Anhydrotetracycline (hydrochloride)

Anhydrotetracycline (hydrochloride) is a tetracycline biosynthetic precursor and a competitive broad-spectrum inhibitor of tetracycline destructase^[1]. Anhydrotetracycline (hydrochloride) is a regulator of the tetracycline repressor (TetR) and reverse tetracycline repressor (revTetR) transcriptional repressors in eukaryotic cells ^[2]. TetR is an effector-regulated DNA binding protein that binds tightly to its palindromic tetO operator DNA in the absence of effectors, thereby blocking transcription of any downstream genes ^[3]. Anhydrotetracycline binds poorly to the 30S ribosomal subunit, so it cannot act as a general translation inhibitor and is a poor antibiotic ^[4].

In vitro, treatment of NIH3T3-HER2 cells with Anhydrotetracycline (hydrochloride) (10 ng/ml) for 3 days resulted in downregulation of the HER2 gene to below the detection limit ^[5]. Treatment of N1 cells with Anhydrotetracycline (hydrochloride) (10-200nM) for 7 days significantly reduced the expression level of the Top10-driven GUS gene^[6].

In vivo, treatment of mice with NIH3T3-HER2 cell-based tumor model with Anhydrotetracycline (hydrochloride) (10mg/kg; s.c.) resulted in tumor regression of more than 95% within 7 days^[5]. Treatment of mice with NIH3T3-HER2 cell-based tumor model with Anhydrotetracycline (hydrochloride) (10mg/kg; i.p.) induced downregulation of ERBB2 mRNA and protein, leading to a rapid reduction in tumor volume^[7].

References:
[1] Markley J L, Fang L, Gasparrini A J, et al. Semisynthetic analogues of anhydrotetracycline as inhibitors of tetracycline destructase enzymes[J]. ACS infectious diseases, 2019, 5(4): 618-633.
[2] Gossen M, Bujard H. Anhydrotetracycline, a novel effector for tetracycline controlled gene expression systems in eukaryotic cells[J]. Nucleic acids research, 1993, 21(18): 4411.
[3] Resch M, Striegl H, Henssler E M, et al. A protein functional leap: how a single mutation reverses the function of the transcription regulator TetR[J]. Nucleic acids research, 2008, 36(13): 4390-4401.
[4] Rasmussen B, Noller H F, Daubresse G, et al. Molecular basis of tetracycline action: identification of analogs whose primary target is not the bacterial ribosome[J]. Antimicrobial agents and chemotherapy, 1991, 35(11): 2306-2311.
[5] Eger K, Hermes M, Uhlemann K, et al. 4-Epidoxycycline: an alternative to doxycycline to control gene expression in conditional mouse models[J]. Biochemical and biophysical research communications, 2004, 323(3): 979-986.
[6] Love J, Allen G C, Gatz C, et al. Differential Top10 promoter regulation by six tetracycline analogues in plant cells[J]. Journal of experimental botany, 2002, 53(376): 1871-1877.
[7] Hermes M, Schormann W, Brulport M, et al. Trastuzumab therapy vs tetracycline controlled ERBB2 downregulation: influence on tumour development in an ERBB2-dependent mouse tumour model[J]. British Journal of Cancer, 2008, 98(9): 1525-1532.

Anhydrotetracycline (hydrochloride)是一种四环素生物合成前体，是竞争性四环素破坏酶广谱抑制剂^[1]。Anhydrotetracycline (hydrochloride)是真核细胞中四环素阻遏蛋白（TetR）和反向四环素阻遏蛋白（revTetR）转录抑制因子的调节因子^[2]。TetR是一种效应子调节的DNA结合蛋白，在没有效应子的情况下与其回文tetO操纵子DNA紧密结合，从而阻断任何下游基因的转录^[3]。Anhydrotetracycline (hydrochloride)与30S核糖体亚基的结合较差， 因此它不能作为一般的翻译抑制剂，并且是一种较差的抗生素^[4]。

在体外，Anhydrotetracycline (hydrochloride)（10 ng/ml）处理NIH3T3-HER2细胞3天，导致了HER2基因下调至检测限以下^[5]。Anhydrotetracycline (hydrochloride)（10-200nM）处理N1细胞7天，显著降低了Top10驱动的GUS基因的表达水平^[6]。

在体内，Anhydrotetracycline (hydrochloride)（10mg/kg；s.c.）治疗基于NIH3T3-HER2细胞的肿瘤模型小鼠，在7天内使肿瘤消退了95%以上^[5]。Anhydrotetracycline (hydrochloride)（10mg/kg；i.p.）治疗基于NIH3T3-HER2细胞的肿瘤模型小鼠，诱导了ERBB2 mRNA和蛋白下调，导致肿瘤体积迅速缩小^[7]。