Daratumumab monotherapy in refractory warm autoimmune hemolytic anemia and cold agglutinin disease.

ABSTRACT: Autoimmune hemolytic anemia (AIHA) is a rare autoantibody-mediated disease. For steroid and/or rituximab-refractory AIHA, there is no consensus on optimal treatment. Daratumumab, a monoclonal antibody targeting CD38, could be beneficial by suppression of CD38+ plasma cells and thus autoantibody secretion. In addition, because CD38 is also expressed by activated T cells, daratumumab may also act via immunomodulatory effects. We evaluated the efficacy and safety of daratumumab monotherapy in an international retrospective study including 19 adult patients with heavily pretreated refractory AIHA. In warm AIHA (wAIHA, n = 12), overall response was 50% with a median response duration of 5.5 months (range, 2-12), including ongoing response in 2 patients after 6 and 12 months. Of 6 nonresponders, 4 had Evans syndrome. In cold AIHA (cAIHA, n = 7) overall hemoglobin (Hb) response was 57%, with ongoing response in 3 of 7 patients. One additional patient with nonanemic cAIHA was treated for severe acrocyanosis and reached a clinical acrocyanosis response as well as a Hb increase. Of 6 patients with cAIHA with acrocyanosis, 4 had improved symptoms after daratumumab treatment. In 2 patients with wAIHA treated with daratumumab, in whom we prospectively collected blood samples, we found complete CD38+ T-cell depletion after daratumumab, as well as altered T-cell subset differentiation and a severely diminished capacity for cell activation and proliferation. Reappearance of CD38+ T cells coincided with disease relapse in 1 patient. In conclusion, our data show that daratumumab therapy may be a treatment option for refractory AIHA. The observed immunomodulatory effects that may contribute to the clinical response deserve further exploration.

Olecranon bursitis caused by Scedosporium apiospermum in a patient treated with CAR-T cells.

Chimeric antigen receptor (CAR-) T cell therapy is a relatively new form of immunotherapy for hematological malignancies. Although patients are at increased risk of infection following CAR-T cell therapy, reports of fungal infections are scarce. We report a case of Scedosporium apiospermum infection causing bursitis of the elbow in a lymphoma patient after treatment with CAR-T cells. The fungal bursitis relapsed under posaconazole treatment, but was cured after surgical extirpation of the bursa.

The opposing effect of hypoxia-inducible factor-2alpha on expression of telomerase reverse transcriptase.

Hypoxia-inducible factor-1alpha (HIF-1alpha) has been implicated in the transcriptional regulation of the telomerase reverse transcriptase (hTERT) gene expression and telomerase activity, essential elements for cellular immortalization and transformation. However, controversial results were obtained in different studies. Moreover, it is totally unclear whether HIF-2alpha, the paralog of HIF-1alpha, plays a role in regulating hTERT expression. In the present study, we found that hypoxic treatment enhanced hTERT mRNA expression and telomerase activity in three renal cell carcinoma (RCC) cell lines with different genetic backgrounds. Both HIF-1alpha and HIF-2alpha were capable of significantly increasing the hTERT promoter activity in these cells. Moreover, depleting HIF-2alpha led to a down-regulation of hTERT mRNA level in RCC A498 cells expressing constitutive HIF-2alpha. It was found that HIF-2alpha bound to the hTERT proximal promoter and enhanced the recruitment of the histone acetyltransferase p300 and histone H3 acetylation locally in A498 cells treated with hypoxia. Increased levels of hTERT mRNA were observed in two of three hypoxia-treated malignant glioma cell lines. However, HIF-1alpha stimulated whereas HIF-2alpha inhibited the hTERT promoter activity in these glioma cell lines. Ectopic expression of HIF-2alpha resulted in diminished hTERT expression in glioma cells. Collectively, HIF-1alpha activates hTERT and telomerase expression in both RCC and glioma cells, and HIF-2alpha enhances hTERT expression in RCC cells, whereas it represses the hTERT transcription in glioma cells. These findings reveal a complex relationship between HIF-1alpha/2alpha and hTERT/telomerase expression in malignant cells, which may have both biological and clinical implications.

The telomerase reverse transcriptase (hTERT) gene is a direct target of the histone methyltransferase SMYD3.

Recent evidence has accumulated that the dynamic histone methylation mediated by histone methyltransferases and demethylases plays key roles in regulation of chromatin structure and transcription. In the present study, we show that SET and MYND domain-containing protein 3 (SMYD3), a histone methyltransferase implicated in oncogenesis, directly trans-activates the telomerase reverse transcriptase (hTERT) gene that is essential for cellular immortalization and transformation. SMYD3 occupies its binding motifs on the hTERT promoter and is required for maintenance of histone H3-K4 trimethylation, thereby contributing to inducible and constitutive hTERT expression in normal and malignant human cells. Knocking down SMYD3 in tumor cells abolished trimethylation of H3-K4, attenuated the occupancy by the trans-activators c-MYC and Sp1, and led to diminished histone H3 acetylation in the hTERT promoter region, which was coupled with down-regulation of hTERT mRNA and telomerase activity. These results suggest that SMYD3-mediated trimethylation of H3-K4 functions as a licensing element for subsequent transcription factor binding to the hTERT promoter. The present findings provide significant insights into regulatory mechanisms of hTERT/telomerase expression; moreover, identification of the hTERT gene as a direct target of SMYD3 contributes to a better understanding of SMYD3-mediated cellular transformation.

Human normal T lymphocytes and lymphoid cell lines do express alternative splicing variants of human telomerase reverse transcriptase (hTERT) mRNA.

Alternative splicing of telomerase reverse transcriptase (hTERT) mRNA is known to contribute to regulation of telomerase activity in normal and cancerous cells, however, previous studies indicated that normal human T and B cells exhibited constitutive expression of full-length hTERT mRNA without splicing variants and that activation of telomerase upon stimulation of the cells was due to the shuttling of hTERT protein from cytoplasm to nucleus [Proc. Natl. Acad. Sci. USA 96 (1999) 5147; J. Immunol. 166 (2001) 4826]. We found that typical variants of hTERT mRNA were widespread in human lymphocyte-derived cell lines and normal stimulated T cells. In activated T cells, induction of the full-length hTERT mRNA was coupled with increased hTERT protein expression and telomerase activity. Collectively, human normal and malignant lymphocytes, like other human cells, express splicing variants of hTERT mRNA and require transcriptional activation of the hTERT gene to acquire telomerase activity.