A novel metric-based approach of scoring early host immune response from oro-nasopharyngeal swabs predicts COVID-19 outcome.

Unpredictable fatal outcome of COVID-19 is attributed to dysregulated inflammation. Impaired early adaptive immune response leads to late-stage inflammatory outcome. The purpose of this study was to develop biomarkers for early detection of host immune impairment at first diagnosis from leftover RNA samples, which may in turn identify high risk patients. Leftover RNA samples of COVID-19 patients at first diagnosis were stored. Following prospective follow-up, the samples were shorted and categorized into outcome groups. Impaired adaptive T cell response (severity score) and Impaired IL-10 response (undetectable IL-10 in the presence of high expression of a representative interferon response gene) were determined by RT-PCR based assay. We demonstrate that a T cell response based 'severity score' comprising rational combination of Ct values of a target genes' signature can predict high risk noncomorbid potentially critical COVID-19 patients with a sensitivity of 91% (95% CI 58.7-99.8) and specificity of 92.6% (95% CI 75.7-99) (AUC:0.88). Although inclusion of comorbid patients reduced sensitivity to 77% (95% CI 54.6-92.2), the specificity was still 94% (95% CI 79.8-99.3) (AUC:0.82). The same for 'impaired IL-10 response' were little lower to predict high risk noncomorbid patients 64.2% (95% CI 35.1-87.2) and 82% (95% CI 65.5-93.2) respectively. Inclusion of comorbid patients drastically reduce sensitivity and specificity51.6% (95% CI 33.1-69.8) and 80.5% (95% CI 64.0-91.8) respectively. As best of our knowledge this is the first demonstration of a metric-based approach showing the 'severity score' as an indicator of early adoptive immune response, could be used as predictor of severe COVID-19 outcome at the time of first diagnosis using the same leftover swab RNA. The work flow could reduce expenditure and reporting time of the prognostic test for an earliest clinical decision ensuring possibility of early rational management.

Clinical Validation of the Somatic FANCD2 Mutation (c.2022-5C>T) as a Novel Molecular Biomarker for Early Disease Progression in Chronic Myeloid Leukemia: A Case-Control Study.

Background: Chronic myeloid leukemia (CML) results from chromosomal translocation t(9;22) leading to the formation of the BCR-ABL fusion oncogene. CML has three stages: the chronic phase (CP), the accelerated phase (AP), and the blast crisis (BC). Tyrosine kinase inhibitors (TKIs) have revolutionized the treatment of CML. TKIs work well in CP-CML, and these patients have a survival rate similar to the normal population, but TKIs are less effective in advanced-phase CML. Even with current advances in treatment, BC-CML patients have an average overall survival of less than a year. Early recognition of CML patients at risk of disease progression can help in timely interventions with appropriate TKIs or other therapeutic modalities. Although some markers of disease progression like BCR-ABL kinase domain, ASXL1, and GATA2 mutations are available, no universal and exclusively specific molecular biomarkers exist to early diagnose CML patients at risk of CML progression for timely therapeutic interventions to delay or minimize blast crisis transformation in CML. A recent study found that all BC-CML patients harbored the FANCD2 (c.2022-5C>T) mutation. Therefore, the current study was designed to detect this FANCD2 mutant in AP-CML (early progression phase) and to clinically validate its potential as a novel molecular biomarker of early CML progression from CP to AP. Methods: Our study comprised 123 CP-CML (control group) and 60 AP-CML patients (experimental group) from 2 oncology centers, from January 2020 to July 2023. Mean hemoglobin level, WBC count, platelet count, treatment type, hepatomegaly, splenomegaly, and survival status of AP-CML patients were significantly different from those of CP-CML patients. However, as these clinical parameters cannot help in the early detection of patients at risk of CML progression, there was a need for a clinically validated biomarker of AP-CML. DNA was extracted from the patients' blood samples, and the FANCD2 gene was sequenced using an Illumina NextSeq500 next-generation sequencer (NGS). Results: The NGS analysis revealed a unique splice-site mutation in the FANCD2 gene (c.2022-5C>T). This mutation was detected in the majority (98.3%) of AP-CML patients but in none of the CP-CML patients or healthy control sequences from genomic databases. The mutation was confirmed by Sanger sequencing. FANCD2 is a member of the Fanconi anemia pathway genes involved in DNA repair and genomic stability, and aberrations of this gene are associated with many cancers. Conclusions: In conclusion, our study shows that the somatic FANCD2 (c.2022-5C>T) mutation is a new molecular biomarker for early CML progression. We recommend further clinical validation of this biomarker in prospective clinical trials.

Venetoclax resistance leads to broad resistance to standard-of-care anti-MM agents, but not to immunotherapies.

ABSTACT: To our knowledge, venetoclax is the first example of personalized medicine for multiple myeloma (MM), with meaningful clinical activity as a monotherapy and in combination in patients with myeloma harboring the t(11:14) translocation. However, despite the high response rates and prolonged progression-free survival, a significant proportion of patients eventually relapse. Here, we aim to study adaptive molecular responses after the acquisition of venetoclax resistance in sensitive t(11:14) MM cell models. We therefore generated single-cell venetoclax-resistant t(11:14) MM cell lines and investigated the mechanisms contributing to resistance as well as the cells' sensitivity to other treatments. Our data suggest that acquired resistance to venetoclax is characterized by reduced mitochondrial priming and changes in B-cell lymphoma-2 (BCL-2) family proteins' expression in MM cells, conferring broad resistance to standard-of-care antimyeloma drugs. However, our results show that the resistant cells are still sensitive to immunotherapeutic treatments, highlighting the need to consider appropriate sequencing of these treatments after venetoclax-based regimens.

ABL1 kinase plays an important role in spontaneous and chemotherapy-induced genomic instability in multiple myeloma.

ABSTRACT: Genomic instability contributes to cancer progression and is at least partly due to dysregulated homologous recombination (HR). Here, we show that an elevated level of ABL1 kinase overactivates the HR pathway and causes genomic instability in multiple myeloma (MM) cells. Inhibiting ABL1 with either short hairpin RNA or a pharmacological inhibitor (nilotinib) inhibits HR activity, reduces genomic instability, and slows MM cell growth. Moreover, inhibiting ABL1 reduces the HR activity and genomic instability caused by melphalan, a chemotherapeutic agent used in MM treatment, and increases melphalan's efficacy and cytotoxicity in vivo in a subcutaneous tumor model. In these tumors, nilotinib inhibits endogenous as well as melphalan-induced HR activity. These data demonstrate that inhibiting ABL1 using the clinically approved drug nilotinib reduces MM cell growth, reduces genomic instability in live cell fraction, increases the cytotoxicity of melphalan (and similar chemotherapeutic agents), and can potentially prevent or delay progression in patients with MM.

Elevated APE1 Dysregulates Homologous Recombination and Cell Cycle Driving Genomic Evolution, Tumorigenesis, and Chemoresistance in Esophageal Adenocarcinoma.

BACKGROUND & AIMS: The purpose of this study was to identify drivers of genomic evolution in esophageal adenocarcinoma (EAC) and other solid tumors. METHODS: An integrated genomics strategy was used to identify deoxyribonucleases correlating with genomic instability (as assessed from total copy number events in each patient) in 6 cancers. Apurinic/apyrimidinic nuclease 1 (APE1), identified as the top gene in functional screens, was either suppressed in cancer cell lines or overexpressed in normal esophageal cells and the impact on genome stability and growth was monitored in vitro and in vivo. The impact on DNA and chromosomal instability was monitored using multiple approaches, including investigation of micronuclei, acquisition of single nucleotide polymorphisms, whole genome sequencing, and/or multicolor fluorescence in situ hybridization. RESULTS: Expression of 4 deoxyribonucleases correlated with genomic instability in 6 human cancers. Functional screens of these genes identified APE1 as the top candidate for further evaluation. APE1 suppression in EAC, breast, lung, and prostate cancer cell lines caused cell cycle arrest; impaired growth and increased cytotoxicity of cisplatin in all cell lines and types and in a mouse model of EAC; and inhibition of homologous recombination and spontaneous and chemotherapy-induced genomic instability. APE1 overexpression in normal cells caused a massive chromosomal instability, leading to their oncogenic transformation. Evaluation of these cells by means of whole genome sequencing demonstrated the acquisition of changes throughout the genome and identified homologous recombination as the top mutational process. CONCLUSIONS: Elevated APE1 dysregulates homologous recombination and cell cycle, contributing to genomic instability, tumorigenesis, and chemoresistance, and its inhibitors have the potential to target these processes in EAC and possibly other cancers.

High-dose melphalan treatment significantly increases mutational burden at relapse in multiple myeloma.

High-dose melphalan (HDM) improves progression-free survival in multiple myeloma (MM), yet melphalan is a DNA-damaging alkylating agent; therefore, we assessed its mutational effect on surviving myeloma cells by analyzing paired MM samples collected at diagnosis and relapse in the IFM 2009 study. We performed deep whole-genome sequencing on samples from 68 patients, 43 of whom were treated with RVD (lenalidomide, bortezomib, and dexamethasone) and 25 with RVD + HDM. Although the number of mutations was similar at diagnosis in both groups (7137 vs 7230; P = .67), the HDM group had significantly more mutations at relapse (9242 vs 13 383, P = .005). No change in the frequency of copy number alterations or structural variants was observed. The newly acquired mutations were typically associated with DNA damage and double-stranded breaks and were predominantly on the transcribed strand. A machine learning model, using this unique pattern, predicted patients who would receive HDM with high sensitivity, specificity, and positive prediction value. Clonal evolution analysis showed that all patients treated with HDM had clonal selection, whereas a static progression was observed with RVD. A significantly higher percentage of mutations were subclonal in the HDM cohort. Intriguingly, patients treated with HDM who achieved complete remission (CR) had significantly more mutations at relapse yet had similar survival rates as those treated with RVD who achieved CR. This similarity could have been due to HDM relapse samples having significantly more neoantigens. Overall, our study identifies increased genomic changes associated with HDM and provides rationale to further understand clonal complexity.

A MIR17HG-derived long noncoding RNA provides an essential chromatin scaffold for protein interaction and myeloma growth.

Long noncoding RNAs (lncRNAs) can drive tumorigenesis and are susceptible to therapeutic intervention. Here, we used a large-scale CRISPR interference viability screen to interrogate cell-growth dependency to lncRNA genes in multiple myeloma (MM) and identified a prominent role for the miR-17-92 cluster host gene (MIR17HG). We show that an MIR17HG-derived lncRNA, named lnc-17-92, is the main mediator of cell-growth dependency acting in a microRNA- and DROSHA-independent manner. Lnc-17-92 provides a chromatin scaffold for the functional interaction between c-MYC and WDR82, thus promoting the expression of ACACA, which encodes the rate-limiting enzyme of de novo lipogenesis acetyl-coA carboxylase 1. Targeting MIR17HG pre-RNA with clinically applicable antisense molecules disrupts the transcriptional and functional activities of lnc-17-92, causing potent antitumor effects both in vitro and in vivo in 3 preclinical animal models, including a clinically relevant patient-derived xenograft NSG mouse model. This study establishes a novel oncogenic function of MIR17HG and provides potent inhibitors for translation to clinical trials.

RAD51 Is Implicated in DNA Damage, Chemoresistance and Immune Dysregulation in Solid Tumors.

BACKGROUND: In normal cells, homologous recombination (HR) is tightly regulated and plays an important role in the maintenance of genomic integrity and stability through precise repair of DNA damage. RAD51 is a recombinase that mediates homologous base pairing and strand exchange during DNA repair by HR. Our previous data in multiple myeloma and esophageal adenocarcinoma (EAC) show that dysregulated HR mediates genomic instability. Purpose of this study was to investigate role of HR in genomic instability, chemoresistance and immune dysregulation in solid tumors including colon and breast cancers. METHODS: The GEO dataset were used to investigate correlation of RAD51 expression with patient survival and expression of various immune markers in EAC, breast and colorectal cancers. RAD51 was inhibited in cancer cell lines using shRNAs and a small molecule inhibitor. HR activity was evaluated using a plasmid-based assay, DNA breaks assessed by evaluating expression of γ-H2AX (a marker of DNA breaks) and p-RPA32 (a marker of DNA end resection) using Western blotting. Genomic instability was monitored by investigating micronuclei (a marker of genomic instability). Impact of RAD51 inhibitor and/or a DNA-damaging agent was assessed on viability and apoptosis in EAC, breast and colon cancer cell lines in vitro and in a subcutaneous tumor model of EAC. Impact of RAD51 inhibitor on expression profile was monitored by RNA sequencing. RESULTS: Elevated RAD51 expression correlated with poor survival of EAC, breast and colon cancer patients. RAD51 knockdown in cancer cell lines inhibited DNA end resection and strand exchange activity (key steps in the initiation of HR) as well as spontaneous DNA breaks, whereas its overexpression increased DNA breaks and genomic instability. Treatment of EAC, colon and breast cancer cell lines with a small molecule inhibitor of RAD51 inhibited DNA breaking agent-induced DNA breaks and genomic instability. RAD51 inhibitor potentiated cytotoxicity of DNA breaking agent in all cancer cell types tested in vitro as well as in a subcutaneous model of EAC. Evaluation by RNA sequencing demonstrated that DNA repair and cell cycle related pathways were induced by DNA breaking agent whereas their induction either prevented or reversed by RAD51 inhibitor. In addition, immune-related pathways such as PD-1 and Interferon Signaling were also induced by DNA breaking agent whereas their induction prevented by RAD51 inhibitor. Consistent with these observations, elevated RAD51 expression also correlated with that of genes involved in inflammation and other immune surveillance. CONCLUSIONS: Elevated expression of RAD51 and associated HR activity is involved in spontaneous and DNA damaging agent-induced DNA breaks and genomic instability thus contributing to chemoresistance, immune dysregulation and poor prognosis in cancer. Therefore, inhibitors of RAD51 have great potential as therapeutic agents for EAC, colon, breast and probably other solid tumors.

A standalone approach to utilize telomere length measurement as a surveillance tool in oral leukoplakia.

Oral squamous cell carcinoma (OSCC) is often preceded by a white patch on a surface of the mouth, called oral leukoplakia (OL). As accelerated telomere length (TL) shortening in dividing epithelial cells may lead to oncogenic transformation, telomere length measurement could serve as a predictive biomarker in OL. However, due to high variability and lack of a universal reference, there has been a limited translational application. Here, we describe an approach of evaluating TL using paired peripheral blood mononuclear cells (PBMC) as an internal reference and demonstrate its translational relevance. Oral brush biopsy and paired venous blood were collected from 50 male OL patients and 44 male healthy controls (HC). Relative TL was measured by quantitative PCR. TL of each OL or healthy sample was normalized to the paired PBMC sample (TL ratio). In OL patients, the mean TL ratio was significantly smaller not only in the patch but also in distal normal oral tissue, relative to healthy controls without a high-risk oral habit. Dysplasia was frequently associated with a subgroup that showed a normal TL ratio at the patch but significantly smaller TL ratio at a paired normal distal site. Our data suggest that evaluation of TL attrition using a paired PBMC sample eliminates the requirement of external reference DNA, makes data universally comparable and provides a useful marker to define high-risk OL groups for follow-up programs. Larger studies will further validate the approach and its broader application in other premalignant conditions.

CLINICAL VALIDATION OF ANKRD36 MUTATIONS AS A NOVEL BIOMARKER FOR MONITORING EARLY PROGRESSION AND TIMELY CLINICAL INTERVENTIONS IN BLAST CRISIS CML.

Background: Chronic Myeloid Leukemia (CML) is initiated in the bone marrow due to the chromosomal translocation t(9;22), resulting in the fusion oncogene BCR-ABL. Tyrosine kinase inhibitors (TKIs) targeting BCR-ABL have transformed fatal CML into an almost curable disease. However, TKIs lose efficacy during disease progression, and the mechanism of CML progression remains to be fully understood. Additionally, common molecular biomarkers for CML progression are lacking. Our studies previously detected ANKRD36 (c.1183_1184 delGC and c.1187_1188 dupTT) associated exclusively with advanced phase CML. However, clinical validation of this finding was pending. Therefore, this study aimed to clinically validate mutated ANKRD36 as a novel biomarker of CML progression. Materials and Methods: The study enrolled 124 patients in all phases of CML, recruited from Mayo Hospital and Hameed Latif Hospital in Lahore, Punjab, between January 2019 and August 2021. All response criteria were adopted from the European LeukemiaNet guideline 2020. Informed consent was obtained from all study subjects. The study was approved by scientific and ethical review committees of all participating centers.Sanger sequencing was employed to detect ANKRD36 mutations in CML patients in accelerated phase (AP) (n=11) and blast crisis (BC) (n=10), with chronic-phase CML (CP-CML) patients as controls (n=103). Samples were processed using Big Dye Terminator Cycle Sequencing Ready Reaction kits and sequenced using ABI Prism 3730 Genetic Analyzer, and sequencing using forward and reverse primers for ANKRD36. Results: During our study, 17% of CML patients progressed to advanced phases AP-CML n=11 (8.9%) and BC-CML n=10 (8.1%). The chronic- and advanced-phase patients showed significant difference with respect to male-to-female ratio, hemoglobin level, WBC count, and platelet count. Sanger sequencing detected ANKRD36 mutations c. 1183 1184 delGC and c. 1187 1185 dupTT exclusively in all AP- and BC-CML patients but in none of the CP-CML patients. Nevertheless, mutations status was not associated with male-to-female ratio, hemoglobin level, WBC count, and platelet count, which makes ANKRD32 as an independent predictor of early and terminal disease progression in CML. Conclusions: The study confirms ANKRD36 as a novel genomic biomarker for early and late CML progression. Further prospective studies should be carried out in this regard. ANKRD36, although fully uncharacterized in humans, shows the highest expression in bone marrow, particularly myeloid cells. Functional integrated genomic studies are recommended to further explore the role of ANKRD36 in the biology and pathogenesis of CML.

Integrated Genomic Analysis Identifies ANKRD36 Gene as a Novel and Common Biomarker of Disease Progression in Chronic Myeloid Leukemia.

Background: Chronic myeloid leukemia (CML) is initiated in bone marrow due to chromosomal translocation t(9;22) leading to fusion oncogene BCR-ABL. Targeting BCR-ABL by tyrosine kinase inhibitors (TKIs) has changed fatal CML into an almost curable disease. Despite that, TKIs lose their effectiveness due to disease progression. Unfortunately, the mechanism of CML progression is poorly understood and common biomarkers for CML progression are unavailable. This study was conducted to find novel biomarkers of CML progression by employing whole-exome sequencing (WES). Materials and Methods: WES of accelerated phase (AP) and blast crisis (BC) CML patients was carried out, with chronic-phase CML (CP-CML) patients as control. After DNA library preparation and exome enrichment, clustering and sequencing were carried out using Illumina platforms. Statistical analysis was carried out using SAS/STAT software version 9.4, and R package was employed to find mutations shared exclusively by all AP-/BC-CML patients. Confirmation of mutations was carried out using Sanger sequencing and protein structure modeling using I-TASSER followed by mutant generation and visualization using PyMOL. Results: Three novel genes (ANKRD36, ANKRD36B and PRSS3) were mutated exclusively in all AP-/BC-CML patients. Only ANKRD36 gene mutations (c.1183_1184 delGC and c.1187_1185 dupTT) were confirmed by Sanger sequencing. Protein modeling studies showed that mutations induce structural changes in ANKRD36 protein. Conclusions: Our studies show that ANKRD36 is a potential common biomarker and drug target of early CML progression. ANKRD36 is yet uncharacterized in humans. It has the highest expression in bone marrow, specifically myeloid cells. We recommend carrying out further studies to explore the role of ANKRD36 in the biology and progression of CML.

IgM-MM is predominantly a pre-germinal center disorder and has a distinct genomic and transcriptomic signature from WM.

Immunoglobulin M (IgM) multiple myeloma (MM) is a rare disease subgroup. Its differentiation from other IgM-producing gammopathies such as Waldenström macroglobulinemia (WM) has not been well characterized but is essential for proper risk assessment and treatment. In this study, we investigated genomic and transcriptomic characteristics of IgM-MM samples using whole-genome and transcriptome sequencing to identify differentiating characteristics from non-IgM-MM and WM. Our results suggest that IgM-MM shares most of its defining structural variants and gene-expression profiling with MM, but has some key characteristics, including t(11;14) translocation, chromosome 6 and 13 deletion as well as distinct molecular and transcription-factor signatures. Furthermore, IgM-MM translocations were predominantly characterized by VHDHJH recombination-induced breakpoints, as opposed to the usual class-switching region breakpoints; coupled with its lack of class switching, these data favor a pre-germinal center origin. Finally, we found elevated expression of clinically relevant targets, including CD20 and Bruton tyrosine kinase, as well as high BCL2/BCL2L1 ratio in IgM-MM, providing potential for targeted therapeutics.

Dysregulated APOBEC3G causes DNA damage and promotes genomic instability in multiple myeloma.

Multiple myeloma (MM) is a heterogeneous disease characterized by significant genomic instability. Recently, a causal role for the AID/APOBEC deaminases in inducing somatic mutations in myeloma has been reported. We have identified APOBEC/AID as a prominent mutational signature at diagnosis with further increase at relapse in MM. In this study, we identified upregulation of several members of APOBEC3 family (A3A, A3B, A3C, and A3G) with A3G, as one of the most expressed APOBECs. We investigated the role of APOBEC3G in MM and observed that A3G expression and APOBEC deaminase activity is elevated in myeloma cell lines and patient samples. Loss-of and gain-of function studies demonstrated that APOBEC3G significantly contributes to increase in DNA damage (abasic sites and DNA breaks) in MM cells. Evaluation of the impact on genome stability, using SNP arrays and whole genome sequencing, indicated that elevated APOBEC3G contributes to ongoing acquisition of both the copy number and mutational changes in MM cells over time. Elevated APOBEC3G also contributed to increased homologous recombination activity, a mechanism that can utilize increased DNA breaks to mediate genomic rearrangements in cancer cells. These data identify APOBEC3G as a novel gene impacting genomic evolution and underlying mechanisms in MM.

Integrated genomics and comprehensive validation reveal drivers of genomic evolution in esophageal adenocarcinoma.

Esophageal adenocarcinoma (EAC) is associated with a marked genomic instability, which underlies disease progression and development of resistance to treatment. In this study, we used an integrated genomics approach to identify a genomic instability signature. Here we show that elevated expression of this signature correlates with poor survival in EAC as well as three other cancers. Knockout and overexpression screens establish the relevance of these genes to genomic instability. Indepth evaluation of three genes (TTK, TPX2 and RAD54B) confirms their role in genomic instability and tumor growth. Mutational signatures identified by whole genome sequencing and functional studies demonstrate that DNA damage and homologous recombination are common mechanisms of genomic instability induced by these genes. Our data suggest that the inhibitors of TTK and possibly other genes identified in this study have potential to inhibit/reduce growth and spontaneous as well as chemotherapy-induced genomic instability in EAC and possibly other cancers.

RAD51 Inhibitor Reverses Etoposide-Induced Genomic Toxicity and Instability in Esophageal Adenocarcinoma Cells.

AIM: In normal cells, homologous recombination (HR) is strictly regulated and precise and plays an important role in preserving genomic integrity by accurately repairing DNA damage. RAD51 is the recombinase which mediates homologous base pairing and strand exchange during DNA repair by HR. We have previously reported that HR is spontaneously elevated (or dysregulated) in esophageal adenocarcinoma (EAC) and contributes to ongoing genomic changes and instability. The purpose of this study was to evaluate the impact of RAD51 inhibitor on genomic toxicity caused by etoposide, a chemotherapeutic agent. METHODS: EAC cell lines (FLO-1 and OE19) were cultured in the presence of RAD51 inhibitor and/or etoposide, and impact on cell viability, apoptosis and genomic integrity/stability investigated. Genomic integrity/stability was monitored by evaluating cells for γ-H2AX (a marker for DNA breaks), phosphorylated RPA32 (a marker of DNA end resection which is a distinct step in the initiation of HR) and micronuclei (a marker of genomic instability). RESULTS: Treatment with etoposide, a chemotherapeutic agent, was associated with marked genomic toxicity (as evident from increase in DNA breaks) and genomic instability in both EAC cell lines. Consistently, the treatment was also associated with apoptotic cell death. A small molecule inhibitor of RAD51 increased cytotoxicity while reducing genomic toxicity and instability caused by etoposide, in both EAC cell lines. CONCLUSION: RAD51 inhibitors have potential to increase cytotoxicity while reducing harmful genomic impact of chemotherapy.

Genome-Wide Somatic Alterations in Multiple Myeloma Reveal a Superior Outcome Group.

PURPOSE: Multiple myeloma (MM) is accompanied by heterogeneous somatic alterations. The overall goal of this study was to describe the genomic landscape of myeloma using deep whole-genome sequencing (WGS) and develop a model that identifies patients with long survival. METHODS: We analyzed deep WGS data from 183 newly diagnosed patients with MM treated with lenalidomide, bortezomib, and dexamethasone (RVD) alone or RVD + autologous stem cell transplant (ASCT) in the IFM/DFCI 2009 study (ClinicalTrials.gov identifier: NCT01191060). We integrated genomic markers with clinical data. RESULTS: We report significant variability in mutational load and processes within MM subgroups. The timeline of observed activation of mutational processes provides the basis for 2 distinct models of acquisition of mutational changes detected at the time of diagnosis of myeloma. Virtually all MM subgroups have activated DNA repair-associated signature as a prominent late mutational process, whereas APOBEC signature targeting C>G is activated in the intermediate phase of disease progression in high-risk MM. Importantly, we identify a genomically defined MM subgroup (17% of newly diagnosed patients) with low DNA damage (low genomic scar score with chromosome 9 gain) and a superior outcome (100% overall survival at 69 months), which was validated in a large independent cohort. This subgroup allowed us to distinguish patients with low- and high-risk hyperdiploid MM and identify patients with prolongation of progression-free survival. Genomic characteristics of this subgroup included lower mutational load with significant contribution from age-related mutations as well as frequent NRAS mutation. Surprisingly, their overall survival was independent of International Staging System and minimal residual disease status. CONCLUSION: This is a comprehensive study identifying genomic markers of a good-risk group with prolonged survival. Identification of this patient subgroup will affect future therapeutic algorithms and research planning.

MUS81 Participates in the Progression of Serous Ovarian Cancer Associated With Dysfunctional DNA Repair System.

Objective: Methyl methanesulfonate ultraviolet sensitive gene clone 81 (MUS81) is a structure-specific endonuclease that plays a pivotal role in the DNA repair system of cancer cells. In this study, we aim to elucidate the potential association between the dysfunction of MUS81 and the progression of Serous Ovarian Cancer (SOC). Methods: To investigate the association between MUS81 and prognosis of SOC, immunohistochemistry technology and qPCR were used to analyze the level of MUS81 expression, and transcriptional profile analysis and protein interaction screening chip were used to explore the MUS81 related signal pathways. Random amplified polymorphic DNA (RAPD) analysis, immunofluorescence and comet assays were further performed to evaluate genomic instability and DNA damage status of transduced SOC cells. Experiments both in vitro and in vivo were conducted to verify the impact of MUS81 silencing on chemotherapeutic drug sensitivity of SOC. Results: The overexpression of MUS81 in SOC tissues was related to poor clinical outcomes. The transcriptional chip data showed that MUS81 was involved in multiple pathways associated with DNA repair. Deficiency of MUS81 intensified the genome instability of SOC cells, promoted the emergence of DSBs and restrained the formation of RAD51 foci in SOC cells with exposure to UV. Furthermore, downregulation of MUS81 enhanced the sensitivity to Camptothecin and Olaparib in SOC cell lines and xenograft model. Conclusions: MUS81 is involved in the progression of SOC and inhibition of MUS81 could augment the susceptibility to chemotherapeutic agents. MUS81 might represent a novel molecular target for SOC chemotherapy.

Deciphering the chronology of copy number alterations in Multiple Myeloma.

Multiple myeloma (MM) and its precursor condition MGUS are characterized by chromosomal aberrations. Here, we comprehensively characterize the order of occurrence of these complex genomic events underlying MM development using 500 MGUS, and MM samples. We identify hyperdiploid MM (HMM) and non-HMM as genomically distinct entities with different evolution of the copy number alterations. In HMM, gains of 9,15 or 19 are the first and clonal events observed as clonal even at MGUS stage. These events are thus early and may underlie initial transformation of normal plasma cells to MGUS cells. However, CNAs may not be adequate for progression to MM except in 15% of the patients in whom the complex subclonal deletion events are observed in MM but not MGUS. In NHMM, besides the driver translocations, clonal deletion of 13 and 1q gain are early events also observed in MGUS. We combined this information to propose a timeline for copy number alteration.