Natural history of clonal haematopoiesis seen in real-world haematology settings.

Recursive partitioning of healthy consortia led to the development of the Clonal Hematopoiesis Risk Score (CHRS) for clonal haematopoiesis (CH); however, in the practical setting, most cases of CH are diagnosed after patients present with cytopenias or related symptoms. To address this real-world population, we characterize the clinical trajectories of 94 patients with CH and distinguish CH harbouring canonical DNMT3A/TET2/ASXL1 mutations alone ('sole DTA') versus all other groups ('non-sole DTA'). TET2, rather than DNMT3A, was the most prevalent mutation in the real-world setting. Sole DTA patients did not progress to myeloid neoplasm (MN) in the absence of acquisition of other mutations. Contrastingly, 14 (20.1%) of 67 non-sole DTA patients progressed to MN. CHRS assessment showed a higher frequency of high-risk CH in non-sole DTA (vs. sole DTA) patients and in progressors (vs. non-progressors). RUNX1 mutation conferred the strongest risk for progression to MN (odds ratio [OR] 10.27, 95% CI 2.00-52.69, p = 0.0053). The mean variant allele frequency across all genes was higher in progressors than in non-progressors (36.9% ± 4.62% vs. 24.1% ± 1.67%, p = 0.0064). This analysis in the post-CHRS era underscores the natural history of CH, providing insight into patterns of progression to MN.

The BRAF oncoprotein functions through the transcriptional repressor MAFG to mediate the CpG Island Methylator phenotype.

Most colorectal cancers (CRCs) containing activated BRAF (BRAF[V600E]) have a CpG island methylator phenotype (CIMP) characterized by aberrant hypermethylation of many genes, including the mismatch repair gene MLH1. MLH1 silencing results in microsatellite instability and a hypermutable phenotype. Through an RNAi screen, here we identify the transcriptional repressor MAFG as the pivotal factor required for MLH1 silencing and CIMP in CRCs containing BRAF(V600E). In BRAF-positive human CRC cell lines and tumors, MAFG is bound at the promoters of MLH1 and other CIMP genes, and recruits a corepressor complex that includes its heterodimeric partner BACH1, the chromatin remodeling factor CHD8, and the DNA methyltransferase DNMT3B, resulting in hypermethylation and transcriptional silencing. BRAF(V600E) increases BRAF/MEK/ERK signaling resulting in phosphorylation and elevated levels of MAFG, which drives DNA binding. Analysis of transcriptionally silenced CIMP genes in KRAS-positive CRCs indicates that different oncoproteins direct the assembly of distinct repressor complexes on common promoters.

Prosurvival kinase PIM2 is a therapeutic target for eradication of chronic myeloid leukemia stem cells.

A major obstacle to curing chronic myeloid leukemia (CML) is the intrinsic resistance of CML stem cells (CMLSCs) to the drug imatinib mesylate (IM). Prosurvival genes that are preferentially expressed in CMLSCs compared with normal hematopoietic stem cells (HSCs) represent potential therapeutic targets for selectively eradicating CMLSCs. However, the discovery of such preferentially expressed genes has been hampered by the inability to completely separate CMLSCs from HSCs, which display a very similar set of surface markers. To overcome this challenge, and to minimize confounding effects of individual differences in gene expression profiles, we performed single-cell RNA-seq on CMLSCs and HSCs that were isolated from the same patient and distinguished based on the presence or absence of BCR-ABL. Among genes preferentially expressed in CMLSCs is PIM2, which encodes a prosurvival serine-threonine kinase that phosphorylates and inhibits the proapoptotic protein BAD. We show that IM resistance of CMLSCs is due, at least in part, to maintenance of BAD phosphorylation by PIM2. We find that in CMLSCs, PIM2 expression is promoted by both a BCR-ABL-dependent (IM-sensitive) STAT5-mediated pathway and a BCR-ABL-independent (IM-resistant) STAT4-mediated pathway. Combined treatment with IM and a PIM inhibitor synergistically increases apoptosis of CMLSCs, suppresses colony formation, and significantly prolongs survival in a mouse CML model, with a negligible effect on HSCs. Our results reveal a therapeutically targetable mechanism of IM resistance in CMLSCs. The experimental approach that we describe can be generally applied to other malignancies that harbor oncogenic fusion proteins or other characteristic genetic markers.

Molecular and immunophenotypic characterization of anal squamous cell carcinoma reveals distinct clinicopathologic groups associated with HPV and TP53 mutation status.

Squamous cell carcinoma (SqCC) is the most common malignancy of the anal canal, where it is strongly associated with HPV infection. Characteristic genomic alterations have been identified in anal SqCC, but their clinical significance and correlation with HPV status, pathologic features, and immunohistochemical markers are not well established. We examined the molecular and clinicopathologic features of 96 HPV-positive and 20 HPV-negative anal SqCC. HPV types included 89 with HPV16, 2 combined HPV16/HPV18, and 5 HPV33. HPV-positive cases demonstrated frequent mutations or amplifications in PIK3CA (30%; p = 0.027) or FBXW7 mutations (10%). HPV-negativity was associated with frequent TP53 (53%; p = 0.00001) and CDKN2A (21%; p = 0.0045) mutations. P16 immunohistochemistry was positive in all HPV-positive cases and 3/20 HPV-negative cases (p < 0.0001; sensitivity: 100%; specificity: 85%) and was associated with basaloid morphology (p = 0.0031). Aberrant p53 immunohistochemical staining was 100% sensitive and specific for TP53 mutation (p < 0.0001). By the Kaplan-Meier method, HPV-negativity, aberrant p53 staining, and TP53 mutation were associated with inferior overall survival (OS) (p < 0.0001, p = 0.0103, p = 0.0103, respectively) and inferior recurrence-free survival (p = 0.133, p = 0.0064, and p = 0.0064, respectively). TP53/p53 status stratified survival probability by HPV status (p = 0.013), with HPV-negative/aberrant p53 staining associated with the worst OS, HPV-positive/wild-type p53 with best OS, and HPV-positive/aberrant p53 or HPV-negative/wild-type p53 with intermediate OS. On multivariate analysis HPV status (p = 0.0063), patient age (p = 0.0054), T stage (p = 0.039), and lymph node involvement (p = 0.044) were independently associated with OS. PD-L1 expression (CPS ≥ 1) was seen in 30% of HPV-positive and 40% of HPV-negative cases, and PD-L1 positivity was associated with a trend toward inferior OS within the HPV-negative group (p = 0.064). Our findings suggest that anal SqCC can be subclassified into clinically, pathologically, and molecularly distinct groups based on HPV and TP53 mutation status, and p16 and p53 immunohistochemistry represent a clinically useful method of predicting these prognostic groups.

CD56 expression in basaloid anal squamous cell carcinoma - A potential diagnostic pitfall.

Anal squamous cell carcinoma (SqCC) is a morphologically heterogeneous entity. Basaloid and non-keratinizing anal SqCC may be confused with other tumors including neuroendocrine carcinoma due to morphologic overlap, and expression of neuroendocrine markers is not well-studied in anal SqCC. Prompted by a case of anal SqCC that was initially misdiagnosed as neuroendocrine carcinoma on the basis of morphology and CD56 expression, we retrospectively examined the expression of neuroendocrine markers CD56, synaptophysin, and chromogranin in 48 cases of basaloid anal SqCC, with clinicopathologic correlation. HPV16 was identified in 46 cases, HPV33 in one case, and one case was HPV-negative. Three (6.3%) cases demonstrated CD56 expression, including two with diffuse and one with focal expression. Two CD56-positive cases demonstrated basaloid morphology with peripheral palisading and the other demonstrated adenoid cystic/cylindroma-like morphology. None of the cases showed significant synaptophysin or chromogranin expression. The three cases expressing CD56 were HPV16-positive, and one demonstrated a CTNNB1 mutation. There was no difference in clinicopathologic features including stage, outcome, or HPV status, between CD56-positive and negative groups. Our findings support that CD56 expression is infrequently expressed in anal SqCC and is not indicative of neuroendocrine differentiation in the absence of expression of more specific neuroendocrine markers such as synaptophysin and chromogranin. Pathologists should be aware that CD56 expression may occur in basaloid anal SqCC and is a diagnostic pitfall due to morphologic overlap with neuroendocrine carcinoma and other tumors including basal cell carcinoma.

Dedifferentiated Carcinoma of the Endometrium Associated With Low-grade Müllerian Adenosarcoma: A Clinicopathologic Case Report Including the Immunohistochemical and Molecular Profile.

Dedifferentiated carcinoma is defined as undifferentiated carcinoma coexisting with a second component of FIGO grade 1 or 2 endometrioid carcinoma. It is a rare entity with highly aggressive behavior. Dedifferentiated carcinoma combined with another primary uterine tumor is even rarer. We describe a case containing 3 different morphologies comprised of a dedifferentiated carcinoma associated with a low-grade endometrioid carcinoma coexisting with a low-grade Müllerian adenosarcoma. We also used targeted genomic analysis to show all 3 components arise from the same founding clone and identify novel mutations that drive tumor progression.

Next-Generation Sequencing to Detect Deletion of RB1 and ERBB4 Genes in Chromophobe Renal Cell Carcinoma: A Potential Role in Distinguishing Chromophobe Renal Cell Carcinoma from Renal Oncocytoma.

Overlapping morphologic, immunohistochemical, and ultrastructural features make it difficult to diagnose chromophobe renal cell carcinoma (ChRCC) and renal oncocytoma (RO). Because ChRCC is a malignant tumor, whereas RO is a tumor with benign behavior, it is important to distinguish these two entities. We aimed to identify genetic markers that distinguish ChRCC from RO by using next-generation sequencing (NGS). NGS for hotspot mutations or gene copy number changes was performed on 12 renal neoplasms, including seven ChRCC and five RO cases. Matched normal tissues from the same patients were used to exclude germline variants. Rare hotspot mutations were found in cancer-critical genes (TP53 and PIK3CA) in ChRCC but not RO. The NGS gene copy number analysis revealed multiple abnormalities. The two most common deletions were tumor-suppressor genes RB1 and ERBB4 in ChRCC but not RO. Fluorescence in situ hybridization was performed on 65 cases (ChRCC, n = 33; RO, n = 32) to verify hemizygous deletion of RB1 (17/33, 52%) or ERBB4 (11/33, 33%) in ChRCC, but not in RO (0/32, 0%). In total, ChRCCs (23/33, 70%) carry either a hemizygous deletion of RB1 or ERBB4. The combined use of RB1 and ERBB4 fluorescence in situ hybridization to detect deletion of these genes may offer a highly sensitive and specific assay to distinguish ChRCC from RO.

Common BRAF(V600E)-directed pathway mediates widespread epigenetic silencing in colorectal cancer and melanoma.

During cancer development, it is well established that many genes, including tumor suppressor genes, are hypermethylated and transcriptionally repressed, a phenomenon referred to as epigenetic silencing. In general, the factors involved in, and the mechanistic basis of, epigenetic silencing during cancer development are not well understood. We have recently described an epigenetic silencing pathway, directed by the oncogenic B-Raf proto-oncogene (BRAF) variant BRAF(V600E), that mediates widespread epigenetic silencing in colorectal cancer (CRC). Notably, the BRAF(V600E) mutation is also present in 50-70% of melanomas. Here, we show that the same pathway we identified in CRC also directs epigenetic silencing of a similar set of genes in BRAF-positive melanoma. In both CRC and melanoma, BRAF(V600E) promotes epigenetic silencing through up-regulation of v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog G (MAFG), a transcriptional repressor with sequence-specific DNA-binding activity. The elevated concentration of MAFG drives DNA binding on the promoter. Promoter-bound MAFG recruits a set of corepressors that includes its heterodimeric partner BTB and CNC homology 1, basic leucine zipper transcription factor 1 (BACH1), the chromatin remodeling factor chromodomain helicase DNA-binding protein 8 (CHD8), and the DNA methyltransferase DNMT3B, resulting in hypermethylation and transcriptional silencing. Our results reveal a common BRAF(V600E)-directed transcriptional regulatory pathway that mediates epigenetic silencing in unrelated solid tumors and provide strong support for an instructive model of oncoprotein-directed epigenetic silencing.

Immunohistochemical loss of 5-hydroxymethylcytosine expression in acute myeloid leukaemia: relationship to somatic gene mutations affecting epigenetic pathways.

AIMS: Genes affecting epigenetic pathways are frequently mutated in myeloid malignancies, including acute myeloid leukaemia (AML). The genes encoding TET2, IDH1 and IDH2 are among the most commonly mutated genes, and cause defective conversion of 5-methylcytosine into 5-hydroxymethylcytosine (5hmC), impairing demethylation of DNA, and presumably serving as driver mutations in leukaemogenesis. The aim of this study was to correlate 5hmC immunohistochemical loss with the mutation status of genes involved in epigenetic pathways in AML. METHODS AND RESULTS: Immunohistochemical staining with an anti-5hmC antibody was performed on 41 decalcified, formalin-fixed paraffin-embedded (FFPE) bone marrow biopsies from patients with AML. Archived DNA was subjected to next-generation sequencing for analysis of a panel of genes, including TET2, IDH1, IDH2, WT1 and DNMT3A. TET2, IDH1, IDH2, WT1 and DNMT3A mutations were found in 46% (19/41) of the cases. Ten of 15 cases (67%) with TET2, IDH1, IDH2 or WT1 mutations showed deficient 5hmC staining, whereas nine of 26 cases (35%) without a mutation in these genes showed loss of 5hmC. It is of note that all four cases with TET2 mutations showed deficient 5hmC staining. CONCLUSIONS: Overall, somatic mutations in TET2, IDH1, IDH2, WT1 and DNMT3A were common in our cohort of AML cases. Immunohistochemical staining for 5hmC was lost in the majority of cases harbouring mutations in these genes, reflecting the proposed relationship between dysfunctional epigenetic pathways and leukaemogenesis.

Mutually exclusive extracellular signal-regulated kinase pathway mutations are present in different stages of multi-focal pulmonary Langerhans cell histiocytosis supporting clonal nature of the disease.

AIMS: Pulmonary Langerhans cell histiocytosis (PLCH) is an idiopathic cigarette smoking-related disorder of the lung. Molecular changes in cellular or fibrotic stages of PLCH have not been investigated. We studied the prevalence of extracellular signal-regulated kinase (ERK) pathway mutations in different PLCH stages and other non-PLCH smoking-related lung diseases. METHODS AND RESULTS: The cohort included 28 PLCH with cellular (n = 10), mixed cellular/fibrotic (n = 4) and fibrotic histology (n = 14). Seven cases had concurrent multi-focal/multi-lobar tumours. Respiratory bronchiolitis interstitial lung disease (RB-ILD, n = 2), desquamative interstitial pneumonia (DIP, n = 4) and mixed RB-ILD/DIP (n = 2) were included for comparison. BRAF(V) (600E) immunohistochemistry, next-generation sequencing (NGS) and peptide nucleic acid (PNA) clamp polymerase chain reaction (PCR) with high analytical sensitivity (<0.1-0.2%) were used to analyse RAS, BRAF and MAP2K1 genes. Of 26 cases with gene mutation data, BRAF(V) (600E) was identified in eight of 12 (67%) cellular cases and in one of 14 (7%) fibrotic cases. MAP2K1 or KRAS mutations were observed in four of 14 (29%) fibrotic cases and three of the 12 (25%) cellular cases. Multi-focal/multi-lobar specimens carried identical BRAF (n = 5) or non-hotspot MAP2K1 (n = 2) mutations. The other smoking-related disorders were negative for mutations. Patients with cellular lesions or BRAF mutation were significantly younger than patients with fibrotic or BRAF wild-type PLCH. CONCLUSION: The presence of identical but mutually exclusive ERK pathway mutations in multi-focal PLCH supports a neoplastic/clonal origin for this disease. Patient age and mutation type differed between cellular and fibrotic histology and may indicate a natural progression or a mutation-specific pathogenicity.

Complete Spontaneous Regression of Merkel Cell Carcinoma After Biopsy: A Case Report and Review of the Literature.

Merkel cell carcinoma (MCC) is a rare primary cutaneous neuroendocrine tumor that typically occurs on the head and neck of the elderly and follows an aggressive clinical course. Merkel cell polyomavirus (MCPyV) has been identified in up to 80% of cases and has been shown to participate in MCC tumorigenesis. Complete spontaneous regression of MCC has been rarely reported in the literature. We describe a case of a 79-year-old man that presented with a rapidly growing, 3-cm mass on the left jaw. An incisional biopsy revealed MCC. Additional health issues were discovered in the preoperative workup of this patient which delayed treatment. One month after the biopsy, the lesion showed clinical regression in the absence of treatment. Wide excision of the biopsy site with sentinel lymph node dissection revealed no evidence of MCC 2 months later. The tumor cells in the patient's biopsy specimen were negative for MCPyV by polymerase chain reaction and immunohistochemistry (CM2B4 antibody, Santa Cruz, CA). The exact mechanism for complete spontaneous regression in MCC is unknown. To our knowledge, only 2 previous studies evaluated the presence of MCPyV by polymerase chain reaction in MCC with spontaneous regression. Whether the presence or absence of MCPyV correlates with spontaneous regression warrants further investigation.

Prevalence of Chlamydia infection detected by immunohistochemistry in patients with anorectal ulcer and granulation tissue.

Anorectal ulcer with granulation tissue is typically associated with left-sided inflammatory bowel disease or infection. Due to emerging cases of Chlamydia proctitis, we aim to investigate the prevalence of Chlamydia infection using immunohistochemistry (IHC) in anorectal biopsies showing ulcer and granulation tissue. Seventy-seven patients including 60 males and 17 females with mean age of 51 years old were retrospectively identified in surgical pathology archives. Chlamydia IHC was validated with a monoclonal antibody on an index who was positive for Chlamydia by rectal swab nucleic acid amplification test (NAAT), then performed on formalin fixed and paraffin embedded (FFPE) tissue sections. Confirmative molecular test using real-time PCR was performed on DNA extractions of 14 IHC-positive and 14 IHC-negative FFPEs, 18 NAAT-positive, and 5 NAAT-negative cytology specimens. Chlamydia IHC showed strong intracytoplasmic or extracellular sphere morphology in 14 of 77 (18.2 %) FFPEs, including 11 of 60 (18.3 %) males and 3 of 17 (17.6 %) females (age 11-84 years). Eight of 14 (57.1 %) Chlamydia-IHC positive patients had known history of STDs, high-risk behavior, or immunosuppressive conditions. One of 14 (7.1 %) IHC-positive FFEP and 15 of 18 (83.3 %) NAAT-positive cytology cases were confirmed by real-time PCR. Chlamydia inclusions were detected in all 4 randomly selected NAAT and PCR-positive cytology specimens by IHC. Our data suggested that Chlamydia infection is more prevalent than we thought in patients with active proctitis and ulceration. Chlamydia IHC may be performed as a screening test in biopsies to facilitate early detection of this treatable proctitis in high-risk population.

Molecular Profiling of Syringocystadenocarcinoma Papilliferum Reveals RAS-Activating Mutations.

CONTEXT.­: Syringocystadenocarcinoma papilliferum (SCACP) is a rare adnexal carcinoma and the malignant counterpart of syringocystadenoma papilliferum (SCAP), which is commonly located on the head and neck and may arise in association with a nevus sebaceus. RAS mutations have been identified in both SCAP and nevus sebaceus. OBJECTIVE.­: To evaluate the clinicopathologic and molecular features of SCACPs, which have not been previously explored. DESIGN.­: We obtained 11 SCACPs from 6 institutions and reviewed the clinicopathologic features. We also performed molecular profiling using next-generation sequencing. RESULTS.­: The cohort comprised 6 women and 5 men with ages ranging from 29 to 96 years (mean, 73.6 years). The neoplasms occurred on the head and neck (n = 8; 73%) and extremities (n = 3; 27%). Three tumors possibly arose in a nevus sebaceus. A total of 4 cases showed at least carcinoma in situ (adenocarcinoma, n = 3; squamous cell carcinoma [SCC], n = 1), and 7 cases were invasive (SCC, n = 5; mixed adenocarcinoma + SCC, n = 2). A total of 8 of 11 cases (73%) had hot spot mutations consisting of HRAS (n = 4), KRAS (n = 1), BRAF (n = 1), TP53 (n = 4), ATM (n = 2), FLT3 (n = 1), CDKN2A (n = 1), and PTEN (n = 1). All 4 cases with HRAS mutations occurred on the head and neck, whereas the KRAS mutation occurred on the extremity. CONCLUSIONS.­: RAS-activating mutations were detected in 50% of the cases, of which most (80%) involved HRAS and occurred on the head and neck, which shows overlapping features with SCAP, supporting that a subset may arise as a result of malignant transformation and likely an early oncogenic event.

Frameshift Mutations in Leukemia-Associated Genes Correlate With Superior Outcomes in Patients Undergoing Allogeneic Stem Cell Transplant for De Novo Acute Myeloid Leukemia.

Background: Allogeneic stem cell transplant (allo-SCT) is a mainstay of treatment for acute myeloid leukemia (AML). Its success depends largely on response of donor T lymphocytes against leukemia cells, known as graft-vs-leukemia (GvL) effect. A key potential driver of GvL is immune response to mutation-derived neoantigens. Previous studies in solid tumors have demonstrated enhanced immunogenicity of frameshift (FS)-derived peptides vs. those from non-synonymous single nucleotide variants (SNVs). We therefore hypothesized that AML cases bearing FS mutations in leukemia-associated genes would be more immunogenic than those with only other types of mutations (non-FS), and thus benefit more from allo-SCT via more robust GvL. Methods: We identified AML patients who had undergone allo-SCT between 2010 and 2022 and had next-generation sequencing data available on diagnostic specimens using a 42-gene hot spot panel. We compared the impact of tumor mutations present at diagnosis on overall survival and relapse-free survival based on FS versus non-FS status. Results: Ninety-five AML allo-SCT patients were identified. We observed superior relapse-free survival (P = 0.038, hazard ratio (HR): 0.24) and borderline superior overall survival (P = 0.058, HR: 0.55) post-transplant in de novo AML patients, who had at least one FS mutation (other than NPM1) in one of the 42 assessed genes versus those with only non-FS mutations. Conclusions: Our findings suggest that FS-mutated AML cases may benefit more from allo-SCT than those with only non-FS mutations, possibly due to increased generation of immunogenic neoepitopes. If validated in an expanded study, incorporation of somatic FS mutation status in AML could improve patient selection algorithms for bone marrow transplant and thereby lead to superior outcomes.

Systemic mastocytosis with associated clonal hematological non-mast cell lineage disease: clinical significance and comparison of chomosomal abnormalities in SM and AHNMD components.

Some patients with systemic mastocytosis have concurrent hematological neoplasms, designated in the World Health Organization (WHO) classification as systemic mastocytosis with associated clonal hematological non-mast cell lineage disease (SM-AHNMD). In this study, we analyzed 29 patients with SM-AHNMD and compared them to 40 patients with pure SM. The AHNMDs were classified as chronic myelomonocytic leukemia (CMML) (n = 10), myelodysplastic syndrome (MDS) (n = 7), myeloproliferative neoplasms (n = 4), B-cell lymphoma/leukemia/plasma cell neoplasms (n = 7), and acute myeloid leukemia (n = 1). Patients with SM-AHNMD were older, more frequently had constitutional symptoms and hematological abnormalities, less often had skin lesions, and had an inferior overall survival compared with pure SM patients (48 months vs. not-reached, P < 0.001). Karyotypic abnormalities were detected in 9/28 (32%) patients with SM-AHNMD but not in pure SM patients (P < 0.001). Combined imaging/ fluorescence-in-situ hybridization performed in four SM-AHNMD cases revealed shared abnormal signals in mast cells and myeloid cells in two patients with SM-CMML and one patient with SM-MDS, but not in the mast cells of a case SM-associated with chronic lymphocytic leukemia with ATM-deletion. Quantitative mutation analysis showed higher levels of mutant KIT D816V in SM-CMML and SM-MDS than in pure SM (P < 0.001). Our data indicate that the SM-AHNMD category in the WHO classification is heterogeneous, including clonally related and unrelated forms of AHNMD. The presentation, treatment, and outcome of patients with SM-AHNMD is often dictated by the type of AHNMD.

Is it a primary or metastatic melanocytic neoplasm of the central nervous system?: A molecular based approach.

Primary melanocytic neoplasms of the central nervous system (CNS) are uncommon and must be distinguished from metastatic lesions as patients with metastatic disease carry a worse prognosis. Therefore, tools to aid in the diagnosis of a primary CNS melanocytic neoplasm would be of clinical utility. Primary CNS melanocytic neoplasms, including uveal melanomas have frequent mutations in GNAQ and GNA11, but are rare in cutaneous and mucosal melanomas. Additionally, primary uveal melanomas often exhibit monosomy 3 conferring an elevated risk of metastasis. We present a 63 year-old male with a melanocytic neoplasm in the thoracic spinal cord. Molecular studies revealed the tumor contained a GNAQ mutation and four-color fluorescent in situ hybridization (FISH) composed of chromosome enumeration probes for 3, 7, 17 and a locus specific probe for 9p21/CDKN2A yielded a normal result (i.e. two copies per cell), favoring a primary versus metastatic melanocytic neoplasm of the CNS. We report a case in which the combination of mutational analysis and FISH aided in identifying the origin of the neoplasm.

Prognostic heterogeneity and clonal dynamics within distinct subgroups of myelodysplastic syndrome and acute myeloid leukemia with TP53 disruptions.

TP53 aberrations constitute the highest risk subset of myelodysplastic neoplasms (MDS) and acute myeloid leukemia (AML). The International Consensus Classification questions the blast threshold between MDS and AML. In this study, we assess the distinction between MDS and AML for 76 patients with TP53 aberrations. We observed no significant differences between MDS and AML regarding TP53 genomics. Median overall survival (OS) was 223 days for the entire group, but prognostic discrimination within subgroups showed the most inferior OS (46 days) for AML with multihit allelic state plus TP53 variant allele frequency (VAF) > 50%. In multivariate analysis, unadjusted Cox models revealed the following variables as independent risk factors for mortality: AML (vs. MDS) (hazard ratio [HR]: 2.50, confidence interval [CI]: 1.4-4.4, p = 0.001), complex karyotype (HR: 3.00, CI: 1.4-6.1, p = 0.003), multihit status (HR: 2.30, CI 1.3-4.2, p = 0.005), and absence of hematopoietic cell transplant (HCT) (HR: 3.90, CI: 1.8-8.9, p = 0.0009). Clonal dynamic modeling showed a significant reduction in TP53 VAF with front-line hypomethylating agents. These findings clarify the impact of specific covariates on outcomes of TP53-aberrant myeloid neoplasms, irrespective of the diagnosis of MDS versus AML, and may influence HCT decisions.

Pathological MAPK activation-mediated lymphatic basement membrane disruption causes lymphangiectasia that is treatable with ravoxertinib.

Lymphangiectasia, an anomalous dilation of lymphatic vessels first described in the 17th century, is frequently associated with chylous effusion, respiratory failure, and high mortality in young patients, yet the underlying molecular pathogenesis and effective treatments remain elusive. Here, we identify an unexpected causal link between MAPK activation and defective development of the lymphatic basement membrane that drives lymphangiectasia. Human pathological tissue samples from patients diagnosed with lymphangiectasia revealed sustained MAPK activation within lymphatic endothelial cells. Endothelial KRASG12D-mediated sustained MAPK activation in newborn mice caused severe pulmonary and intercostal lymphangiectasia, accumulation of chyle in the pleural space, and complete lethality. Pathological activation of MAPK in murine vasculature inhibited the Nfatc1-dependent genetic program required for laminin interactions, collagen crosslinking, and anchoring fibril formation, driving defective development of the lymphatic basement membrane. Treatment with ravoxertinib, a pharmacological inhibitor of MAPK, reverses nuclear-to-cytoplasmic localization of Nfatc1, basement membrane development defects, lymphangiectasia, and chyle accumulation, ultimately improving survival of endothelial KRAS mutant neonatal mice. These results reveal defective lymphatic basement membrane assembly and composition as major causes of thoracic lymphangiectasia and provide a potential treatment.