Chromosome Band 11q23 Research Articles

5G2 mutant mice model loss of a commonly deleted segment of chromosome 7q22 in myeloid malignancies

Monosomy 7 and del(7q) are among the most common and poorly understood genetic alterations in myelodysplastic neoplasms and acute myeloid leukemia. Chromosome band 7q22 is a minimally deleted segment in myeloid malignancies with a del(7q). However, the rarity of “second hit” mutations supports the idea that del(7q22) represents a contiguous gene syndrome. We generated mice harboring a 1.5 Mb germline deletion of chromosome band 5G2 syntenic to human 7q22 that removes Cux1 and 27 additional genes. Hematopoiesis is perturbed in 5G2+/del mice but they do not spontaneously develop hematologic disease. Whereas alkylator exposure modestly accelerated tumor development, the 5G2 deletion did not cooperate with KrasG12D, NrasG12D, or the MOL4070LTR retrovirus in leukemogenesis. 5G2+/del mice are a novel platform for interrogating the role of hemopoietic stem cell attrition/stress, cooperating mutations, genotoxins, and inflammation in myeloid malignancies characterized by monosomy 7/del(7q).

A cryptic EWSR1::DDIT3 fusion in myxoid liposarcoma: Potential pitfalls with FISH and cytogenetics.

Myxoid liposarcoma (MLS) is molecularly characterized by fusions involving the DDIT3 gene in chromosome band 12q13; the fusion partner is FUS in band 16p11 in 90-95% of the cases and EWSR1 in band 22q12 in the remaining 5-10%. Hence, molecular studies, often fluorescence in situ hybridization (FISH) for DDIT3 rearrangement, are useful for establishing a correct diagnosis. Although all MLS tumors should have DDIT3 fusions, it is important to be aware of reasons for potential false-negative results. We here present a case of MLS that was negative for FISH for DDIT3, that showed an unexpected t(11;22) at G-banding, but that displayed a characteristic EWSR1::DDIT3 fusion at RNA-sequencing. The results suggest that neoplasia-associated fusions that, due to the transcriptional orientations of the two genes involved, cannot arise through only two double-strand breaks are more likely to be associated with negative FISH-findings and unexpected karyotypes.

Identification of Novel Cell Surface Therapeutic Targets for KMT2A-Rearranged Acute Myeloid Leukemia

INTRODUCTION. Acute myeloid leukemia (AML) is a highly heterogeneous disease that represents 80% of adult leukemias and is characterized by chromosomal rearrangements such as translocations and inversions, as well as genetic mutations. These genomic alterations affect the normal maturation process of myeloid precursor cells and lead to their clonal expansion in the bone marrow and peripheral blood. Genetic aberrations define AML subgroups, which are associated with risk stratification (favorable, intermediate or adverse risk) and prognosis. One AML subgroup, KMT2A-rearranged AML (KMT2Ar), is driven by chromosomal translocations involving the lysine methyltransferase 2A gene (KMT2A) located on chromosome band 11q23.3, leading to gene fusions with over 94 distinct partners. KMT2A rearrangements are present in 5 to 10% of adult AML. Patients with t(9;11)(p21.3;q23.3) translocation generating the MLLT3::KMT2A fusion are classified in the intermediate cytogenetic risk group according to the 2022 ELN genetic risk classification, but KMT2A fusions with other translocation partners are included in the adverse risk group with current therapies. Ongoing clinical trials for KMT2Ar AML focus mainly on chemical inhibitors (DOT1L and Menin inhibitors) that target and disrupt KMT2A fusion protein complexes, with encouraging results. Precision medicine, such as antibody-based immunotherapy, also represents a promising approach for the treatment of KMT2Ar AML, but is currently underexploited due to the limited availability of targetable surface antigens. To overcome this issue, we analyzed the cell surface proteome (surfaceome) of primary human KMT2Ar AML samples to identify novel cell surface antigens with high therapeutic potential. METHODS AND RESULTS. We used a proteomics-based approach to analyze the surfaceome of 100 primary human AML specimens from 13 different AML subgroups, including 13 KMT2Ar AML samples. Briefly, we combined cell surface biotinylation with streptavidin affinity purification to enrich cell extracts with cell surface proteins, which were then analyzed by LC-MS/MS to obtain an overview of primary AML sample surfaceomes. Differential surfaceome analyses were performed to identify surface proteins selectively expressed by KMT2Ar AML specimens, and proteins with low expression in normal hematopoietic cells and normal tissues, as determined by expression analyses using publicly available datasets, were further selected. Using this approach, we identified several potential KMT2Ar AML surface antigens, including CD93, reported to be expressed by KMT2Ar AML stem cells. We then analyzed surface expression of these antigens by flow cytometry using a collection of 112 primary human AML specimens including 10 KMT2Ar AML samples. Results indicate that in the majority of KMT2Ar AML samples analyzed, more than 90% of blasts express selected antigens, suggesting that targeting these antigens with immunotherapeutic approaches could potentially lead to the elimination of most AML cells. Furthermore, flow cytometry data showed specificity of antigen expression for KMT2Ar AML samples, opening new therapeutic opportunities for this AML subgroup. Finally, single cell RNA sequencing of 8 primary KMT2Ar AML specimens was performed to further analyze the expression profile of these antigens in patient samples. AML specimens harboring the MLLT3::KMT2A fusion expressed these antigens in the vast majority of blast cells, confirming the homogeneous expression profile of identified antigens and the high interest to target these antigens for KMT2Ar AML. CONCLUSIONS Using a cell surface proteomics-based approach for the analysis of KMT2Ar AML surfaceome, we identified several antigens for this AML subgroup with high therapeutic potential. These antigens are very specific for KMT2Ar AML and are expressed homogeneously in primary human KMT2Ar AML samples, suggesting that targeting these antigens represents a promising therapeutic strategy for these patients. Several immunotherapeutic approaches to target these antigens are currently being developed and tested by our group, with the hope of identifying novel therapeutic strategies for the treatment of KMT2Ar AML.

315 (PB095) - Cisplatin resistance of TMEM16A overexpressing head and neck tumors and cancer models is driven by increased lysosomal flux and reversed by Hydroxychloroquine

Background. Squamous cell carcinoma of the head and neck (SCCHN) is the sixth deadliest tumor worldwide. Despite the advances in therapies, this disease has low cure rates. Approximately 30% of SCCHN tumors amplify chromosomal band 11q13, which harbors the ANO1 gene that encodes the calcium-activated chloride ion channel TMEM16A. TMEM16A overexpression in SCCHN is associated with poor response to cisplatin and reduced survival. The mechanisms of cisplatin resistance in TMEM16A-overexpressing tumors are unclear; finding them would suggest new combinatorial therapy targets for SCCHN.

Lysosomal inhibition sensitizes TMEM16A-expressing cancer cells to chemotherapy

Squamous cell carcinoma of the head and neck (SCCHN) is a devastating disease that continues to have low cure rates despite the recent advances in therapies. Cisplatin is the most used chemotherapy agent, and treatment failure is largely driven by resistance to this drug. Amplification of chromosomal band 11q13 occurs in ∼30% of SCCHN tumors. This region harbors the ANO1 gene that encodes the TMEM16A ion channel, which is responsible for calcium-activated chloride transport in epithelial tissues. TMEM16A overexpression is associated with cisplatin resistance, and high TMEM16A levels correlate with decreased survival. However, the mechanistic underpinning of this effect remains unknown. Lysosomal biogenesis and exocytosis have been implicated in cancer because of their roles in the clearance of damaged organelles and exocytosis of chemotherapeutic drugs and toxins. Here, we show that TMEM16A overexpression promotes lysosomal biogenesis and exocytosis, which is consistent with the expulsion of intracellular cisplatin. Using a combination of genetic and pharmacologic approaches, we find that TMEM16A promotes lysosomal flux in a manner that requires reactive oxygen species, TRPML1, and the activation of the β-catenin–melanocyte-inducing transcription factor pathway. The lysosomal inhibitor hydroxychloroquine (HCQ) synergizes with cisplatin in killing SCCHN cells in vitro. Using a murine model of SCCHN, we show that HCQ and cisplatin retard the growth of cisplatin-resistant patient-derived xenografts in vivo. We propose that TMEM16A enables cell survival by the up-regulation of lysosomal sequestration and exocytosis of the cytotoxic drugs. These results uncover a model of treatment for resistance in cancer, its reversal, and a role for TMEM16A.

The role of delta-like non-canonical Notch ligand 1 (DLK1) in cancer.

Delta-like non-canonical Notch ligand 1 (DLK1) is a cleavable single-pass transmembrane protein and a member of the Notch/Delta/Serrate family. It is paternally expressed and belongs to a group of imprinted genes located on chromosome band 14q32 in humans and 12qF1 in mice. DLK1 is expressed in many human tissues during embryonic development but in adults expression is low and is mostly restricted to (neuro)endocrine tissues and other immature stem/progenitor cells (notably hepatoblasts). However, DLK1 is expressed at a high frequency in many common malignancies (liver, breast, brain, pancreas, colon and lung). More recently, high levels of expression have been identified in endocrine-related cancers such as ovarian and adrenocortical carcinoma. There is growing evidence that DLK1 expression in cancer is associated with worse prognosis and that DLK1 may be a marker of cancer stem cells. Although the exact mechanism through which DLK1 functions is not fully understood, it is known to maintain cells in an undifferentiated phenotype and has oncogenic properties. These effects are partly exacted through interaction with the Notch signalling pathway. In this review, we have detailed the functional role of DLK1 within physiology and malignancy and posited a mechanism for how it exacts its oncogenic effects. In describing the expression of DLK1 in cancer and in healthy tissue, we have highlighted the potential for its use both as a biomarker and as a potential therapeutic target.

Retinoblastoma and mosaic 13q deletion: a case report

BackgroundPatients with 13q-syndrome are at risk of retinoblastoma when the RB1 gene, located in the chromosomal band 13q14.2, is deleted. This syndrome is frequently associated with congenital malformations and developmental delay, although these signs could be mild. Mosaic 13q-deletion patients have been previously reported in the literature; their phenotype is variable, and they may not be recognized.Case presentationRetinoblastoma diagnosed in a child with 13q-mosaicism confirmed in blood, oral mucosa, healthy retina and retinoblastoma. A second RB1 hit is present exclusively in the retinoblastoma sample (RB1 c.958C>T p.Arg320Ter). Other detected molecular events in retinoblastoma are 6p12.3pter gain and 6q25.3qter loss. Clinical examination is unremarkable except for clinodactyly of the right fifth finger.Discussion and conclusionsWe describe a case of mosaic 13q deletion syndrome affected by retinoblastoma. Molecular data obtained from the tumor analysis are similar to previous data available about this malignancy. High clinical suspicion is essential for an adequate diagnosis of mosaic cases.

Non-randomness distribution of micro-RNAs on human chromosomes

BackgroundMicro-RNA (miRNA) is one of the non-coding RNAs that exist in human genome. miRNAs play an important role in the expression of target genes. Several studies have indicated that organization of human genome is not random. In order to investigate the distribution of miRNAs on human chromosomes, the present study was carried out.ResultsUsing the data from miRBase database, we found 1913 loci coding for miRNAs (MIRs). Human chromosome bands 1p36, 1q22, 1q24, 2q13, 2q35, 3p21, 6p21, 7q22, 8p23, 8q24, 9q22, 9q34, 11q12-q13, 12q13, 14q32, 16p13, 16q24, 17p13, 17q11, 17q21, 17q25, 19p13, 19q13, 20q13, 21p11, 22q13, and Xq26-q28 were significantly bearing higher number of MIRs. The 14q32 and 19q13 with 4.11 and 3.59 MIRs per mega-base pair, respectively, were the most MIR-richest human chromosomal bands. The number of MIRs on chromosomal bands significantly decreased as a function of distance from telomere (r = − 0.949, df = 5, P = 0.001).ConclusionsOur current data suggest that MIRs are not randomly distributed on human genomes.

A Distinct Oncofetal B-Cell Transcriptional Program Is Activated in Infant Acute Myeloid Leukemia and Reveals Novel Therapeutic Strategies

Infants with acute leukemia present special challenges, due in part to the unusual features of their disease. A clonal oncofusion protein is detectable in the vast majority of infants presenting with acute leukemia, whether myeloid (AML), lymphoblastic (ALL), or mixed-phenotype (MPAL). Many recurrent oncofusion proteins seen in this age group involve the KMT2A (formerly MLL, Mixed Lineage Leukemia) gene, located on chromosomal band 11q23, which is notorious for both lineage switching and poor clinical prognosis. Other fusions involve components of the nuclear pore complex (NUP98), chromatin modifiers (KAT6A), or lineage-associated pioneer factors (such as MYB and GLIS2). This diversity of oncofusion proteins has hampered efforts to develop targeted agents, and over half of infant cases eventually succumb to their disease. Previous work showed that infant ALL is distinct from childhood or adult ALL. Here we show that infant AML is similarly distinct from AML in other age groups, and this difference yields transformative opportunities to repurpose existing targeted and immune therapies. mRNA sequencing (RNAseq) analysis of over 1500 participants in Children's Oncology Group clinical trials revealed hundreds of infant-AML-specific transcripts, many associated with early B cell development. 18 of the affected genes form a canonical B-cell regulatory circuit, including the epigenetic regulators BRD4 and POU2AF1, and their onco-fetal targets LIN28B and IGF2BP3. These four genes also host hypo-methylated super-enhancers in most infant AML cases, but in few older patients. The MYC proto-oncogene is directly implicated in a regulatory loop involving microRNA let7a-2, which is expressed in a mutually exclusive manner with its target and regulator LIN28B (Fig 1A). By contrast, the WT1 gene, abundantly expressed in over 90% of AML cases from other age groups, is transcribed at low levels in most infant AML cases. Single-sample gene set enrichment analysis revealed that most cases of infant AML bear more similarity to infant B-cell ALL or MPAL than to AML in older children, adolescents, and adults (Fig 1B). Higher-order chromatin conformation further emphasizes these differences. The MYC proto-oncogene, a critical node in the gene regulatory network (Fig. 1A) observed across infant acute leukemia cases, is routinely amplified via trisomy 8 in this age group (and in fact all childhood leukemia). The IGF2BP3 gene, another critical node (Fig. 1A), is rarely deleted in infants (deletions of 7q tend to spare 7p15, and complex karyotype cases often retain two or more copies on derivative chromosomes). The immature humoral and adaptive immune system in infants complicates treatment in this age group, but also yields metabolic vulnerabilities which can be exploited to break the nearly ubiquitous positive feedback loop described herein. These age-specific differences suggest that alternative therapeutic targets and modalities are warranted not only in infant AML, but across infant acute leukemia more generally. The relatively small numbers of infant acute leukemia cases by lineage, along with common mechanisms that appear to sustain most cases of infant acute leukemia, and frequent clinical reports of lineage switching in infants regardless of morphology at presentation, all support this contention. Our results indicate that combination therapies coupling experimental ages (such as DFMO, which targets metabolic regulation of MYC, LIN28B, and let-7a via ornithine decarboxylase, and has shown substantial efficacy in MYCN-amplified neuroblastoma) with already approved agents such as BET inhibitors (which target BRD4) and blinatumomab (targeting CD19 as a canonical B-cell antigen) may be warranted. These immune-targeting combinations represent promising and potentially lineage-agnostic approach to improving outcomes for infants with acute leukemia, a group of patients for whom novel therapeutic strategies are desperately needed, and for whom less toxic, more effective, targeted combination therapies may spare a lifetime of late effects. Disclosures Pardo: Hematologics, Inc: Employment. Kaeding:Celgene: Employment. Loken:Hematologics, Inc: Employment, Equity Ownership. Farrar:Novartis: Research Funding.

Chromosome band 11q23 deletion predicts poor prognosis in bone marrow metastatic neuroblastoma patients without MYCN amplification

BackgroundInterphase fluorescence in situ hybridization (FISH) of bone marrow cells has been confirmed to be a direct and valid method to assess the v-myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog (MYCN) amplification in patients with bone marrow metastatic neuroblastoma. MYCN amplification alone, however, is insufficient for pretreatment risk stratification. Chromosome band 11q23 deletion has recently been included in the risk stratification of neuroblastoma. In the present study, we aimed to evaluate the biological characteristics and prognostic impact of 11q23 deletion and MYCN amplification in patients with bone marrow metastatic neuroblastoma.MethodsWe analyzed the MYCN and 11q23 statuses of 101 patients with bone marrow metastatic neuroblastoma using interphase FISH of bone marrow cells. We specifically compared the biological characteristics and prognostic impact of both aberrations.ResultsMYCN amplification and 11q23 deletion were seen in 12 (11.9%) and 40 (39.6%) patients. The two markers were mutually exclusive. MYCN amplification occurred mainly in patients with high lactate dehydrogenase (LDH) and high neuron-specific enolase (NSE) levels (both P < 0.001), and MYCN-amplified patients had more events (tumor relapse, progression, or death) than MYCN-normal patients (P = 0.004). 11q23 deletion was associated only with age (P = 0.001). Patients with MYCN amplification had poorer outcomes than those with normal MYCN (3-year event-free survival [EFS] rate: 8.3 ± 8.0% vs. 43.8 ± 8.5%, P < 0.001; 3-year overall survival [OS] rate: 10.4 ± 9.7% vs. 63.5% ± 5.7%, P < 0.001). 11q23 deletion reflected a poor prognosis only for patients with normal MYCN (3-year EFS rate: 34.3 ± 9.5% vs. 53.4 ± 10.3%, P = 0.037; 3-year OS rate: 42.9 ± 10.4% vs. 75.9 ± 6.1%, P = 0.048). Those with both MYCN amplification and 11q23 deletion had the worst outcome (P < 0.001).ConclusionsChromosome band 11q23 deletion predicts poor prognosis only in bone marrow metastatic neuroblastoma patients without MYCN amplification. Combined assessment of the two markers was much superior to single-marker assessment in recognizing the patients at a high risk of disease progression.

Homogeneously staining region (hsr) on chromosome 11 is highly specific for KMT2A amplification in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS)

AML and MDS are most common myeloid neoplasms that affect mainly older patients. Overexpression of certain proto-oncogenes plays an indispensable role in tumorigenesis and overexpression can be a consequence of gene rearrangement, amplification and/or mutation. Rearrangement and amplification of KMT2A located at chromosome band 11q23 is a well-characterized genetic driver in a subset of AML/MDS cases and is associated with a poor prognosis. The presence of homogeneously staining regions (hsr) also has been correlated with amplification of specific proto-oncogenes. In this study, we correlated hsr(11)(q23) with KMT2A in a large cohort of AML/MDS (n = 54) patients. We identified 37 patients with hsr(11)(q23) in the setting of AML (n = 27) and MDS (n = 10). All patients showed a complex karyotype including 12 cases with monosomy 17. KMT2A FISH analysis was available for 35 patients which showed KMT2A amplification in all patients. Among control cases with hsr involving chromosomes other than 11q [non-11q hsr, n = 17], FISH analysis for KMT2A was available in 10 cases and none of these cases showed KMT2A amplification (p = 0.0001, Fisher's exact test, two-tailed). Mutational analysis was performed in 32 patients with hsr(11)(q23). The most common mutated gene was TP53 (n = 29), followed by DNMT3A (n = 4), NF1 (n = 4), and TET2 (n = 3). Thirty (83%) patients died over a median follow-up of 7.6 months (range, 0.4–33.4). In summary, hsr(11)(q23) in AML/MDS cases is associated with a complex karyotype, monosomy 17, KMT2A amplification, and TP53 mutation.

Abstract 5: Pre-clinical development of CDK9 inhibitor - LS007 for the treatment of acute myeloid leukemia

Abstract Acute myeloid leukemia (AML) is the most common form of acute leukaemia with an increasing incidence and dismal long-term prognosis with age. The most aggressive subtype, MLL-AML, is characterized by translocations of the mixed-lineage leukaemia gene (MLL) in chromosome band 11q23 and resistance to conventional chemotherapy with a short overall survival of only 9 months. Cyclin-dependent 9 (CDK9) activity is required for MLL-driven oncogenic transcription. Inhibition of CDK9 directly targets the transcriptional machinery required for MLL-fusion proteins (e.g. MLL-AF4, MLL-AF9 and MLL-ENL), thereby leading to apoptosis. We have developed a highly effective drug candidate - LS007 that inhibited CDK9 with a Ki = 4 nM. It suppressed proliferation and induced apoptosis in a range of AML cell lines and patient samples. LS007 was highly efficacious against several AML cell line xenografts and patient-derived xenografts, and well tolerated in mice after oral administration. In a Molm-13 xenograft model, after 21 days of orally daily dosing at 25 mg/kg and 50 mg/kg LS007 inhibited tumor growth (p &amp;lt; 0.001) with T/C = 44 % and 18 %, respectively. Similarly, in a HL-60 xenograft model, LS007 exhibited a significant tumor growth inhibition at 45 mg/kg and 60 mg/kg daily by oral administration with T/C = 40 % and 20 %, respectively. The pharmacokinetic profiles of LS007 was further assessed in the cynomolgus monkeys. After oral doses at the dose of 2, 4 and 8 mg/kg, LS007 was detected in the plasma with the maximum concentration (Cmax) from 55.9 to 279.5 ng/mL and the area under the plasma drug concentration-time graph (AUC) up to 1862.7 h·ng/mL. The mean terminal half-life (T 1/2) was around 3 h. LS007 possessed the favorable pharmacokinetic profiles with oral bioavailability F = 56% in the cynomolgus monkey (at dose 4 mg/kg), suggesting its therapeutic potential as an orally bioavailable anti-AML agent. The GLP- toxicological studies of LS007 in the cynomolgus monkey were completed, and the maximum tolerated doses were identified, which were used to estimate the safe starting dose for the Phase I clinical trials. With its excellent anti-cancer efficacy along with appreciable absorption and safety profiles, LS007 offers a very exciting prospect as a clinical development candidate. The application of Investigational New Drug (IND) has been approved allowing the commencement of clinical trials of LS007 in patients with AML. Citation Format: Wang Hui, Wang Shudong. Pre-clinical development of CDK9 inhibitor - LS007 for the treatment of acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5.

Abstract 1811: The long noncoding RNA KIAA0125 is aberrantly expressed in ameloblastomas

Abstract Long noncoding RNAs (lncRNAs) are transcribed RNA molecules that can interact with DNA or RNA, transcription factors, histones and other chromatin modifying proteins, affecting the expression level of a broad spectrum of genes. Because of their tissue-specific expression characteristics, lncRNAs hold strong promise as novel biomarkers and therapeutic targets for diseases. High-density whole-genome microarray analysis in ameloblastomas, an aggressive odontogenic tumor, and in adenomatoid odontogenic tumor (AOT), an indolent one, showed a high frequency of copy number alteration at chromosome band 14q32.33, which encompasses the lncRNA gene KIAA0125. To understand molecular mechanisms associated with their biological behavior and considering the copy-number gains observed at 14q32.33, we aimed to investigate the expression levels of the lncRNA KIAA0125 in ameloblastomas and in AOTs. The University Ethics Committee approved this study and patients signed informed consent. Thirteen frozen samples were included: five solid/multicystic ameloblastomas, four AOT, and four dental follicles. qPCR reactions were performed in triplicates and the relative changes in gene expression were obtained using the 2^-ΔΔCt formula. The reference gene HPRT1 was used for normalization and the human keratinocyte HaCat cell line was used as calibrator. Differences in the relative changes in gene expression between two groups of samples were assessed and the significance level was set at 0.05. All samples expressed more KIAA0125 than the calibrator. The ameloblastoma group showed higher expression levels of KIAA0125 when compared to dental follicles (p=0.042), while there was no difference between the expression in ameloblastomas and AOT (p&amp;gt;0.05). The expression levels of KIAA0125 in AOT were not different from that of the dental follicle. Our findings suggest that lncRNA KIAA0125 is likely involved in ameloblastoma pathobiology. LncRNAs hold strong promise as therapeutic targets, and experimental validation of this lncRNA functions may lead to tailored therapies targeting KIAA0125 in extensive and recurrent ameloblastomas. Citation Format: Silvia F. Sousa, Marina G. Diniz, Josiane A. França, Fabricio A. Vilas-Boas, Fabricio T. Souza, George A. Calin, Ricardo S. Gomez, Carolina C. Gomes. The long noncoding RNA KIAA0125 is aberrantly expressed in ameloblastomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1811.

Genomic instability in a chronic lymphocytic leukemia patient with mono-allelic deletion of the DLEU and RB1 genes

BackgroundThe most frequent cytogenetic abnormality detected in chronic lymphocytic leukemia (CLL) patients is the presence of a deletion within the chromosome band 13q14. Deletions can be heterogeneous in size, generally encompassing the DLEU1 and DLEU2 genes (minimal deleted region), but at times also including the RB1 gene. The latter, larger type of deletions are associated with worse prognosis.Genomic instability is a characteristic of most cancers and it has been observed in CLL patients mainly associated with telomere shortening.Case presentationCytogenetic and fluorescence in situ hybridization studies of a CLL patient showed a chromosomal translocation t(12;13)(q15;q14), a mono-allelic 13q14 deletion encompassing both the DLEU and RB1 genes, and genomic instability manifested as chromosomal breaks, telomeric associations, binucleated cells, nucleoplasmic bridges, and micronucleated cells.In conclusion, our CLL patient showed genomic instability in conjunction with a 13q14 deletion of approximately 2.6 megabase pair involving the DLEU and RB1 genes, as well as other genes with potential for producing genomic instability due to haploinsufficiency.

Deletion of the murine ortholog of the 8q24 gene desert has anti-cancer effects in transgenic mammary cancer models

BackgroundThe gene desert on human chromosomal band 8q24 harbors multiple genetic variants associated with common cancers, including breast cancer. The locus, including the gene desert and its flanking genes, MYC, PVT1 and FAM84B, is also frequently amplified in human breast cancer. We generated a megadeletion (MD) mouse model lacking 430-Kb of sequence orthologous to the breast cancer-associated region in the gene desert. The goals were to examine the effect of the deletion on mammary cancer development and on transcript level regulation of the candidate genes within the locus.MethodsThe MD allele was engineered using the MICER system in embryonic stem cells and bred onto 3 well-characterized transgenic models for breast cancer, namely MMTV-PyVT, MMTV-neu and C3(1)-TAg. Mammary tumor growth, latency, multiplicity and metastasis were compared between homozygous MD and wild type mice carrying the transgenes. A reciprocal mammary gland transplantation assay was conducted to distinguish mammary cell-autonomous from non-mammary cell-autonomous anti-cancer effects. Gene expression analysis was done using quantitative real-time PCR. Chromatin interactions were evaluated by 3C. Gene-specific patient outcome data were analysed using the METABRIC and TCGA data sets through the cBioPortal website.ResultsMice homozygous for the MD allele are viable, fertile, lactate sufficiently to nourish their pups, but maintain a 10% lower body weight mainly due to decreased adiposity. The deletion interferes with mammary tumorigenesis in mouse models for luminal and basal breast cancer. In the MMTV-PyVT model the mammary cancer-reducing effects of the allele are mammary cell-autonomous. We found organ-specific effects on transcript level regulation, with Myc and Fam84b being downregulated in mammary gland, prostate and mammary tumor samples. Through analysis using the METABRIC and TCGA datasets, we provide evidence that MYC and FAM84B are frequently co-amplified in breast cancer, but in contrast with MYC, FAM84B is frequently overexpressed in the luminal subtype, whereas MYC activity affect basal breast cancer outcomes.ConclusionDeletion of a breast cancer-associated non-protein coding region affects mammary cancer development in 3 transgenic mouse models. We propose Myc as a candidate susceptibility gene, regulated by the gene desert locus, and a potential role for Fam84b in modifying breast cancer development.

NOTCH1 Signaling Is Activated in CLL By Mutations of FBXW7 and Low Expression of USP28 at 11q23

NOTCH1 Signaling Is Activated in CLL By Mutations of FBXW7 and Low Expression of USP28 at 11q23

Establishment and characterization of a novel vincristine-resistant diffuse large B-cell lymphoma cell line containing the 8q24 homogeneously staining region.

Chromosome band 8q24 is the most frequently amplified locus in various types of cancers. MYC has been identified as the primary oncogene at the 8q24 locus, whereas a long noncoding gene, PVT1, which lies adjacent to MYC, has recently emerged as another potential oncogenic regulator at this position. In this study, we established and characterized a novel cell line, AMU‐ML2, from a patient with diffuse large B‐cell lymphoma (DLBCL), displaying homogeneously staining regions at the 8q24 locus. Fluorescence in situ hybridization clearly detected an elevation in MYC copy numbers corresponding to the homogenously staining region. In addition, a comparative genomic hybridization analysis using high‐resolution arrays revealed that the 8q24 amplicon size was 1.4 Mb, containing the entire MYC and PVT1 regions. We also demonstrated a loss of heterozygosity for TP53 at 17p13 in conjunction with a TP53 frameshift mutation. Notably, AMU‐ML2 cells exhibited resistance to vincristine, and cell proliferation was markedly inhibited by MYC‐shRNA‐mediated knockdown. Furthermore, genes involved in cyclin D, mTOR, and Ras signaling were downregulated following MYC knockdown, suggesting that MYC expression was closely associated with tumor cell growth. In conclusion, AMU‐ML2 cells are uniquely characterized by homogenously staining regions at the 8q24 locus, thus providing useful insights into the pathogenesis of DLBCL with 8q24 abnormalities.

An update of knowledge on cortactin as a metastatic driver and potential therapeutic target in oral squamous cell carcinoma

Cortactin is a protein encoded by the CTTN gene, localized on chromosome band 11q13. As a result of the amplification of this band, an important event in oral carcinogenesis, CTTN is also usually amplified, promoting the frequent overexpression of cortactin. Cortactin enhances cell migration in oral cancer, playing a key role in the regulation of filamentous actin and of protrusive structures (invadopodia and lamellipodia) on the cell membrane that are necessary for the acquisition of a migratory phenotype. We also analyze a series of emerging functions that cortactin may exert in oral cancer (cell proliferation, angiogenesis, regulation of exosomes, and interactions with the tumor microenvironment). We review its molecular structure, its most important interactions (with Src, Arp2/3 complex, and SH3-binding partners), the regulation of its functions, and its specific oncogenic role in oral cancer. We explore the mechanisms of its overexpression in cancer, mainly related to genetic amplification. We analyze the prognostic implications of the oncogenic activation of cortactin in potentially malignant disorders and in head and neck cancer, where it appears to be relevant in the development of lymph node metastasis. Finally, we discuss its usefulness as a therapeutic target and suggest future research lines.

Epigallocatechin-3-gallate improves cardiac hypertrophy and short-term memory deficits in a Williams-Beuren syndrome mouse model.

Williams-Beuren syndrome (WBS) is a neurodevelopmental disorder caused by a heterozygous deletion of 26–28 genes at chromosome band 7q11.23. The complete deletion (CD) mouse model mimics the most common deletion found in WBS patients and recapitulates most neurologic features of the disorder along with some cardiovascular manifestations leading to significant cardiac hypertrophy with increased cardiomyocytes’ size. Epigallocatechin-3-gallate (EGCG), the most abundant catechin found in green tea, has been associated with potential health benefits, both on cognition and cardiovascular phenotypes, through several mechanisms. We aimed to investigate the effects of green tea extracts on WBS-related phenotypes through a phase I clinical trial in mice. After feeding CD animals with green tea extracts dissolved in the drinking water, starting at three different time periods (prenatal, youth and adulthood), a set of behavioral tests and several anatomical, histological and molecular analyses were performed. Treatment resulted to be effective in the reduction of cardiac hypertrophy and was also able to ameliorate short-term memory deficits of CD mice. Taken together, these results suggest that EGCG might have a therapeutic and/or preventive role in the management of WBS.

MYC-containing amplicons in acute myeloid leukemia: genomic structures, evolution, and transcriptional consequences

Double minutes (dmin), homogeneously staining regions, and ring chromosomes are vehicles of gene amplification in cancer. The underlying mechanism leading to their formation as well as their structure and function in acute myeloid leukemia (AML) remain mysterious. We combined a range of high-resolution genomic methods to investigate the architecture and expression pattern of amplicons involving chromosome band 8q24 in 23 cases of AML (AML-amp). This revealed that different MYC-dmin architectures can coexist within the same leukemic cell population, indicating a step-wise evolution rather than a single event origin, such as through chromothripsis. This was supported also by the analysis of the chromothripsis criteria, that poorly matched the model in our samples. Furthermore, we found that dmin could evolve toward ring chromosomes stabilized by neocentromeres. Surprisingly, amplified genes (mainly PVT1) frequently participated in fusion transcripts lacking a corresponding DNA template. We also detected a significant overexpression of the circular RNA of PVT1 (circPVT1) in AML-amp cases versus AML with a normal karyotype. Our results show that 8q24 amplicons in AML are surprisingly plastic DNA structures with an unexpected association to novel fusion transcripts and circular RNAs.