Cytokines in AIDS and Cancer

我们利用先前开发的RNA优化技术优化IL-15细胞因子的表达，并证明我们可以在小鼠和猕猴的DNA传递后过量产生生物活性细胞因子。这一时期的一个重要结论是，表达异二聚体IL-15的载体的DNA递送导致细胞因子的全系统活性水平和NK细胞和T细胞的增殖增加。因此，DNA注射现在是一种实用的方法来传递IL-15在人。我们探索了IL-15的生物学特性，并表明只有在同一细胞中与所谓的IL-15受体- α共表达才能有效地生产IL-15。我们还发现，当与IL-15受体α共表达时，第二种形式的IL-15 （SSP IL-15）在人类和啮齿动物中被鉴定为细胞内或核IL-15，也能有效地从细胞分泌出来。这些结果为IL-15的生物学和调控提供了新的思路，为该细胞因子的高效生产和临床应用提供了方法。已经构建了可过量产生可溶性生物活性IL-15/IL-15受体α异源二聚体的细胞系，并用于生产在体内发现的真正生物活性形式的IL-15。从过量生产的人类细胞中纯化的IL-15被注射到小鼠体内，并显示出生物活性。对灵长类动物的广泛研究表明，异二聚体IL-15具有良好的药代动力学和药效学特性。异二聚体IL-15的半衰期延长与其他细胞因子形成鲜明对比，并与该细胞因子对免疫系统的稳态作用一致。异二聚体IL-15已被广泛表征，它是重度糖基化的。IL-15因其能够刺激淋巴细胞（包括CD8和NK细胞）的生长、活化和存活而引起人们的兴趣。因此，IL-15已被考虑用于癌症免疫治疗和过继转移后支持细胞毒性细胞克隆的生长。IL-15的其他用途包括淋巴细胞减少，NK细胞转移后支持NK细胞生长和活化，以及作为疫苗佐剂。我们已经证明，注射IL-15可以加速细胞毒性药物治疗后淋巴细胞减少的小鼠淋巴细胞的恢复。我们利用优化后的表达载体在动物体内表达IL-12细胞因子。高效的表达导致了生物活性水平的提高，从而增加了DNA接种后的免疫应答，从而成为我们疫苗的重要分子佐剂。本工作建立了优化IL-12家族细胞因子（IL-12、IL-23、IL-27、IL-35）表达的方法。DNA传递后IL-27的有效表达证明了IL-2在消除小鼠神经母细胞瘤转移中的协同作用。

We have used the previously developed technologies of RNA optimization to optimize expression of IL-15 cytokine, and have shown that we can over-produce bioactive cytokine after DNA delivery in mice and macaques. An important conclusion this period is that DNA delivery of vectors expressing heterodimeric IL-15 leads to systemically active levels of cytokine and the increased proliferation of NK and T cells. Therefore, DNA injection is now a practical method for the delivery of IL-15 in humans. We explored the biology of IL-15 and showed that efficient production of IL-15 is possible only by co-expression in the same cell with the so-called IL-15 Receptor-alpha. We also showed that a second form of IL-15 (SSP IL-15) previously identified in humans and rodents as intracellular or nuclear IL-15 is also efficiently secreted from the cells when co-expressed with the IL-15 Receptor alpha. These results shed new light in the biology and regulation of IL-15 and provide methods for the efficient production and clinical application of this cytokine. Cell lines overproducing soluble bioactive IL-15/IL-15 Receptor alpha heterodimers have been constructed and were used for the production of the authentic bioactive form of IL-15 found in the body. IL-15 purified from over-producing human cells was injected in mice and shown to be bioactive. Extensive studies in primates have shown that heterodimeric IL-15 has favorable pharmacokinetic and pharmacodynamic properties. The extended half life of heterodimeric IL-15 is in sharp contrast to other cytokines and is in agreement with the homeostatic role of this cytokine for the immune system. Heterodimeric IL-15 has been extensively characterized and it is heavily glycosylated. IL-15 is of interest due to its ability to stimulate the growth, activation and survival of lymphocytes, including CD8 and NK cells. Thus, IL-15 has been considered for cancer immunotherapy and for support of the growth of cytotoxic cell clones after adoptive transfer. Other proposed uses of IL-15 are in lymphopenia, in supporting NK cell growth and activation after NK transfer, and as vaccine adjuvant. We have shown that IL-15 injection accelerates the recovery of lymphocytes in mice rendered lymphopenic after treatment with cytotoxic drugs. We have used optimized expression vectors to express IL-12 cytokine in animals. Efficient expression results in bioactive levels, which increase immune response after DNA vaccination, thus becoming important molecular adjuvant for our vaccines. This work established methods to optimize expression of the IL-12 family of cytokines (IL-12, IL-23, IL-27, IL-35). Efficient expression of IL-27 after DNA delivery demonstrated synergy with IL-2 in the elimination of neuroblastoma metastases in mice.