In-vitro analysis of the effects of TA65 and danazol on the proliferation and telomerase activity of T lymphocytes in bone marrow failure syndromes

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• 作者：Dr Janine Collins ; MRCP^a ; ^{janine@cantab.net" class="auth_mail" title="E-mail the corresponding author} ; Hemanth Tummala ; PhD^a ; Laura Collopy ; MSc^a ; Tom Vulliamy ; PhD^a ; Prof Inderjeet Dokal ; FMedSci^a
• 刊名：The Lancet
• 出版年：2016
• 出版时间：25 February 2016
• 年：2016
• 卷：387
• 期：supp_S1
• 页码：S29
• 全文大小：60 K

文摘

The bone marrow failure syndromes are a diverse group of rare genetic conditions. Mutations in telomere maintenance genes cause a large proportion of cases of dyskeratosis congenita. Our aim was to determine whether TA65, marketed as a telomerase activator, can correct this defect in vitro. We also investigated danazol, which improves peripheral blood counts in dyskeratosis congenita and Fanconi's anaemia by an unknown mechanism.

Methods

We isolated T lymphocytes from the peripheral blood of 13 patients with bone marrow failure (seven dyskeratosis congenita, two Fanconi's anaemia, four uncharacterised) and ten age-matched controls. All 23 T lymphocyte cultures were incubated with TA65 (0·5 μg/mL and 0·05 μg/mL). Lymphocytes from nine patients and five controls were also incubated with danazol (dose range 0·048 ng/mL to 3 μg/mL). Cell proliferation was measured by neutral red assay (days 6 and 8) and nucleocounter (day 8). Telomerase activity of cells from six patients with dyskeratosis congenita was measured by TRAP (telomeric repeat amplification protocol) assay on day 8.

Findings

We observed increased proliferation of T lymphocytes after treatment with TA65 0·5μg/mL and danazol 6 ng/mL (starting cell count 1 × 10⁶, mean final count 7·64 × 10⁶ with TA65 and 7·66 × 10⁶ with danazol vs 7·31 × 10⁶ with dimethyl sulfoxide [DMSO] only, as the control condition). Dyskeratosis congenita cells showed the most consistent results with TA65 treatment, with increased growth of five out of six cultures and overall relative proliferation of 1·033 (95% CI 1·001–1·064). Danazol improved cell growth most markedly in patients with Fanconi's anaemia (1·4 times increased proliferation over DMSO control). There was 1·23 times increased telomerase activity of T lymphocytes from patients with dyskeratosis congenita treated with TA65 0·5 μg/mL compared with DMSO controls. Treatment with danazol 1·2 ng/mL resulted in 1·45 times increased telomerase activity.

Interpretation

This is the first time, to our knowledge, that these drugs have been studied in vitro in patients with bone marrow failure. Our results show a trend to increased proliferation of T lymphocytes from patients with different bone marrow failure syndromes treated with TA65 and danazol. In addition, our data show increased telomerase activity in dyskeratosis congenita cells treated with these drugs. Our study is limited by small patient numbers from this rare heterogeneous bone marrow failure group. However, these preliminary results provide an exciting basis for further studies to help establish the clinical utility of these drugs.

Funding

Barts Charity (Academic Clinical Fellowship Support Grant).