To examine the effect of
(
) on the microenvironment of colonic neoplasms and the expression of inflammatory mediators and microRNAs (miRNAs).
Levels of
DNA, cytokine gene mRNA (
,
,
,
,
,
and
), and ...potentially interacting miRNAs (miR-21-3p, miR-22-3p, miR-28-5p, miR-34a-5p, miR-135b-5p) were measured by quantitative polymerase chain reaction (qPCR) TaqMan
assays in DNA and/or RNA extracted from the disease and adjacent normal fresh tissues of 27 colorectal adenoma (CRA) and 43 colorectal cancer (CRC) patients.
mutations were detected by direct sequencing and microsatellite instability (MSI) status by multiplex PCR. Cytoscape v3.1.1 was used to construct the postulated miRNA:mRNA interaction network.
Overabundance of
in neoplastic tissue compared to matched normal tissue was detected in CRA (51.8%) and more markedly in CRC (72.1%). We observed significantly greater expression of
,
,
, and miR-135b in CRA lesions and
,
,
,
, miR-34a and miR-135b in CRC tumours compared to their respective normal tissues. Only two transcripts for miR-22 and miR-28 were exclusively downregulated in CRC tumour samples. The mRNA expression of
,
,
and miR-22 was positively correlated with
quantification in CRC tumours. The mRNA expression of miR-135b and
was inversely correlated. The miRNA:mRNA interaction network suggested that the upregulation of miR-34a in CRC proceeds
a
/
-dependent response to
. Finally,
mutations were more frequently observed in CRC samples infected with
and were associated with greater expression of miR-21 in CRA, while
was upregulated in MSI-high CRC.
Our findings indicate that
is a risk factor for CRC by increasing the expression of inflammatory mediators through a possible miRNA-mediated activation of
/
.
NTRK1, 2, and 3 fusions are important therapeutic targets for NSCLC patients, but their prevalence in South American admixed populations needs to be better explored. NTRK fusion detection in small ...biopsies is a challenge, and distinct methodologies are used, such as RNA-based next-generation sequencing (NGS), immunohistochemistry, and RNA-based nCounter. This study aimed to evaluate the frequency and concordance of positive samples for NTRK fusions using a custom nCounter assay in a real-world scenario of a single institution in Brazil. Out of 147 NSCLC patients, 12 (8.2%) cases depicted pan-NTRK positivity by IHC. Due to the absence of biological material, RNA-based NGS and/or nCounter could be performed in six of the 12 IHC-positive cases (50%). We found one case exhibiting an NTRK1 fusion and another an NTRK3 gene fusion by both RNA-based NGS and nCounter techniques. Both NTRK fusions were detected in patients diagnosed with lung adenocarcinoma, with no history of tobacco consumption. Moreover, no concomitant EGFR, KRAS, and ALK gene alterations were detected in NTRK-positive patients. The concordance rate between IHC and RNA-based NGS was 33.4%, and between immunohistochemistry and nCounter was 40%. Our findings indicate that NTRK fusions in Brazilian NSCLC patients are relatively rare (1.3%), and RNA-based nCounter methodology is a suitable approach for NRTK fusion identification in small biopsies.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Colorectal cancer (CRC) is a leading cause of cancer death worldwide. Microsatellite instability (MSI) is a genetic pathway leading to CRC, associated with particular clinicopathological features, ...and recently a major biomarker of immunotherapy response. There is little information the frequency MSI among Brazilian CRC patients, and it is still debatable the ideal methodology for MSI screening in countries with limited resources. We proposed to evaluate MSI by molecular and immunohistochemistry (IHC) methods, to compare both methodologies and also to assess the inclusion of a novel microsatellite marker,
(T17). The molecular MSI evaluation was performed using a PCR-multiplex panel in a total of 1013 CRC patients. Mismatch repair (MMR) proteins (MLH1, MSH2, MSH6 and PMS2) expression were evaluated by IHC.
(T17) marker was analyzed by fragment analysis. Molecularly, 89.5% of cases were MSI-negative and 10.5% were MSI-positive. The IHC showed that 88.9% of cases exhibited MMR-proficient status, 10.2% were MMR-deficient and 0.9% was inconclusive. Genotyping of the
(T17) in 106 MSI-positive and 215 MSI-negative cases showed its alteration only among the MSI-positive cases. We observed agreement (0.956, Kappa Test) between both molecular and IHC methodologies, with only eight discordant results, and in this subset of cases the
(T17) corroborate the molecular findings. This study suggests the use of molecular assays over IHC for MSI analysis and proposes the inclusion
(T17) marker as a complementary analysis in discordant cases.
Microbial diversity has been pointed out as a major factor in the development and progression of colorectal cancer (CRC). We sought to explore the richness and abundance of the microbial community of ...a series of colorectal tumor samples treated at Barretos Cancer Hospital, Brazil, through 16S rRNA sequencing. The presence and the impact of
Fusobacterium nucleatum
(
Fn
) DNA in CRC prognosis was further evaluated by qPCR in a series of 152 CRC cases. An enrichment for potentially oncogenic bacteria in CRC was observed, with
Fusobacterium
being the most abundant genus in the tumor tissue. In the validation dataset,
Fn
was detected in 35/152 (23.0%) of fresh-frozen tumor samples and in 6/57 (10.5%) of paired normal adjacent tissue, with higher levels in the tumor (
p
= 0.0033).
Fn
DNA in the tumor tissue was significantly associated with proximal tumors (
p
= 0.001), higher depth of invasion (
p
= 0.014), higher clinical stages (
p
= 0.033), poor differentiation (
p
= 0.011), MSI-positive status (
p
< 0.0001), BRAF mutated tumors (
p
< 0.0001), and the loss of expression of mismatch-repair proteins MLH1 (
p
< 0.0001), MSH2 (
p
= 0.003), and PMS2 (
p
< 0.0001). Moreover, the presence of
Fn
DNA in CRC tissue was also associated with a worse patient cancer-specific survival (69.9 vs. 82.2% in 5 years;
p
= 0.028) and overall survival (63.5 vs. 76.5%;
p
= 0.037). Here we report, for the first time, the association of
F. nucleatum
presence with important clinical and molecular features in a Brazilian cohort of CRC patients. Tumor detection and classification based on the gut microbiome might provide a promising approach to improve the prediction of patient outcome.
•CD276 (B7-H3) gene expression is associated with BCG unresponseviness in NMIBC patients.•IDO1 expression is a marker of worse prognosis in NMIBC.•MSI and PD-L1 expressions are not associated with ...responsiveness of BCG vaccine in NMIBC patients.
One-hundred-six patients diagnosed with non-muscle invasive bladder cancer and treated with intravesical BCG were included and divided into two groups, BCG-responsive (n = 47) and -unresponsive (n = 59). Immunohistochemistry was used to evaluate PD-L1 expression and MSI was assessed by a commercial multiplex PCR kit. The mRNA expression profile of 15 immune checkpoints was performed using the nCounter technology. For in silico validation, two distinct cohorts sourced from the Gene Expression Omnibus (GEO) database were used.
Among the 106 patients, only one (<1 %) exhibited MSI instability. PD-L1 expression was present in 9.4 % of cases, and no association was found with BCG-responsive status. We found low gene expression of canonic actionable immune checkpoints PDCD1 (PD-1), CD274 (PD-L1), and CTLA4, while high expression was observed for CD276 (B7-H3), CD47, TNFRSF14, IDO1 and PVR (CD155) genes. High IDO1 expression levels was associated with worst overall survival. The PDCD1, CTLA4 and TNFRSF14 expression levels were associated with BCG responsiveness, whereas TIGIT and CD276 were associated with unresponsiveness. Finally, CD276 was validated in silico cohorts.
In NMIBC, MSI is rare and PD-L1 expression is present in a small subset of cases. Expression levels of PDCD1, CTLA4, TNFRSF14, TIGIT and CD276 could constitute predictive biomarkers of BCG responsiveness.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Rosette-forming glioneuronal tumor (RGNT) of the IV ventricle is a rare and recently recognized brain tumor entity. It is histologically composed by two distinct features: a glial component, ...resembling pilocytic astrocytoma, and a component forming neurocytic rosettes and/or perivascular rosettes. Herein, we describe a 33-year-old man with RGNT arising in the spinal cord. Following an immunohistochemistry validation, we further performed an extensive genomic analysis, using array-CGH (aCGH), whole exome and cancer-related hotspot sequencing, in order to better understand its underlying biology. We observed the loss of 1p and gain of 1q, as well as gain of the whole chromosomes 7, 9 and 16. Local amplifications in 9q34.2 and 19p13.3 (encompassing the gene SBNO2) were identified. Moreover, we observed focal gains/losses in several chromosomes. Additionally, on chromosome 7, we identified the presence of the KIAA1549:BRAF gene fusion, which was further validated by RT-PCR and FISH. Across all mutational analyses, we detected and validated the somatic mutations of the genes MLL2, CNNM3, PCDHGC4 and SCN1A. Our comprehensive molecular profiling of this RGNT suggests that MAPK pathway and methylome changes, driven by KIAA1549:BRAF fusion and MLL2 mutation, respectively, could be associated with the development of this rare tumor entity.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Colorectal cancer (CRC) is the third most frequent and the second deadliest cancer worldwide. The ethnic structure of the population has been gaining prominence as a cancer player. The purpose of ...this study was to determine the genetic ancestry of Brazilian CRC patients. Moreover, we intended to interrogate its impact on patients' clinicopathological features.
Retrospective observational cohort study with 1,002 patients with CRC admitted from 2000 to 2014 at Barretos Cancer Hospital. Following tumor DNA isolation, genetic ancestry was assessed using a specific panel of 46 ancestry informative markers. Survival rates were obtained by the Kaplan-Meier method, and the log-rank test was used to compare the survival curves. Multivariable Cox proportional regression models were used to estimate hazard ratios (HRs).
We observed considerable admixture in the genetic composition, with the following average proportions: European 74.2%, African 12.7%, Asian 6.5%, and Amerindian 6.6%. The multivariate analysis for cancer-specific survival showed that clinical stage, lymphovascular invasion, and the presence of recurrence were associated with an increased relative risk of death from cancer (
< 0.05). High African proportion was associated with younger age at diagnosis, while high Amerindian proportion was associated with the mucinous histological subtype.
This represents the larger assessment of genetic ancestry in a population of Brazilian patients with CRC. Brazilian CRC patients exhibited similar clinicopathological features as described in Western countries.
Genetic ancestry components corroborated the significant admixture, and importantly, patients with high African proportion develop cancer at a younger age.
Targeted and immunotherapies are currently moving toward early-stage settings for patients with non-small cell lung cancer (NSCLC). Predictive biomarkers data are scarce in this scenario. We aimed to ...describe the frequency of EGFR mutations and PD-L1 expression levels in early-stage non-squamous patients with NSCLC from a large, single Brazilian oncology center.
We retrospectively evaluated patients with NSCLC diagnosed at an early-stage (IB to IIIA-AJCC seventh edition) at Barretos Cancer Hospital (n = 302). EGFR mutational status was assessed in FFPE tumor tissues using distinct methodologies (NGS, Cobas, or Sanger sequencing). PD-L1 expression was evaluated by immunohistochemistry (clone 22C3) and reported as Tumor Proportion Score (TPS), categorized as <1%, 1-49%, and ≥50%. We evaluated the association between EGFR mutational status and PD-L1 expression with sociodemographic and clinicopathological parameters by Fisher's test, qui-square test, and logistic regression. Survival analysis was assessed by the Kaplan-Meier method and Cox regression model.
EGFR mutations were detected in 17.3% (n = 48) of cases and were associated with female sex, never smokers, and longer overall and event-free survival. PD-L1 positivity was observed in 36.7% (n = 69) of cases TPS 1-49% n = 44(23.4%); TPS ≥50% n = 25(13.3%). PD-L1 positivity was associated with smoking, weight loss, and higher disease stages (IIB/IIIA).
The frequencies of EGFR mutations and PD-L1 positivity were described for early-stage non-squamous patients with NSCLC. These results will be essential for guiding treatment strategies with the recent approvals of osimertinib and immunotherapy in the adjuvant setting.
The molecular profile of endometrial cancer has become an important tool in determining patient prognosis and their optimal adjuvant treatment. In addition to The Cancer Genome Atlas (TCGA), simpler ...tools have been developed, such as the Proactive Molecular Risk Classifier for Endometrial Cancer (ProMisE). We attempted to determine a genetic signature to build a recurrence risk score in patients diagnosed with low- and intermediate-risk endometrial cancer.
A case-control study was conducted. The eligible patients were women diagnosed with recurrence low- and intermediate-risk endometrial cancer between January 2009 and December 2014 at a single institution; the recurrence patients were matched to two nonrecurrence patients with the same diagnosis by age and surgical staging. Following RNA isolation of 51 cases, 17 recurrence and 34 nonrecurrence patients, the expression profile was determined using the
, which contains 770 genes.
The expression profile was successfully characterized in 49/51 (96.1%) cases. We identified 12 genes differentially expressed between the recurrence and nonrecurrence groups. The ROC curve for each gene was generated, and all had AUCs higher than 0.7. After backward stepwise logistic regression, four genes were highlighted:
. The recurrence risk score was calculated, leading to a ROC curve of the 4-gene model with an AUC of 0.93, sensitivity of 100%, and specificity of 72.7%.
We identified a four-gene signature that may be associated with recurrence in patients with low- and intermediate-risk endometrial cancer. This finding suggests a new prognostic factor in this poorly explored group of patients with endometrial cancer.
Pyknons are specific human/primate-specific DNA motifs at least 16 nucleotides long that are repeated in blocks in intergenic and intronic regions of the genome and can be located in a new class of ...non-coding RNAs of variable length. Recent studies reported that pyknon deregulation could be involved in the carcinogenesis process, including colorectal cancer. We evaluated the expression profile of a set of 12 pyknons in a set of molecularly characterized colorectal cancer (CRC) patients. The pyknons (
PYK10
,
PYK14
,
PYK17
,
PYK26
,
PYK27
,
PYK40
,
PYK41
,
PYK42
,
PYK43
,
PYK44
,
PYK83
, and
PYK90
) expression was determined by qRT-PCR. A pilot analysis of 20 cases was performed, and consistent results were obtained for
PYK10
,
PYK17
,
PYK42
,
PYK44
, and
PYK83
. Further, the expression of the selected pyknons was evaluated in 73 CRC cases. Moreover, in 52 patients, we compared the expression profile in both tumor and normal tissues. All five pyknons analyzed showed significantly lower expression levels in the tumor compared to normal tissue. It was observed an association between expression of
PYK10
with
TP53
mutations (
p
= 0.029),
PYK17
to histologic grade (
p
= 0.035), and
PYK44
to clinical staging (
p
= 0.016). Moreover, levels of
PYK44
were significantly associated with the patient's poor overall survival (
p
= 0.04). We reported the significant downregulation of pyknons motifs in tumor tissue compared with the normal counterpart, and the association of lower
PYK44
expression with worse patient outcome. Further studies are needed to extend and validate these findings and determine the clinical-pathological impact.
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