The Brigham and Women's Hospital and the Tübingen cutaneous squamous cell carcinoma (SCC) stratification systems propose different criteria from the American Joint Committee on Cancer, eighth ...edition. Our group identified prognostic subgroups within T3 stage according to the American Joint Committee on Cancer eighth edition's classification, the most common classification for high-risk cutaneous SCCs.
To compare the performance and prognostic accuracy of these staging systems in a subset of high-risk cutaneous SCCs.
Homogeneity, monotonicity, and McNemar tests for pairwise comparisons were assessed. Distinctiveness and relative risk of poor outcome were calculated by stage. Prognostic accuracy was compared with respect to quality (Akaike and Bayesian information criteria), concordance (Harrell C-index and Gönen and Heller concordance probability estimate), and predictive accuracy (sensitivity, specificity, negative predictive value, positive predictive value, and global accuracy).
The Brigham and Women's Hospital and Salamanca systems were more distinctive, homogeneous, and monotonic than the Tübingen system. The Tübingen system was the most specific, whereas the Salamanca and Brigham and Women's Hospital systems were more sensitive. Negative predictive value was high in all 3 systems, but positive predictive value and accuracy were low overall.
Alternative staging systems may partially overcome the heterogeneity and low prognostic accuracy of the American Joint Committee on Cancer, eighth edition and enable high-risk cutaneous SCCs to be stratified more reliably, but their prognostic accuracy is still low. Considering the accumulation of risk factors may improve high-risk cutaneous SCC risk stratification.
Graft-versus-host disease (GvHD) remains the major obstacle to successful allogeneic hematopoietic stem cell transplantation, despite of the immunosuppressive regimens administered to control T cell ...alloreactivity. PI3K/AKT/mTOR pathway is crucial in T cell activation and function and, therefore, represents an attractive therapeutic target to prevent GvHD development. Recently, numerous PI3K inhibitors have been developed for cancer therapy. However, few studies have explored their immunosuppressive effect.
The effects of a selective PI3K inhibitor (BKM120) and a dual PI3K/mTOR inhibitor (BEZ235) on human T cell proliferation, expression of activation-related molecules, and phosphorylation of PI3K/AKT/mTOR pathway proteins were analyzed. Besides, the ability of BEZ235 to prevent GvHD development in mice was evaluated.
Simultaneous inhibition of PI3K and mTOR was efficient at lower concentrations than PI3K specific targeting. Importantly, BEZ235 prevented naïve T cell activation and induced tolerance of alloreactive T cells, while maintaining an adequate response against cytomegalovirus, more efficiently than BKM120. Finally, BEZ235 treatment significantly improved the survival and decreased the GvHD development in mice.
These results support the use of PI3K inhibitors to control T cell responses and show the potential utility of the dual PI3K/mTOR inhibitor BEZ235 in GvHD prophylaxis.
The search for new drugs that control the continuous relapses of multiple myeloma is still required. Here, we report for the first time the potent antimyeloma activity of amiloride, an old ...potassium-sparing diuretic approved for the treatment of hypertension and edema due to heart failure.
Myeloma cell lines and primary samples were used to evaluate cytotoxicity of amiloride.
studies were carried out in a xenograft mouse model. The mechanisms of action were investigated using RNA-Seq experiments, qRT-PCR, immunoblotting, and immunofluorescence assays.
Amiloride-induced apoptosis was observed in a broad panel of multiple myeloma cell lines and in a xenograft mouse model. Moreover, amiloride also had a synergistic effect when combined with dexamethasone, melphalan, lenalidomide, and pomalidomide. RNA-Seq experiments showed that amiloride not only significantly altered the level of transcript isoforms and alternative splicing events, but also deregulated the spliceosomal machinery. In addition, disruption of the splicing machinery in immunofluorescence studies was associated with the inhibition of myeloma cell viability after amiloride exposure. Although amiloride was able to induce apoptosis in myeloma cells lacking p53 expression, activation of p53 signaling was observed in wild-type and mutated
cells after amiloride exposure. On the other hand, we did not find a significant systemic toxicity in mice treated with amiloride.
Overall, our results demonstrate the antimyeloma activity of amiloride and provide a mechanistic rationale for its use as an alternative treatment option for relapsed multiple myeloma patients, especially those with 17p deletion or
mutations that are resistant to current therapies.
.
In the last few years, nivolumab has become the standard of care for advanced-stage lung cancer patients. Unfortunately, up to 60% of patients do not respond to this treatment. In our study, we ...identified variations in gene expression related to primary resistance to immunotherapy. Bronchoscopy biopsies were obtained from advanced non-small cell lung cancer (NSCLC) patients previously characterized as responders or non-responders after nivolumab treatment. Ten tumor biopsies (from three responders and seven non-responders) were analyzed by the differential expression of 760 genes using the NanoString nCounter platform. These genes are known to be involved in the response to anti-PD1/PD-L1 therapy. All the patients were treated with nivolumab. Examining the dysregulated expression of 24 genes made it possible to predict the response to nivolumab treatment. Supervised analysis of the gene expression profile (GEP) revealed that responder patients had significantly higher levels of expression of
and
than did non-responder patients. In contrast,
and
genes had significantly lower expression levels than non-responder patients. These findings were validated as predictive biomarkers in an independent series of 201 patients treated with nivolumab (22 hepatocellular carcinomas, 14 non-squamous cell lung carcinomas, 5 head and neck squamous cell carcinomas, 1 ureter/renal pelvis carcinoma, 120 melanomas, 4 bladder carcinomas, 31 renal cell carcinomas, and 4 squamous cell lung carcinomas). ROC curve analysis showed that the expression levels of
, and
were the best predictors of response to nivolumab. Further, 13/24 genes showed an adverse impact on overall survival (OS) in an independent, large series of patients with NSCLC (2166 cases). In summary, we found a strong association between the global GEP of advanced NSCLC and the response to nivolumab. The classification of NSCLC patients based on GEP enabled us to identify those patients who genuinely benefited from treatment with immune checkpoint inhibitors (ICIs). We also demonstrated that abnormal expression of most of the markers comprising the genomic signature has an adverse influence on OS, making them significant markers for therapeutic decision-making. Additional prospective studies in larger series of patients are required to confirm the clinical utility of these biomarkers.
The new eighth edition of the American Joint Committee on Cancer staging system (AJCC-8) incorporates changes regarding cutaneous squamous cell carcinoma (CSCC).
We aimed to compare the AJCC-8 ...staging system with the previous seventh edition of the AJCC staging system (AJCC-7) and the Brigham and Women's Hospital (BWH) alternative staging system to identify their usefulness and the utility of their risk factors in defining prognostic groups in CSCC.
A series of 186 CSCCs of the head and neck were retrospectively collected. All 3 staging systems were compared from the standpoint of their ability to predict poor prognosis. Binary logistic regression models were built to determine which risk factors were most relevant.
Poor prognosis was mainly associated with stage T2 of the AJCC-7, with stages T2b/T3 of the BWH system, and with stage T3 of the AJCC-8. The AJCC-8 and the BWH staging systems displayed overlap with each another in predicting poor prognosis, and both were superior to the AJCC-7. The new risk factors incorporated into the AJCC-8 and the poor degree of differentiation were independently associated with poor outcome.
Retrospective study and few cases with bone invasion.
The AJCC-8 is more distinctive, monotonous, and homogeneous than the AJCC-7 and shows some overlap with the BWH system in stratification of tumors.
Dysregulation of one of the three D-cyclin genes has been observed in virtually all multiple myeloma tumors. The mechanisms by which CCND2 is upregulated in a set of multiple myeloma are not ...completely deciphered. We investigated the role of post-transcriptional regulation through the interaction between miRNAs and their binding sites at 3'UTR in CCND2 overexpression in multiple myeloma.
Eleven myeloma cell lines and 45 primary myeloma samples were included in the study. Interactions between miRNAs deregulated in multiple myeloma and mRNA targets were analyzed by 3'UTR-luciferase plasmid assay. The presence of CCND2 mRNA isoforms different in length was explored using qRT-PCR, Northern blot, mRNA FISH, and 3' rapid amplification of cDNA ends (RACE)-PCR.
We detected the presence of short CCND2 mRNA, both in the multiple myeloma cell lines and primary cells. The results obtained by 3'RACE experiments revealed that changes in CCND2 3'UTR length are explained by alternative polyadenylation. The luciferase assays using plasmids harboring the truncated CCND2 mRNA strongly confirmed the loss of miRNA sites in the shorter CCND2 mRNA isoform. Those multiple myelomas with greater abundance of the shorter 3'UTR isoform were associated with significant higher level of total CCND2 mRNA expression. Furthermore, functional analysis showed significant CCND2 mRNA shortening after CCND1 silencing and an increased relative expression of longer isoform after CCND1 and CCND3 overexpression, suggesting that cyclin D1 and D3 could regulate CCND2 levels through modifications in polyadenylation-cleavage reaction.
Overall, these results highlight the impact of CCND2 3'UTR shortening on miRNA-dependent regulation of CCND2 in multiple myeloma.
FAM46C, frequently mutated in multiple myeloma (MM), has recently been shown to encode a non‐canonical poly(A) polymerase (ncPAP). However, its target mRNAs and its role in MM pathogenesis remain ...mostly unknown. Using CRISPR‐Cas9 technology and gene expression analysis, we found that the inactivation of FAM46C in MM down‐regulates immunoglobulins (Igs) and several mRNAs encoding ER‐resident proteins, including some involved in unfolded protein response and others that affect glycosylation. Interestingly, we show that FAM46C expression is induced during plasma cell (PC) differentiation and that Ig mRNAs encoding heavy and light chains are substrates of the ncPAP, as revealed by poly(A) tail‐length determination assays. The absence of the ncPAP results in Ig mRNA poly(A) tail‐shortening, leading to a reduction in mRNA and protein abundance. On the other hand, loss of FAM46C up‐regulates metastasis‐associated lncRNA MALAT1 and results in a sharp increase in the migration ability. This phenotype depends mainly on the activation of PI3K/Rac1 signalling, which might have significant therapeutic implications. In conclusion, our results identify Ig mRNAs as targets of FAM46C, reveal an important function of this protein during PC maturation to increase antibody production and suggest that its role as a tumour suppressor might be related to the inhibition of myeloma cell migration.
Protein analysis in bone marrow samples from patients with multiple myeloma has been limited by the low concentration of proteins obtained after CD138
cell selection. A novel approach based on ...capillary nano-immunoassay could make it possible to quantify dozens of proteins from each myeloma sample in an automated manner. Here we present a method for the accurate and robust quantification of the expression of multiple proteins extracted from CD138-purified multiple myeloma samples frozen in RLT Plus buffer, which is commonly used for nucleic acid preservation and isolation. Additionally, the biological and clinical value of this analysis for a panel of 12 proteins essential to the pathogenesis of multiple myeloma was evaluated in 63 patients with newly diagnosed multiple myeloma. The analysis of the prognostic impact of
/Cereblon and
/Ikaros mRNA/protein showed that only the protein levels were able to predict progression-free survival of patients; mRNA levels were not associated with prognosis. Interestingly, high levels of Cereblon and Ikaros proteins were associated with longer progression-free survival only in patients who received immunomodulatory drugs and not in those treated with other drugs. In conclusion, the capillary nano-immunoassay platform provides a novel opportunity for automated quantification of the expression of more than 20 proteins in CD138
primary multiple myeloma samples.
Excessive alcohol consumption impairs the immune system, induces oxidative stress, and triggers the activation of peripheral blood (PB) monocytes, thereby contributing to alcoholic liver disease ...(ALD). We analyzed the M1/M2 phenotypes of circulating classical monocytes and macrophage-derived monocytes (MDMs) in excessive alcohol drinkers (EADs). PB samples from 20 EADs and 22 healthy controls were collected for isolation of CD14+ monocytes and short-term culture with LPS/IFNγ, IL4/IL13, or without stimulation. These conditions were also used to polarize MDMs into M1, M2, or M0 phenotypes. Cytokine production was assessed in the blood and culture supernatants. M1/M2-related markers were analyzed using mRNA expression and surface marker detection. Additionally, the miRNA profile of CD14+ monocytes was analyzed. PB samples from EADs exhibited increased levels of pro-inflammatory cytokines. Following short-term culture, unstimulated blood samples from EADs showed higher levels of soluble TNF-α and IL-8, whereas monocytes expressed increased levels of surface TNF-α and elevated mRNA expression of pro-inflammatory cytokines and inducible nitric oxide synthase. MDMs from EADs showed higher levels of TNF-α and CD206 surface markers and increased IL-10 production. LPS/IFNγ induced higher mRNA expression of Nrf2 only in the controls. miRNA analysis revealed a distinctive miRNA profile that is potentially associated with liver carcinogenesis and ALD through inflammation and oxidative stress. This study confirms the predominantly pro-inflammatory profile of PB monocytes among EADs and suggests immune exhaustion features in MDMs.
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