5q Deletion Research Articles

Characterizing ATM Aberrations in Chronic Lymphocytic Leukemia (CLL): Prognostic Implications and Sensitivity to PARP Inhibition

Introduction ATM mutations (muts) frequently occur in CLL, yet their influence on ATM protein function and DNA break repair is poorly elucidated. Our prior study demonstrated functional hypomorphism of the rare germline variant L2307F in ATM and a higher occurrence of 11q deletions (del(11q)) with ATM germline variants in CLL (Lampson et al, JCO 2023). However, the role of other ATM germline variants in association with del(11q) or somatic muts remains unknown. Here, we analyzed the characteristics of CLL patients (pts) with ATM aberrancies and evaluated ATM function in primary CLL cells. Methods We retrospectively analyzed 278 CLL pts with ATM aberrancy, evaluated by clinical targeted sequencing from 2014 to 2019, as per our recent study (Lampson et al, JCO 2023). ATM, KAP1, and H2AX phosphorylation levels were examined after etoposide or irradiation (IR) exposure, by western blot or flow cytometry, in primary CLL cells with or without ATM aberrancy. We assessed the inhibitory effect of talazoparib, a PARP inhibitor, on CLL proliferation over 14 days using flow cytometry and Cell Trace Violet dye. Results We identified 294 ATM variants, 146 unique (germline N=74, somatic N=72). In CLL pts, 58 had dual ATM allele loss (biallelic group), including del(11q) together with only germline ATM variant(s) (N=29), only somatic mut(s) (N=18), or both (N= 11). 168 pts had ATM muts or variants without del(11q), including only germline (N=137), somatic (N=19), or both (N=12) ( ATM group), and 52 pts had del(11q) alone (11q group). Complex karyotype increased with ATM allele deficiency, particularly with 5+ abnormalities (complex karyotype 5+, biallelic vs 11q vs ATM: 22.6% vs 10.6% vs 5.8%, P=0.002). Unmutated IGHV was significantly higher in the groups with del(11q) versus ATM alone (biallelic vs 11q vs ATM: 85.1% vs 87.8% vs 53.9%, p<0.001). Interestingly, when we compared IGHV status in the ATM alone group with or without somatic ATM muts, unmutated IGHV significantly increased in those with somatic muts ( ATM-somatic vs ATM-germline alone, 81.8% vs 43.6%, p<0.001), and karyotypic complexity tended to be higher in the somatic ATM mut group (P=0.077). This might suggest that pts with ATM somatic muts have clinical characteristics and pathogenicity more similar to del(11q) as compared to those with germline muts. SF3B1 muts were more common in those with del(11q) (biallelic vs 11q vs ATM: 20.7% vs 26.9% vs 12.5%, P=0.036). For functional evaluation, we compared the five groups of primary CLL samples: del(11q) with somatic ATM mut with/without germline (11q+ ATM-S), del(11q) alone with no ATM muts (11q), somatic ATM muts without del(11q) with/without germline ( ATM-S), germline ATM muts alone ( ATM-G), and no ATM aberrancies (wild type, WT). Western blot demonstrated significantly lower phosphorylated protein levels (pATM, pKAP1, γH2AX) in response to DNA damage with etoposide in ATM aberrancy groups compared to WT (WT vs 11q, vs ATM-S, vs 11q+ ATM-S, all P<0.001 except for WT vs 11q in γH2AX, P<0.05) (Figure 1A). Flow cytometry also showed significantly lower phosphorylation levels of ATM after IR in 11q, ATM-S and 11q+ ATM-S groups compared to WT, while ATM-G group didn't show significance. To assess ATM deficient cell sensitivity to talazoparib, we compared proliferation inhibition across groups cocultured with talazoparib for 14 days. The 11q+ ATM-S group exhibited significantly greater sensitivity to talazoparib at lower dosage compared to the WT (talazoparib 0.1 μM, P<0.05; 0.5 μM, P=0.098) (Figure 1B). The monoallelic ATM deficient groups (del11q and ATM-G) tended to have lower proliferation, but not significantly. Higher concentrations of talazoparib (2.5μM) strongly inhibited proliferation of all CLLs including WT. Conclusions Pts with ATM somatic aberrancy including del11q had worse associated prognostic factors including unmutated IGHV and complex karyotype. Somatic events showed greater functional impact than some germline variants. CLL cells with ATM aberrancies had ATM pathway dysfunction, and those with biallelic ATM aberrancies were more sensitive to the PARP inhibitor, talazoparib, than WT. This work could potentially lead to additional avenues for treatment based on differential sensitivities of CLLs with ATM aberrancy. Further studies are needed to characterize specific ATM germline variants with or without somatic ATM events.

TP53 Gene allelic State in Myelodysplastic Syndromes (MDS) with Isolated 5q Deletion

Background & aim: The mutational status of TP53 gene is a significant prognostic factor in MDS, with two-thirds of patients harboring TP53 multihit alterations which are associated with poor outcomes. Approximately 20% of patients with MDS with isolated 5q deletion (MDS-del5q) exhibit TP53 aberrations, though their characteristics and prognostic impact have not been fully elucidated. In this study, we aimed to analyze the characteristics and impact of TP53 allelic state in patients with MDS-del5q. Methods: We conducted an international multicenter study in de novo MDS-del5q patients according to WHO 2017 classification. Clinical and biological characteristics, including the genetic profile were collected at diagnosis. Genetic profiling included conventional G-banding, TP53 deletion by FISH and/or SNP-arrays (for TP53 deletion and copy number neutral loss of heterozygosity). Also, the presence of mutations in myeloid-related genes was assessed by NGS or Sanger. Variant filtering and categorization were performed according to the Spanish Guidelines. Patients were classified as TP53 wild-type (TP53-wt), TP53-monoallelic, and TP53-multihit as proposed by Bernard et al. 2020. Clinical and molecular variables were evaluated for associations with acute myeloid leukemia (AML) evolution and overall survival (OS). In addition, a prognostic model to predict risk of AML evolution was developed using variables selected by the LASSO-cox method with minimum lambda. To calculate the points for each variable, a multivariate competitive risk model (Fine-Gray model) was fitted. Evolution to AML was used as the event and death as the competitive event. Statistical analysis was performed by R (4.2.2). Results: We included 682 patients with MDS-del5q (Table 1). Median follow-up was 66.8 months (CI95% 61.6-75.2). TP53 mutations were evaluated by NGS in 92.2% (n=629) and by Sanger in 7.8% (n=53) of patients. Overall, 18.7% (n=128) of MDS-del5q presented TP53 mutations. After integrating mutational data (FISH and SNP-arrays), 72.7% (n=93) of TP53-mutated patients were classified as TP53-monoallelic, whereas 27.3% (n=31) as TP53-multihit (4.5% of the whole cohort). Other recurrent mutations were SF3B1 (21%), DNMT3A (18%), TP53 (18%), TET2 (14%), ASXL1 (10%), CSNK1A1 (6%) and JAK2 (6%). Only bone marrow blasts differ between the TP53-wt and TP53-mutated groups ( p=0.031). Furthermore, only gender and VAF were different between TP53-monoallelic and TP53-multihit, being the number and type of co-occurring mutations similar among all subgroups (Table 1). Median OS of the whole cohort was 73.8 months (CI95% 63.7-83.4), and 77.3 months (CI95% 66.5-85.3) for MDS-del5q TP53-wt and 104 months (CI95% 56.9-89.4) for MDS-del5q TP53-mutated ( p=0.3). Similarly, there was no significant differences in the risk to AML evolution between TP53-wt and TP53-mutated patients (AML evolution at 60 months of 19.9% and 29.7%, respectively; p=0.307). Of note, however, the TP53-multihit group presented worse prognosis compared to TP53-monoallelic and TP53-wt patients (median OS of 55.2, 73.2 and 77.3 months, and AML evolution at 60 months of 40.4%, 25.5% and 19.9% in TP53-multihit, TP53-monoallelic and TP53-wt MDS-del5q) [Figure 1]. Interestingly, TP53-monoallelic MDS with a TP53 VAF >20% showed similar prognosis to TP53-multihit patients (median OS of 43.7 and 55.2 months, and AML evolution at 60 months of 36.7% and 40.4%, respectively). Finally, a risk score (MDS-del5q score) for AML evolution was developed with five variables, each assigned a different weight based on the regression coefficients: additional chromosomal abnormality, 2 points; TP53-multihit, 2 points; BM blast >2%, 2 points; platelets ≤ 100x10 9/L, 3 points, and SF3B1-mutation, 1 point. Three risk-groups were defined: Low, intermediate, and high-risk for ≤1, 2, or ≥3 points, respectively, with an AML evolution at 60 months of 11.5%, 23.3% and 43.7%, respectively; p <0.05) Conclusions: In contrast to previous findings in MDS without del5q, our series of MDS-del5q, the longest described to date, reveals that multihit TP53 status is uncommon in MDS-del5q. However, multihit alterations have prognostic significance in this subgroup. Finally, the MDS-del5q score allows to stratify patients into three distinct risk groups for AML evolution. These findings could hold considerable implications for guiding treatment decisions.

Neoplastic Monocyte-Derived Fibrocytes Induce Bone Marrow Fibrosis in Patients with Primary Myelofibrosis

Progressive bone marrow (BM) fibrosis is a defining feature of primary myelofibrosis (PMF). The extent of BM fibrosis has been closely associated with the clinical manifestations of advanced disease, including anemia and thrombocytopenia, rendering it an important prognostic indicator. Mesenchymal stromal cells (MSCs) have long been regarded as the exclusive inducer of BM fibrosis. However, using xenograft and transgenic animal models, we and other investigators demonstrated that neoplastic monocyte-derived fibrocytes independently contribute to the induction of BM fibrosis in PMF. Notably, our data revealed that PMF fibrocytes but not MSCs, carry pro-fibrotic signaling pathways that are absent in healthy individuals' fibrocytes or MSCs. Despite these findings, the precise contribution of BM fibrocytes and MSCs to the induction of BM fibrosis in patients with PMF remains elusive. To determine which of two subsets of collagen-producing cells are the main contributors to the induction of BM fibrosis in PMF, we analyzed BM biopsies of patients with PMF and assessed whether their distribution spatially correlated with areas of fibrosis. We analyzed formalin-fixed paraffin-embedded BM trephine biopsies obtained from previously untreated patients with PMF (n = 85) and age-matched healthy donors (n = 6). Using a 7-plexed fluorescence immunohistochemistry (IHC) in conjunction with hematoxylin/eosin, Masson's trichrome, and Gomorri's silver stains we analyzed those BM biopsy specimens. To ensure comprehensive assessment, we imaged serial whole-tissue sections from each patient at 400x resolution, meticulously aligning and carefully correcting for bone trabeculae and any artifacts. To quantitate the spatial distribution of individual cells, collagen fibers, and reticulin fibers, we developed a set of analytical algorithms based on a U-Net neural network in Visiopharm software. Our analysis revealed a 6-fold higher density of fibrocytes, defined as CD45 +/CD68 +/procollagen type I + cells, in the BM of patients with PMF ( P=0.0003) compared to normal controls. By contrast, the number of MSCs, defined as CD45 -/CD90 +/CD105 + cells, in the PMF BM was not different from the number of MSCs in normal BM. After dividing the area of each BM biopsy into a 100 μm regular hexagonal grid, the observed spatial distribution of fibrocytes strongly and positively correlated with the areas covered by reticulin and collagen fibers ( P<0.0001 in both). Furthermore, spatial analysis showed that increased densities of fibrocytes were associated with an increased number of fibrotic cellular zones in the BM tissue of PMF patients. To investigate the clonality of PMF BM fibrocytes, we conducted DNA in situ hybridization (ISH) analysis, employing two sets of locus-specific probe pairs. The DNA-ISH analysis provided valuable insights, revealing that fibrocytes obtained from PMF patients with trisomy 9 or 20q deletion carried the same corresponding cytogenetic abnormalities, indicating that PMF fibrocytes indeed originate from the neoplastic PMF clone. Because increased peripheral blood monocyte counts, commonly detected in patients with PMF, are associated with a poor prognosis, we sought to determine the potential correlation between BM fibrocyte and circulating monocyte counts. As expected, we found a significant positive correlation between BM fibrocyte counts and the percentage of circulating monocytes ( P=0.001). Moreover, we also found that fibrocyte numbers correlated with the levels of the plasma cytokines pentraxin 3, known to induce the differentiation of monocytes into fibrocytes, and matrix metalloproteinase 2, as assessed by using magnetic immunoassay, suggesting that neoplastic fibrocytes induce BM fibrosis in PMF and that fibrocyte quantification could serve as a valuable indicator of disease severity and prognosis. Taken together, our quantitative imaging analysis of BM biopsy tissue from treatment naïve PMF patients revealed that neoplastic fibrocytes, rather than MSCs, are the main contributors to the induction of BM fibrosis in PMF. Our findings underscore the importance of BM fibrocytes in the pathogenesis of PMF and warrant further investigation to identify novel therapeutic agents that will either inhibit fibrocyte differentiation or block their pro-fibrotic function.

Radiogenomics Provides Insights into Gliomas Demonstrating Single-Arm 1p or 19q Deletion.

IDH-mutant gliomas are further divided on the basis of 1p/19q status: oligodendroglioma, IDH-mutant and 1p/19q-codeleted, and astrocytoma, IDH-mutant (without codeletion). Occasionally, testing may reveal single-arm 1p or 19q deletion (unideletion), which remains within the diagnosis of astrocytoma. Molecular assessment has some limitations, however, raising the possibility that some unideleted tumors could actually be codeleted. This study assessed whether unideleted tumors had MR imaging features and survival more consistent with astrocytomas or oligodendrogliomas. One hundred twenty-one IDH-mutant grade 2-3 gliomas with 1p/19q results were identified. Two neuroradiologists assessed the T2-FLAIR mismatch sign and calcifications, as differentiators of astrocytomas and oligodendrogliomas. MR imaging features and survival were compared among the unideleted tumors, codeleted tumors, and those without 1p or 19q deletion. The cohort comprised 65 tumors without 1p or 19q deletion, 12 unideleted tumors, and 44 codeleted. The proportion of unideleted tumors demonstrating the T2-FLAIR mismatch sign (33%) was similar to that in tumors without deletion (49%; P = .39), but significantly higher than codeleted tumors (0%; P = .001). Calcifications were less frequent in unideleted tumors (0%) than in codeleted tumors (25%), but this difference did not reach statistical significance (P = .097). The median survival of patients with unideleted tumors was 7.8 years, which was similar to that in tumors without deletion (8.5 years; P = .72) but significantly shorter than that in codeleted tumors (not reaching median survival after 12 years; P = .013). IDH-mutant gliomas with single-arm 1p or 19q deletion have MR imaging appearance and survival that are similar to those of astrocytomas without 1p or 19q deletion and significantly different from those of 1p/19q-codeleted oligodendrogliomas.

Clinicopathological profile of myelodysplastic syndrome (MDS) with monosomy 7 and deletion 7q: An institute experience

Objective Assessment of clinicopathological and bone marrow parameters in Myelodysplastic Syndrome (MDS) with monosomy 7 and deletion (del) 7q and their prognostic stratification. Material and Methods Retrospective observational study of MDS patients with monosomy 7 and deletion (del) 7q was conducted from January 2013 to August 2021. Demographic, clinical, and hematological variables were acquired apart from cytogenetic analysis and karyotyping. Prognostic International Prostate Symptom Score (IPSS) risk stratification was performed. Results 110 patients of MDS underwent cytogenetics study, 8 patients had monosomy 7, and 17 patients had del 7q. The median age group for both subsets was 51–54 years. Both groups showed male predominance. In monosomy 7 MDS, severe anemia was more profound (87%) in comparison to del 7q (53%). Absolute neutrophil count (ANC) of <800/cubic mm was found equally in both groups. 88% of both the subsets had platelet count <50 thousand/liter with higher Lactate Dehydrogenase (LDH) in the del 7q group (81.25%). About 50% of MDS cases with monosomy 7 and 37.5% of del 7q cases had excess blasts of > 5%. Based on the Revised International Prognostic Scoring System (IPSS-R), 75% of patients in both subsets had a high and very high-risk category. Progression to Acute myeloid leukemia (AML) was more common in monosomy 7 than in del 7q (23% vs 24 %). Conclusion Early age of presentation with predominance in men was noted in both the groups. The IPSS-R score was more valid in determining the risk category for predicting the course of these patients rather than considering cytogenetic type alone. However, more cases need to be analyzed to validate our findings.

CCDC103 as a Prognostic Biomarker Correlated with Tumor Progression and Immune Infiltration in Glioma.

The Coiled-coil domain-containing proteins (CCDCs) are expressed in many cancers, but the role of Coiled-coil domain-containing protein 103 (CCDC103) in cancers remains unclear. Further investigations are necessary to ascertain its diagnostic significance and understand its biological function in cancers. This study aims to elucidate the biological functionalities of CCDC103 in glioma and evaluate the correlation between CCDC103 expression with glioma progression. Clinical data on glioma patients were acquired from The Cancer Genome Atlas (TCGA), the Chinese Glioma Genome Atlas (CGGA), and the Gene Expression Omnibus (GEO). The evaluation encompassed the examination of correlations between CCDC103 expression, pathological characteristics, and clinical outcomes. Furthermore, the analysis included the assessment of the correlations between CCDC103 expression and immune cell infiltration as well as glioma progression. Gliomas have higher levels of CCDC103 expression than the para-carcinoma tissues. Poorer prognosis, unfavorable histological characteristics, the absence of IDH gene mutations, and the absence of chromosome 1p and 19q deletions were all associated with higher expression of CCDC103 in gliomas. In addition to patient age, tumor grade, the absence of IDH mutations, and the absence of chromosome 1p and 19q deletions, univariate and multivariate Cox analyses showed that CCDC103 expression was independently prognostic of overall survival, disease-free survival, and progression-free survival in patients with glioma. Furthermore, tumor infiltration of B cells, neutrophils, macrophages, and dendritic cells were all linked with elevated expression of CCDC103. High CCDC103 expression was linked to immune response-related signaling pathways and cell proliferation, according to gene set enrichment analysis (GSEA). Notably, the knockdown of CCDC103 in glioma cell lines resulted in a significant reduction in cell proliferation and migration. The correlation between CCDC103 expression and both glioma progression and immune cell infiltration implies that CCDC103 expression holds promise as a valuable prognostic biomarker for glioma.

Primary Plasma Cell Leukaemia with Lambda Light Chain secretion and Chromosomal abnormalities presenting as hyperleukocytosis - A case report.

Primary plasma cell leukaemia (pPCL) is a rare and lethal form of plasma cell dyscrasia that can occur either de novo (primary) or as a leukemic transformation of refractory or relapsed multiple myeloma. We report a case of pPCL with lambda light chain disease and multiple chromosomal abnormalities in a 75-year-old female who presented with generalized weakness. Her peripheral blood smear showed hyperleukocytosis with 78% circulating plasma cells. Serum protein electrophoresis (SPE) does not reveal the M band and serum immunofixation (SI) light chain analysis in serum showed an elevated lambda light chain. Skeletal radiography and PET-CT (positron emission tomography—computerized tomography) did not reveal any lytic lesions. Immunophenotypic studies reveal a CD38-positive population with negativity for T cells, B cells, myeloid markers, and a negative CD45. Fluorescent in situ hybridization (FISH) analysis results disclosed the deletion of 13q14.3 and t(11; 14). This case highlighted the usefulness of the peripheral smear findings and high fluorescent lymphocyte activity, which directed the pathologist to initiate workup for pPCL, and comprehensive laboratory biochemical and radiological evaluation further helps to confirm and differentiate pPCL from multiple myeloma. Bangladesh Journal of Medical Science Vol. 22 No. 04 October’23 Page : 932-936

The transcriptomic landscape of normal and ineffective erythropoiesis at single-cell resolution.

The anemias of myelodysplastic syndrome (MDS) and Diamond Blackfan anemia (DBA) are generally macrocytic and always reflect ineffective erythropoiesis yet result from diverse genetic mutations. To delineate shared mechanisms that lead to cell death, we studied the fate of single erythroid marrow cells from individuals with DBA or MDS-5q. We defined an unhealthy (vs healthy) differentiation trajectory using transcriptional pseudotime and cell surface proteins. The pseudotime trajectories diverge immediately after cells upregulate transferrin receptor (CD71), import iron, and initiate heme synthesis, although cell death occurs much later. Cells destined to die express high levels of heme-responsive genes, including ribosomal protein and globin genes, whereas surviving cells downregulate heme synthesis and upregulate DNA damage response, hypoxia, and HIF1 pathways. Surprisingly, 24%± 12% of cells from control subjects follow the unhealthy trajectory, implying that heme might serve as a rheostat directing cells to live or die. When heme synthesis was inhibited with succinylacetone, more DBA cells followed the healthy trajectory and survived. We also noted high numbers of messages with retained introns that increased as erythroid cells matured, confirmed the rapid cycling of colony forming unit-erythroid, and demonstrated that cell cycle timing is an invariant property of differentiation stage. Including unspliced RNA in pseudotime determinations allowed us to reliably align independent data sets and accurately query stage-specific transcriptomic changes. MDS-5q (unlike DBA) results from somatic mutation, so many normal (unmutated) erythroid cells persist. By independently tracking erythroid differentiation of cells with and without chromosome 5q deletions, we gained insight into why 5q+ cells cannot expand to prevent anemia.

Genetic Aspects of Conjunctival Melanoma: A Review.

Conjunctival melanoma (CM) is a rare but aggressive cancer. Over the past decade, molecular studies using rapidly advancing technologies have increasingly improved our understanding of CM genetics. CMs are mainly characterized by dysregulated MAPK and PI3K/AKT/mTOR pathways, driven by commonly mutated (BRAF, NRAS, NF1) or less commonly mutated (KIT, PTEN) genes. Another group of genes frequently mutated in CMs include TERT and ATRX, with known roles in telomere maintenance and chromatin remodeling/epigenetic regulation. Uveal melanoma-related genes (BAP1, SF3B1, GNAQ/11) can also be mutated in CMs, albeit infrequently. Additional CM-related mutated genes have increasingly been identified using more comprehensive genetic analyses, awaiting further confirmation in additional/larger studies. As a tumor arising in a partly sun-exposed mucosal tissue, CM exhibits a distinct genomic profile, including the frequent presence of an ultraviolet (UV) signature (and high mutational load) and also the common occurrence of large structural variations (distributed across the genome) in addition to specific gene mutations. The knowledge gained from CM genetic studies to date has led to new therapeutic avenues, including the use of targeted and/or immuno-therapies with promising outcomes in several cases. Accordingly, the implementation of tumor genetic testing into the routine clinical care of CM patients holds promise to further improve and personalize their treatments. Likewise, a growing knowledge of poor prognosis-associated genetic changes in CMs (NRAS, TERT, and uveal melanoma signature mutations and chromosome 10q deletions) may ultimately guide future strategies for prognostic testing to further improve clinical outcomes (by tailoring surveillance and considering prophylactic treatments in patients with high-risk primary tumors).

Acute myeloid leukemia with LRRFIP1::FGFR1 rearrangement and a complex karyotype

We report a case of a 20-year-old man who presented with splenomegaly, hyperleukocytosis, anemia, and thrombocytopenia. A diagnosis of acute myeloid leukemia (AML) with LRRFIP1::FGFR1 rearrangement with complex karyotype was determined. Chromosome analysis showed a male karyotype: 46,XY,i(1)(q10),t(2;8)(q37;p11.2),der(5)t(1;5) (p22;q13)[17]46,XY[3]. Fluorescence in situ hybridization (FISH) analysis using the Cytocell FGFR1 break apart/amplification probe detected FGFR1 rearrangement with t(2:8) in 126/200 cells analyzed. Other FISH probes including 1p36/ 1q25 probes, del(5q) deletion probe, TLX3 break apart probe, and PDGFRB break apart probe were also utilized to confirm the other karyotypic abnormalities. Next-generation sequencing (NGS) SureSelectXT Custom DNA Target Somatic Detection detected RUNX1 gene mutation. NGS Archer FusionPlex (RNA) confirmed the LRRFIP1::FGFR1 rearrangement. This is the second reported case of AML with LRRFIP1::FGFR1 rearrangement and the first with a complex karyotype.

Comprehensive molecular characterization of adenoid cystic carcinoma reveals tumor suppressors as novel drivers and prognostic biomarkers.

Adenoid cystic carcinoma (ACC) is a MYB-driven head and neck malignancy with high rates of local recurrence and distant metastasis and poor long-term survival. New effective targeted therapies and clinically useful biomarkers for patient stratification are needed to improve ACC patient survival. Here, we present an integrated copy number and transcriptomic analysis of ACC to identify novel driver genes and prognostic biomarkers. A total of 598 ACCs were studied. Clinical follow-up was available from 366 patients, the largest cohort analyzed to date. Copy number losses of 1p36 (70/492; 14%) and of the tumor suppressor gene PARK2 (6q26) (85/343; 25%) were prognostic biomarkers; patients with concurrent losses (n = 20) had significantly shorter overall survival (OS) than those with one or no deletions (p < 0.0001). Deletion of 1p36 independently predicted short OS in multivariate analysis (p = 0.02). Two pro-apoptotic genes, TP73 and KIF1B, were identified as putative 1p36 tumor suppressor genes whose reduced expression was associated with poor survival and increased resistance to apoptosis. PARK2 expression was markedly reduced in tumors with 6q deletions, and PARK2 knockdown increased spherogenesis and decreased apoptosis, indicating that PARK2 is a tumor suppressor in ACC. Moreover, analysis of the global gene expression pattern in 30 ACCs revealed a transcriptomic signature associated with short OS, multiple copy number alterations including 1p36 deletions, and reduced expression of TP73. Taken together, the results indicate that TP73 and PARK2 are novel putative tumor suppressor genes and potential prognostic biomarkers in ACC. Our studies provide new important insights into the pathogenesis of ACC. The results have important implications for biomarker-driven stratification of patients in clinical trials. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Evaluation of chromosome 1p/19q deletion by Fluorescence in Situ Hybridization (FISH) as prognostic factors in malignant glioma patients on treatment with alkylating chemotherapy

BackgroundEither deletion or co-deletion of chromosomal arms 1p or 19q is a characteristic and early genetic event in oligodendroglial tumors that is associated with a better prognosis and enhanced response to therapy. Information of 1p/19q status is now regarded as the standard of care when managing oligodendroglial tumors for therapeutic options in anticipation of the increased survival and progression-free survival times associated with it. Keeping this in view, we first time attempted to establish the FISH based detection of 1p/19q deletion in glioma tissue samples to evaluate its role and involvement in the disease. MethodOverall 39 glioma cases of different histologies were evaluated by fluorescence in situ hybridization (FISH) technique using specific FISH probes with Olympus BX43 fluorescent microscope to detect chromosomes 1p and 19q or co-deletions therein. ResultsOf the 39 glioma samples, overall 27 (69.2%) were found to have deletion either in 1p, 19q or both. Deletions were observed in 23.0%, 7.6% and 38.4% in 1p, 19q and 1p/19q co-deletions respectively. Overall oligidendrioglioma presented with 53.8% (21 of 39) deletions, astrocytoma group showed 12.8% and GBM accounted for 2.5% deletions. Overall survival and disease free survival was seen significantly better in oligidendrioglioma and astrocytoma with deleted tumors as compared to non-deleted ones (p<0.05). ConclusionAllelic losses on 1p and 19q, either discretely or shared, were more frequent in classic oligodendrogliomas than in either astrocytoma or Glioblastoma with better survival and response to therapy.

Molecular characteristics in Chinese with chronic lymphocytic leukemia by next-generation sequencing: A single-center retrospective analysis.

Although the prevalence of Asian chronic lymphocytic leukemia (CLL) patients is not as high as that of Caucasians, there are more atypical CLLs in Asia whose genetic characteristics and their clinical significance are distinct and remain unclear. A retrospective analysis of 85 CLL samples in our center was conducted from 2019 to 2022. We used next-generation sequencing with a 172 gene panel to explore the multi-gene mutational data and the mutational status of immunoglobulin heavy variable (IGHV) gene. MYD88 (20.0%) was the most frequently mutated gene, much higher than in Europe, followed in order by TP53 (18.8%), NOTCH1 (14.1%), IGLL5 (11.8%), and DNMT3A (8.2%). In addition, the incidence of ATM and SF3B1 mutations was relatively lower in our centre compared to Europe. Mutated (M)-IGHV patients were more likely to have a cooccurrence of MYD88 mutation, while complex karyotype and DNMT3A mutation were more common in the unmutated (U)-IGHV group. MYD88 mutated CLL was characterized by prevalence in young males in high-risk staging, with isolated 13q deletion and concomitant mutation of IGLL5. CLL patients with MYD88 and TP53 mutation showed an unfavorable prognosis. These results would be valuable in helping to understand the characteristics and significance of cytogenetic genetics in Chinese patients with CLL.

Lenalidomide treatment of Japanese patients with myelodysplastic syndromes with 5q deletion: a post-marketing surveillance study

Lenalidomide was approved in Japan for the treatment of patients with myelodysplastic syndromes associated with a 5q deletion (del 5q-MDS) in August 2010. A post-marketing surveillance (PMS) study enrolled 173 patients with del 5q-MDS who started lenalidomide treatment between August 2010 and September 2011 (mean ± standard deviation [SD] age 72.4 ± 9.0 years) and observed for up to 6 cycles or 6 months. Adverse drug reactions (ADRs) and serious ADRs were reported in 78.0% and 50.9% of patients. The most commonly observed ADRs were thrombocytopenia or platelet count decreased (46.2%), neutropenia or neutrophil count decreased (42.2%), and rash (23.1%). Of 114 patients who were red blood cell transfusion-dependent at baseline, 39 (34.2%) achieved transfusion independence during lenalidomide treatment. Of 173 patients, 19 (11.0%) had confirmed acute myeloid leukemia (AML) progression during the study. Moreover, long-term follow-up (3 years) was available for 68 of the 173 patients, of whom 12 (17.6%) progressed to AML during the additional period. This PMS study investigated the safety and effectiveness of lenalidomide in patients with del 5q-MDS. No new safety concerns were noted in routine clinical use in Japan and no evidence was found for an increased risk of AML progression following lenalidomide treatment.

APC-Related Phenotypes and Intellectual Disability in 5q Interstitial Deletions: A New Case and Review of the Literature

The 5q deletion syndrome is a relatively rare condition caused by the monoallelic interstitial deletion of the long arm of chromosome 5. Patients described in literature usually present variable dysmorphic features, behavioral disturbance, and intellectual disability (ID); moreover, the involvement of the APC gene (5q22.2) in the deletion predisposes them to tumoral syndromes (Familial Adenomatous Polyposis and Gardner syndrome). Although the development of gastrointestinal tract malignancies has been extensively described, the genetic causes underlying neurologic manifestations have never been investigated. In this study, we described a new patient with a 19.85 Mb interstitial deletion identified by array-CGH and compared the deletions and the phenotypes reported in other patients already described in the literature and the Decipher database. Overlapping deletions allowed us to highlight a common region in 5q22.1q23.1, identifying KCNN2 (5q22.3) as the most likely candidate gene contributing to the neurologic phenotype.

Incidence and prognostic impact of U2AF1 mutations and other gene alterations in myelodysplastic neoplasms with isolated 20q deletion.

In myelodysplastic neoplasms (MDS), the 20q deletion [del(20q)] is a recurrent chromosomal abnormality that it has a high co-occurrence with U2AF1 mutations. Nevertheless, the prognostic impact of U2AF1 in these MDS patients is uncertain and the possible clinical and/or prognostic differences between the mutation type and the mutational burden are also unknown. Our study analyzes different molecular variables in 100 MDS patients with isolated del(20q). We describe the high incidence and negative prognostic impact of U2AF1 mutations and other alterations such as in ASXL1 gene to identify prognostic markers that would benefit patients to receive earlier treatment.

Prevalence of chromosomal abnormalities detected by fluorescence in situ hybridization in patients with chronic lymphocytic leukemia at Alexandria Main University Hospital

Abstract Background Understanding the genetic basis of chronic lymphocytic leukemia (CLL) will help in risk stratification and a better therapeutic strategy. We aimed to evaluate the frequency of chromosomal abnormalities using fluorescence in situ hybridization (FISH) panel at our institution compared with other studies. Patients and methods and results A CLL FISH panel that included P53/ATM Probe Combination and D13S319/13qter/12cen Deletion Enumeration Probe was analyzed in 100 newly diagnosed patients with CLL. TP53 deletion was the most prevalent aberration, which is in contrast with other Middle Eastern countries, where deletion 13q14 was the commonest aberration among patients. Conclusion The heterogeneity of CLL clinical course is possibly explained by underlying molecular factors that affect prognosis including data from FISH probes. It is proposed that these abnormalities should be investigated at the time of diagnosis to better understand the disease outcome and prognosis.

Management of Patients with Lower-Risk Myelodysplastic Neoplasms (MDS).

Myelodysplastic neoplasms (MDS) are a heterogenous group of clonal hematologic disorders characterized by morphologic dysplasia, ineffective hematopoiesis, and cytopenia. In the past year, the classification of MDS has been updated in the 5th edition of the World Health Organization (WHO) Classification of Haematolymphoid Tumours and the International Consensus Classification (ICC) of Myeloid Neoplasms and Acute Leukemia with incorporation of morphologic, clinical, and genomic data. Furthermore, the more comprehensive International Prognostic Scoring System-Molecular (IPSS-M) allows for improved risk stratification and prognostication. These three developments allow for more tailored therapeutic decision-making in view of the expanding treatment options in MDS. For patients with lower risk MDS, treatment is aimed at improving cytopenias, usually anemia. The recent approval of luspatercept and decitabine/cedazuridine have added on to the current armamentarium of erythropoietic stimulating agents and lenalidomide (for MDS with isolated deletion 5q). Several newer agents are being evaluated in phase 3 clinical trials for this group of patients, such as imetelstat and oral azacitidine. This review provides a summary of the classification systems, the prognostic scores and clinical management of patients with lower risk MDS.

Search, Look, and See; Late Recognised Hypereosinophilic Syndrome with Deletion (4) (q12)

The hypereosinophilic syndrome (HES) is a group of rare disorders characterized by persistently high peripheral blood eosinophiles (≥ 1.5x109/L), and related signs or symptoms of organ involvement without secondary causes. Eosinophilia with recurrent genetic abnormalities (PDGFRA/B, FGFR1) comprises a minority of these patients. In this report, we aimed to point out a case with 4q12 deletion whose diagnosis and treatment were delayed for quite a while. The patient was followed for bronchial asthma for a long time and the recognition of hypereosinophilia yielded a suspicion for HES / Chronic eosinophilic leukemia (CEL). During the initial part of his diagnostic evaluation, there was an unawareness of the cryptic deletion which was a target for tyrosine kinases. The symptoms resolved and complete cytogenetic response was achieved with 100 mg imatinib continuing for 57 months.

Abstract A004: Investigating the function of ZFHX3 in hormone sensitive prostate cancer initiation and treatment resistance

Abstract Background: The zinc-finger homeobox ZFXH3 (also known as ATBF1 for AT motif-binding factor 1) is a transcription factor that is frequently downregulated or deleted in human prostate cancers. In a recent clinical trial (NCT02430480), patients with localized prostate cancer (PCa) received six months of neoadjuvant androgen deprivation therapy (ADT) with 10.8mg of the GnRH agonist goserelin every 12 weeks and 160mg of oral enzalutamide daily before a radical prostatectomy (RP). Throughout the six months, the patients had two multiparametric MRIs (mpMRI) at baseline and prior to surgery, along with targeted biopsies of the tumor extracted. DNA from laser capture microdissection (LCM) biopsy and RP tissues was extracted for whole exome sequencing with somatic mutation calling and copy number analysis. The most common alteration in all residual tumor samples shared a single-copy loss or deletion of chromosome 16q overlapping with ZFHX3. We hypothesized that ZFHX3 is necessary for the proper development of normal human prostates and drives tumor-suppressive activity. Methods: To mimic the reduced expression of ZFHX3 observed in patient samples, we first surveyed ZFHX3 expression by Western blotting and real-time qPCR and then developed shRNA knockdowns in androgen-sensitive (22Rv1 and LNCaP) and insensitive (C4-2) PCa cell lines. These cell lines were challenged by dose-response curve studies utilizing standard fetal bovine serum (FBS) growth media with the androgen receptor (AR) axis inhibitors abiraterone and enzalutamide. Continued treatment conditions included charcoal dextran-stripped fetal calf serum (CSS) to simulate androgen deprivation supplemented with either abiraterone or enzalutamide. Cell proliferation was determined at the end of the five-day dose-response curve treatment studies using the CellTiter Glo 2.0 cell viability assay. Results and Conclusions: In the three PCa cell lines with ZFHX3 knockdown, we observed increased sensitivity to AR-targeting drugs with a lower IC50 dose. In addition, ZFHX3 knockdown cell lines exhibited increased cell proliferation compared to scrambled control and parental cell lines using dose-response time course studies. These preliminary results form the scientific premise for this work and substantiate ongoing efforts to expand these assays to more cell lines and treatments. Our future efforts will provide mechanistic insights into the functions of ZFHX3 through chromatin immunoprecipitation sequencing with antibodies against AR and FOXA1 and the expansion of ZFHX3 knockdown into additional cell lines to investigate the transcriptional activity of multiple hormonal signaling pathways. We will further explore genomic interactions between the effects of ZFHX3 reduction and alterations to commonly-mutated genes in PCa, such as PTEN, TP53, and SPOP. Findings in this study will seek to determine how ZFHX3 exerts its tumor suppressor function and its role in the development and progression of prostate cancer. Citation Format: Isaiah M. King, John R. Bright, Nicholas T. Terrigino, Scott Wilkinson, Adam G. Sowalsky. Investigating the function of ZFHX3 in hormone sensitive prostate cancer initiation and treatment resistance [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr A004.