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1.
Hepatol Commun ; 8(4)2024 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-38551383

RESUMO

BACKGROUND: Biliary dysplasia, a precursor of cholangiocarcinoma (CCA), is a common complication of primary sclerosing cholangitis. Patients with high-grade dysplasia (HGD) or early CCA who have received oncological treatment are candidates for liver transplantation. The preoperative diagnosis of CCA or HGD is challenging, and the sensitivity of biliary brush cytology (BC) is limited. METHODS: By using next-generation sequencing (NGS), we retrospectively analyzed archived tissue samples (n=62) obtained from explanted liver tissue and CCA samples to identify oncogenic mutations that occur during primary sclerosing cholangitis carcinogenesis. BC samples were prospectively collected from patients with primary sclerosing cholangitis (n=97) referred for endoscopic retrograde cholangiography to measure the diagnostic utility of NGS combined with BC compared with traditional cytology alone. RESULTS: Mutations in KRAS, GNAS, FLT3, RNF43, TP53, ATRX, and SMAD4 were detected in archived CCA or HGD samples. KRAS, GNAS, TP53, CDKN2A, FBXW7, BRAF, and ATM mutations were detected in prospectively collected brush samples from patients with histologically verified CCA or HGD. One patient with low-grade dysplasia in the explanted liver had KRAS and GNAS mutations in brush sample. No mutations were observed in brush samples or archived tissues in liver transplantation cases without biliary neoplasia. While KRAS mutations are common in biliary neoplasms, they were also observed in patients without biliary neoplasia during surveillance. CONCLUSIONS: In summary, NGS of BC samples increased the sensitivity of detecting biliary neoplasia compared with traditional cytology. Performing NGS on BC samples may help diagnose HGD or early CCA, benefiting the timing of liver transplantation.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Colangite Esclerosante , Humanos , Estudos Retrospectivos , Estudos Prospectivos , Colangite Esclerosante/complicações , Colangite Esclerosante/diagnóstico , Colangite Esclerosante/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Neoplasias dos Ductos Biliares/diagnóstico , Neoplasias dos Ductos Biliares/genética , Colangiopancreatografia Retrógrada Endoscópica/efeitos adversos , Colangiocarcinoma/diagnóstico , Colangiocarcinoma/genética , Ductos Biliares Intra-Hepáticos , Sequenciamento de Nucleotídeos em Larga Escala
2.
Cancers (Basel) ; 16(5)2024 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-38473410

RESUMO

The primary tumor location (PTL) is associated with the phenotype, metastatic sites, mutations, and outcomes of metastatic colorectal cancer (mCRC) patients, but this has mostly been studied according to sidedness (right vs. left sided). We studied right colon vs. left colon vs. rectal PTL in a real-life study population (n = 1080). Health-related quality of life (HRQoL) was assessed multi-cross-sectionally with QLQ-C30, QLQ-CR29, EQ-5D, and 15D. A chi-square, Kaplan-Meier, and Cox regression were used to compare the groups. The PTL was in the right colon in 310 patients (29%), the left colon in 396 patients (37%), and the rectum in 375 patients (35%). The PTL was associated with distinct differences in metastatic sites during the disease trajectory. The resectability, conversion, and resection rates were lowest in the right colon, followed by the rectum, and were highest in the left colon. Overall survival was shortest for right colon compared with left colon or rectal PTL (median 21 vs. 35 vs. 36 months), with the same trends after metastasectomy or systemic therapy only. PTL also remained statistically significant in a multivariable model. The distribution of symptoms varied according to PTL, especially between the right colon (with general symptoms of metastases) and rectal PTL (with sexual- and bowel-related symptoms). mCRC, according to PTL, behaves differently regarding metastatic sites, resectability of the metastases, outcomes of treatment, and HRQoL.

3.
J Clin Oncol ; 42(12): 1378-1390, 2024 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-38232336

RESUMO

PURPOSE: Clinical relapse is the major threat for patients with myelodysplastic syndrome (MDS) undergoing hematopoietic stem-cell transplantation (HSCT). Early detection of measurable residual disease (MRD) would enable preemptive treatment and potentially reduced relapse risk. METHODS: Patients with MDS planned for HSCT were enrolled in a prospective, observational study evaluating the association between MRD and clinical outcome. We collected bone marrow (BM) and peripheral blood samples until relapse, death, or end of study 24 months after HSCT. Patient-specific mutations were identified with targeted next-generation sequencing (NGS) panel and traced using droplet digital polymerase chain reaction (ddPCR). RESULTS: Of 266 included patients, estimated relapse-free survival (RFS) and overall survival (OS) rates 3 years after HSCT were 59% and 64%, respectively. MRD results were available for 221 patients. Relapse was preceded by positive BM MRD in 42/44 relapses with complete MRD data, by a median of 71 (23-283) days. Of 137 patients in continuous complete remission, 93 were consistently MRD-negative, 39 reverted from MRD+ to MRD-, and five were MRD+ at last sampling. Estimated 1 year-RFS after first positive MRD was 49%, 39%, and 30%, using cutoff levels of 0.1%, 0.3%, and 0.5%, respectively. In a multivariate Cox model, MRD (hazard ratio [HR], 7.99), WHO subgroup AML (HR, 4.87), TP53 multi-hit (HR, 2.38), NRAS (HR, 3.55), and acute GVHD grade III-IV (HR, 4.13) were associated with shorter RFS. MRD+ was also independently associated with shorter OS (HR, 2.65). In a subgroup analysis of 100 MRD+ patients, presence of chronic GVHD was associated with longer RFS (HR, 0.32). CONCLUSION: Assessment of individualized MRD using NGS + ddPCR is feasible and can be used for early detection of relapse. Positive MRD is associated with shorter RFS and OS (ClinicalTrials.gov identifier: NCT02872662).


Assuntos
Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Síndromes Mielodisplásicas , Humanos , Doença Enxerto-Hospedeiro/etiologia , Transplante de Células-Tronco Hematopoéticas/métodos , Leucemia Mieloide Aguda/genética , Síndromes Mielodisplásicas/terapia , Recidiva Local de Neoplasia/genética , Neoplasia Residual/genética , Prognóstico , Estudos Prospectivos , Recidiva
4.
Int J Cancer ; 154(3): 488-503, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-37724848

RESUMO

BRAF-V600E mutation (mt) is a strong negative prognostic and predictive biomarker in metastatic colorectal cancer (mCRC). Non-V600Emt, designated atypical BRAFmt (aBRAFmt) are rare, and little is known about their frequency, co-mutations and prognostic and predictive role. These were compared between mutational groups of mCRC patients collected from three Nordic population-based or real-world cohorts. Pathology of aBRAFmt was studied. The study included 1449 mCRC patients with 51 (3%) aBRAFmt, 182 (13%) BRAF-V600Emt, 456 (31%) RAS&BRAF wild-type (wt) and 760 (52%) RASmt tumours. aBRAFmt were seen in 2% of real-world and 4% of population-based cohorts. Twenty-six different aBRAFmt were detected, 11 (22%) class 2 (serrated adenocarcinoma in 2/9 tested), 32 (64%) class 3 (serrated in 15/25) and 4 (8%) unclassified. aBRAFmt patients were predominantly male, had more rectal primaries, less peritoneal metastases, deficient mismatch repair in one (2%), and better survival after metastasectomy (89% 5-year overall survival [OS]-rate) compared with BRAF-V600Emt. aBRAFmt and BRAF-V600Emt had poorer performance status and received fewer treatment lines than RAS&BRAFwt and RASmt. OS among aBRAFmt (median 14.4 months) was longer than for BRAF-V600Emt (11.2 months), but shorter than for RAS&BRAFwt (30.5 months) and RASmt (23.4 months). Addition of bevacizumab trended for better OS for the aBRAFmt. Nine patients with aBRAFmt received cetuximab/panitumumab without response. aBRAFmt represents a distinct subgroup differing from other RAS/BRAF groups, with serrated adenocarcinoma in only half. OS for patients with aBRAFmt tumours was slightly better than for BRAF-V600Emt, but worse than for RASmt and RAS&BRAFwt. aBRAFmt should not be a contraindication for metastasectomy.


Assuntos
Adenocarcinoma , Neoplasias do Colo , Neoplasias Colorretais , Neoplasias Retais , Humanos , Masculino , Feminino , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Colorretais/patologia , Mutação
5.
Cancers (Basel) ; 15(14)2023 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-37509233

RESUMO

Aberrant glycosylation affects cancer progression and immune evasion. Approximately 15% of colorectal cancers (CRCs) demonstrate microsatellite instability (MSI) and display major differences in outcomes and therapeutic responses, as compared to corresponding microsatellite stable (MSS) tumors. We compared the N-glycan profiles of stage II and IV MSI CRC tumors, further subdivided into BRAFV600E wild-type and mutated subgroups (n = 10 in each subgroup), with each other and with those of paired non-neoplastic mucosal samples using mass spectrometry. Further, the N-glycans of BRAFV600E wild-type stage II MSI tumors were compared to corresponding MSS tumors (n = 9). Multiple differences in N-glycan profiles were identified between the MSI CRCs and control tissues, as well as between the stage II MSI and MSS samples. The MSI CRC tumors showed a lower relative abundance of high-mannose N-glycans than did the control tissues or the MSS CRCs. Among MSI CRC subgroups, acidic N-glycans showed tumor stage and BRAF mutation status-dependent variation. Specifically, the large, sulfated/phosphorylated, and putative terminal N-acetylhexosamine-containing acidic N-glycans differed between the MSI CRC subgroups, showing opposite changes in stages II and IV, when comparing BRAF mutated and wild-type tumors. Our results show that molecular subgroups of CRC exhibit characteristic glycan profiles that may explain certain carcinogenic properties of MSI tumors.

9.
Acta Oncol ; 61(10): 1263-1267, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-36307938

RESUMO

BACKGROUND: Detectable circulating tumor DNA (ctDNA) has been associated with worse prognosis in melanoma patients. MATERIAL AND METHODS: We studied plasma ctDNA as a prognostic biomarker in 19 patients with metastatic melanoma and a detectable tumor mutation (13 BRAF, 5 NRAS, and 1 KRAS). Patients had received chemotherapy, interferon-alpha, and vemurafenib in a prospective clinical trial. Mutant allele frequency (MAF %) was determined with droplet digital PCR from pretreatment and sequential plasma samples. RESULTS: Higher pretreatment plasma ctDNA levels (MAF ≥3%) and detectable plasma ctDNA levels (MAF >0%) at the time of radiologically confirmed best objective response were associated with poor prognosis even when accounting for other relevant prognostic factors including performance status, tumor mutation, metastasis stage, and lactate dehydrogenase levels in multivariable analysis. CONCLUSION: Higher pretreatment plasma ctDNA levels and sustained detectable plasma ctDNA levels during treatment indicated poor prognosis in metastatic melanoma patients.


Assuntos
DNA Tumoral Circulante , Melanoma , Segunda Neoplasia Primária , Humanos , Biomarcadores , Biomarcadores Tumorais/genética , DNA Tumoral Circulante/genética , Melanoma/tratamento farmacológico , Melanoma/genética , Melanoma/patologia , Mutação , Prognóstico , Estudos Prospectivos , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/genética
10.
Br J Cancer ; 127(4): 686-694, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35610367

RESUMO

BACKGROUND: Outcomes after metastasectomy for metastatic colorectal cancer (mCRC) vary with RAS and BRAF mutational status, but their effects on resectability and conversion rates have not been extensively studied. METHODS: This substudy of the prospective RAXO trial included 906 patients recruited between 2011 and 2018. We evaluated repeated centralised resectability assessment, conversion/resection rates and overall survival (OS), according to RAS and BRAF status. RESULTS: Patients included 289 with RAS and BRAF wild-type (RAS and BRAFwt), 529 with RAS mutated (RASmt) and 88 with BRAF mutated (BRAFmt) mCRC. Metastatic prevalence varied between the RAS and BRAFwt/RASmt/BRAFmt groups, for liver (78%/74%/61%), lung (24%/35%/28%) and peritoneal (15%/15%/32%) metastases, respectively. Upfront resectability (32%/29%/15%), conversion (16%/13%/7%) and resection/local ablative therapy (LAT) rates (45%/37%/17%) varied for RASa and BRAFwt/RASmt/BRAFmt, respectively. Median OS for patients treated with resection/LAT (n = 342) was 83/69/30 months, with 5-year OS-rates of 67%/60%/24%, while systemic therapy-only patients (n = 564) had OS of 29/21/15 months with 5-year OS-rates of 11%/6%/2% in RAS and BRAFwt/RASmt/BRAFmt, respectively. Resection/LAT was associated with improved OS in all subgroups. CONCLUSIONS: There were significant differences in resectability, conversion and resection/LAT rates according to RAS and BRAF status. OS was also significantly longer for RAS and BRAFwt versus either mutant. Patients only receiving systemic therapy had poorer long-term survival, with variation according to molecular status. CLINICAL TRIAL REGISTRATION: NCT01531621/EudraCT2011-003158-24.


Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Metastasectomia , Neoplasias Retais , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/cirurgia , Humanos , Mutação , Estudos Prospectivos , Proteínas Proto-Oncogênicas B-raf/genética
12.
Front Oncol ; 12: 826073, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35251991

RESUMO

BACKGROUND: KRAS mutations, present in over 40% of metastatic colorectal cancer (mCRC), are negative predictive factors for anti-EGFR therapy. Mutations in KRAS-G12C have a cysteine residue for which drugs have been developed. Published data on this specific mutation are conflicting; thus, we studied the frequency and clinical characteristics in a real-world and population-based setting. METHODS: Patients from three Nordic population-based cohorts and the real-life RAXO-study were combined. RAS and BRAF tests were performed in routine healthcare, except for one cohort. The dataset consisted of 2,559 patients, of which 1,871 could be accurately classified as KRAS, NRAS, and BRAF-V600E. Demographics, treatments, and outcomes were compared using logistic regression. Overall survival (OS) was estimated with Kaplan-Meier, and differences were compared using Cox regression, adjusted for baseline factors. RESULTS: The KRAS-G12C frequency was 2%-4% of all tested in the seven cohorts (mean 3%) and 4%-8% of KRAS mutated tumors in the cohorts (mean 7%). Metastasectomies and ablations were performed more often (38% vs. 28%, p = 0.040), and bevacizumab was added more often (any line 74% vs. 59%, p = 0.007) for patients with KRAS-G12C- vs. other KRAS-mutated tumors, whereas chemotherapy was given to similar proportions. OS did not differ according to KRAS mutation, neither overall (adjusted hazard ratio (HR) 1.03; 95% CI 0.74-1.42, reference KRAS-G12C) nor within treatment groups defined as "systemic chemotherapy, alone or with biologics", "metastasectomy and/or ablations", or "best supportive care", RAS and BRAF wild-type tumors (n = 548) differed similarly to KRAS-G12C, as to other KRAS- or NRAS-mutated (n = 66) tumors. CONCLUSIONS: In these real-life and population-based cohorts, there were no significant differences in patient characteristics and outcomes between patients with KRAS-G12C tumors and those with other KRAS mutations. This contrasts with the results of most previous studies claiming differences in many aspects, often with worse outcomes for those with a KRAS-G12C mutation, although not consistent. When specific drugs are developed, as for this mutation, differences in outcome will hopefully emerge.

13.
Virchows Arch ; 480(4): 807-817, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35237889

RESUMO

Gene fusions can act as oncogenic drivers and offer targets for cancer therapy. Since fusions are rare in colorectal cancer (CRC), their universal screening seems impractical. Our aim was to investigate gene fusions in 62 CRC cases with deficient MLH1 (dMLH1) and BRAFV600E wild-type (wt) status from a consecutive real-life series of 2079 CRCs. First, gene fusions were analysed using a novel FusionPlex Lung v2 RNA-based next-generation sequencing (NGS) panel, and these results were compared to a novel Idylla GeneFusion assay and pan-TRK immunohistochemistry (IHC). NGS detected seven (7/62, 11%) NTRK1 fusions (TPM3::NTRK1, PLEKHA6::NTRK1 and LMNA::NTRK1, each in two cases, and IRF2BP2::NTRK1 in one case). In addition, two ALK, four RET and seven BRAF fusions were identified. Idylla detected seven NTRK1 expression imbalances, in line with the NGS results (overall agreement 100%). Furthermore, Idylla detected the two NGS-identified ALK rearrangements as one specific ALK fusion and one ALK expression imbalance, whilst only two of the four RET fusions were discovered. However, Idylla detected several expression imbalances of ALK (n = 7) and RET (n = 1) that were found to be fusion negative with the NGS. Pan-TRK IHC showed clearly detectable, fusion partner-dependent staining patterns in the seven NTRK1 fusion cases. Overall agreement for pan-TRK antibody clone EPR17341 was 98% and for A7H6R 100% when compared to the NGS. Of the 62 CRCs, 43 were MLH1 promoter hypermethylated (MLH1ph) and 39 were RASwt. All fusion cases were both MLH1ph and RASwt. Our results show that kinase fusions (20/30, 67%) and most importantly targetable NTRK1 fusions (7/30, 23%) are frequent in CRCs with dMLH1/BRAFV600Ewt/MLH1ph/RASwt. NGS was the most comprehensive method in finding the fusions, of which a subset can be screened by Idylla or IHC, provided that the result is confirmed by NGS.


Assuntos
Neoplasias Colorretais , Receptor trkA , Neoplasias Colorretais/genética , Fusão Gênica , Rearranjo Gênico , Humanos , Imuno-Histoquímica , Proteína 1 Homóloga a MutL/genética , Mutação/genética , Receptor trkA/genética
14.
Hemasphere ; 6(3): e701, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35233509

RESUMO

In adult patients, the treatment outcome of acute lymphoblastic leukemia (ALL) remains suboptimal. Here, we used an ex vivo drug testing platform and comprehensive molecular profiling to discover new drug candidates for B-ALL. We analyzed sensitivity of 18 primary B-ALL adult patient samples to 64 drugs in a physiological concentration range. Whole-transcriptome sequencing and publicly available expression data were used to examine gene expression biomarkers for observed drug responses. Apoptotic modulators targeting BCL2 and MDM2 were highly effective. Philadelphia chromosome-negative (Ph-) samples were sensitive to both BCL2/BCL-W/BCL-XL-targeting agent navitoclax and BCL2-selective venetoclax, whereas Ph-positive (Ph+) samples were more sensitive to navitoclax. Expression of BCL2 was downregulated and BCL-W and BCL-XL upregulated in Ph+ ALL compared with Ph- samples, providing elucidation for the observed difference in drug responses. A majority of the samples were sensitive to MDM2 inhibitor idasanutlin. The regulatory protein MDM2 suppresses the function of tumor suppressor p53, leading to impaired apoptosis. In B-ALL, the expression of MDM2 was increased compared with other hematological malignancies. In B-ALL cell lines, a combination of BCL2 and MDM2 inhibitor was synergistic. In summary, antiapoptotic proteins including BCL2 and MDM2 comprise promising targets for future drug studies in B-ALL.

15.
Blood Cancer Discov ; 2(3): 238-249, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34661156

RESUMO

In myelodysplastic syndrome (MDS) and myeloproliferative neoplasm (MPN), bone marrow (BM) histopathology is assessed to identify dysplastic cellular morphology, cellularity, and blast excess. Yet, other morphologic findings may elude the human eye. We used convolutional neural networks to extract morphologic features from 236 MDS, 87 MDS/MPN, and 11 control BM biopsies. These features predicted genetic and cytogenetic aberrations, prognosis, age, and gender in multivariate regression models. Highest prediction accuracy was found for TET2 [area under the receiver operating curve (AUROC) = 0.94] and spliceosome mutations (0.89) and chromosome 7 monosomy (0.89). Mutation prediction probability correlated with variant allele frequency and number of affected genes per pathway, demonstrating the algorithms' ability to identify relevant morphologic patterns. By converting regression models to texture and cellular composition, we reproduced the classical del(5q) MDS morphology consisting of hypolobulated megakaryocytes. In summary, this study highlights the potential of linking deep BM histopathology with genetics and clinical variables. SIGNIFICANCE: Histopathology is elementary in the diagnostics of patients with MDS, but its high-dimensional data are underused. By elucidating the association of morphologic features with clinical variables and molecular genetics, this study highlights the vast potential of convolutional neural networks in understanding MDS pathology and how genetics is reflected in BM morphology. See related commentary by Elemento, p. 195.


Assuntos
Síndromes Mielodisplásicas , Doenças Mieloproliferativas-Mielodisplásicas , Medula Óssea/patologia , Humanos , Aprendizado de Máquina , Mutação/genética , Síndromes Mielodisplásicas/diagnóstico , Doenças Mieloproliferativas-Mielodisplásicas/genética
16.
Virchows Arch ; 479(3): 471-479, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33755781

RESUMO

Universal testing of microsatellite instability (MSI) is recommended for colorectal cancer (CRC) and endometrial cancer (EC) to screen for Lynch syndrome and to aid in assessing prognosis and optimal treatment. We compared the performance of Idylla MSI test to immunohistochemistry (IHC) of mismatch repair (MMR) proteins in consecutive series of 100 CRC and 108 EC samples, as well as in retrospective series of 28 CRC and 33 EC specimens with known deficient MMR protein expression. The concordance between the Idylla test and IHC was 100% in all CRC samples (n=128) but lower in EC samples (87.2%; n=141). In the EC samples, sensitivity of Idylla test was 72.7% and specificity 100%. EC MSI/dMMR agreement was 85.4% for MLH1, 87.5% for MSH2, and only 35.3% for MSH6. When we analyzed 14 EC samples that were discrepant, i.e., dMMR using IHC and microsatellite stable using Idylla, with microsatellite markers BAT25 and BAT26, we found four cases to be replication error (RER) positive. All RER positive cases were deficient for MSH6 protein expression. We also re-analyzed EC samples with variable tumor cellularity to determine the limit of detection of the Idylla test and found that a 30% or higher tumor cellularity is required. We conclude that Idylla MSI test offers a sensitive and specific method for CRC diagnostics but is less sensitive in EC samples especially in the case of MSH6 deficiency.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Reparo de Erro de Pareamento de DNA , Detecção Precoce de Câncer , Neoplasias do Endométrio/genética , Instabilidade de Microssatélites , Reação em Cadeia da Polimerase , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Neoplasias Colorretais/química , Neoplasias Colorretais/patologia , Enzimas Reparadoras do DNA/análise , Neoplasias do Endométrio/química , Neoplasias do Endométrio/patologia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Mutação , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Estudos Retrospectivos
17.
Glycobiology ; 31(3): 211-222, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33539510

RESUMO

Pseudomyxoma peritonei (PMP) is a highly mucinous adenocarcinoma growing in the peritoneal cavity and most commonly originating from the appendix. Glycans play an important role in carcinogenesis, and glycosylation is altered in malignant diseases, including PMP. We have previously demonstrated that fucosylation of N-glycans is increased in PMP, but we did not observe modulation of overall sialylation. As sialic acids can be attached to the rest of the glycan via α2,3- or α2,6-linkage, we have now analyzed the linkage patterns of sialic acids in tissue specimens of normal appendices, low-grade appendiceal mucinous neoplasms (LAMN), low-grade (LG) PMP and high-grade (HG) PMP. For the linkage analysis, the enzymatically released acidic N-glycans were first treated with ethyl esterification or α2,3-sialidase digestion followed by MALDI-TOF mass spectrometry. Significant increase in the relative abundance of α2,6-sialylated and decrease in α2,3-sialylated N-glycans was observed in PMP tumors as compared to the normal appendices (P < 0.025). More specifically, increased α2,6-sialylation (P < 0.05) and decreased α2,3-sialylation (P < 0.01) were detected in afucosylated and monofucosylated N-glycans of PMPs, whereas the less abundant multifucosylated glycans, containing terminal fucose, demonstrated increased α2,3-sialylation (P < 0.01). Importantly, the increase in α2,6-sialylation was also detected between PMP and the appendiceal precursor lesion LAMN (P < 0.01). The identified glycosylation alterations produce ligands for sialic acid-binding immunoglobulin-like lectins (Siglecs) and sialofucosylated glycans binding selectins, which play a role in the peritoneal dissemination and progression of the disease.


Assuntos
Adenocarcinoma Mucinoso/química , Polissacarídeos/metabolismo , Pseudomixoma Peritoneal/química , Ácidos Siálicos/metabolismo , Adenocarcinoma Mucinoso/metabolismo , Adenocarcinoma Mucinoso/patologia , Humanos , Polissacarídeos/química , Polissacarídeos/isolamento & purificação , Pseudomixoma Peritoneal/metabolismo , Pseudomixoma Peritoneal/patologia , Ácidos Siálicos/química , Ácidos Siálicos/isolamento & purificação
18.
Leukemia ; 35(7): 1964-1975, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33168949

RESUMO

The oncogenic protein Bcr-Abl has two major isoforms, p190Bcr-Abl and p210Bcr-Abl. While p210Bcr-Abl is the hallmark of chronic myeloid leukemia (CML), p190Bcr-Abl occurs in the majority of Philadelphia-positive acute lymphoblastic leukemia (Ph + ALL) patients. In CML, p190Bcr-Abl occurs in a minority of patients associating with distinct hematological features and inferior outcomes, yet the pathogenic role of p190Bcr-Abl and potential targeting therapies are largely uncharacterized. We employed next generation sequencing, phospho-proteomic profiling, and drug sensitivity testing to characterize p190Bcr-Abl in CML and hematopoietic progenitor cell line models (Ba/f3 and HPC-LSK). p190Bcr-Abl CML patients demonstrated poor response to imatinib and frequent mutations in epigenetic modifiers genes. In contrast with p210Bcr-Abl, p190Bcr-Abl exhibited specific transcriptional upregulation of interferon, interleukin-1 receptor, and P53 signaling pathways, associated with hyperphosphorylation of relevant signaling molecules including JAK1/STAT1 and PAK1 in addition to Src hyperphosphorylation. Comparable to p190Bcr-Abl CML patients, p190Bcr-Abl cell lines demonstrated similar transcriptional and phospho-signaling signatures. With the drug sensitivity screening we identified targeted drugs with specific activity in p190Bcr-Abl cell lines including IAP-, PAK1-, and Src inhibitors and glucocorticoids. Our results provide novel insights into the mechanisms underlying the distinct features of p190Bcr-Abl CML and promising therapeutic targets for this high-risk patient group.


Assuntos
Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Transdução de Sinais/genética , Idoso , Idoso de 80 Anos ou mais , Animais , Linhagem Celular Tumoral , Feminino , Perfilação da Expressão Gênica/métodos , Glucocorticoides/genética , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Oncogenes/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Proteômica/métodos , Transcrição Gênica/genética , Regulação para Cima/genética
19.
Haematologica ; 105(12): 2757-2768, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33256375

RESUMO

Common variable immunodeficiency and other late-onset immunodeficiencies often co-manifest with autoimmunity and lymphoproliferation. The pathogenesis of most cases is elusive, as only a minor subset harbors known monogenic germline causes. The involvement of both B and T cells is however implicated. To study whether somatic mutations in CD4+ and CD8+ T cells associate with immunodeficiency, we recruited 17 patients and 21 healthy controls. Eight patients had late-onset common variable immunodeficiency and nine patients other immunodeficiency and/or severe autoimmunity. In total, autoimmunity occurred in 94% and lymphoproliferation in 65%. We performed deep sequencing of 2533 immune-associated genes from CD4+ and CD8+ cells. Deep T-cell receptor beta sequencing was used to characterize CD4+ and CD8+ T-cell receptor repertoires. The prevalence of somatic mutations was 65% in all immunodeficiency patients, 75% in common variable immunodeficiency and 48% in controls. Clonal hematopoiesis-associated variants in both CD4+ and CD8+ cells occurred in 24% of immunodeficiency patients. Results demonstrated mutations in known tumor suppressors, oncogenes, and genes that are critical for immune- and proliferative functions, such as STAT5B (two patients), C5AR1 (two patients), KRAS (one patient), and NOD2 (one patient). Additionally, as a marker of T-cell receptor repertoire perturbation, common variable immunodeficiency patients harbored increased frequencies of clones with identical complementarity determining region 3 sequences despite unique nucleotide sequences when compared to controls. In conclusion, somatic mutations in genes implicated for autoimmunity and lymphoproliferation are common in CD4+ and CD8+ cells of patients with immunodeficiency. They may contribute to immune dysregulation in a subset of immunodeficiency patients.


Assuntos
Síndromes de Imunodeficiência , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Regiões Determinantes de Complementaridade/genética , Humanos , Mutação , Receptores de Antígenos de Linfócitos T alfa-beta/genética
20.
PLoS One ; 15(11): e0239819, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33237900

RESUMO

Circulating tumor DNA (ctDNA) is released from cancer cells and oncogenic mutations in ctDNA can be measured from plasma samples. Droplet digital PCR (ddPCR) is a sensitive and specific method for the detection of mutations in ctDNA. We analyzed serial plasma samples (n = 80) from ten metastatic colorectal cancer (mCRC) patients with a known KRAS mutation in their primary tumor. The patients were undergoing oncological treatment with bevacizumab in combination with alternating capecitabine and oxaliplatin or irinotecan. Baseline ddPCR KRAS mutation allele frequency (MAF) values ranged from 0% to 63%. The first radiologic response evaluation criteria in solid tumors (RECIST) evaluation was performed 45-63 days after the initiation of treatment, and by this time three patients had an undetectable level of KRAS mutation, one had a MAF value of 0.5%, and one had a MAF value of 3% that had been reduced by 95% from the baseline value. In three of these patients the RECIST assessment was stable disease and in two partial response. In seven patients, ddPCR MAF values increased before radiological disease progression or death, while one patient remained disease-free with an undetectable KRAS mutation level. Next, we analyzed all available plasma samples with the Idylla ctKRAS system (n = 60), and found that the overall degree of agreement between ddPCR and Idylla was almost perfect (kappa value = 0.860). We used next-generation sequencing (NGS) to detect treatment-induced mutations in the last serial plasma sample of each patient, but were unable to find any new mutations when compared to the primary tumor. This study shows that ddPCR and Idylla are equally efficient for the detection of KRAS mutations in the liquid biopsies from mCRC patients and that ctDNA may indicate the disappearance of treatment responsive KRAS positive mCRC clones and serve as an early sign of disease progression.


Assuntos
Biomarcadores Tumorais/genética , DNA Tumoral Circulante/genética , Neoplasias do Colo/genética , Análise Mutacional de DNA/métodos , Proteínas Proto-Oncogênicas p21(ras)/genética , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Bevacizumab/uso terapêutico , Capecitabina/uso terapêutico , DNA Tumoral Circulante/sangue , Neoplasias do Colo/tratamento farmacológico , Feminino , Frequência do Gene , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Irinotecano/uso terapêutico , Biópsia Líquida/métodos , Masculino , Pessoa de Meia-Idade , Oxaliplatina/uso terapêutico , Reação em Cadeia da Polimerase/métodos
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