Pan-TRK immunohistochemistry as a tool in the screening for NTRK gene fusions in cancer patients

Therapy with TRK inhibitors is a tumor-agnostic treatment directed against specific molecular changes rather than cancer type. NTRK fusions are rare in most prevalent cancers, accounting for less than 0.5% of cases. However, there is a group of rare cancers in which NTRK fusion is more prevalent. These include secretory carcinoma of the breast and salivary gland, childhood sarcomas, such as infantile fibrosarcoma, and cellular and mixed congenital mesoblastic nephroblastoma. The most common rearrangement pertains to NTRK3 and the most common fusion gene is ETV6. Identifying patients with NTRK gene fusions who would likely benefit from targeted therapy with TRK inhibitors requires practical diagnostic tools and an appropriate management strategy of diagnostic trajectory. The fusions can be detected by molecular biology techniques or pan-TRK immunohistochemistry. The latter detects NTRK1/2/3 gene fusions independently of the resulting fusion gene but does not determine which of them has been rearranged or what the fusion partner is. The sensitivity and specificity of the method reach 97% and 100%, respectively. Other advantages include the relatively low cost, short duration of examination, and broad accessibility of immunohistochemistry laboratories. These characteristics make this method a useful screening tool for detecting patients with NTRK gene fusions

SARS-CoV-2 infects an <I>in vitro</I> model of the human developing pancreas through endocytosis

Recent studies showed that SARS-CoV-2 can infect adult human pancreas and trigger pancreatic damage. Here, using human fetal pancreas samples and 3D differentiation of human pluripotent cells into pancreatic endocrine cells, we determined that SARS-CoV-2 receptors ACE2, TMPRSS2, and NRP1 are expressed in precursors of insulin-producing pancreatic β-cells, rendering them permissive to SARS-CoV-2 infection. We also show that SARS-CoV-2 enters and undergoes efficient replication in human multipotent pancreatic and endocrine progenitors in vitro. Moreover, we investigated mechanisms by which SARS-CoV-2 enters pancreatic cells, and found that ACE2 mediates the entry, while NRP1 and TMPRSS2 do not. Surprisingly, we found that in pancreatic progenitors, SARS-CoV-2 enters cells via cathepsin-dependent endocytosis, which is a different route than in respiratory tract. Therefore, pancreatic spheroids might serve as a model to study candidate drugs for endocytosis-mediated viral entry inhibition and to investigate whether SARS-CoV-2 infection may affect pancreas development, possibly causing lifelong health consequences

Advanced adenoid cystic carcinoma (ACC) is featured by SWI/SNF chromatin remodeling complex aberrations

Abstract Purpose Adenoid cystic carcinoma (ACC) is a rare neurotropic cancer with slow progression occurring in salivary glands and less frequently in other body parts. ACC is featured by hyperchromatic nuclei and various mutations in genes encoding chromatin-related machineries. The ACC treatment is mainly limited to the radical surgery and radiotherapy while the chemotherapy remains ineffective. As the knowledge about molecular basis of ACC development is limited, we investigated here the molecular features of this disease. Patients and methods This study included 50 patients with ACC. Transcript profiling of available ACC samples vs normal salivary gland tissue, quantitative real-time PCR (qRT-PCR) transcript level measurements and the immunohistochemistry (IHC) for SWI/SNF chromatin remodeling complex (CRC) subunits and androgen receptor on surgery-derived paraffinembedded samples were performed. Results Transcriptomic study followed by Gene Ontology classification indicated alteration of chromatin-related processes, including downregulated transcript levels of main SWI/SNF CRC subunits and elevated expression of BRM ATPase-coding SMARCA2 gene in ACC. Subsequent IHC indicated broad accumulation of BRM ATPase and several SWI/SNF subunits, suggesting affected control of their protein level in ACC. The IHC revealed ectopic, heterogeneous expression of androgen receptor (AR) in some ACC cells. Conclusions Our study indicated that ACC features aberrant expression of genes controlling chromatin status and structure. We found that the balance between SWI/SNF classes is moved towards the BRM ATPase-containing complex in ACC. As BRM is known to be involved in chemoresistance in cancer cells, this observation may be the likely explanation for ACC chemoresistance

Are Elevated Levels of IGF-1 Caused by Coronary Arteriesoclerosis?: Molecular and Clinical Analysis

The importance of insulin-like growth factor-1 (IGF-1) in coronary artery disease (CAD) due to wide range of its biological effects and its therapeutic potential, has already been described. Our aim was to evaluate possible influence of IGF-1 serum level changes on coronary atherosclerosis. In case of existence of such association our further aim was to verify and explain this phenomenon by examination of promoter P1 of IGF-1gene and receptor gene for IGF-1. The study was performed in 101 consecutive patients undergo for routine coronary angiography. Quantitative and qualitative assessment of coronary atherosclerosis was performed respectively by estimation of the number of culprit lesions in coronary arteries and by Gensini score calculation. IGF-1, IGFBP3 and plasma lipoproteins were measured in all patients. In addition, we evaluated DNA from 101 patients, isolated from blood cells, which was amplified by using PCR with sophisticated primers for P1 promoter of IGF-1 gene and IGF-1 receptor gene, then analyzed utilizing SSCP technique and automatically sequenced. We observed significant increase of serum IGF-1 levels in patients with “3 vessel disease” and with high score in Gensini scale when compared to those without any narrowing lesions in coronary arteries and 0 Gensini score (in group with 3 vessel disease 215.0 ± 71.3 versuss 176.7 ± 34.2 ng/ml p = 0.04 and with high Gensini score 231.4 ± 59.3 versus 181.0 ± 37.8 ng/ml p = 0.01).We found different genotypes for five P1 promoter polymorphisms of IGF-1 gene (RS35767, RS5742612, RS228837, RS11829693, RS17879774). There were no significant associations between the observed single nucleotide polymorphism (SNP) and coronary atherosclerosis nor with levels of circulating IGF-1. We found no structural polymorphism in receptor gene for IGF-1 nor in its extracellular domain(exon 2–4) nor in internal domain (exon 16–21). The effect of increased IGF-1 serum level in our study was probably independent from structural polymorphism in promoter P1 for IGF-1 or in receptor gene for IGF-1

Antiviral innate immune response in non-myeloid cells is augmented by chloride ions via an increase in intracellular hypochlorous acid levels

Abstract Phagocytes destroy ingested microbes by producing hypochlorous acid (HOCl) from chloride ions (Cl−) and hydrogen peroxide within phagolysosomes, using the enzyme myeloperoxidase. HOCl, the active ingredient in bleach, has antibacterial/antiviral properties. As myeloperoxidase is needed for HOCl production, non-myeloid cells are considered incapable of producing HOCl. Here, we show that epithelial, fibroblast and hepatic cells have enhanced antiviral activity in the presence of increasing concentrations of sodium chloride (NaCl). Replication of enveloped/non-enveloped, DNA (herpes simplex virus-1, murine gammaherpesvirus 68) and RNA (respiratory syncytial virus, influenza A virus, human coronavirus 229E, coxsackievirus B3) viruses are inhibited in a dose-dependent manner. Whilst treatment with sodium channel inhibitors did not prevent NaCl-mediated virus inhibition, a chloride channel inhibitor reversed inhibition by NaCl, suggesting intracellular chloride is required for antiviral activity. Inhibition is also reversed in the presence of 4-aminobenzoic hydrazide, a myeloperoxidase inhibitor, suggesting epithelial cells have a peroxidase to convert Cl− to HOCl. A significant increase in intracellular HOCl production is seen early in infection. These data suggest that non-myeloid cells possess an innate antiviral mechanism dependent on the availability of Cl− to produce HOCl. Antiviral activity against a broad range of viral infections can be augmented by increasing availability of NaCl

Immunohistochemia pan-TRK jako narzędzie w badaniach przesiewowych fuzji genów NTRK u chorych na nowotwory

Therapy with TRK inhibitors is a tumor-agnostic treatment directed against specific molecular changes rather than a type of cancer. NTRK fusions are rare in most prevalent cancers, accounting for less than 0.5% of cases. However, there is a group of rare cancers in which NTRK fusion is more prevalent. These include secretory carcinoma of the breast and salivary gland, childhood sarcomas, such as infantile fibrosarcoma, and cellular and mixed congenital mesoblastic nephroblastoma. The most common rearrangement pertains to NTRK3 and the most common fusion gene is ETV6. Identifying patients with NTRK gene fusions who would likely benefit from targeted therapy with TRK inhibitors requires practical diagnostic tools and an appropriate management strategy of diagnostic trajectory. The fusions can be detected by molecular biology techniques or pan-TRK immunohistochemistry. The latter detects NTRK1/2/3 gene fusions independently of the resulting fusion gene but does not determine which of them has been rearranged or what the fusion partner is. The sensitivity and the specificity of the method reach 97% and 100%, respectively. Other advantages include the relatively low cost, short duration of examination, and broad accessibility of immunohistochemistry laboratories. These characteristics make this method a useful screening tool for detecting patients with NTRK gene fusions.Stosowanie inhibitorów TRK należy do leczenia agnostycznego, co oznacza wykorzystwanie leków ukierunkowanych przeciw konkretnym zmianom molekularnym niezależnie od rodzaju nowotworu. Wśród rozpowszechnionych nowotworów fuzje NTRK występują bardzo rzadko (poniżej 0,5% przypadków). Istnieje jednak grupa rzadkich nowotworów, w których fuzje NTRK pojawiają się częściej. Należą do niej rak wydzielniczy piersi i ślinianki, mięsaki wieku dziecięcego (włókniakomięsak typu niemowlęcego oraz komórkowy i mieszany wrodzony nerczak mezoblastyczny). Najczęściej rearanżacje dotyczą genu NTRK3, a najczęstszym genem fuzyjnym jest ETV6. Identyfikacja chorych z fuzjami genów NTRK, którzy potencjalnie mogliby odnieść korzyści z leczenia inhibitorami TRK, wymaga skutecznych narzędzi diagnostycznych i odpowiedniej strategii postępowania. Fuzje genów NTRK mogą być wykrywane metodami biologii molekularnej lub metodą immunohistochemiczną pan-TRK. Druga — z wymienionych — ścieżka diagnostyczna umożliwia wykrywanie fuzji genów NTRK1/2/3 niezależnie od powstałego genu fuzyjnego, jednak bez określenia, który gen uległ rearanżacji oraz jaki jest gen fuzyjny. Czułość metody sięga 97%, natomiast swoistość wynosi 100%. Innymi zaletami metody jest stosunkowo niski koszt, krótki czas wykonania badania oraz dostępność pracowni immunohistochemicznych. Z uwagi na powyższe cechy metoda ta może być użyta w badaniach przesiewowych w kierunku wykrycia fuzji genów NTRK

Aberrant Activity of Histone–Lysine N-Methyltransferase 2 (KMT2) Complexes in Oncogenesis

KMT2 (histone-lysine N-methyltransferase subclass 2) complexes methylate lysine 4 on the histone H3 tail at gene promoters and gene enhancers and, thus, control the process of gene transcription. These complexes not only play an essential role in normal development but have also been described as involved in the aberrant growth of tissues. KMT2 mutations resulting from the rearrangements of the KMT2A (MLL1) gene at 11q23 are associated with pediatric mixed-lineage leukemias, and recent studies demonstrate that KMT2 genes are frequently mutated in many types of human cancers. Moreover, other components of the KMT2 complexes have been reported to contribute to oncogenesis. This review summarizes the recent advances in our knowledge of the role of KMT2 complexes in cell transformation. In addition, it discusses the therapeutic targeting of different components of the KMT2 complexes