Meister B, Fink F M, Hittmair A, Marth C, Widschwendter M
Department of Pediatrics, University of Innsbruck, Austria.
Anticancer Res. 1998 May-Jun;18(3A):1777-86.
Retinoids modulate several cell functions and especially inhibit the growth of tumor cells. Their biological activity is mediated by retinoic acid receptors (RARs), of which three subtypes (alpha, beta, gamma) have been identified. In human neuroblastoma (NB) reduced endogenous RAR-gamma expression was suggested to diminish the sensitivity for retinoids, to promote proliferation, and to contribute to the malignant phenotype. To correlate receptor selectivity with in vitro activity, we analysed the effect of six synthetic retinoids with selectivity for human RAR-alpha/beta/gamma on the human LAN-5 NB cell line and compared it with the natural compound all-trans-retinoic acid (ATRA). Apoptosis was determined by flow-cytometry using terminal-deoxynucleotidyl transferase to end-label DNA fragments in situ in apoptotic cells. The antagonist for RAR-beta/gamma CD2665 as well as the selective agonists for RAR-alpha CD336 and RAR-beta CD2019 were less effective in growth inhibition than ATRA. In contrast, the synthetic RAR-gamma selective agonists CD437 and CD2325 induced a concentration- and time-dependent antiproliferative effect, which was similar or even more pronounced than ATRA. In contrast to ATRA, the adition of CD437 and CD2325 did not induce morphological changes typical of NB cell maturation but resulted in morphological features consistent with the occurrence of programmed cell death. Flow-cytometric analysis showed that in contrast to ATRA the addition of CD 437 and CD 2325 results in progressive time-dependent increase of apoptotic cells (25.9% and 57.7% after 72 hours). In conclusion, our study demonstrates RAR-gamma selectively binding retinoids dramatically suppress NB cell growth, primarily by inducing programmed cell death rather than by cell differentiation. Since advanced or disseminated NB tumors endogenously express low levels of RAR-gamma and lack of apoptosis is involved in tumor progression, RAR-gamma selectively binding retinoids may be more appropriate retinoids for clinical trials in NB.
维甲酸可调节多种细胞功能，尤其能抑制肿瘤细胞的生长。其生物活性由维甲酸受体（RARs）介导，已鉴定出三种亚型（α、β、γ）。在人类神经母细胞瘤（NB）中，内源性RAR-γ表达降低被认为会降低对维甲酸的敏感性，促进增殖，并导致恶性表型。为了将受体选择性与体外活性相关联，我们分析了六种对人类RAR-α/β/γ具有选择性的合成维甲酸对人类LAN-5 NB细胞系的影响，并将其与天然化合物全反式维甲酸（ATRA）进行比较。通过使用末端脱氧核苷酸转移酶原位标记凋亡细胞中的DNA片段，采用流式细胞术测定凋亡情况。RAR-β/γ拮抗剂CD2665以及RAR-α选择性激动剂CD336和RAR-β选择性激动剂CD2019在生长抑制方面比ATRA效果更差。相比之下，合成的RAR-γ选择性激动剂CD437和CD2325诱导了浓度和时间依赖性的抗增殖作用，其作用与ATRA相似甚至更显著。与ATRA不同，添加CD437和CD2325并未诱导NB细胞成熟的典型形态变化，而是导致了与程序性细胞死亡发生一致的形态特征。流式细胞术分析表明，与ATRA不同，添加CD437和CD2325会导致凋亡细胞随时间逐渐增加（72小时后分别为25.9%和57.7%）。总之，我们的研究表明，选择性结合维甲酸的RAR-γ主要通过诱导程序性细胞死亡而非细胞分化来显著抑制NB细胞生长。由于晚期或播散性NB肿瘤内源性表达低水平的RAR-γ，且肿瘤进展涉及凋亡缺失，选择性结合维甲酸的RAR-γ可能是NB临床试验中更合适的维甲酸。
