Somatic mutations in clonally expanded cytotoxic T lymphocytes in patients with newly diagnosed rheumatoid arthritis.

Somatic mutations contribute to tumorigenesis. Although these mutations occur in all proliferating cells, their accumulation under non-malignant conditions, such as in autoimmune disorders, has not been investigated. Here, we show that patients with newly diagnosed rheumatoid arthritis have expanded CD8+ T-cell clones; in 20% (5/25) of patients CD8+ T cells, but not CD4+ T cells, harbour somatic mutations. In healthy controls (n=20), only one mutation is identified in the CD8+ T-cell pool. Mutations exist exclusively in the expanded CD8+ effector-memory subset, persist during follow-up, and are predicted to change protein functions. Some of the mutated genes (SLAMF6, IRF1) have previously been associated with autoimmunity. RNA sequencing of mutation-harbouring cells shows signatures corresponding to cell proliferation. Our data provide evidence of accumulation of somatic mutations in expanded CD8+ T cells, which may have pathogenic significance for RA and other autoimmune diseases.

Immunosuppressive therapy of LGL leukemia: prospective multicenter phase II study by the Eastern Cooperative Oncology Group (E5998).

Failure to undergo activation-induced cell death due to global dysregulation of apoptosis is the pathogenic hallmark of large granular lymphocyte (LGL) leukemia. Consequently, immunosuppressive agents are rational choices for treatment. This first prospective trial in LGL leukemia was a multicenter, phase 2 clinical trial evaluating methotrexate (MTX) at 10 mg/m(2) orally weekly as initial therapy (step 1). Patients failing MTX were eligible for treatment with cyclophosphamide at 100 mg orally daily (step 2). The overall response in step 1 was 38% with 95% confidence interval (CI): 26 and 53%. The overall response in step 2 was 64% with 95% CI: 35 and 87%. The median overall survival for patients with anemia was 69 months with a 95% CI lower bound of 46 months and an upper bound not yet reached. The median overall survival for patients with neutropenia has not been reached 13 years from study activation. Serum biomarker studies confirmed the inflammatory milieu of LGL but were not a priori predictive of response. We identify a gene expression signature that correlates with response and may be STAT3 mutation driven. Immunosuppressive therapies have efficacy in LGL leukemia. Gene signature and mutational profiling may be an effective tool in determining whether MTX is an appropriate therapy.

Crackles in patients with fibrosing alveolitis, bronchiectasis, COPD, and heart failure.

We have studied the crackling lung sounds of ten patients with cryptogenic fibrosing alveolitis, ten with bronchiectasis, ten with chronic obstructive pulmonary disease, and ten with heart failure by analyzing frequency, waveform, and timing of crackles. The upper frequency limit of inspiratory sounds was higher in CFA than in COPD or in HF. The period of crackling was shorter in COPD than in CFA or BE. Inspiratory crackling terminated significantly earlier in COPD than in CFA, BE, or HF. The initial deflection width and the two-cycle duration of the expanded waveforms of crackles were smaller in CFA than in BE, COPD, or HF. The largest deflection width was smaller in CFA than in BE, HF, or COPD and smaller in BE than in HF. The results indicate that crackling lung sounds in different diseases have distinctive features and that their analysis can be of diagnostic value.

Validated method for automatic detection of lung sound crackles.

Crackling lung sounds are associated with many pulmonary diseases. Their occurrence reflects the quality and the severity of the disease. An automatic method for crackle detection is developed, based on analysing the spectral stationarity of the lung sound. The method is validated by studying the crackles of 20 adult patients; 10 with fibrosing alveolitis (FA) and 10 with bronchiectasis (BE). The number of crackles detected by the automatic method in inspiratory cycles is compared to the number of crackles counted from time-expanded waveforms by two expert observers. The total number of inspiratory cycles studied is 117 and that of crackles 1064. The method has a sensitivity of 89 per cent and a positive predictivity of 88 per cent for patients with FA, and 80 per cent and 83 per cent respectively, for patients with BE. The linear correlation coefficients between the numbers of crackles counted by the automatic method and by the observers is 0.86 (p less than 0.001) for the patients with FA and 0.93 (p less than 0.001) for the patients with BE. The values refer to whole inspiratory cycles. The new automatic method seems reliable enough for clinical and scientific purposes. It enables a rapid and objective analysis of large materials with crackling lung sounds.

Novel somatic mutations in large granular lymphocytic leukemia affecting the STAT-pathway and T-cell activation.

T-cell large granular lymphocytic (T-LGL) leukemia is a clonal disease characterized by the expansion of mature CD3+CD8+ cytotoxic T cells. It is often associated with autoimmune disorders and immune-mediated cytopenias. Our recent findings suggest that up to 40% of T-LGL patients harbor mutations in the STAT3 gene, whereas STAT5 mutations are present in 2% of patients. In order to identify putative disease-causing genetic alterations in the remaining T-LGL patients, we performed exome sequencing from three STAT mutation-negative patients and validated the findings in 113 large granular lymphocytic (LGL) leukemia patients. On average, 11 CD8+ LGL leukemia cell-specific high-confidence nonsynonymous somatic mutations were discovered in each patient. Interestingly, all patients had at least one mutation that affects either directly the STAT3-pathway (such as PTPRT) or T-cell activation (BCL11B, SLIT2 and NRP1). In all three patients, the STAT3 pathway was activated when studied by RNA expression or pSTAT3 analysis. Screening of the remaining 113 LGL leukemia patients did not reveal additional patients with same mutations. These novel mutations are potentially biologically relevant and represent rare genetic triggers for T-LGL leukemia, and are associated with similar disease phenotype as observed in patients with mutations in the STAT3 gene.