LINC01432 binds to CELF2 in newly diagnosed multiple myeloma promoting short progression-free survival to standard therapy.

Multiple Myeloma (MM) is a highly prevalent and incurable form of cancer that arises from malignant plasma cells, with over 35,000 new cases diagnosed annually in the United States. While there are a growing number of approved therapies, MM remains incurable and nearly all patients will relapse and exhaust all available treatment options. Mechanisms for disease progression are unclear and in particular, little is known regarding the role of long non-coding RNAs (lncRNA) in mediating disease progression and response to treatment. In this study, we used transcriptome sequencing to compare newly diagnosed MM patients who had short progression- free survival (PFS) to standard first-line treatment (PFS < 24 months) to patients who had prolonged PFS (PFS > 24 months). We identified 157 differentially upregulated lncRNAs with short PFS and focused our efforts on characterizing the most upregulated lncRNA, LINC01432 . We investigated LINC01432 overexpression and CRISPR/Cas9 knockdown in MM cell lines to show that LINC01432 overexpression significantly increases cell viability and reduces apoptosis, while knockdown significantly reduces viability and increases apoptosis, supporting the clinical relevance of this lncRNA. Next, we used individual-nucleotide resolution cross-linking immunoprecipitation with RT-qPCR to show that LINC01432 directly interacts with the RNA binding protein, CELF2. Lastly, we showed that LINC01432 -targeted locked nucleic acid antisense oligonucleotides reduce viability and increases apoptosis. In summary, this fundamental study identified lncRNAs associated with short PFS to standard NDMM treatment and further characterized LINC01432, which inhibits apoptosis. Key points: lncRNA expression was found to be dysregulated in patients with short PFS to standard multiple myeloma therapy. LINC01432 -bound CELF2 inhibits apoptosis.

A single-cell atlas characterizes dysregulation of the bone marrow immune microenvironment associated with outcomes in multiple myeloma.

Multiple Myeloma (MM) remains incurable despite advances in treatment options. Although tumor subtypes and specific DNA abnormalities are linked to worse prognosis, the impact of immune dysfunction on disease emergence and/or treatment sensitivity remains unclear. We established a harmonized consortium to generate an Immune Atlas of MM aimed at informing disease etiology, risk stratification, and potential therapeutic strategies. We generated a transcriptome profile of 1,149,344 single cells from the bone marrow of 263 newly diagnosed patients enrolled in the CoMMpass study and characterized immune and hematopoietic cell populations. Associating cell abundances and gene expression with disease progression revealed the presence of a proinflammatory immune senescence-associated secretory phenotype in rapidly progressing patients. Furthermore, signaling analyses suggested active intercellular communication involving APRIL-BCMA, potentially promoting tumor growth and survival. Finally, we demonstrate that integrating immune cell levels with genetic information can significantly improve patient stratification.

A Single Reference Interval for Interpreting Serum Free Light Chains across Patients with Varying Renal Function.

BACKGROUND: Serum free light chain (sFLC) assays are interpreted using a sFLC-ratio-based reference interval (manufacturer's interval) that was defined using a cohort of healthy patients. However, renal impairment elevates the sFLC-ratio, leading to a high false positive rate when using the manufacturer's interval. Prior studies have developed renal-specific reference intervals; however, this approach has not been widely adopted due to practical limitations. Thus, there remains a critical need for a renally robust sFLC interpretation method. METHODS: Retrospective data mining was used to define patient cohorts that reflect the spectrum of renal function seen in clinical practice. Two new reference intervals, one based on the sFLC-ratio and one based on a novel principal component analysis (PCA)-based metric, were developed for the FREELITE assay (Binding Site) on the Roche Cobas c501 instrument (Roche). RESULTS: Compared to the manufacturer's reference interval, both new methods exhibited significantly lower false positive rates and greater robustness to renal function while maintaining equivalent sensitivity for monoclonal gammopathy (MG) diagnosis. While not significantly different, the point estimate for sensitivity was highest for the PCA-based approach. CONCLUSION: Renally robust sFLC interpretation using a single reference interval is possible given a reference cohort that reflects the variation in renal function observed in practice. Further studies are needed to achieve sufficient power and determine if the novel PCA-based metric offers superior sensitivity for MG diagnosis. These new methods offer the practical advantages of not requiring an estimated glomerular filtration rate result or multiple reference intervals, thereby lowering practical barriers to implementation.

Single-Cell Discovery and Multiomic Characterization of Therapeutic Targets in Multiple Myeloma.

Multiple myeloma (MM) is a highly refractory hematologic cancer. Targeted immunotherapy has shown promise in MM but remains hindered by the challenge of identifying specific yet broadly representative tumor markers. We analyzed 53 bone marrow (BM) aspirates from 41 MM patients using an unbiased, high-throughput pipeline for therapeutic target discovery via single-cell transcriptomic profiling, yielding 38 MM marker genes encoding cell-surface proteins and 15 encoding intracellular proteins. Of these, 20 candidate genes were highlighted that are not yet under clinical study, 11 of which were previously uncharacterized as therapeutic targets. The findings were cross-validated using bulk RNA sequencing, flow cytometry, and proteomic mass spectrometry of MM cell lines and patient BM, demonstrating high overall concordance across data types. Independent discovery using bulk RNA sequencing reiterated top candidates, further affirming the ability of single-cell transcriptomics to accurately capture marker expression despite limitations in sample size or sequencing depth. Target dynamics and heterogeneity were further examined using both transcriptomic and immuno-imaging methods. In summary, this study presents a robust and broadly applicable strategy for identifying tumor markers to better inform the development of targeted cancer therapy. SIGNIFICANCE: Single-cell transcriptomic profiling and multiomic cross-validation to uncover therapeutic targets identifies 38 myeloma marker genes, including 11 transcribing surface proteins with previously uncharacterized potential for targeted antitumor therapy.

Development and Validation of a Short Questionnaire Assessing the Behavior of Local Food Procurement in Quebec, Canada.

Background: Very few validated instruments, particularly screening tools applicable to large-cohort studies, are available to assess the behavior of local food procurement. Objective: The aim was to develop and validate a short questionnaire that measures local food procurement in a sample of French-speaking adults from Quebec, Canada, and to assess the association between local food-procurement behavior and diet quality. Methods: A comprehensive questionnaire developed previously to measure local food procurement [Locavore-Index (Locavore-I)] was simplified through a series of steps that included face-validity, exploratory factor analysis, and reliability testing (internal consistency). Construct validity of the resulting short Locavore-I Short Form (Locavore-I-SF) was examined in a sample of 299 adults (85% women) from the Quebec City metropolitan community. Results: The Locavore-I-SF comprises 12 questions that measure the frequency of short food supply chain use (self-production, farmers' markets, and community-supported agriculture box scheme) for 3 locally produced foods (carrot, tomato, and lettuce) as well as the geographical origin of those 3 foods. The Locavore-I-SF, which is scored on a 12-point scale, had a high internal consistency (Cronbach ɑ: 0.74). The Locavore-I-SF scores were strongly correlated with the reference scores obtained from the Locavore-I from which it was developed (r = 0.84, P < 0.0001). Locavore-I-SF scores also correlated (r = 0.50, P < 0.0001) with the geographical origin of foods measured by pictures of food labels taken by participants. Higher Locavore-I-SF scores were associated with behaviors consistent with eating local foods, such as gardening (vs. not gardening; mean ± SEM difference: 2.3 ± 0.4 points; P < 0.0001) and not being preoccupied by the foods' appearance standards (vs. being preoccupied; 1.4 ± 0.4 points; P = 0.0002). Finally, the Locavore-I-SF scores were weakly associated with the Healthy Eating Food Index-2019 score (B = 0.05 ± 0.02; P = 0.02). Conclusions: The Locavore-I-SF, a short questionnaire based on 3 locally produced foods in Quebec, measures the behavior of local food procurement with good reliability and acceptable validity metrics.

Challenges and Strategies to Adapt the Provision of Support Services to Older Adults and Caregivers During the COVID-19 Pandemic: The Perspective of Community Organizations.

This study documented the provision of services and issues experienced by community organizations supporting older adults and caregivers in the province of Quebec during the coronavirus disease (COVID-19) pandemic, as well as promising strategies to adapt the provision of services in this context. A cross-sectional electronic survey using open- and closed-ended questions was conducted in July 2020. Almost three-quarters of the 307 respondents (71.4%) reported having maintained services at least partially throughout the lockdown, and the majority (85.3%) adapted their services. Among key challenges, participants reported difficulties identifying and supporting older adults at greater risk of vulnerability (54.8%), managing health risks for service users (60.2%), and recruiting volunteers (59.5%). Promising strategies included strategies to reach out to older adults and understand their needs (e.g., systematic phone calls) in addition to direct interventions supporting them (e.g., activities promoting social ties); implementing prevention and protection measures; accessing and using technologies; human resources management (e.g., recruiting new volunteers); finding financial support for their organization; developing intersectoral partnerships (e.g., multisectoral crisis cell); and promoting a positive view of older adults. The integration of multiple perspectives from different stakeholders may help identify strategies potentially transferable to other crises in order to meet older adults' needs.

A comparison of scoring models for computerised mental health screening for federal prison inmates.

BACKGROUND: There are high rates of mental disorder in correctional environments, so effective mental health screening is needed. Implementation of the computerised mental health screen of the Correctional Service of Canada has led to improved identification of offenders with mental health needs but with high rates of false positives. AIMS: The goal of this study is to evaluate the use of an iterative classification tree (ICT) approach to mental health screening compared with a simple binary approach using cut-off scores on screening tools. METHODS: A total of 504 consecutive admissions to federal prison completed the screen and were also interviewed by a mental health professional. Relationships between screening results and more extended assessment and clinical team discussion were tested. RESULTS: The ICT was more parsimonious in identifying probable 'cases' than standard binary screening. ICT was also highly accurate at detecting mental health needs (AUC=0.87, 95% CI 0.84-0.90). The model identified 118 (23.4%) offenders as likely to need further assessment or treatment, 87% of whom were confirmed cases at clinical interview. Of the 244 (48.4%) offenders who were screened out, only 9% were clinically assessed as requiring further assessment or treatment. Standard binary screening was characterised by more false positives and a comparable false negative rate. CONCLUSIONS: The use of ICTs to interpret screening data on the mental health of prisoners needs further evaluation in independent samples in Canada and elsewhere. This first evaluation of the application of such an approach offers the prospect of more effective and efficient use of the scarce resource of mental health services in prisons. Although not required, the use of computers can increase the ease of implementing an ICT model.

Murine models of human acute myeloid leukemia.

Primary human AML cells can be isolated and studied in vitro, but many experimental questions can only be addressed using in vivo models. In particular, tractable animal models are needed to test novel therapies. The genetic complexity of human AML poses significant challenges for the generation of reliable animal models. The hematopoietic systems of both zebrafish ( Danio rerio) and Drosophila have been well characterized ( reviewed in [5, 31]) . Both organisms are well suited to forward genetics mutagenesis screens. Although this approach has been useful for identification of mutants with hematopoietic phenotypes ( e.g., cloche), the impact on cancer biology and hematopoietic malignancies in particular has been limited. A zebrafish model of acute lymphoblastic leukemia has been generated [37] and Drosophila models have shed light on the biology of epithelial tumors ( reviewed in [60]). Nonetheless, in vivo modeling of human AML relies most heavily on mice. Most cellular, molecular, and developmental features of the hematopoietic system are well conserved across mammalian species. The availability of the human and mouse genome sequences and the capability of manipulating the mouse genome make mice the most valuable model organism for AML research. Mice have additional practical value because they have a short reproductive cycle and are relatively inexpensive to house.

NK lytic-associated molecule, involved in NK cytotoxic function, is an E3 ligase.

NK lytic-associated molecule (NKLAM) is a protein involved in the cytolytic function of NK cells and CTLs. It has been localized to the cytolytic granules in NK cells and is up-regulated when cells are exposed to cytokines IL-2 or IFN-beta. We report in this study that NKLAM contains a cysteine-rich really interesting new gene (RING) in between RING-RING domain, and that this domain possesses strong homology to the RING domain of the known E3 ubiquitin ligase, Dorfin. To determine whether NKLAM functions as an E3 ligase, we performed coimmunoprecipitation binding assays with ubiquitin conjugates (Ubcs) UbcH7, UbcH8, and UbcH10. We demonstrated that both UbcH7 and UbcH8 bind to full-length NKLAM. We then performed a similar binding assay using endogenous NKLAM and UbcH8 expressed by human NK clone NK3.3 to show that the protein interaction occurs in vivo. Using the yeast two-hybrid system, we identified uridine kinase like-1 (URKL-1) protein as a substrate for NKLAM. We confirmed that NKLAM and URKL-1 interact in mammalian cells by using both immunoprecipitation and confocal microscopy. We demonstrated decreased protein expression and enhanced ubiquitination of URKL-1 in the presence of NKLAM. These data indicate that NKLAM is a RING finger protein that binds Ubcs and has as one of its substrates, URKL-1, thus defining this cytolytic protein as an E3 ubiquitin ligase.