Novel Spirocyclic Dimer, SpiD3, Targets Chronic Lymphocytic Leukemia Survival Pathways with Potent Preclinical Effects.

Chronic lymphocytic leukemia (CLL) cell survival and growth is fueled by the induction of B-cell receptor (BCR) signaling within the tumor microenvironment (TME) driving activation of NFκB signaling and the unfolded protein response (UPR). Malignant cells have higher basal levels of UPR posing a unique therapeutic window to combat CLL cell growth using pharmacologic agents that induce accumulation of misfolded proteins. Frontline CLL therapeutics that directly target BCR signaling such as Bruton tyrosine kinase (BTK) inhibitors (e.g., ibrutinib) have enhanced patient survival. However, resistance mechanisms wherein tumor cells bypass BTK inhibition through acquired BTK mutations, and/or activation of alternative survival mechanisms have rendered ibrutinib ineffective, imposing the need for novel therapeutics. We evaluated SpiD3, a novel spirocyclic dimer, in CLL cell lines, patient-derived CLL samples, ibrutinib-resistant CLL cells, and in the Eµ-TCL1 mouse model. Our integrated multi-omics and functional analyses revealed BCR signaling, NFκB signaling, and endoplasmic reticulum stress among the top pathways modulated by SpiD3. This was accompanied by marked upregulation of the UPR and inhibition of global protein synthesis in CLL cell lines and patient-derived CLL cells. In ibrutinib-resistant CLL cells, SpiD3 retained its antileukemic effects, mirrored in reduced activation of key proliferative pathways (e.g., PRAS, ERK, MYC). Translationally, we observed reduced tumor burden in SpiD3-treated Eµ-TCL1 mice. Our findings reveal that SpiD3 exploits critical vulnerabilities in CLL cells including NFκB signaling and the UPR, culminating in profound antitumor properties independent of TME stimuli. SIGNIFICANCE: SpiD3 demonstrates cytotoxicity in CLL partially through inhibition of NFκB signaling independent of tumor-supportive stimuli. By inducing the accumulation of unfolded proteins, SpiD3 activates the UPR and hinders protein synthesis in CLL cells. Overall, SpiD3 exploits critical CLL vulnerabilities (i.e., the NFκB pathway and UPR) highlighting its use in drug-resistant CLL.

Clinicomycological Study of the Spectrum of Pulmonary Aspergillosis at a Tertiary Care Hospital in Central India.

Knowing the spectrum, prevalence, and modes of diagnosis of pulmonary aspergillosis (PA) will be beneficial to clinicians for its early diagnosis and management. This study aims to estimate the prevalence, spectrum, and role of serological tests and radiological findings in the diagnosis of PA. A total of 150 patients were suspected of having PA after obtaining relevant clinical history and radiological imaging. The patients were grouped into each spectrum of PA as invasive PA (IPA), chronic necrotizing PA (CNPA), aspergilloma, allergic bronchopulmonary aspergillosis (ABPA) based on predisposing factors, clinical and radiological findings, and the guidelines of the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG). Samples (bronchoalveolar lavage (BAL), sputum, blood) were collected from these patients and processed in a microbiology lab. BAL and sputum were subjected to microscopy by potassium hydroxide mount, calcofluor white mount, and culture. The serum was separated from blood by centrifugation and subjected to specific serological tests based on the spectrum of PA that the patient was suspected to have. For IPA, serum and BAL galactomannan antigen enzyme-linked immunosorbent assay (ELISA) was performed. For CNPA and aspergilloma, the anti-Aspergillus IgG antibody ELISA was performed. For ABPA, the tests performed were total immunoglobulin E (IgE) ELISA, Aspergillus fumigatus-specific IgE ELISA, and anti-Aspergillus immunoglobulin G (IgG) antibody ELISA. After compiling the clinical, radiological, culture, and serological findings, patients were diagnosed to have a particular spectrum of PA. The prevalence of IPA was 1.4%, CNPA was 4%, ABPA was 3.2%, and aspergilloma was 2.9%. CNPA was the predominant spectrum of PA in our study. Culture positivity for Aspergillus species was seen the highest in aspergilloma patients, followed by IPA, ABPA, and CNPA patients. A. fumigatus was the most common causative agent of PA, except for IPA for which Aspergillus flavus was the most common causative. Aspergillus niger and Aspergillus terreus were less the frequent causes of PA. A combination of radiological, microbiological, and serological tests along with clinical correlation is needed to confirm the diagnosis of PA.

Chromatin alternates between A and B compartments at kilobase scale for subgenic organization.

Nuclear compartments are prominent features of 3D chromatin organization, but sequencing depth limitations have impeded investigation at ultra fine-scale. CTCF loops are generally studied at a finer scale, but the impact of looping on proximal interactions remains enigmatic. Here, we critically examine nuclear compartments and CTCF loop-proximal interactions using a combination of in situ Hi-C at unparalleled depth, algorithm development, and biophysical modeling. Producing a large Hi-C map with 33 billion contacts in conjunction with an algorithm for performing principal component analysis on sparse, super massive matrices (POSSUMM), we resolve compartments to 500 bp. Our results demonstrate that essentially all active promoters and distal enhancers localize in the A compartment, even when flanking sequences do not. Furthermore, we find that the TSS and TTS of paused genes are often segregated into separate compartments. We then identify diffuse interactions that radiate from CTCF loop anchors, which correlate with strong enhancer-promoter interactions and proximal transcription. We also find that these diffuse interactions depend on CTCF's RNA binding domains. In this work, we demonstrate features of fine-scale chromatin organization consistent with a revised model in which compartments are more precise than commonly thought while CTCF loops are more protracted.