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Objective It is debatable whether BCR-ABL1 transcript type has an impact on outcome of treatment of patients with CML, and it is not widely studied whether body weight influences response to treatment. In this study, we tried to find out if any of these factors has an impact on response to treatment and outcome. Methodology We conducted a retrospective analysis of the files of 79 patients being treated in our center for CML with known BCR-ABL1 breakpoints, and patients’ management and response assessment was done based on ELN 2013 guidelines. The analysis was performed based on two main groups, obese vs. normal BMI, and then based on BCR-ABL1 transcripts: e13a2 vs. e14a2. Cumulative incidence of MMR, CCyR, and DMR were estimated using the Kaplan–Meier survival curve method, and comparisons between groups were performed by the Log-rank/Gray test methods. Results/conclusion In the patient-cohort studied, there was no statistically significant difference in molecular response between patients with CML based on body weight or transcript type although patients in the obesity group achieved higher and faster MMR with no statistical significance.

MiRNA Epitranscriptomics has placed a new layer of complexity in the cancer field. Despite the fast-growing interest in miRNA editing and shifted miRNA isoforms, a simultaneous study of both modifications in cancer is still missing. Here, we concurrently profiled multiple miRNA modifications, including A-to-I RNA editing and shifted miRNA isoforms, in >13K adult and pediatric tumor samples across 38 distinct cancer cohorts from The Cancer Genome Atlas and The Therapeutically Applicable Research to Generate Effective Treatments datasets. We investigated the differences among canonical miRNAs and the wider miRNAome in terms of expression, clustering, dysregulation, and prognostic standpoint. The combination of canonical miRNAs/miRNA isoforms boosted the quality of clustering results, outlining unique cohorts’ clinical-pathological features. We described modified miRNAs showing opposite dysregulation with respect to their canonical counterparts in cancer, potentially impacting their targetome and function. The abundance of expressed miRNA isoforms directly impacted the activation/deactivation of critical carcinogenesis pathways. Finally, we experimentally validated unique targeting for a shifted and edited miRNA isoform. Our findings outlined once more the importance of going beyond the well-established paradigm of one-mature-miRNA per miRNA arm to elucidate novel mechanisms related to cancer progression.

Simple Summary Adenosine to inosine (A-to-I) editing is a type of RNA editing where individual adenosines are enzymatically converted into inosines. A-to-I RNA editing plays an important role in cancer biology. Several studies have demonstrated that A-to-I editing of microRNAs (miRNAs) very often affect miRNA function as oncosuppressors or oncogenes, hence showing clinical relevance. Hence, A-to-I miRNA editing has been suggested as a potential diagnostic and prognostic tool in the monitoring of cancer patients. Nevertheless, the process of identifying and characterizing miRNA editing events in tumor samples still presents several challenges. In this review, we outline molecular aspects linked to miRNA A-to-I editing and retrace methods and approaches dedicated to detection of editing sites and functional characterization of edited miRNAs in cancer. Abstract Adenosine to inosine (A-to-I) editing consists of an RNA modification where single adenosines along the RNA sequence are converted into inosines. Such a biochemical transformation is catalyzed by enzymes belonging to the family of adenosine deaminases acting on RNA (ADARs) and occurs either co- or post-transcriptionally. The employment of powerful, high-throughput detection methods has recently revealed that A-to-I editing widely occurs in non-coding RNAs, including microRNAs (miRNAs). MiRNAs are a class of small regulatory non-coding RNAs (ncRNAs) acting as translation inhibitors, known to exert relevant roles in controlling cell cycle, proliferation, and cancer development. Indeed, a growing number of recent researches have evidenced the importance of miRNA editing in cancer biology by exploiting various detection and validation methods. Herein, we briefly overview early and currently available A-to-I miRNA editing detection and validation methods and discuss the significance of A-to-I miRNA editing in human cancer.

Background: It has been reported that general adiposity in adulthood and early adulthood, and greater height may increase the risk of almost all types of lympho-haematopoietic cancers while a study done in MD Anderson found that obesity and adult weight gain are independent risk factors for CML however no study evaluated the role of obesity in the disease progression while more studies investigate the impact of translocation types. Method: We conducted a retrospective analysis of the files of 37 patients being treated in our center for CML in chronic phase (CMP-CP) with known BCR-ABL1 breakpoints, Results: patients’ management and response assessment was done based on ELN 2013 guidelines. Analysis is done based on two main groups, obese vs normal BMI, and then based on BCR-ABL1 transcripts: e13a2 vs e14a2. Although the number of cases is limited, in the patient-cohort studied an e14a2 BCR-ABL1 transcript type / normal body weight was associated with an inferior outcome when compared to e13a2 transcript / obesity groups Conclusion: our finding suggest the need to enlarge the series to better evaluate a potential role of altered metabolism and/or specific transcripts in the response to TKI in CML.

Double knockout of the two miR-15/16 loci in mouse resulted in the development of acute myeloid leukemia (AML). This result suggested that, at least, a fraction of human AMLs could be due to a similar mechanism. We analyzed the role of the two miR-15/16 clusters in 93 myelodysplastic syndrome (MDS) patients divided in three subgroups: patients with MDS, patients with MDS before transforming into AML (MDS-T), and patients with AML evolving from MDS (MDS-AML). Then, we tested 139 AML cases and 14 different AML cell lines by assessing microRNA (miRNA) expression, target protein expression, genetic loss, and silencing. MDS-T and MDS-AML patients show a reduction of the expression of miR-15a/-15b/-16 compared to MDS patients. Each miRNA can significantly predict MDS and MDS-T groups. Then, 79% of primary AMLs show a reduced expression of miR-15a and/or miR-15b. The expression of miR-15a/-15b/-16 significantly stratified AML patients in two prognostic classes. Furthermore, 40% of AML cell lines showed a combined loss of the expression of miR-15a/-15b and overexpression of their direct/indirect targets. As potential mechanisms involved in the silencing of the two miR-15/16 loci, we identified a genetic loss of miR-15a and miR-15b and silencing of these two loci by methylation. We identified a potential driver oncogenic role in the loss of expression of both miR-15/16 clusters in the progression of MDS into AML and in AML pathogenesis. The stratification of AML patients, based on miR-15/16 expression, can lead to targeted and combination therapies for the treatment of this incurable disease.

Introduction The hallmark of CML is BCR-ABL1 (breakpoint cluster region gene-Abelson murine leukemia viral oncogene homolog 1) on Philadelphia chromosome, which is the result of a reciprocal translocation between the long arms of chromosomes 9 and 22 (t[9;22][q34;q11]) [1]. Chromosome 22 breakpoints influence the BCR portions preserved in the BCL-ABL1 fusion mRNA and protein and are mainly localized to one of three BCRs, namely major-BCR (M-BCR), minor BCR (m-BCR) and micro-BCR (µ-BCR). In comparison, breaks in chromosome 9 arise most frequently by alternative splicing of the two first ABL1 exons, and can also be generated in a large genetic region, upstream of exon Ib at the 5' end, or downstream of exon Ia at the 3' end. In the majority of CML cases, the breakpoint lies within the M-BCR and gives rise to e13a2 or e14a2 fusion mRNAs (previously denoted as b2a2 and b3a2) and a p210BCR-ABL fusion protein [2]. [3] Methodology We conducted a retrospective analysis of the files of 79 patients being treated in our center for CML with known BCR-ABL1 breakpoints; there were few more patients with known transcript type but excluded because either travelled immediately on diagnosis or had a failure due to confirmed compliance issues. Patients' management and response assessment was done based on ELN 2013 guidelines. The analysis is done based on two main groups, obese versus normal BMI, and then based on BCR-ABL1 transcripts: e13a2 versus e14a2. Ethical approval was obtained from Medical Research Center for Hamad Medical Corporation (MRC-01-18-337). Results Patients included 62 males (78.5%) and 17 females (21.5%) with the mean age at diagnosis 38.8±11.8 years (median, 38; range 21 to 69 years). The characteristics (demographics, anthropometric, hematological and clinico-pathological) of the patients and their association with transcript types and obesity are summarized in Table 1. Patient outcomes, cytogenetic and molecular responses The median follow-up was 30 months (range 6 to 196 months) and 38 months (range 3 to 192 months) in normal weight and obesity groups, respectively. The median follow-up was 28 months (range 3 to 196 months) and 39 months (range 10 to 192 months) in e14a2 and e13a2 patients, respectively. A total of 22 patients distributed among different groups ended up leaving the country (censored) after a variable duration of follow-up (6 - 196 months), 18 of them CML-CP, and 4 CML-AP. 3 patients died in our cohort, all of them had e14a2 transcript, one of them was in the normal weight/BMI group, two were in the obesity group. In e14a2 group, more patients were on imatinib at the time of analysis (15 (39.5%) vs 7 (17.1%) in e13a2 group, p = 0.026). The percentage of patients of had to switch TKI was similar in both groups (47.4% vs 53.7%, p = 0.576). However, less patients in e14a2 group had to switch TKI because of failure/progression (10 (55.6%) vs 17 (77.3%), p = 0.145); however, this didn't translate into a significant difference of achieving MMR at 1 year, where in e14a2 group, 10 patients achieved MMR at 1 year (31.3%), same as in e13a2 group (10 patients = 29.3%) p 0.331 (all shown in table 1). When comparing long-term outcomes, there was also no significant difference between groups based on transcript type with regards to MMR (44.7% vs 46.3% in e14a2 vs e13a2 respectively) or DMR (26.3% vs 22% respectively) as shown in figure. In the obesity group, there were 2 patients using ponatinib due to T315I mutation, compared to none in normal weight group. However, there were no significant differences in TKI used, switch of TKI, or reason for switch. Same applies for achieving MMR at 1 year, as 11 patients in the obesity group achieved MMR (28.2%) compared to 9 patients in normal weight group (33.3%), p = 0.778 (as shown in table 1). Regarding the long-term outcomes, more patients in the obesity group achieved MMR (53.2%) compared to normal weight group (34.3%), and this response was faster, but not statistically significant. This difference was less clear with regards to DMR (25.5% in the obesity group compared to 21.9% in normal weight group) as shown in figure. Conclusion In the patient-cohort studied there were no significant differences in molecular response based on transcript type or body weight/BMI. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Loss of miR-15/16 is the most common genetic lesion in chronic lymphocytic leukemia (CLL), promoting overexpression of BCL2, which factors in leukemia pathogenesis. Indeed, an inhibitor of Bcl2, venetoclcax, is highly active in the treatment of patients with CLL. However, single-agent venetoclcax fails to eradicate minimal residual disease in most patients. Accordingly, we were interested in other genes that may be regulated by miR-15/16, which may target other drivers in CLL. We found that miR-15/16 targets ROR1, which encodes an onco-embryonic surface protein expressed on the CLL cells of over 90% of patients, but not on virtually all normal postpartum tissues. CLL with high-level expression of ROR1 also have high-level expression of Bcl2, but low-to-negligible miR-15/16. Moreover, CLL cases with high-level ROR1 have deletion(s) at the chromosomal location of the genes encoding miR-15/16 (13q14) more frequently than cases with low-to-negligible ROR1, implying that deletion of miR-15/16 may promote overexpression of ROR1, in addition to BCL2. ROR1 is a receptor for Wnt5a, which can promote leukemia-cell proliferation and survival, and can be targeted by cirmtuzumab, a humanized anti-ROR1 mAb. We find that this mAb can enhance the in vitro cytotoxic activity of venetoclcax for CLL cells with high-level expression of ROR1, indicating that combining these agents, which target ROR1 and Bcl2, may have additive, if not synergistic, activity in patients with this disease.

Background Protein tyrosine phosphatase receptor gamma (PTPRG) is a ubiquitously expressed member of the protein tyrosine phosphatase family known to act as a tumor suppressor gene in many different neoplasms with mechanisms of inactivation including mutations and methylation of CpG islands in the promoter region. Although a critical role in human hematopoiesis and an oncosuppressor role in chronic myeloid leukemia (CML) have been reported, only one polyclonal antibody (named chPTPRG) has been described as capable of recognizing the native antigen of this phosphatase by flow cytometry. Protein biomarkers of CML have not yet found applications in the clinic, and in this study, we have analyzed a group of newly diagnosed CML patients before and after treatment. The aim of this work was to characterize and exploit a newly developed murine monoclonal antibody specific for the PTPRG extracellular domain (named TPγ B9-2) to better define PTPRG protein downregulation in CML patients. Methods TPγ B9-2 specifically recognizes PTPRG (both human and murine) by flow cytometry, western blotting, immunoprecipitation, and immunohistochemistry. Results Co-localization experiments performed with both anti-PTPRG antibodies identified the presence of isoforms and confirmed protein downregulation at diagnosis in the Philadelphia-positive myeloid lineage (including CD34+/CD38bright/dim cells). After effective tyrosine kinase inhibitor (TKI) treatment, its expression recovered in tandem with the return of Philadelphia-negative hematopoiesis. Of note, PTPRG mRNA levels remain unchanged in tyrosine kinase inhibitors (TKI) non-responder patients, confirming that downregulation selectively occurs in primary CML cells. Conclusions The availability of this unique antibody permits its evaluation for clinical application including the support for diagnosis and follow-up of these disorders. Evaluation of PTPRG as a potential therapeutic target is also facilitated by the availability of a specific reagent capable to specifically detect its target in various experimental conditions. Electronic supplementary material The online version of this article (doi:10.1186/s13045-017-0494-z) contains supplementary material, which is available to authorized users.

Chronic lymphocytic leukemia (CLL) is the most common human leukemia, and dysregulation of tRNA-derived short noncoding RNA (tsRNA) (tRF-1) expression is an accompanying event in the development of this disease. tsRNAs are fragments originating from the 3′ end of tRNA precursors and do not contain mature tRNA sequences. In contrast to tsRNAs, mature tRFs (tRF-3s, tRF-5s, and internal tRFs) are produced from mature tRNA sequences and are redundant fragments. We investigated tsRNA expression in CLL and determined tsRNA signatures in indolent CLL and aggressive CLL vs. normal B cells. We noticed that both ts-43 and ts-44 are derived from distinct genes of pre-tRNA His , and are down-regulated in CLL 3- to 5-fold vs. normal B cells. Thus, we investigated expression levels of tRF-5 fragments from tRNA His in CLL samples and healthy controls, and determined that such fragments are down-regulated by 5-fold in CLLs vs. normal controls. Given these results, we investigated the expression of all mature tRFs in CLLs vs. normal controls. We found a drastic dysregulation of the expression of mature tRFs in CLL. In aggressive CLL, for the top 15 up-regulated fragments, linear fold change varied from 2,053- to 622-fold. For the top 15 down-regulated fragments in CLL, linear fold change varied from 314- to 52-fold. In addition, 964 mature tRFs were up-regulated at least 2-fold in CLL, while 701 fragments were down-regulated at least 2-fold. Similar results were obtained for indolent CLL. Our results suggest that mature tRFs may have oncogenic and/or tumor suppressor function in CLL.

Introduction Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm characterized by the dysregulated production and uncontrolled proliferation of mature and maturing granulocytes with fairly normal differentiation. The hallmark of CML is BCR-ABL1 (breakpoint cluster region gene-Abelson murine leukemia viral oncogene homolog 1) on Philadelphia chromosome, which is the result of a reciprocal translocation between the long arms of chromosomes 9 and 22 (t[9;22][q34;q11]). With rare exceptions, breaks in chromosome 22 localize to one of three BCRs and determine the portions of BCR retained in the BCR-ABL1 fusion mRNA and protein. In contrast, the chromosome 9 breaks can occur over a large genetic region, 5′ of ABL1 exon Ib, 3′ of ABL1 exon Ia, or most commonly between the two alternative first ABL1 exons. In an overwhelming majority of CML patients, the break occurs in the major BCR (M-BCR), generating e13a2 or e14a2 fusion mRNAs and a p210BCR-ABL fusion protein. p230BCR-ABL, the largest of the fusion proteins, corresponds to a break in the micro BCR (μ-BCR), an e19a2 fusion mRNA, and is associated with neutrophilic predominance and possibly less aggressive disease. Molecular monitoring of BCR-ABL1 transcript levels following treatment with tyrosine kinase inhibitors (TKIs) is central to the effective clinical management of patients with CML. BCR-ABL1 transcripts measured at standardized time points is used to define responses at key milestones in treatment allowing early signs of poor adherence or resistance to treatment to be detected and allow for early, effective clinical intervention. Objective The aim of this study is to evaluate response to treatment with standard dose TKI in obese and non-obese CML patients together with BCR-ABL1 transcript type Methodology A retrospective analysis of clinicopathological variables and response to treatment was performed for 37 chronic phase CMLs to compare, obese vs normal weight, and BCR-ABL1 transcript type determined at diagnosis. Patients' management and response assessment was done based on ELN 2013 guidelines. Response to treatment was assessed using RT-qPCR analysis of blood calculated on the International Scale (IS). Various statistical methods used, all Statistical analyses were done using statistical packages SPSS 22.0 (SPSS Inc. Chicago, IL) and Epi-info (Centers for Disease Control and Prevention, Atlanta, GA) software. Results The study cohort included 26 males (70.3%) and 11 females (29.7%) with mean age at diagnosis 36.8 years. 59.5% (n=22) expressed an e14a2 transcript, and 40.5% (n=15) an e13a2 transcript, most patients were started on imatinib, then switched either due to toxicity or failure. Median follow-up was 18 months for both transcript types. WBC, platelet counts, spleen size and Sokal scores at diagnosis, both median and Inter-quartile range (IQR) were observed to be higher in e14a2 compared to e13a2 transcript group, and, lower in obese patients compared patients with normal weight. At one year, patients with e13a2 transcript had higher percentage of CCyR (or better) 60% (95% CI 36.6, 80.3%) compared to e14a2 group 46.7% (95% CI 24.8, 69.9%), however this difference was statistically insignificant (odds ratio =1.71, 95% CI 0.40, 7.29; P=0.464) Overall, there was higher and faster achievement of CCyR and MMR in patients with e13a2 transcript compared to e14a2 transcript, and in the obese vs normal-weight patients. Patients in e13a2 group and obesity group had a lower rate of treatment failure and fewer indications to switch TKI. Of note MMR was observed to be significantly higher in patients of African origin (n=10) compared to patients with Asian ethnicity (50% vs 16%; P=0.038), which could be reflect differences in disease biology. Conclusion In the patient cohort studied an e14a2 BCR-ABL1 transcript type / normal body weight was associated with an inferior outcome when compared to e13a2 transcript / obesity groups Disclosures No relevant conflicts of interest to declare.

Chronic lymphocytic leukemia (CLL) is the most common adult leukemia. It is characterized by the accumulation of CD19+/CD5+ lymphocytes and can have variable outcomes. Richter syndrome (RS) is a lethal complication in CLL patients that results in aggressive B-cell lymphomas, and there are no tests to predict its occurrence. Because alterations in microRNA expression can predict the development and progression of several cancers, we investigated whether dysregulation of specific microRNAs can predict RS in CLL patients. Thus, we compared microRNA expression levels in samples from 49 CLL patients who later developed RS with samples from 59 CLL patients who did not. We found that high expression of miR-125a-5p or low expression of miR-34a-5p can predict ∼50% of RS with a false positive rate of ∼9%. We found that CLL patients predicted to develop RS show either an increase of miR-125a-5p expression (∼20-fold) or a decrease of miR-34a-5p expression (∼21-fold) compared with CLL patients that are not predicted to develop RS. Thus, miR-125a-5p and miR-34a-5p can be valuable predictor markers of RS and have the potential to provide physicians with information that can indicate the best therapeutic strategy for CLL patients.

Small, noncoding RNAs are short untranslated RNA molecules, some of which have been associated with cancer development. Recently we showed that a class of small RNAs generated during the maturation process of tRNAs (tRNA-derived small RNAs, hereafter “tsRNAs”) is dysregulated in cancer. Specifically, we uncovered tsRNA signatures in chronic lymphocytic leukemia and lung cancer and demonstrated that the ts-4521/3676 cluster (now called “ts-101” and “ts-53,” respectively), ts-46, and ts-47 are down-regulated in these malignancies. Furthermore, we showed that tsRNAs are similar to Piwi-interacting RNAs (piRNAs) and demonstrated that ts-101 and ts-53 can associate with PiwiL2, a protein involved in the silencing of transposons. In this study, we extended our investigation on tsRNA signatures to samples collected from patients with colon, breast, or ovarian cancer and cell lines harboring specific oncogenic mutations and representing different stages of cancer progression. We detected tsRNA signatures in all patient samples and determined that tsRNA expression is altered upon oncogene activation and during cancer staging. In addition, we generated a knocked-out cell model for ts-101 and ts-46 in HEK-293 cells and found significant differences in gene-expression patterns, with activation of genes involved in cell survival and down-regulation of genes involved in apoptosis and chromatin structure. Finally, we overexpressed ts-46 and ts-47 in two lung cancer cell lines and performed a clonogenic assay to examine their role in cell proliferation. We observed a strong inhibition of colony formation in cells overexpressing these tsRNAs compared with untreated cells, confirming that tsRNAs affect cell growth and survival.

In lung cancer, the survival of patients with the same clinical stage varies widely for unknown reasons. In this two-phase study, we examined the hypothesis that germline variations influence the survival of patients with lung adenocarcinoma. First, we analyzed existing genotype and clinical data from 289 UK-resident patients with lung adenocarcinoma, identifying 86 single nucleotide polymorphisms (SNPs) that associated with survival (p

MicroRNAs-221 and -222 are highly upregulated in several solid tumors, including melanomas. We demonstrate that the proto-oncogene ETS-1, involved in the pathogenesis of cancers of different origin, is a transcriptional regulator of miR-222 by direct binding to its promoter region. Differently from 293FT cells or early stage melanomas, where unphosphorylated ETS-1 represses miR-222 transcription, in metastatic melanoma the constitutively Thr-38 phosphorylated fraction of ETS-1 induces miR-222. Despite its stepwise decreased expression along with melanoma progression, the oncogenic activity of ETS-1 relies on its RAS/RAF/ERK-dependent phosphorylation status more than on its total amount. To close the loop, we demonstrate ETS-1 as a direct target of miR-222, but not miR-221, showing the novel option of their uncoupled functions. In addition, a spatial redistribution of ETS-1 protein from the nucleus to the cytoplasm is also evidenced in advanced melanoma cells. Finally, in vivo studies confirmed the contribution of miR-222 to the increased invasive potential obtained by ETS- silencing.

Background: Chronic Myelocytic Leukemia (CML) is a clonal myeloproliferative disorder characterized by constitutive phosphorylation of Protein Tyrosine kinases (PTKS) that continuously activates multiple proliferative and antiapoptotic signaling pathways. Protein Tyrosine Phosphatases (PTPs) on the other hand is potential natural inhibitory mechanism for regulating the tyrosine kinase activities in which phosphorylation is reciprocally controlled and maintained in equilibrium state by PTKs and PTPs. As a member of PTPs family, Protein Tyrosine Phosphatase Receptor Gamma (PTPRG) was found to act as a tumor suppressor gene. This negative regulatory mechanism of PTPRG was observed to be down-regulated and disabled in CML and one of the possible mechanisms that alter the negative regulatory effect of PTPs is mutations. Several mutations have been identified in PTPs in many different leukemias such as Acute Myeloid Leukemia (AML), Juvenile MyeloMonocytic Leukemia (JMML), Myelodysplasic Syndrome (MDS), B-cell Acute Lymphoblastic Leukemia (B-ALL) and these mutations are associated with hyper-cellular proliferation, disease progression and poor outcome. However, relatively little is known about PTPRG mutations and no studies on CML are available in the literature while mutations inBCR-ABL1tyrosine kinase have been extensively characterized. Thus, understanding the role of PTPRG in antagonizing the PTK phosphorylation of BCR-ABL1 will be important to determine its role in CML development and progression. Aim: 1) To identify potential genetic alterations causing inactivation of PTPRG and 2) correlate the PTPRG findings with patients' response to the Tyrosine kinase Inhibitors. Methods: 16 CML patients, 9 from Qatar and 7 from Italy respectively, were studied for PTPRG mutations by exome sequencing. Custom primers were designed for Human PTPRG gene (5 Kb of exonic region of interest) using Ion AmpliSeq Designer. Target regions were enriched and amplified for the 16 DNA samples using Ion AmpliSeq Library kit 2.0. The amplicons were partially digested with FuPa reagent and phosphorylated prior to ligation of Ion Xpress Barcode Adapters followed by cleanup using HighPrep reagent. The adapter ligated molecules were enriched with adapter specific primers using a limited cycle PCR followed by a cleanup using HighPrep reagent. The final libraries were quantified on Qubit Flurometer using Qubit dsDNA HS Assay Kit and Agilent Bioanalyzer using Agilent High Sensitivity DNA Kit. All samples were pooled according to the concentrations on the Bioanalyzer and loaded on Ion 318TM Chip kit V2 to be sequenced on Ion Personal Genome Machine (PGM) system. European Leukemia Net (ELN) 2013 criteria were employed to assess the response/resistance of patients to treatment. Responses are defined at the hematological, cytogenetic and molecular levels. Patients response was classified into optimal and failure Results: Four mutations/variants were identified in PTPRG genes, three were missense Y92H, G574S, S561Y and 1 was frameshift Y285fs in the 16 CML patients. PTPRG Y92H was identified in 5 (1 Homozygous and 4 heterozygous alleles) patients and the 5 patient failed the Imatinib Mesylate (IM) treatment. On the other hand, The PTPRG G574S was identified in 6 (2 homozygous and 4 heterozygous alleles) patients. Out of the 6 patients, 4 were classified as failure to the treatment and 2 responded optimally. In addition, the PTPRG S561Y and Y285fs were identified on 1 and 3 patients respectively and these patients responded optimally to IM treatment. Discussion and Conclusions: This is the first prospective pilot study to investigate PTPRG gene mutations as underlying mechanism to explain treatment failure. Our preliminary data showed that the identified variant PTPRG Y92H might be associated with IM failure although it has been reported as Single Nucleotide Polymorphisms (SNPs) (rs62620047) and this could be attributed that some polymorphisms might behave like a mutation. On the other hand, PTPRG G574S variant (rs2292245) showed various clinical outcomes regardless to its allele zygosity as 67% (4/6) of patients failed the TKIs treatment. From the results of our pilot study we recommend carrying out PTPRG sequencing in a significantly larger cohort of patients to further explore and pinpoint the crucial mutations that can be correlated with CML resistance/response to treatment. Disclosures No relevant conflicts of interest to declare.

Background Apoptosis is a critical biological phenomenon, executed under the guidance of the Apoptotic Machinery (AM), which allows the physiologic elimination of terminally differentiated, senescent or diseased cells. Because of its relevance to BioMedicine, we have sought to obtain a detailed characterization of AM Omics in Homo sapiens, namely its Genomics and Evolution, Transcriptomics, Proteomics, Interactomics, Oncogenomics, and Pharmacogenomics. Methods This project exploited the methodology commonly used in Computational Biology (i.e., mining of many omics databases of the web) as well as the High Throughput biomolecular analytical techniques. Results In Homo sapiens AM is comprised of 342 protein-encoding genes (possessing either anti- or pro-apoptotic activity, or a regulatory function) and 110 MIR-encoding genes targeting them: some have a critical role within the system (core AM nodes), others perform tissue-, pathway-, or disease-specific functions (peripheral AM nodes). By overlapping the cancer type-specific AM mutation map in the fourteen most frequent cancers in western societies (breast, colon, kidney, leukaemia, liver, lung, neuroblastoma, ovary, pancreas, prostate, skin, stomach, thyroid, and uterus) to their transcriptome, proteome and interactome in the same tumour type, we have identified the most prominent AM molecular alterations within each class. The comparison of the fourteen mutated AM networks (both protein- as MIR-based) has allowed us to pinpoint the hubs with a general and critical role in tumour development and, conversely, in cell physiology: in particular, we found that some of these had already been used as targets for pharmacological anticancer therapy. For a better understanding of the relationship between AM molecular alterations and pharmacological induction of apoptosis in cancer, we examined the expression of AM genes in K562 and SH-SY5Y after anticancer treatment. Conclusion We believe that our data on the Apoptotic Machinery will lead to the identification of new cancer genes and to the discovery of new biomarkers, which could then be used to profile cancers for diagnostic purposes and to pinpoint new targets for pharmacological therapy. This approach could pave the way for future studies and applications in molecular and clinical Medicine with important perspectives both for Oncology as for Regenerative Medicine.