Recent developments in experimental and computational chemistry have identified a rapidly growing class of nucleophilic aromatic substitutions that proceed by concerted (cSNAr) rather than classical, ...two‐step, SNAr mechanisms. Whereas traditional SNAr reactions require substantial activation of the aromatic ring by electron‐withdrawing substituents, such activating groups are not mandatory in the concerted pathways.
Concerted or stepwise? A class of nucleophilic aromatic substitutions has been developed that proceeds by concerted (cSNAr) rather than classical, two‐step, SNAr mechanisms. Whereas traditional SNAr reactions require substantial activation of the aromatic ring by electron‐withdrawing substituents, such activating groups are not mandatory in the concerted pathways.
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The majority of deaths from breast cancer occur following the development of metastatic disease, a process inhibited by β-blockers in preclinical studies. This phase II randomized controlled trial ...evaluated the effect of preoperative β-blockade with propranolol on biomarkers of metastatic potential and the immune cell profile within the primary tumor of patients with breast cancer.
In this triple-blind placebo-controlled clinical trial, 60 patients were randomly assigned to receive an escalating dose of oral propranolol (
= 30; 80-160 mg daily) or placebo (
= 30) for 7 days prior to surgery. The primary endpoint investigated the effect of propranolol on prometastatic and proinflammatory gene expression within the primary tumor.
Propranolol downregulated primary tumor expression of mesenchymal genes (
= 0.002) without affecting epithelial gene expression (
= 0.21). Bioinformatic analyses implicated downregulation of Snail/Slug (
= 0.03), NF-κB/Rel (
< 0.01), and AP-1 (
< 0.01) transcription factors in structuring the observed transcriptome alterations, and identified changes in intratumoral neutrophil, natural killer cell, and dendritic cell recruitment (all
< 0.01). Patients with clinical evidence of drug response (lowered heart rate and blood pressure) demonstrated elevated tumor infiltration of CD68
macrophages and CD8
T cells.
One week of β-blockade with propranolol reduced intratumoral mesenchymal polarization and promoted immune cell infiltration in early-stage surgically-resectable breast cancer. These results show that β-blockade reduces biomarkers associated with metastatic potential, and support the need for larger phase III clinical trials powered to detect the impact of β-blockade on cancer recurrence and survival.
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In recent years anatomical pathology has been revolutionised by the incorporation of molecular findings into routine diagnostic practice, and in some diseases the presence of specific molecular ...alterations are now essential for diagnosis. Spatial transcriptomics describes a group of technologies that provide up to transcriptome‐wide expression profiling while preserving the spatial origin of the data, with many of these technologies able to provide these data using a single tissue section. Spatial transcriptomics allows expression profiling of highly specific areas within a tissue section potentially to subcellular resolution, and allows correlation of expression data with morphology, tissue type and location relative to other structures. While largely still research laboratory‐based, several spatial transcriptomics methods have now achieved compatibility with formalin‐fixed paraffin‐embedded tissue (FFPE), allowing their use in diagnostic tissue samples, and with further development potentially leading to their incorporation in routine anatomical pathology practice. This mini review provides an overview of spatial transcriptomics methods, with an emphasis on platforms compatible with FFPE tissue, approaches to assess the data and potential applications in anatomical pathology practice.
This mini review provides an overview of spatial transcriptomics methods, with an emphasis on platforms compatible with FFPE tissue, approaches to assess the data and potential applications in anatomical pathology practice.
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The immune microenvironment of breast ductal carcinoma
(DCIS) has yet to be fully explored, and the relationship of immune cells to genetic features of DCIS is unknown.
We quantified tumor associated ...lymphocytes (TIL) and evaluated PD-L1 protein levels by immunohistochemistry in a cohort of pure DCIS (138 and 79 cases, respectively), some of which had copy number (
= 55) and mutation data (
= 20).
TILs were identified in the stroma surrounding DCIS (119/138, 86%) and present at a median TIL score of 5% (range, 0%-90%). Most DCIS were negative for tumor cell PD-L1 staining (89%), but 25% of cases were positive for immune cell staining. We observed that, as in invasive breast cancer, TILs and PD-L1 positivity were significantly greater in high-grade (
= 0.002/0.035), ER-negative (
= 0.02/0.02), and
-amplified tumors (
< 0.001/0.048). Comedo necrosis was significantly positively associated with TILs (
< 0.0001) but not with PD-L1. The TILs score was significantly higher in cases with
mutation (
= 0.03) but not with
or
mutation. In the cases with copy number data, both the fraction of the genome altered and the number of telomeric imbalances were significantly positively correlated with TILs (both
< 0.001). This result strongly contrasted with invasive breast cancer data, where aneuploidy was not correlated to TIL levels.
Although a small cohort, our data suggest a preliminary model by which the progression of DCIS to invasive carcinoma may involve an altered relationship of tumor copy number with the immune microenvironment, possibly by the immunoediting of the tumor.
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Immune checkpoint inhibitors have emerged as a potent new class of anticancer therapy. They have changed the treatment landscape for a range of tumors, particularly those with a high mutational load. ...To date, however, modest results have been observed in breast cancer, where tumors are rarely hypermutated. Because
-associated tumors frequently exhibit a triple-negative phenotype with extensive lymphocyte infiltration, we explored their mutational load, immune profile, and response to checkpoint inhibition in a
-deficient tumor model.
-mutated triple-negative breast cancers (TNBCs) exhibited an increased somatic mutational load and greater numbers of tumor-infiltrating lymphocytes, with increased expression of immunomodulatory genes including
(
) and
, when compared to TNBCs from
-wild-type patients. Cisplatin treatment combined with dual anti-programmed death-1 and anti-cytotoxic T lymphocyte-associated antigen 4 therapy substantially augmented antitumor immunity in
-deficient mice, resulting in an avid systemic and intratumoral immune response. This response involved enhanced dendritic cell activation, reduced suppressive FOXP3
regulatory T cells, and concomitant increase in the activation of tumor-infiltrating cytotoxic CD8
and CD4
T cells, characterized by the induction of polyfunctional cytokine-producing T cells. Dual (but not single) checkpoint blockade together with cisplatin profoundly attenuated the growth of
-deficient tumors in vivo and improved survival. These findings provide a rationale for clinical studies of combined immune checkpoint blockade in
-associated TNBC.
Ductal carcinoma in situ (DCIS) accounts for ~20–25% of breast cancers. While DCIS is not life‐threatening, it may progress to invasive carcinoma over time, and treatment intended to prevent invasive ...progression may itself cause significant morbidity. Accurate risk assessment is therefore necessary to avoid over‐ or undertreatment of an individual patient. In this review we will outline the evidence for current management of DCIS, discuss approaches to DCIS risk assessment and challenges facing identification of novel DCIS biomarkers.
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A protocol for regio‐controlled hydromagnesiation of 1,3‐enynes was developed using magnesium hydride that is generated in situ by solvothermal treatment of sodium hydride (NaH) and magnesium iodide ...(MgI2) in THF. The resulting allenylmagnesium species could be converted into tri‐ and tetra‐substituted allenes by subsequent treatment with various carbon‐ and silicon‐based electrophiles with the aid of CuCN as a catalyst.
Regio‐controlled hydromagnesiation of 1,3‐enynes was enabled by magnesium hydride. Subsequent downstream alkylation and silylation allowed for construction of tri‐ and tetra‐substituted allenes.
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Clinical specimens undergoing diagnostic molecular pathology testing are fixed in formalin due to the necessity for detailed morphological assessment. However, formalin fixation can cause major ...issues with molecular testing, as it causes DNA damage such as fragmentation and non-reproducible sequencing artefacts after PCR amplification. In the context of massively parallel sequencing (MPS), distinguishing true low frequency variants from sequencing artefacts remains challenging. The prevalence of formalin-induced DNA damage and its impact on molecular testing and clinical genomics remains poorly understood.
The Cancer 2015 study is a population-based cancer cohort used to assess the feasibility of mutational screening using MPS in cancer patients from Victoria, Australia. While blocks were formalin-fixed and paraffin-embedded in different anatomical pathology laboratories, they were centrally extracted for DNA utilising the same protocol, and run through the same MPS platform (Illumina TruSeq Amplicon Cancer Panel). The sequencing artefacts in the 1-10% and the 10-25% allele frequency ranges were assessed in 488 formalin-fixed tumours from the pilot phase of the Cancer 2015 cohort. All blocks were less than 2.5 years of age (mean 93 days).
Consistent with the signature of DNA damage due to formalin fixation, many formalin-fixed samples displayed disproportionate levels of C>T/G>A changes in the 1-10% allele frequency range. Artefacts were less apparent in the 10-25% allele frequency range. Significantly, changes were inversely correlated with coverage indicating high levels of sequencing artefacts were associated with samples with low amounts of available amplifiable template due to fragmentation. The degree of fragmentation and sequencing artefacts differed between blocks sourced from different anatomical pathology laboratories. In a limited validation of potentially actionable low frequency mutations, a NRAS G12D mutation in a melanoma was shown to be a false positive.
These findings indicate that DNA damage following formalin fixation remains a major challenge in laboratories working with MPS. Methodologies that assess, minimise or remove formalin-induced DNA damaged templates as part of MPS protocols will aid in the interpretation of genomic results leading to better patient outcomes.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
PD-1 immune checkpoint blockade provides significant clinical benefits for melanoma patients. We analyzed the somatic mutanomes and transcriptomes of pretreatment melanoma biopsies to identify ...factors that may influence innate sensitivity or resistance to anti-PD-1 therapy. We find that overall high mutational loads associate with improved survival, and tumors from responding patients are enriched for mutations in the DNA repair gene BRCA2. Innately resistant tumors display a transcriptional signature (referred to as the IPRES, or innate anti-PD-1 resistance), indicating concurrent up-expression of genes involved in the regulation of mesenchymal transition, cell adhesion, extracellular matrix remodeling, angiogenesis, and wound healing. Notably, mitogen-activated protein kinase (MAPK)-targeted therapy (MAPK inhibitor) induces similar signatures in melanoma, suggesting that a non-genomic form of MAPK inhibitor resistance mediates cross-resistance to anti-PD-1 therapy. Validation of the IPRES in other independent tumor cohorts defines a transcriptomic subset across distinct types of advanced cancer. These findings suggest that attenuating the biological processes that underlie IPRES may improve anti-PD-1 response in melanoma and other cancer types.
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•High mutational loads may predict better survival, but not response to anti-PD-1•BRCA2 mutations are enriched in melanomas responsive to anti-PD-1•A transcriptional signature is related to innate anti-PD-1 resistance (IPRES)•IPRES defines a transcriptomic subset across distinct types of advanced cancer
High mutational loads are associated with improved survival in melanoma patients but are not predictive of response to anti-PD-1 therapy, suggesting that other genomic and non-genomic features also contribute to response patterns on PD-1 checkpoint blockade therapy.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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