEGF receptor (VEGFR) 1, and VEGFR2 using automatic immunostaining in a series of 298 patients with early-stage NSCLC recruited as part of the multicenter European Early Lung Cancer Detection Group project. A score measuring the VEGF signaling pathway was calculated by adding the tumor cell expression value of VEGF and its two receptors. The results were validated in two additional independent cohorts of patients with NSCLC.
Results: The combination of high VEGF, VEGFR1, and VEGFR2 protein expression was associated with lower risk of disease progression in early SCC (univariate analysis, P = .008; multivariate analysis, hazard ratio, 0.62; 95% CI, 0.42 to 0.92; P = .02). The results were validated in two independent patient cohorts, confirming the favorable prognostic value of high VEGF signaling score in early lung SCC.
Conclusion: Our results clearly indicate that the combination of high expression of the three key elements in the VEGF pathway is associated with a good prognosis in patients with early SCC but not in patients with ADC.
Revista:
MOLECULAR CANCER
ISSN:
1476-4598
Año:
2010
Vol.:
9
N°:
320
Págs.:
1 - 14
Background: Different isoforms of VEGF-A (mainly VEGF(121), VEGF(165) and VEGF(189)) have been shown to display particular angiogenic properties in the generation of a functional tumor vasculature. Recently, a novel class of VEGF-A isoforms, designated as VEGF(xxx)b, generated through alternative splicing, have been described. Previous studies have suggested that these isoforms may inhibit angiogenesis. In the present work we have produced recombinant VEGF(121/165)b proteins in the yeast Pichia pastoris and constructed vectors to overexpress these isoforms and assess their angiogenic potential. Results: Recombinant VEGF(121/165)b proteins generated either in yeasts or mammalian cells activated VEGFR2 and its downstream effector ERK1/2, although to a lesser extent than VEGF(165). Furthermore, treatment of endothelial cells with VEGF(121/165)b increased cell proliferation compared to untreated cells, although such stimulation was lower than that induced by VEGF165. Moreover, in vivo angiogenesis assays confirmed angiogenesis stimulation by VEGF(121/165)b isoforms. A549 and PC 3 cells overexpressing VEGF(121)b or VEGF(165)b (or carrying the PCDNA3.1 empty vector, as control) and xenotransplanted into nude mice showed increased tumor volume and angiogenesis compared to controls. To assess whether the VEGF(xxx)b isoforms are differentially expressed in tumors compared to healthy tissues, immunohistochemical analysis was conducted on a breast cancer tissue microarray. A significant increase (p < 0.05) in both VEGF(xxx)b and total VEGF-A protein expression in infiltrating ductal carcinomas compared to normal breasts was observed. A positive significant correlation (r = 0.404, p = 0.033) between VEGF(xxx)b and total VEGF-A was found. Conclusions: Our results demonstrate that VEGF(121/165)b are not anti-angiogenic, but weakly angiogenic isoforms of VEGF-A. In addition, VEGF(xxx)b isoforms are up-regulated in breast cancer in comparison with non malignant breast tissues. These results are to be taken into account when considering a possible use of VEGF(121/165)b-based therapies in patients.
