TORNADO-seq: A Protocol for High-Throughput Targeted RNA-seq-Based Drug Screening in Organoids.

Organoids are 3D ex vivo cell aggregates derived from primary tissue and shown to closely recapitulate tissue homeostasis. Organoids deliver certain advantages compared to 2D cell lines and mouse models, especially in drug-screening studies and translational research projects. The application of organoids in the research field is fast-emerging and new techniques for organoid manipulation are constantly developing.

IFNγ-induced stem-like state of cancer cells as a driver of metastatic progression following immunotherapy.

Despite the remarkable success of immune checkpoint blockade (ICB) therapy, most cancer patients still do not respond. We now find that immunotherapy can induce stem-like properties in tumors. Using mouse models of breast cancer, we observe that cancer stem cells (CSCs) show not only enhanced resistance to T cell cytotoxicity, but that interferon gamma (IFNγ) produced by activated T cells directly converts non-CSCs to CSCs.

Platform combining statistical modeling and patient-derived organoids to facilitate personalized treatment of colorectal carcinoma.

Background: We propose a new approach for designing personalized treatment for colorectal cancer (CRC) patients, by combining ex vivo organoid efficacy testing with mathematical modeling of the results.

Methods: The validated phenotypic approach called Therapeutically Guided Multidrug Optimization (TGMO) was used to identify four low-dose synergistic optimized drug combinations (ODC) in 3D human CRC models of cells that are either sensitive or resistant to first-line CRC chemotherapy (FOLFOXIRI). Our findings were obtained using second order linear regression and adaptive lasso.

National Marrow Donor Program-Sponsored Multicenter, Phase II Trial of HLA-Mismatched Unrelated Donor Bone Marrow Transplantation Using Post-Transplant Cyclophosphamide.

Purpose: Hematopoietic cell transplantation (HCT) is curative for hematologic disorders, but outcomes are historically inferior when using HLA-mismatched donors. Despite unrelated donor registries listing > 38 million volunteers, 25%-80% of US patients lack an HLA-matched unrelated donor, with significant disparity across ethnic groups. We hypothesized that HCT with a mismatched unrelated donor (MMUD) using post-transplant cyclophosphamide (PTCy), a novel strategy successful in overcoming genetic disparity using mismatched related donors, would be feasible and increase access to HCT.

High-content, targeted RNA-seq screening in organoids for drug discovery in colorectal cancer.

Organoids allow the recapitulation of intestinal homeostasis and cancerogenesis in vitro; however, RNA sequencing (RNA-seq)-based methods for drug screens are missing. We develop targeted organoid sequencing (TORNADO-seq), a high-throughput, high-content drug discovery platform that uses targeted RNA-seq to monitor the expression of large gene signatures for the detailed evaluation of cellular phenotypes in organoids. TORNADO-seq is a fast, highly reproducible time- and cost-effective ($5 per sample) method that can probe cell mixtures and their differentiation state in the intestinal system.

Genomic Instability Profiles at the Single Cell Level in Mouse Colorectal Cancers of Defined Genotypes.

The genomes of many human CRCs have been sequenced, revealing a large number of genetic alterations. However, the molecular mechanisms underlying the accumulation of these alterations are still being debated. In this study, we examined colorectal tumours that developed in mice with , , and targetable alleles.

Distinct genetic pathways define pre-malignant versus compensatory clonal hematopoiesis in Shwachman-Diamond syndrome.

To understand the mechanisms that mediate germline genetic leukemia predisposition, we studied the inherited ribosomopathy Shwachman-Diamond syndrome (SDS), a bone marrow failure disorder with high risk of myeloid malignancies at an early age. To define the mechanistic basis of clonal hematopoiesis in SDS, we investigate somatic mutations acquired by patients with SDS followed longitudinally. Here we report that multiple independent somatic hematopoietic clones arise early in life, most commonly harboring heterozygous mutations in EIF6 or TP53.

Phase II Trial of Costimulation Blockade With Abatacept for Prevention of Acute GVHD.

Purpose: Severe (grade 3-4) acute graft-versus-host disease (AGVHD) is a major cause of death after unrelated-donor (URD) hematopoietic cell transplant (HCT), resulting in particularly high mortality after HLA-mismatched transplantation. There are no approved agents for AGVHD prevention, underscoring the critical unmet need for novel therapeutics. ABA2 was a phase II trial to rigorously assess safety, efficacy, and immunologic effects of adding T-cell costimulation blockade with abatacept to calcineurin inhibitor (CNI)/methotrexate (MTX)-based GVHD prophylaxis, to test whether abatacept could decrease AGVHD.

Single-Cell Studies of Intestinal Stem Cell Heterogeneity During Homeostasis and Regeneration.

Single-cell RNA-sequencing (scRNA-seq) provides a unique opportunity to study heterogeneous cell populations within tissues, including the intestinal epithelium, to gain detailed molecular insights into their biology. Many new putative markers of intestinal stem cells and their progeny have been described using single-cell transcriptomics, which has contributed to the identification of novel subpopulations of mature cell types and insight into their developmental trajectories. This approach has revealed tremendous cellular heterogeneity within the intestinal epithelium that is concordant with its diverse and multifaceted functions.

Subsequent neoplasms and late mortality in children undergoing allogeneic transplantation for nonmalignant diseases.

We examined the risk of subsequent neoplasms (SNs) and late mortality in children and adolescents undergoing allogeneic hematopoietic cell transplantation (HCT) for nonmalignant diseases (NMDs). We included 6028 patients (median age, 6 years; interquartile range, 1-11; range, <1 to 20) from the Center for International Blood and Marrow Transplant Research (1995-2012) registry. Standardized mortality ratios (SMRs) in 2-year survivors and standardized incidence ratios (SIRs) were calculated to compare mortality and SN rates with expected rates in the general population.

Impact of autologous blood transfusion after bone marrow harvest on unrelated donor's health and outcome: a CIBMTR analysis.

Pre-harvest autologous blood collection from bone marrow (BM) donors is performed to meet potential post-operative transfusion needs. This study examines the impact of autologous blood transfusion on BM donor's health and safety. The study included first-time unrelated BM donors from the United States whose BM harvest was facilitated by the National Marrow Donor Program (NMDP) centers between 2006 and 2017.

Clinical predictors of delayed engraftment in autologous hematopoietic cell transplant recipients.

Objective/background: Clinical predictors of delayed engraftment following autologous hematopoietic cell transplantation (AHCT) are poorly described in the literature. The purpose of this study was to identify pretransplant characteristics contributing to delayed engraftment (DE) following AHCT.

Methods: A retrospective, single institution study of 1162 consecutive patients undergoing AHCT from January 1996 to August 2016 was studied for DE.

A genomics-informed computational biology platform prospectively predicts treatment responses in AML and MDS patients.

Patients with myelodysplastic syndromes (MDS) or acute myeloid leukemia (AML) are generally older and have more comorbidities. Therefore, identifying personalized treatment options for each patient early and accurately is essential. To address this, we developed a computational biology modeling (CBM) and digital drug simulation platform that relies on somatic gene mutations and gene CNVs found in malignant cells of individual patients.

Association of Antiepileptic Medications with Outcomes after Allogeneic Hematopoietic Cell Transplantation with Busulfan/Cyclophosphamide Conditioning.

High-dose busulfan (BU) followed by high-dose cyclophosphamide (CY) before allogeneic hematopoietic cell transplantation (HCT) has long been used as treatment for hematologic malignancies. Administration of phenytoin or newer alternative antiepileptic medications (AEMs) prevents seizures caused by BU. Phenytoin induces enzymes that increase exposure to active CY metabolites in vivo, whereas alternative AEMs do not have this effect.

The Concentration of Total Nucleated Cells in Harvested Bone Marrow for Transplantation Has Decreased over Time.

Bone marrow (BM) is an essential source of hematopoietic stem cell grafts for many allogeneic hematopoietic cell transplant (HCT) recipients, including adult patients (for specific diseases and transplantation strategies) and the majority of pediatric recipient. However, since the advent of granulocyte colony-stimulating factor-mobilized peripheral blood stem cell (PBSC) grafts, there has been a significant decrease in the use of BM in HCT, thought to be due mainly to the increased logistical challenges in harvesting BM compared with PBSCs, as well as generally no significant survival advantage of BM over PBSCs. The decreased frequency of collection has the potential to impact the quality of BM harvests.

Effect of Conditioning Regimen Dose Reduction in Obese Patients Undergoing Autologous Hematopoietic Cell Transplantation.

Data are limited on whether to adjust high-dose chemotherapy before autologous hematopoietic cell transplant (autoHCT) in obese patients. This study explores the effects of dose adjustment on the outcomes of obese patients, defined as body mass index (BMI) ≥ 30 kg/m. Dose adjustment was defined as a reduction in standard dosing ≥20%, based on ideal, reported dosing and actual weights.

Health related quality of life for multiple myeloma patients according to treatment strategy after autologous stem cell transplant: a cross-sectional study using EORTC, EQ-5D and MY-20 scales.

Maintenance (MT) may be prescribed after autologous stem cell transplant (ASCT) but there are often concerns about the impact on quality of life (QoL). QoL was compared between baseline patients (30-100 days post-ASCT and had not commenced MT); MT patients (>100 days post-ASCT and receiving MT), and no MT (>100 days post-ASCT and not receiving MT). Patients completed the EuroQoL five dimension (EQ-5D), the European Organization for Research and Treatment of Cancer QoL Questionnaire Core 30 (EORTC QLQ-C30), and the QoL Questionnaire Myeloma 20 module (QLQ-MY20).