Immunophenotypic characterization of leukemic stem cells in acute myeloid leukemia using single tube 10-colour panel by multiparametric flow cytometry: Deciphering the spectrum, complexity and immunophenotypic heterogeneity.

INTRODUCTION: Despite extensive research, comprehensive characterization of leukaemic stem cells (LSC) and information on their immunophenotypic differences from normal haematopoietic stem cells (HSC) is lacking. Herein, we attempted to unravel the immunophenotypic (IPT) characteristics and heterogeneity of LSC using multiparametric flow cytometry (MFC) and single-cell sequencing. MATERIALS AND METHODS: Bone marrow aspirate samples from patients with acute myeloid leukaemia (AML) were evaluated using MFC at diagnostic and post induction time points using a single tube-10-colour-panel containing LSC-associated antibodies CD123, CD45RA, CD44, CD33 and COMPOSITE (CLL-1, TIM-3, CD25, CD11b, CD22, CD7, CD56) with backbone markers that is, CD45, CD34, CD38, CD117, sCD3. Single-cell sequencing of the whole transcriptome was also done in a bone marrow sample. RESULTS: LSCs and HSCs were identified in 225/255 (88.2%) and 183/255 (71.6%) samples, respectively. Significantly higher expression was noted for COMPOSITE, CD45RA, CD123, CD33, and CD44 in LSCs than HSCs (p < 0.0001). On comparing the LSC specific antigen expressions between CD34+ (n = 184) and CD34- LSCs (n = 41), no difference was observed between the groups. More than one sub-population of LSC was demonstrated in 4.4% of cases, which further revealed high concordance between MFC and single cell transcriptomic analysis in one of the cases displaying three LSC subpopulations by both methods. CONCLUSION: A single tube-10-colour MFC panel is proposed as an easy and reproducible tool to identify and discriminate LSCs from HSCs. LSCs display both inter- and intra-sample heterogeneity in terms of antigen expressions, which opens the facets for single cell molecular analysis to elucidate the role of subpopulations of LSCs in AML progression.

Emtricitabine/tenofovir alafenamide usage in a Scottish sexual health service providing HIV pre-exposure prophylaxis: A service evaluation.

Background: In this service evaluation we evaluate the usage of emtricitabine and tenofovir alafenamide (FTC-TAF) in a large Scottish sexual health service providing Human Immunodeficiency Virus (HIV) Pre-Exposure Prophylaxis (PrEP) over a one-year period (May 2022-May 2023).Purpose: We evaluated the use of FTC-TAF as it is 30 times more expensive per 30 tablet supply than emtricitabine/tenofovir disoproxil (FTC-TDF). We sought to establish: - Number of patients initiated on FTC-TAF - Reasons for FTC-TAF initiation - Adherence to FTC-TAF. Results: Out of a total cohort of 1744 patients on HIV PrEP as of May 2023, seven patients (0.4%) had been initiated on FTC-TAF. The remainder (99.6%) were taking FTC-TDF. The majority of patients (n = 6) were initiated on FTC-TAF due to renal reasons, one patient was initiated FTC-TAF for bone mineral density concerns.Conclusions: All of the patients met eligibility criteria. In terms of adherence, three patients were regular attenders, two patients used FTC-TAF sporadically, and two had initiated and subsequently discontinued FTC-TAF altogether at the time of data analysis. FTC-TAF utilisation was lower than anticipated, initiated apropriately, and followed similar adherence patterns to FTC-TDF users. This will be helpful for financial forecasting and in the development of services where FTC-TAF is newly commissioned.

Early Postoperative Seizures Following Awake Craniotomy and Functional Brain Mapping for Lesionectomy.

OBJECTIVE: Awake craniotomy with electrocorticography (ECoG) and direct electrical stimulation (DES) facilitates lesionectomy while avoiding adverse effects. Early postoperative seizures (EPS), occurring within 7 days following surgery, can lead to morbidity. However, risk factors for EPS after awake craniotomy including clinical and ECoG data are not well defined. METHODS: We retrospectively studied the incidence and risk factors of EPS following awake craniotomy for lesionectomy, and report short-term outcomes between January 1, 2020, and December 31, 2022. RESULTS: We included 138 patients (56 female) who underwent 142 awake craniotomies, average age was 50.78 ± 15.97 years. Eighty-eight (63.7%) patients had a preoperative history of tumor-related epilepsy treated with antiseizure medication (ASM), 12 (13.6%) with drug-resistance. All others (36.3%) received ASM prophylaxis with levetiracetam perioperatively and continued for 14 days. An equal number of cases (71) each utilized a novel circle grid or strip electrodes for ECoG. There were 31 (21.8%) cases of intraoperative seizures, 16 with EPS (11.3%). Acute abnormality on early postoperative neuroimaging (P = 0.01), subarachnoid hemorrhage (P = 0.01), young age (P = 0.01), and persistent postoperative neurologic deficits (P = 0.013) were associated with EPS. Acute abnormality on neuroimaging remained significant in multivariate analysis. Outcomes during hospitalization and early outpatient follow up were worse with EPS. CONCLUSIONS: We report novel findings using ECoG and clinical features to predict EPS, including acute perioperative brain injury, persistent postoperative deficits and young age. Given worse outcomes with EPS, clinical indicators for EPS should alert clinicians of potential need for early postoperative EEG monitoring and perioperative ASM adjustment.

Enhanced sensitivity of electrocorticography during awake craniotomy using a novel circular grid electrode.

PURPOSE: Awake craniotomy with intraoperative functional brain mapping (FBM) bedside neurological testing is an important technique used to optimize resective brain surgeries near eloquent cortex. Awake craniotomy performed with electrocorticography (ECoG) and direct electrical stimulation (DES) for FBM can delineate eloquent cortex from lesions and epileptogenic regions. However, current electrode technology demonstrates spatial limitations. Our group has developed a novel circular grid with the goal of improving spatial recording of ECoG to enhance detection of ictal and interictal activity. METHODS: This retrospective study was approved by the institutional review board at Mayo Clinic Florida. We analyzed patients undergoing awake craniotomy with ECoG and DES and compared ECoG data obtained using the 22 contact circular grid to standard 6 contact strip electrode. RESULTS: We included 144 cases of awake craniotomy with ECoG, 73 using circular grid and 71 with strip electrode. No significant differences were seen regarding preoperative clinical and demographic data, duration of ECoG recording (p = 0.676) and use of DES (p = 0.926). Circular grid was more sensitive in detecting periodic focal epileptiform discharges (PFEDs) (p = 0.004), PFEDs plus (p = 0.032), afterdischarges (ADs) per case (p = 0.022) at lower minimum (p = 0.012) and maximum (p < 0.0012) intensity stimulation, and seizures (p = 0.048). PFEDs (p < 0.001), PFEDs plus (p < 0.001), and HFOs (p < 0.001) but not ADs (p = 0.255) predicted electrographic seizures. CONCLUSION: We demonstrate higher sensitivity in detecting ictal and interictal activity on ECoG during awake craniotomy with a novel circular grid compared to strip electrode, likely due to better spatial sampling during ECoG. We also found association between PFEDs and intraoperative seizures.

Pearls & Oy-sters: Harnessing New Diagnostic and Therapeutic Approaches to Treat a Patient With Genetic Drug-Resistant Focal Epilepsy.

Focal cortical dysplasia (FCD) is a congenital developmental malformation and is one of the leading causes of drug-resistant focal epilepsy (DRFE). Although focal epilepsies traditionally have been regarded as acquired disorders, increasing evidence suggests a substantial genetic contribution to the pathogenesis of focal structural epilepsies, including FCDs. Variations in the Dishevelled, Egl-10, and domain-containing protein 5 (DEPDC5) have recently emerged as a causative gene mutation in familial focal epilepsies associated with FCD type 2a, including bottom-of-sulcus dysplasia (BOSD). We present the case of a 20-year-old man with DRFE, positive for DEPDC5 c.1555C>T (p.GIn519*) heterozygous pathogenic variant. Initial 3T brain MRI was unrevealing, but subsequent 7T MRI including 7T edge-enhancing gradient echo revealed a left superior frontal sulcus BOSD concordant with the electroclinical data. The patient underwent treatment with MR-guided laser interstitial thermal ablation of the left frontal BOSD without intracranial EEG monitoring (skipped candidate), resulting in a seizure-free outcome of 9 months since the last follow-up. Our case highlights the real-world application of summative information obtained through advancements in epilepsy genetic testing, minimally invasive surgeries, and ultra-high field MRI, allowing us to provide a safe and effective treatment for a patient with a genetic DRFE.

Integrated single-cell transcriptome analysis of CD34 + enriched leukemic stem cells revealed intra- and inter-patient transcriptional heterogeneity in pediatric acute myeloid leukemia.

To gain insights into the idiosyncrasies of CD34 + enriched leukemic stem cells, we investigated the nature and extent of transcriptional heterogeneity by single-cell sequencing in pediatric AML. Whole transcriptome analysis of 28,029 AML single cells was performed using the nanowell cartridge-based barcoding technology. Integrated transcriptional analysis identified unique leukemic stem cell clusters of each patient and intra-patient heterogeneity was revealed by multiple LSC-enriched clusters differing in their cell cycle processes and BCL2 expression. All LSC-enriched clusters exhibited gene expression profile of dormancy and self-renewal. Upregulation of genes involved in non-coding RNA processing and ribonucleoprotein assembly were observed in LSC-enriched clusters relative to HSC. The genes involved in regulation of apoptotic processes, response to cytokine stimulus, and negative regulation of transcription were upregulated in LSC-enriched clusters as compared to the blasts. Validation of top altered genes in LSC-enriched clusters confirmed upregulation of TCF7L2, JUP, ARHGAP25, LPAR6, and PRDX1 genes, and serine/threonine kinases (STK24, STK26). Upregulation of LPAR6 showed trend towards MRD positive status (Odds ratio = 0.126; 95% CI = 0.0144-1.10; p = 0.067) and increased expression of STK26 significantly correlated with higher RFS (HR = 0.231; 95% CI = 0.0506-1.052; p = 0.04). Our findings addressed the inter- and intra-patient diversity within AML LSC and potential signaling and chemoresistance-associated targets that warrant investigation in larger cohort that may guide precision medicine in the near future.

Dosimetric effects of repeat computed tomography scan during radiotherapy planning in esophagus carcinoma.

Aim of Study: The aim was to assess the potential reduction in the doses to organs at risk (OARs) and target organ volume by doing replanning on repeat computed tomography (CT) scan during the 4th week of radiation therapy (RT). Materials and Methods: Twenty-four histologically proven patients of inoperable esophagus carcinoma were studied. All patients received induction chemotherapy followed by concurrent chemotherapy and radiotherapy. CT simulation with proper immobilization was done, and images were transferred to the treatment planning system. Delineation of target volumes and OARs was done, and two plans were generated for 60 Gy in 30 fractions and 40 Gy in 20 fractions with intensity-modulated RT keeping the doses to OARs within the tolerance limits. Replanning for 20 Gy in 10 fractions was done on repeat CT scan during the 4th week of radiotherapy treatment, and potential reduction in doses to OARs and target organ volume was assessed. Results: A total of 24 cases were analyzed for the adaptive plan with the coverage of the 95% prescription isodose for planning target volume. Statistical analysis was done by t-test. The difference in the doses received by the OARs was analyzed and was seen that due to re CT scan, the doses were reduced to the left lung V20 (mean 19.23 Gy vs. 17.35 Gy) and Dmean (mean 16.03 Gy vs. 14.25 Gy), right lung V20 (mean 18.38 Gy vs. 16.66 Gy) and Dmean (mean 15.70 Gy vs. 13.97 Gy), heart V25 (mean 38.72 Gy vs. 35.32 Gy) and Dmean (mean 26.40 Gy vs. 22.74 Gy), and spine 1% volume (mean 36.54 Gy vs. 33.39 Gy) and Dmax (mean 39.81 Gy vs. 34.34 Gy), gross tumor volume (GTV) (mean 67.37 cm 3 vs. 24.58 cm 3) and were all significantly smaller for the adaptive plan. Conclusion: By doing adaptive radiotherapy in the 4th week of treatment using repeat CT scan, along with the response evaluation, there is a significant reduction in the volume of GTV, and replanning of treatment on repeat CT scan also helps us in reducing doses to the OARs resulting in reduced toxicity.

Use of misoprostol before tandem application for intracavitary brachytherapy in patients of carcinoma cervix: An institution study.

Aim: To investigate the ease of tandem application and external os identification by giving sublingual misoprostol before initiation of intracavitary brachytherapy in cancer cervix patients. Materials and Methods: 36 patients with cervical cancer stage IIIB which were supposed to undergo intracavitary brachytherapy(ICBT) were randomly divided into 2 subgroups, group A patients receiving misoprostol and group B not receiving misoprostol.Misoprostol 400 mcg was given sublingually 3 hrs prior to the procedure. The efficacy of the drug was measured as per the ease of identification of os and easier tandem application and amount of bleeding during procedure. Results: Application of tandem and identification of external os was easier and amount of bleeding was also less in patients that were administered sublingual misoprostol. Conclusion: Sublingual Misoprostol given before ICBT helps in cervical ripening and thus leads to easier os recognition and central tandem application and reduce overall anaesthesia time.

Quadruple metachronous primary cancer in a single patient: A rare case report.

Multiple primary cancer is a condition where multiple occurrences of different malignancies occur in the same individual. As there is a rise in the long-term survival of patients, multiple primary cancer is now not a rare entity. To see four different tumors in the same patient is very rare, and here, we report the case of a 60-year-old female patient with quadruple primary cancer of bilateral breast, esophagus, and sarcoma of the leg.

Leukemic stem cell signatures in Acute myeloid leukemia- targeting the Guardians with novel approaches.

Acute myeloid leukemia is an aggressive hematopoietic stem cell malignancy with poor outcomes despite the available treatment options including standard chemotherapy, selective targeted therapy and stem cell transplantation. Approximately ~30-40% of AML patients are refractory to initial therapy or succumb to relapse. Induction failure result from inherent resistance to chemotherapy, which is primarily driven by the chemo-resistant residual leukemic stem cells (LSC) that lead to disease progression and recurrence. The rarity and lack of universal surface markers for the identification and isolation of AML LSC renders a major challenge. Therefore, a perpetual quest for novel markers to characterize LSC and design anti-LSC therapies is ongoing. The evolving technologies from high-throughput bulk cell sequencing to high-dimensional single cell analysis has begun to decode the cellular hierarchies and dysregulated transcriptional networks in AML. These inherent properties of LSC as well as cross-talk with the extrinsic bone marrow microenvironmental milieu induce a conducive environment for leukemogenesis by secretion of various cytokines, chemokines and growth factors that shield LSC against conventional chemotherapy. To overcome these barriers, novel approaches of intratumoural delivery that focus on immune-mediated eradication by inducing microenvironmental changes within the tumour as well as avoid systemic toxicity seem encouraging. Selective targeting of LSC and their protective bone marrow niche holds immense potential as a promising therapeutic strategy for AML. Novel multimodal anti-LSC therapies are being explored that can overcome chemo-resistance and immune escape combined with reduced toxicity and sustained delivery may improve remission and survival rates in AML patients and decrease relapse.

Autophagy in acute myeloid leukemia: a paradoxical role in chemoresistance.

Autophagy is a lysosomal degradation pathway that is constitutively active in almost every cell of our body at basal level. This self-eating process primarily serves to remove superfluous constituents of the cells and recycle the degraded products. Autophagy plays an essential role in cell homeostasis and can be enhanced in response to stressful conditions. Impairment in the regulation of the autophagic pathway is implicated in pathological conditions such as neurodegeneration, cardiac disorders, and cancer. However, the role of autophagy in cancer initiation and development is controversial and context-dependent. Evidence from various studies has shown that autophagy serves dual purpose and may assist in cancer progression or suppression. In the early stages of cancer initiation, autophagy acts as a quality control mechanism and prevents cancer development. When cancer is established and progresses to a later stage, autophagy helps in the survival of these cells through adaptation to stresses, including exposure to anti-cancer drugs. In this review, we highlight various studies on autophagic pathways and describe the role of autophagy in cancer, specifically acute myeloid leukemia (AML). We also discuss the prognostic significance of autophagy genes involved in AML leukemogenesis and implications in conferring resistance to chemotherapy.

Drug-Eluting Stents: Their Preventative/Prophylactic Role Against Gemcitabine Induced Acute Coronary Syndrome.

Acute coronary syndrome (ACS), a subdivision of ischemic cardiac disease, is the sudden occlusion of coronary vessels that results in decreased blood supply to heart muscles and possible infarction. Though some of the etiologies are hypertension, hyperlipidemia, diabetes mellitus, and tobacco; certain types of chemotherapies play a major role. Percutaneous coronary intervention (PCI) has shown lifesaving results via drug-eluting stent (DES) deployment into occluded vessels. In this study, DES utilization among patients receiving chemotherapy will be assessed to observe if it provides any prevention against ACS. Articles were systematically screened in three databases such as PubMed, PubMed Central (PMC), and Medical Literature Analysis and Retrieval System Online (MEDLINE) using keywords and Medical Subject Heading (MeSH) terms for applicable articles. Additionally, a few relevant articles from the Cochrane Library, Molecular Diversity Preservation International (MDPI), and The New England Journal of Medicine were also used. Inclusion/exclusion criteria were applied post article screening via title and abstracts. Quality appraisal check was done using the Scale for the Assessment of Narrative Review Articles (SANRA) checklist, A Measurement Tool to Assess Systematic Reviews (AMSTAR) checklist, Cochrane bias assessment tool, and Joanna Briggs Institute (JBI) checklist. Ten related studies were strictly reviewed. DES did not appear to play a preventable role against ACS during chemotherapy as no study was found assessing DES prophylactically and its efficacy in cancer patients. Future clinical trials on DES prophylactic use might be beneficial to evaluate if ACS adversities of chemotherapy can be prevented. This review is of significant benefit as cardiovascular adversities would not impede chemotherapy efficacy as cardiac adversities would not be part of the equation.