Translating whole-exome sequencing (WES) for prospective clinical use may have an impact on the care of patients with cancer; however, multiple innovations are necessary for clinical implementation. ...These include rapid and robust WES of DNA derived from formalin-fixed, paraffin-embedded tumor tissue, analytical output similar to data from frozen samples and clinical interpretation of WES data for prospective use. Here, we describe a prospective clinical WES platform for archival formalin-fixed, paraffin-embedded tumor samples. The platform employs computational methods for effective clinical analysis and interpretation of WES data. When applied retrospectively to 511 exomes, the interpretative framework revealed a 'long tail' of somatic alterations in clinically important genes. Prospective application of this approach identified clinically relevant alterations in 15 out of 16 patients. In one patient, previously undetected findings guided clinical trial enrollment, leading to an objective clinical response. Overall, this methodology may inform the widespread implementation of precision cancer medicine.
Translating whole exome sequencing (WES) for prospective clinical use may impact the care of cancer patients; however, multiple innovations are necessary for clinical implementation. These include: ...(1) rapid and robust WES from formalin-fixed paraffin embedded (FFPE) tumor tissue, (2) analytical output similar to data from frozen samples, and (3) clinical interpretation of WES data for prospective use. Here, we describe a prospective clinical WES platform for archival FFPE tumor samples. The platform employs computational methods for effective clinical analysis and interpretation of WES data. When applied retrospectively to 511 exomes, the interpretative framework revealed a “long tail” of somatic alterations in clinically important genes. Prospective application of this approach identified clinically relevant alterations in 15/16 patients. In one patient, previously undetected findings guided clinical trial enrollment leading to an objective clinical response. Overall, this methodology may inform the widespread implementation of precision cancer medicine.
Nicotinamide adenine dinucleotide (NAD
+
) is an essential co-factor for cellular metabolism and serves as a substrate in enzymatic processes. NAD
+
is produced by
de novo
synthesis or salvage ...pathways in nearly all bacterial species.
Haemophilus influenzae
lacks the capacity for
de novo
synthesis, so it is dependent on import of NAD
+
from the external environment or salvage biosynthetic pathways for recycling of NAD
+
precursors and breakdown products. However, the actual sources of NAD
+
utilized by
H
.
influenzae
in the respiratory tract are not well defined. In this study, we found that a variety of bacteria, including species found in the upper airway of humans, released NAD
+
that was readily detectable in extracellular culture fluid, and which supported growth of
H
.
influenzae in vitro
. By contrast, certain strains of
Streptococcus pyogenes
(group A streptococcus or GAS) inhibited growth of
H
.
influenzae in vitro
by secreting NAD
+
-glycohydrolase (NADase), which degraded extracellular NAD
+
. Conversely, GAS strains that lacked enzymatically active NADase released extracellular NAD
+
, which could support
H
.
influenzae
growth. Our results suggest that many bacterial species, including normal flora of the upper airway, release NAD
+
into the environment. GAS is distinctive in its ability to both release and degrade NAD
+
. Thus, colonization of the airway with
H
.
influenzae
may be promoted or restricted by co-colonization with GAS in a strain-specific manner that depends, respectively, on release of NAD
+
or secretion of active NADase. We suggest that, in addition to its role as a cytotoxin for host cells, NADase may serve a separate function by restricting growth of
H
.
influenzae
in the human respiratory tract.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
PD-1 blockade has transformed the management of advanced clear cell renal cell carcinoma (ccRCC), but the drivers and resistors of the PD-1 response remain incompletely elucidated. Here, we analyzed ...592 tumors from patients with advanced ccRCC enrolled in prospective clinical trials of treatment with PD-1 blockade by whole-exome and RNA sequencing, integrated with immunofluorescence analysis, to uncover the immunogenomic determinants of the therapeutic response. Although conventional genomic markers (such as tumor mutation burden and neoantigen load) and the degree of CD8
T cell infiltration were not associated with clinical response, we discovered numerous chromosomal alterations associated with response or resistance to PD-1 blockade. These advanced ccRCC tumors were highly CD8
T cell infiltrated, with only 27% having a non-infiltrated phenotype. Our analysis revealed that infiltrated tumors are depleted of favorable PBRM1 mutations and enriched for unfavorable chromosomal losses of 9p21.3, as compared with non-infiltrated tumors, demonstrating how the potential interplay of immunophenotypes with somatic alterations impacts therapeutic efficacy.
Summary
Background
The efficacy and safety of certolizumab pegol (CZP) in moderate‐to‐severe Crohn's disease were demonstrated in two 26‐week double‐blind studies (PRECiSE 1 & 2).
Aim
To report the ...safety and efficacy outcomes of long‐term, CZP therapy from PRECiSE 3, in which patients received treatment up to 7 years treatment.
Methods
Patients completing PRECiSE 1 or 2 were eligible to enter PRECiSE 3 in which they received CZP 400 mg, open‐label, every 4 weeks (without additional induction therapy) for up to 7 years, for up to 91 doses from study start. Safety (adverse events, including infections and malignancies) and efficacy (Harvey–Bradshaw Index, faecal calprotectin, C‐reactive protein) were prospectively monitored. Remission was analysed using observed cases, last observation carried forward imputation and nonresponder imputation.
Results
A total of 595 patients entered the study; 117 (20%) completed 7 years. Discontinuation rates were 29.2%, 13.6%, 16.1%, 7.9%, 5.0%, 4.5% and 3.9% (years 1–7 respectively). During 1920 patient‐years of exposure to CZP, no new safety signals were observed. Incidence rates (new cases/100 patient‐years) for serious infections and malignant neoplasms were 4.37 and 1.06 respectively. No lymphoproliferative malignancies were reported. Clinical remission rates were ≥68% at each year (observed cases); rates by last observation carried forward and nonresponder imputation were 58% and 45% at year 1, 56% and 26% at year 3 and 55% and 13% at year 7 respectively.
Conclusion
Certolizumab pegol was well tolerated in the long‐term treatment of Crohn's disease, with sustained remission in some patients continuing in the study for up to 7 years. ClinicalTrials.gov identifier NCT00552058.
As researchers begin probing deep coverage sequencing data for increasingly rare mutations and subclonal events, the fidelity of next generation sequencing (NGS) laboratory methods will become ...increasingly critical. Although error rates for sequencing and polymerase chain reaction (PCR) are well documented, the effects that DNA extraction and other library preparation steps could have on downstream sequence integrity have not been thoroughly evaluated. Here, we describe the discovery of novel C > A/G > T transversion artifacts found at low allelic fractions in targeted capture data. Characteristics such as sequencer read orientation and presence in both tumor and normal samples strongly indicated a non-biological mechanism. We identified the source as oxidation of DNA during acoustic shearing in samples containing reactive contaminants from the extraction process. We show generation of 8-oxoguanine (8-oxoG) lesions during DNA shearing, present analysis tools to detect oxidation in sequencing data and suggest methods to reduce DNA oxidation through the introduction of antioxidants. Further, informatics methods are presented to confidently filter these artifacts from sequencing data sets. Though only seen in a low percentage of reads in affected samples, such artifacts could have profoundly deleterious effects on the ability to confidently call rare mutations, and eliminating other possible sources of artifacts should become a priority for the research community.
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