Multiple Recipients Research Articles

A public key-based encryption and signature verification model for secured image transmission in network

Image transmission is a challenging task due to the large file size and the security issues. Many existing systems on medical image transmission use symmetric key cryptography where the key is shared to multiple recipients leading to security leakage. This has been addressed by applying public key encryption schemes. However, a secured communication technique which uses compression, user authentication and encryption/decryption is more powerful than the existing systems on medical image transmission. Therefore, a new secured transmission algorithm which performs compression using singular value decomposition (SVD), user authentication using digital signatures and encryption/decryption using AES and RSA algorithms for image encryption and Hill-Cipher for data encryption which are coordinated using intelligent agents for communication and rules for decision making is proposed in this paper. The major advantages of the proposed model include fast transmission, increase in security and non-loss compression and decompression and intelligent decision making on medical image transmission.

A locus on chromosome 5 shows African ancestry–limited association with alloimmunization in sickle cell disease

AbstractRed blood cell (RBC) transfusion remains a critical therapeutic intervention in sickle cell disease (SCD); however, the apparent propensity of some patients to regularly develop RBC alloantibodies after transfusion presents a significant challenge to finding compatible blood for so-called alloimmunization responders. Predisposing genetic loci have long been thought to contribute to the responder phenomenon, but to date, no definitive loci have been identified. We undertook a genome-wide association study of alloimmunization responder status in 267 SCD multiple transfusion recipients, using genetic estimates of ancestral admixture to bolster our findings. Analyses revealed single nucleotide polymorphisms (SNPs) on chromosomes 2 and 5 approaching genome-wide significance (minimum P = 2.0 × 10−8 and 8.4 × 10−8, respectively), with local ancestry analysis demonstrating similar levels of admixture in responders and nonresponders at implicated loci. Association at chromosome 5 was nominally replicated in an independent cohort of 130 SCD transfusion recipients, with meta-analysis surpassing genome-wide significance (rs75853687, Pmeta = 6.6 × 10−9), and this extended to individuals forming multiple (>3) alloantibodies (Pmeta = 9.4 × 10−5). The associated variant is rare outside of African populations, and orthogonal genome-wide haplotype analyses, contingent on local ancestry, revealed genome-wide significant sharing of a ∼60-kb haplotype of African ancestry at the chromosome 5 locus (Bayes Factor = 4.95). This locus overlaps a putative cis-acting enhancer predicted to regulate transcription of ADRA1B and the lncRNA LINC01847, both members of larger ontologies associated with immune regulation. Our findings provide potential insights to the pathophysiology underlying the development of alloantibodies and implicate non-RBC ancestry-limited loci in the susceptibility to alloimmunization.

Long-term colonisation with donor bacteriophages following successful faecal microbial transplantation

BackgroundFaecal microbiota transplantation (FMT) is used in the treatment of recurrent Clostridium difficile infection. Its success is typically attributed to the restoration of a diverse microbiota. Viruses (including bacteriophages) are the most numerically dominant and potentially the most diverse members of the microbiota, but their fate following FMT has not been well studied.ResultsWe studied viral transfer following FMT from 3 donors to 14 patients. Recipient viromes resembled those of their donors for up to 12 months. Tracking individual bacteriophage colonisation revealed that engraftment of individual bacteriophages was dependent on specific donor-recipient pairings. Specifically, multiple recipients from a single donor displayed highly individualised virus colonisation patterns.ConclusionsThe impact of viruses on long-term microbial dynamics is a factor that should be reviewed when considering FMT as a therapeutic option.

Incidence of new-onset diabetes mellitus and association with mortality in childhood solid organ transplant recipients: a population-based study.

Precise estimates of the long-term risk of new-onset diabetes and its impact on mortality among transplanted children are not known. We conducted a cohort study comparing children undergoing solid organ (kidney, heart, liver, lung and multiple organ) transplant (n = 1020) between 1991 and 2014 with healthy non-transplanted children (n = 7 134 067) using Ontario health administrative data. Outcomes included incidence of diabetes among transplanted and non-transplanted children, the relative hazard of diabetes among solid organ transplant recipients, overall and at specific intervals posttransplant, and mortality among diabetic transplant recipients. During 56019824 person-years of follow-up, the incidence rate of diabetes was 17.8 [95% confidence interval (CI) 15-21] and 2.5 (95% CI 2.5-2.5) per 1000 person-years among transplanted and non-transplanted children, respectively. The transplant cohort had a 9-fold [hazard ratio (HR) 8.9; 95% CI 7.5-10.5] higher hazard of diabetes compared with those not transplanted. Risk was highest within the first year after transplant (HR 20.7; 95% CI 15.9-27.1), and remained elevated even at 5 and 10 years of follow-up. Lung and multiple organ recipients had a 5-fold (HR 5.4; 95% CI 3.0-9.8) higher hazard of developing diabetes compared with kidney transplant recipients. Transplant recipients with diabetes had a three times higher hazard of death compared with those who did not develop diabetes (HR 3.3; 95% CI 2.3-4.8). The elevated risk of diabetes in transplant recipients persists even after a decade, highlighting the importance of ongoing surveillance. Diabetes after transplantation increases the risk of mortality among childhood transplant recipients.

The fate of hepatocyte cell line derived from a liver injury model with long-term in vitro passage

Orthotopic Liver Transplantation (OLT) is the therapy of choice for the treatment of end-stage liver disease, but the severe shortage of organ donors, complex and expensive surgical procedure and increased mortality of prospective organ recipient limit the use of OLT. To overcome this problem the technique of hepatocyte transplantation has been considered as an alternative to OLT. Hepatocyte transplantation is less invasive, cost effective cryo-preservable and can be distributed from single donor to multiple recipients. In this study we have established and characterize the hepatocyte cell line possessing the morphological and functional characteristics of hepatocytes from chemically injured liver. Hence we hypothesized that the hepatocyte cell line derived from liver injury can be used for hepatocyte transplantation. To induce the priming of hepatocyte, mice were fed with 3,5-diethoxycarbonyl-1,4-dihydro-collidine (DDC) diet for 3 weeks and hepatocytes were obtained by two step collagenase perfusion method, hepatocytes hence obtained was expended by >300 passages and tested for various hepatocyte specific functions. The cell line derived from liver injury model retains morphological and functional characteristics of hepatocytes in long-term in vitro culture. These cells transplanted to mice showed significant survival rate. The Hepatocyte cell line derived from liver injury model were used for hepatocyte transplantation and showed the significantly higher survival rate.

CRISPR/Cas9-Based Cellular Engineering for Targeted Gene Overexpression.

Gene and cellular therapies hold tremendous promise as agents for treating genetic disorders. However, the effective delivery of genes, particularly large ones, and expression at therapeutic levels can be challenging in cells of clinical relevance. To address this engineering hurdle, we sought to employ the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system to insert powerful regulatory elements upstream of an endogenous gene. We achieved robust activation of the COL7A1 gene in primary human umbilical cord blood CD34+ hematopoietic stem cells and peripheral blood T-cells. CD34+ cells retained their colony forming potential and, in a second engineering step, we disrupted the T-cell receptor complex in T-cells. These cellular populations are of high translational impact due to their engraftment potential, broad circulatory properties, and favorable immune profile that supports delivery to multiple recipients. This study demonstrates the feasibility of targeted knock in of a ubiquitous chromatin opening element, promoter, and marker gene that doubles as a suicide gene for precision gene activation. This system merges the specificity of gene editing with the high level, sustained gene expression achieved with gene therapy vectors. We predict that this design concept will be highly transferrable to most genes in multiple model systems representing a facile cellular engineering platform for promoting gene expression.

Using Texting for Clinical Communication in Surgery: A Survey of Academic Staff Surgeons.

Text messaging has become ubiquitous and is being increasingly used within the health care system. The purpose of this study was to understand texting practices for clinical communication among staff surgeons at a large academic institution. Staff surgeons in 4 subspecialties (vascular, plastics, urology, and general surgery) were surveyed electronically. A total of 62 surgeons from general surgery (n = 33), vascular surgery (n = 6), plastic surgery (n = 13), and urology (n = 10) completed the study (response rate 30%). When conveying urgent patient-related information, staff surgeons preferred directly calling other staff surgeons (61.5%) and trainees (58.8%). When discussing routine patient information, staff surgeons used email to reach other staff surgeons (54.9%) but preferred texting (62.7%) for trainees. The majority of participants used texting because it is fast (65.4%), convenient (69.2%) and allows transmitting information to multiple recipients simultaneously (63.5%). Most felt that texting enhances patient care (71.5%); however, only half believed that it enhanced trainees' educational experiences. The majority believed that texting identifiable patient information breaches patient confidentiality. Our data showed high adoption of text messaging for clinical communication among surgeons, particularly with trainees. The majority of surgeons acknowledge security concerns inherent in texting for patient care. Existing mobile communication platforms fail to meet the needs of academic surgeons. Further research should include guidelines related to texting in clinical practice, educational implications of texting, and technologies to better meet the needs of clinicians working in an academic surgical settings.

The Relation Between Multiple Informal Caregiving Roles and Subjective Physical and Mental Health Status Among Older Adults: Do Racial/Ethnic Differences Exist?

The present study examined whether race/ethnicity moderated the relation between type of caregiving role (none, one, or multiple care recipients) and subjective physical and mental health among older adults. The sample was drawn from the 2009 California Health Interview Survey. Racially/ethnically diverse adults aged 55 and older (n = 24,241) were categorized into 3 groups by caregiving roles: noncaregivers (n = 18,626; referent), caregivers with a single caregiving role (n = 4,023), and caregivers with multiple caregiving roles (n = 1,772). A 2-way analysis of covariance was conducted to test main and interaction effects. After adjustment for covariates, noncaregivers reported significantly worse self-rated health and lower psychological distress than caregivers with any type of role. The interaction between race/ethnicity and caregiving roles was significant only for self-rated health (p < .05). Blacks with multiple caregiving roles had poorer self-rated health than those with a single caregiving role and better self-rated health than noncaregivers, whereas other racial/ethnic groups with multiple caregiving roles had better self-rated health compared to both noncaregivers and those with a single caregiving role. Our sensitivity analysis showed that controlling caregiving-related variables present only among caregivers eliminated the differences in self-rated health between the two types of caregivers. Findings suggest that caregivers report better self-rated health than noncaregivers and that the relation of multiple caregiving roles with self-rated health differs by race/ethnicity, with blacks differing from other racial/ethnic groups. This implies that caregivers experience gain, or are selected into the role of caregiving by virtue of having good health.

Posttransplant Lymphoproliferative Disorder in Adults Receiving Kidney Transplantation in British Columbia: A Retrospective Cohort Analysis.

Background:Posttransplant lymphoproliferative disorder (PTLD) is a major complication following kidney transplantation.Objective:We undertook this study to characterize PTLD in kidney transplant patients in British Columbia with regard to incidence, patient and graft survival, histological subtypes, treatment modalities, and management of immunosuppression.Design:Retrospective cohort analysis.Setting:British Columbia.Patients:All adult patients who underwent kidney transplantation in British Columbia between January 1, 1996, and December 31, 2012, were included. Patients less than 18 years of age at the time of first transplant and multiple organ transplant recipients were excluded from analysis.Measurements:Patients with lymphoproliferative disorders that occurred subsequent to kidney transplantation were considered to have developed PTLD.Methods:Cases of PTLD were identified by cross-referencing data abstracted from the provincial transplant agency’s clinical database with the provincial cancer agency’s lymphoma registry. Patients were followed up for the development of PTLD until December 31, 2012, and for outcomes of death and graft failure until December 31, 2014. Data collection was completed via an electronic chart review.Results:Of 2217 kidney transplant recipients, 37 (1.7%) developed PTLD. Nine cases were early-onset PTLD, occurring within 1 year of transplant; of these cases, 6 were known/presumed Epstein-Barr virus mismatch, compared with only 2 of 28 late-onset cases. Patient survival for early-onset PTLD was 100% at 2 years post diagnosis. Late-onset PTLD had survival rates of 71.4% and 67.9% at 1 and 2 years, respectively. PTLD was associated with significantly decreased patient survival (P = .031) and graft survival (uncensored for death, P = .017), with median graft survival of PTLD and non-PTLD patients being 9.5 and 16 years, respectively. Immunosuppressant therapy was reduced in the majority of patients; additional therapies included rituximab monotherapy, CHOP-R, radiation, and surgery.Limitations:Limitations to this study include its retrospective nature and the unknown adherence of patients to prescribed immunosuppressant regimens. In addition, cumulative doses of immunosuppression received and the degree of immunosuppression reduction for PTLD management were not effectively captured.Conclusions:The incidence of PTLD in British Columbia following kidney transplantation was low and consistent with rates reported in the literature. The incidence of late-onset PTLD and its association with reduced patient and graft survival warrant further analysis of patients’ long-term immunosuppression.

Analysis of Economic Development Impact of Remittances on Recipient (Zimbabwe) and Remitting (South Africa) Countries

The importance of remittances to recipient economies has been greatly researched and there is a general consensus around their continued significance to these countries. However, their impact on the development of the receiving economies remains a subject of much debate among academics and policy makers. There is even greater dearth of academic research and debate around the impact of remittances on the sending economies. Unlike Foreign Direct Investment (FDI) flows, whose impact on the economies can be closely correlated to the economies’ outputs, remittances are micro-payments fragmented to multiple recipients, from multiple individual senders with different motives and for a multitude of uses. The researcher modified The Newtonian Gravity model first adapted by a Dutch economist, Timbergen as the first published proponent of the Newtonian gravity model application in analysis of financial flows. The model was applied to remittance flows between the sender and recipient countries, and assess the economic impact on the two economies. The key corridor of the research was between Zimbabwe and South Africa, which represents one of the biggest regional remittances flow corridors in Africa. The investigation revealed that remittances not only had a significant impact on recipient economies, but showed a negative correlation with Zimbabwe’s GDP in particular. Outflows of remittances proved to have very little impact on the sending country, South Africa. On further examination of the other countries studied, distance from the main remitting country had a negative correlation with remittances flows. Economic impairment of receiving countries increased their dependency on the remittances flows, and the funds were not directed at activities that directly contributed to GDP growth of recipient countries.

Seroprevalence of Transfusion Transmitted Infections among blood donors in a Tertiary care Hospital in Andhra Pradesh

Background: Blood transfusion is a life-saving procedure as even a single unit of blood and its components can save multiple recipients in need. Though it’s a safe procedure, the risk of transmission of infections like HIV, syphilis, hepatitis B and hepatitis C are still seen. The objective of this study was to determine the frequency of HBsAg, HCV, HIV, Syphilis and Malaria in various ABO and Rh (D) blood groups donors. Methods: A retrospective study was performed from January 2014 to May 2017 in a blood bank of a tertiary care hospital. A total of 8260 blood units which were collected from both voluntary and replacement donors during the study period were included. Complete donor’s demography and screening status for TTI’s were analysed. Results: Out of 8260 blood donors, 232(2.8%) were positive for TTIs. The overall prevalence of HIV, HBV, HCV and Syphilis was found to be 0.06%, 2.1 % , 0.15% and 0.4% respectively. The highest percentage of prevalence was observed for HBV, followed by syphilis, HCV and HIV. Higher seroprevalence of TTI’s were noted in replacement donors and younger age group between 18 to 30 years. Conclusion: Following strict donor eligibility criteria and effecting high sensitive screening methods can bring down the prevalence of TTI’s. Proper treatment and follow-up counselling can help in preventing further transmission of infections in the community. DOI: 10.21276/APALM.1674

Improving Environmental Quality Through Aid: An Experimental Analysis of Aid Structures With Heterogeneous Agents

Environmental aid has grown significantly over the past decade with aid allocation mechanisms varying greatly and little guidance as to which should be preferred. To increase aid effectiveness, some have argued that potential recipients should compete for aid. We experimentally test this proposition against a baseline treatment where groups of heterogeneous agents receive aid by improving environmental quality. With the first mechanism, aid is allocated based on the improvement of environmental quality by each group with the potential of multiple groups receiving aid. In the second mechanism, aid is competitively allocated only to the single group that has initiated the greatest improvement of environmental quality. Surprisingly, the average level of environmental progress is lower in a competitive setting than in the setting with the potential for multiple recipients. Underscoring this result is our finding that environmental progress is stable in the baseline but begins high and falls quickly with repetitions in the competition. Although the initial level of progress is higher on average in the competitive setting, this mechanism generates the highest variance which could be catastrophic to sensitive ecosystems. These findings have direct implications on which mechanism the granting agency should use dependent upon their specific objectives.

Effects of maternal obesity on Wharton\u2019s Jelly mesenchymal stromal cells

We investigated whether maternal metabolic environment affects mesenchymal stromal/stem cells (MSCs) from umbilical cord’s Wharton’s Jelly (WJ) on a molecular level, and potentially render them unsuitable for clinical use in multiple recipients. In this pilot study on umbilical cords post partum from healthy non-obese (BMI = 19–25; n = 7) and obese (BMI ≥ 30; n = 7) donors undergoing elective Cesarean section, we found that WJ MSC from obese donors showed slower population doubling and a stronger immunosuppressive activity. Genome-wide DNA methylation of triple positive (CD73+CD90+CD105+) WJ MSCs found 67 genes with at least one CpG site where the methylation difference was ≥0.2 in four or more obese donors. Only one gene, PNPLA7, demonstrated significant difference on methylome, transcriptome and protein level. Although the number of analysed donors is limited, our data suggest that the altered metabolic environment related to excessive body weight might bear consequences on the WJ MSCs.

Shall I call, text, post it online or just tell it face-to-face? How and why Flemish adolescents choose to share their emotions on- or offline

Social sharing of emotions is a frequently used emotion regulation strategy. This study adds to the emotion regulation literature and the affordances of technologies perspective by providing a better understanding of with whom adolescents share emotions on- and offline, how they do this and why they use certain modes. In-depth interviews with 22 Flemish adolescents (aged 14–18) show that these youngsters share almost all experienced emotions, often with multiple recipients and using multiple communication modes. Although they mostly prefer sharing emotions face-to-face, they also share by texting, calling, or posting something on Twitter, Facebook, Snapchat, or Instagram. Our respondents generally make a more or less conscious decision about what and how to share. The valence, type, and intensity of the emotion, the affordances of the mode, social norms, and impression management concerns influence this decision.

Abstract 1010: Genomic analysis of recurrent ovarian cancer in patient-derived xenografts treated with platinum and taxane chemotherapy

Abstract Introduction: Many women with advanced ovarian cancer (OC) are initially sensitive to platinum and taxane chemotherapy, but subsequently develop recurrent disease which is often incurable. OC chemotherapy resistance mechanisms are incompletely understood. We used patient-derived xenografts (PDX) to model OC recurrence following chemotherapy and to analyze genomic evolution of recurrent tumors. Approach: We leveraged PDX derived from human OC ascites that accurately represent high-grade serous OC. We used two PDX models: PDX-A derived from a chemotherapy-naive patient with WT BRCA1/2; PDX-B derived from a chemotherapy-resistant patient with a BRCA1 mutation. Using luciferase-labeled tumor cells, we generated intraperitoneal disease in multiple recipients. We treated cohorts of 5-10 animals with vehicle, carboplatin, paclitaxel, or carboplatin + paclitaxel (C/T) for 3 cycles followed by observation with bioluminescence imaging of tumor burden. Results: While vehicle- and paclitaxel-treated animals exhibited rapid disease outgrowth, carboplatin- and C/T-treated animals showed disease regression, followed by recurrence after 90-300 days. Median survival was significantly increased in carboplatin- and C/T-treated animals. Recurrent tumors were histologically similar to vehicle-treated tumors. We performed whole-exome sequencing and whole-transcriptome sequencing on ascites cells from 4 animals in each treatment group. We used a decontamination algorithm to remove stromal mouse reads from our tumor samples, resulting in a highly pure (over 99%) human tumor sequence data. Mutations, insertions/deletions, and copy number alterations were identified and compared among treatment groups. Known TP53 mutations were present at 100% frequency across samples. Average mutation frequencies were similar across all groups. We identified several mutations and copy number variants present in the carboplatin or C/T recurrent tumors that were distinct from the vehicle groups, including putative alterations in chemoresistance pathways such as DNA repair (BRCA1/2), apoptosis (BCL2L1), drug transport (ABC transporters), and PI3K signaling (EEF2K). In model PDX-A, all four carboplatin and one C/T treated recurrent tumors exhibited a similar copy-number profile, suggestive of subclonal expansion, which included an increase in WT copies of PTEN relative to the vehicle tumors. Phylogenetic analysis and whole-transcriptome analysis are ongoing. Conclusions: Our study models recurrent ovarian cancer in PDX and identifies potential features of in vivo chemotherapy resistance. Citation Format: Elizabeth Stover, Ali Amin-Mansour, Sangeetha Palakurthi, Sanam Dharma, Qing Zeng, Shan Zhou, Palak Desai, Levi Garraway, Ursula Matulonis, Joyce Liu. Genomic analysis of recurrent ovarian cancer in patient-derived xenografts treated with platinum and taxane chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1010. doi:10.1158/1538-7445.AM2017-1010

Pressured Sexting and Revenge Porn in a Sample of Massachusetts Adolescents

Digital communications are largely used for positive interactions but can also be a vehicle for harassment. Previous research has made is clear that sexting occurs, at times, because of peer pressure. This study examined pressured sexting and the unauthorized release of images in a cross-sectional sample studied in 2013-15. The convenience sample examined 1,320 students in Massachusetts. Over the years, more students admitted to sexting, but fewer reported any degree of pressure to sext. More than a third of sexters in 2014 and almost half of sexters in 2015 reported that the picture had been released without their consent. Interestingly, this did not seem to occur primarily within established relationships; instead, it seemed to target most often sexters who declined to date someone. Unauthorized distribution was related to several risk factors, including younger-aged sexters, those who sexted to multiple recipients, and those who were pressured into sexting initially.

Welfare environmentality and REDD+ incentives in Indonesia

ABSTRACTThis paper applies the concept of welfare environmentality to analyse Indonesia’s emerging national and project-based incentive frameworks, a key component to the climate programme reducing emissions from deforestation and forest degradation (REDD+). The paper adapts governmentality theory to explore the rationale and design of various incentive instruments, including government institutions to disburse financial payments and co-benefits to multiple recipients, as well as a local demonstration project in Central Kalimantan Province. These REDD+ incentives are often conflated with the neoliberalisation of the climate agenda, focused on the adoption of market instruments and the commodification of forest carbon. However, REDD+ incentives in Indonesia include diverse policy mechanisms and encompass multiple objectives – such as the delivery of social services and employment schemes aimed at improving community livelihoods. These incentives employ a welfare environmentality, where government agencies and their partners deliver certain rights and socio-economic security for communities in return for adopting practices that improve carbon and forest management. The application of welfare environmentality shows how incentive frameworks operate as a state intervention designed to restructure relations between people and environmental resources.

The value of variety and scarcity across development

An important task that children face is determining the value of items, and two possible cues to value include scarcity and variety. In three studies with 289 children aged 4–12years and 148 adults, we examined the use of these cues to guide choices when making selections among items. At all ages, participants typically preferred varied sets for themselves and others. In contrast, scarce items were rarely preferred to abundant items. However, when in the context of multiple recipients, participants selected scarce and varied items more when items were maximally scarce. Our results suggest that the preference for variety is early emerging, whereas the preference for scarce items is context dependent.

Single-Cell Analysis of Clonal Dynamics in Childhood ALL Reveals a Key Role for Transcriptional Intratumor Heterogeneity in Driving Resistance to Chemotherapy

Tumors exhibit intratumor heterogeneity in both genetic make-up and phenotypic traits such as morphology, cell cycle status and gene expression. Although heterogeneity has been long recognized, its biological and clinical significance is not fully understood. To determine the relative contribution of genetic versus epigenetic heterogeneity to therapeutic resistance and relapse in childhood acute lymphoblastic leukemia (cALL), we have developed a mouse model that affords "real-time" longitudinal analysis of subclonal dynamics.The mouse model allows the engraftment of primary leukemic cells from an individual patient into multiple xenograft recipients, half of which are then treated with a combination of vincristine and dexamethasone. By assessing the reproducibility of independent outcomes, one can distinguish between deterministic and stochastic mechanisms of selection during therapy. We have used automated multicolor fluorescence in situ hybridization (mFISH) for the most frequent drivers of the leukemia (ETV6, RUNX1, PAX5, P16) to track the fate of individual genetic subclones. By comparing clonal composition in control and treated mice pre- and post-chemotherapy we demonstrate that, while treatment of sensitive ALLs results in a striking reduction in leukemic burden, the overall extent of genetic diversity (measured by mean of the Shannon index of diversity) is unaffected. This suggests that resistance in ALL may be largely independent of genetic variegation. In light of recent data demonstrating aberrant RAG activation in ETV6-RUNX1 leukemia, we reasoned that our results could reflect convergent evolution. Because RAG mediated deletions recurrently target the same key B-cell pathways, it is possible that genetically distinct clones could display convergent phenotypes. To test this hypothesis, and extend our mutational analysis to whole genome resolution, we performed single-cell whole genome sequencing. We sequenced matching diagnostic and relapse samples, and further validated our mFISH data. This reveals unprecedented levels of intratumor heterogeneity, with individual cells carrying on average 37 copy number variations (CNVs), a large number of which are private. By resolving the genomic breakpoints of lesions to known drivers of disease we formally proved that convergent evolution can happen. Thus, we identified clones carrying the same lesion but distinct breakpoints, highlighting their independent origins.Functional analysis based on limiting dilution secondary transplantation assays showed that chemotherapy enriches for cells with tumor propagating potential. Transcriptome analysis (RNAseq) of treated and untreated cells revealed that the former have distinct signatures shared amongst metastatic sites. Chemo-resistant cells displayed differences in cell cycle status and transcription rate, as well as differential activation of key signaling pathways (MAPK, Rap1 and Jak-Stat), and expression of cell adhesion molecules. Through gene set enrichment analysis we observed that resistant cells have features characteristic of more primitive hematopoietic cells, including concomitant up-regulation of hematopoietic stem cells and lympho-myeloid progenitors markers. Interestingly, following secondary transplantation global gene expression of treated cells largely reverted to a treatment-naïve signature.Overall this analysis provides novel insight into the contribution of genetic and epigenetic heterogeneity to resistance to cytotoxic chemotherapy. In the case of cALL phenotypic heterogeneity appears to play a larger role than genetic diversity, particularly with regards to cell cycle state and developmental stage. Our data suggest that resistance is driven by a pre-existing population of immature cells with a distinct global gene expression signature and higher tumor propagating potential, and that genetic heterogeneity in cALL arises through independent acquisition of CNVs affecting the same gene or genes within the same pathway, thereby driving convergent phenotypic evolution. Such a view is consistent with the observation that distinct subclones appear to respond to chemotherapy-derived environmental pressure in a similar manner. DisclosuresNo relevant conflicts of interest to declare.

Hematopoietic stem cell heterogeneity is determined at the clonal level

Recent studies suggest that hematopoietic stem cells (HSCs) heterogeneously supply blood cells after transplantation. Little is known about how this heterogeneity is regulated. Here, we used a genetic barcode system to track the blood production of individual HSC clones during serial transplantation in mice. HSC clones were marked with unique genetic barcodes prior to their transplantation into primary recipient mice. Four months later, we recovered HSCs from each primary recipient and transplanted them into multiple secondary recipient mice. Blood cells were sampled and analyzed four months after each transplantation. We found that fewer HSC clones supplied blood cells in the secondary recipients than in the primary recipients but that their clonal expansion had significantly increased. Surprisingly, many clones found in the peripheral blood of the secondary recipients were not present in the peripheral blood of the primary recipients. Most of these clones were activated in multiple secondary recipients and were biased towards myeloid lineages. Strikingly, HSCs derived from the same clone exhibited consistent clonal expansion and lineage bias characteristics across different secondary recipients. However, these differentiation characteristics changed between the primary and secondary recipients. For example, lineage-balanced HSC clones in the primary recipients mostly converted to lymphoid bias in the secondary recipients, and myeloid-biased clones in the primary recipients were more likely to become lineage balanced in the secondary recipients. Overall, our data suggest that HSCs derived from the same clone exhibit similar blood production characteristics, which are altered in a predictable way upon transplantation. These findings suggest that the clonal-level characteristics of HSC blood production may be exploited in bone marrow transplantation therapy. For example, select HSC clones can be transplanted to improve the efficacy of this treatment. Better therapeutic outcomes may also be achieved by matching donor HSC clones to patient specific requirements.