RESUMEN
Extramedullary disease (EMD) is a high-risk feature of multiple myeloma (MM) and remains a poor prognostic factor even in the era of novel immunotherapies. Here we applied spatial transcriptomics (tomo-seq [n=2] and 10X Visium [n=12]), and single-cell RNA sequencing (scRNAseq [n=3]) to a set of 14 EMD biopsies to dissect the three-dimensional architecture of tumor cells and their microenvironment. Overall, the infiltrating immune and stromal cells showed both intra- and inter-patient variation with no uniform distribution over the lesion. We observed substantial heterogeneity at the copy number level within plasma cells, including the emergence of new subclones in circumscribed areas of the tumor, consistent with genomic instability. We further identified spatial expression differences of GPRC5D and TNFRSF17, two important antigens for bispecific antibody therapy. EMD masses were infiltrated by various immune cells, including T-cells. Notably, exhausted TIM3+/PD-1+ T-cells diffusely co-localized with MM cells, whereas functional and activated CD8+ T-cells showed a focal infiltration pattern along with M1 macrophages in otherwise tumor-free regions. This segregation of fit and exhausted T-cells was resolved in the case of response to T-cell engaging bispecific antibodies. MM cells and microenvironment cells were embedded in a complex network that influenced immune activation and angiogenesis, and oxidative phosphorylation represented the major metabolic program within EMD lesions. In summary, spatial transcriptomics has revealed a multicellular ecosystem in EMD with checkpoint inhibition and dual targeting as potential new therapeutic avenues.
RESUMEN
Biomarkers for cytopenias following CAR T-cell treatment in relapsed/refractory (RR) multiple myeloma (MM) are not completely defined. We prospectively analysed 275 sequential peripheral blood (PB) samples from 58 RRMM patients treated with BCMA-targeted CAR T cells, and then divided them into three groups: (i) baseline (before leukapheresis), (ii) ≤day+30, and (iii) >day+30 after CAR T-cell therapy. We evaluated laboratory data and performed flow cytometry to determine the (CAR) T-cell subsets. Baseline hyperferritinaemia was a risk factor for long-lasting grade ≥3 anaemia (r = 0.47, p < 0.001) and thrombocytopenia (r = 0.44, p = 0.002) after CAR T-cell therapy. Low baseline haemoglobin (Hb) and PLT were associated with long-lasting grade ≥3 anaemia (r = -0.56, p < 0.001) and thrombocytopenia (r = -0.44, p = 0.002) respectively. We observed dynamics of CAR-negative T-cell subsets following CAR T-cell infusion. In the late phase after CAR T-cell therapy (>day+30), CD4Tn frequency correlated with anaemia (r = 0.41, p = 0.0014) and lymphocytopenia was related to frequencies of CD8+ T cells (r = 0.72, p < 0.001) and CD8Teff (r = 0.64, p < 0.001). CD4Tcm frequency was correlated with leucocytopenia (r = -0.49, p < 0.001). In summary, preexisting cytopenias and hyperferritinaemia indicated long duration of grade ≥3 post-CAR T-cell cytopenias. Prolonged cytopenia may be related to immune remodelling with a shift in the CAR-negative T-cell subsets following CAR T-cell therapy.
RESUMEN
In pancreatic ductal adenocarcinoma (PDAC), endogenous MYC is required for S-phase progression and escape from immune surveillance. Here we show that MYC in PDAC cells is needed for the recruitment of the PAF1c transcription elongation complex to RNA polymerase and that depletion of CTR9, a PAF1c subunit, enables long-term survival of PDAC-bearing mice. PAF1c is largely dispensable for normal proliferation and regulation of MYC target genes. Instead, PAF1c limits DNA damage associated with S-phase progression by being essential for the expression of long genes involved in replication and DNA repair. Surprisingly, the survival benefit conferred by CTR9 depletion is not due to DNA damage, but to T-cell activation and restoration of immune surveillance. This is because CTR9 depletion releases RNA polymerase and elongation factors from the body of long genes and promotes the transcription of short genes, including MHC class I genes. The data argue that functionally distinct gene sets compete for elongation factors and directly link MYC-driven S-phase progression to tumor immune evasion.
Asunto(s)
Fenómenos Bioquímicos , Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Proteínas Proto-Oncogénicas c-myc , Animales , Ratones , Carcinoma Ductal Pancreático/patología , Proliferación Celular , ARN Polimerasas Dirigidas por ADN/metabolismo , Evasión Inmune , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas c-myc/metabolismoRESUMEN
Pseudo-progression and flare-up phenomena constitute a novel diagnostic challenge in the follow-up of patients treated with immune-oncology drugs. We present a case study on pulmonary flare-up after Idecabtagen Vicleucel (Ide-cel), a BCMA targeting CAR T-cell therapy, and used single-cell RNA-seq (scRNA-seq) to identify a Th17.1 driven autoimmune mechanism as the biological underpinning of this phenomenon. By integrating datasets of various lung pathological conditions, we revealed transcriptomic similarities between post CAR T pulmonary lesions and sarcoidosis. Furthermore, we explored a noninvasive PET based diagnostic approach and showed that tracers binding to CXCR4 complement FDG PET imaging in this setting, allowing discrimination between immune-mediated changes and true relapse after CAR T-cell treatment. In conclusion, our study highlights a Th17.1 driven autoimmune phenomenon after CAR T, which may be misinterpreted as disease relapse, and that imaging with multiple PET tracers and scRNA-seq could help in this diagnostic dilemma.
Asunto(s)
Mieloma Múltiple , Receptores Quiméricos de Antígenos , Sarcoidosis , Humanos , Antígeno de Maduración de Linfocitos B , Inflamación/metabolismo , Mieloma Múltiple/patología , Recurrencia Local de Neoplasia/metabolismo , Sarcoidosis/metabolismo , Linfocitos T , Células Th17RESUMEN
The ATP-dependent nucleosome remodeler Mi-2/CHD4 broadly modulates chromatin landscapes to repress transcription and to maintain genome integrity. Here we use individual nucleotide resolution crosslinking and immunoprecipitation (iCLIP) to show that Drosophila Mi-2 associates with thousands of mRNA molecules in vivo. Biochemical data reveal that recombinant dMi-2 preferentially binds to G-rich RNA molecules using two intrinsically disordered regions of unclear function. Pharmacological inhibition of transcription and RNase digestion approaches establish that RNA inhibits the association of dMi-2 with chromatin. We also show that RNA inhibits dMi-2-mediated nucleosome mobilization by competing with the nucleosome substrate. Importantly, this activity is shared by CHD4, the human homolog of dMi-2, strongly suggesting that RNA-mediated regulation of remodeler activity is an evolutionary conserved mechanism. Our data support a model in which RNA serves to protect actively transcribed regions of the genome from dMi-2/CHD4-mediated establishment of repressive chromatin structures.
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Proteínas de Drosophila , Nucleosomas , Adenosina Trifosfatasas/metabolismo , Animales , Autoantígenos/metabolismo , Cromatina/metabolismo , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Nucleosomas/metabolismo , ARN/metabolismoRESUMEN
T cell-engaging immunotherapies exert unprecedented single-agent activity in multiple myeloma (MM), thereby putting a yet unexplored selective pressure on the clonal architecture. In this study, we report on homozygous BCMA (TNFRSF17) gene deletion after BCMA-targeting T cell-redirecting bispecific antibody therapy in a heavily pretreated MM patient. Loss of BCMA protein expression persisted over subsequent relapses, with no response to treatment with anti-BCMA antibody drug conjugate. In light of the multiple alternative targets that are emerging in addition to BCMA, we extended our analyses to delineate a more complete picture of genetic alterations that may have an impact on immunotherapy targets in MM. We performed whole-genome sequencing and RNA sequencing in 100 MM patients (50 were newly diagnosed; 50 were relapsed/refractory) and identified a significant proportion of patients with aberrations in genes encoding immunotherapy targets; GPRC5D ranked first with 15% heterozygous deletions, followed by CD38 (10%), SDC1 (5%), and TNFRSF17 (4%). Notably, these heterozygous deletions did not lower the expression levels of respective genes, but they may represent a first hit that drives the acquisition of homozygous deletions and subsequent antigen-loss relapse upon targeted immunotherapy. In summary, we show preexisting vulnerability in genes encoding immunotargets before and homozygous deletions after T cell-engaging immunotherapy.
Asunto(s)
Anticuerpos Biespecíficos , Mieloma Múltiple , Antígeno de Maduración de Linfocitos B , Humanos , Inmunoterapia , Mieloma Múltiple/genética , Mieloma Múltiple/terapia , Linfocitos TRESUMEN
BACKGROUND AND PURPOSE: To the best of our knowledge, no information is available regarding the treatment of vulvovaginal candidiasis in gynecological practices. The goal of this study was to analyze the prevalence of vulvovaginal candidiasis (VVC) and the drugs prescribed for the treatment of this condition in women followed in gynecological practices in Germany. MATERIALS AND METHODS: All the women followed in 262 gynecological practices between November 2014 and October 2016 were included in this study. The first outcome was the prevalence of patients diagnosed with VVC during this period. The second outcome was the prevalence of women with VVC who received an appropriate vaginal or systemic antimycotic prescription within 30 days after their first VVC diagnosis. Covariables included the use of gynecological/systemic antibiotics, consumption of oral/vaginal contraceptives, cancer, pregnancy, diabetes, and psychiatric diseases including depression, anxiety, and adjustment and somatoform disorders. RESULTS: Between 2014 and 2016, 954,186 women were followed in gynecological practices, and 50,279 (5.3%) women were diagnosed with VVC during the same period. The use of gynecological antibiotics (OR=2.88), systemic antibiotics (OR=1.45), oral contraceptives (OR=1.74), and vaginal contraceptives (OR=1.84) were associated with an increase in the risk of VVC diagnosis. Cancer (OR=1.20) and pregnancy (OR=1.59) were additional risk factors. Approximately 75% of women diagnosed with VVC received an antimycotic prescription. The three most frequently prescribed drugs were clotrimazole (72%), fluconazole (14%), and nystatin (6%). CONCLUSION: More than 5% of women were diagnosed with VVC and the majority of them received an appropriate prescription.