Clinical-scale isolation of the total Aspergillus fumigatus-reactive T-helper cell repertoire for adoptive transfer

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• 作者：Petra Bacher¹ ; ^&lowast ; ; Andrea Jochheim-Richter² ; ^&lowast ; ; Nadine Mockel-Tenbrink³ ; Olaf Kniemeyer⁴ ; Eva Wingenfeld² ; Regina Alex³ ; Alice Ortigao² ; Darja Karpova² ; Thomas Lehrnbecher⁵ ; Andrew J. Ullmann⁶ ; Axel Hamprecht⁷ ; Oliver Cornely⁸ ; Axel A. Brakhage⁹ ; Mario Assenmacher³ ; Halvard Bonig² ; ¹⁰ ; ¹¹ ; ^&lowast ; ; ^{h.boenig@blutspende.de" class="auth_mail" title="E-mail the corresponding author} ; ^{hbonig@uw.edu" class="auth_mail" title="E-mail the corresponding author} ; Alexander Scheffold¹ ; ¹² ; ^&lowast ;
• 关键词：aspergillosis ; cell therapy ; GMP-compliant protocol ; prospective isolation ; T-helper cells
• 刊名：Cytotherapy
• 出版年：2015
• 出版时间：October 2015
• 年：2015
• 卷：17
• 期：10
• 页码：1396-1405
• 全文大小：1433 K

文摘

Evidence of the criticality of the adaptive immune response for controlling invasive aspergillosis has been provided. This observation is supported by the fact that invasive aspergillosis, a grave complication of allogeneic stem cell transplantation, occurs long after myeloid reconstitution in patients with low T-cell engraftment and/or on immunosuppressants. Adoptive T-cell transfer might be beneficial, but idiosyncrasies of Aspergillus fumigatus and the anti-Aspergillus immune response render established selection technologies ineffective.

Methods

We developed a Good Manufacturing Practice (GMP)-compliant protocol for preparation of A. fumigatus–specific CD4+ cells by sequentially depleting regulatory and cytotoxic T cells, activating A. fumigatus-specific T-helper cells with GMP-grade A. fumigatus lysate, and immuno-magnetically isolating them via the transiently up-regulated activation marker, CD137.

Results

In 13 full-scale runs, we demonstrate robustness and feasibility of the approach. From 2 × 10⁹ peripheral blood mononuclear cells, we isolated 27 × 10³–318 × 10³Aspergillus-specific T-helper cells. Frequency among total T cells was increased, on average, by 200-fold. Specific studies indicate specificity and functionality: After non-specific in vitro expansion and re-stimulation with different antigens, we observed strong cytokine responses to A. fumigatus and some other fungi including Candida albicans, but none to unrelated antigens.

Discussion

Our technology isolates naturally occurring Aspergillus-specific T-helper cells within 2 days of identifying the clinical indication. Rapid adoptive transfer of Aspergillus-specific T cells may be quite feasible; the clinical benefit remains to be demonstrated. A manufacturing license as an advanced-therapy medicinal product was received and a clinical trial in post-transplantation invasive aspergillosis patients approved. The product is dosed at 5 × 10E3/kg T cells (single intravenous injection), of which at least 10% must be A. fumigatus–specific.