Goebel E A, Davidson B L, Graham S M, Kern J A
Otolaryngol Head Neck Surg. 1998 Oct;119(4):331-6. doi: 10.1016/S0194-5998(98)70073-7.
Gene transfer offers the possibility of novel therapies for head and neck squamous cell carcinoma (HNSCC). To this end, we demonstrate that a replication deficient adenovirus vector (Ad.RSVlacZ) can efficiently transduce foreign genes into human HNSCC cell lines in vitro, and that adenoviral mediated transfer of herpes simplex virus thymidine kinase (Ad.RSVtk) followed by exposure to ganciclovir results in tumor cell killing in vitro and in vivo. Exposure to Ad.RSVlacZ resulted in lacZ expression at multiplicities of infection (MOIs) of 10 and 100 for the cell lines HEp-2 and FaDu, respectively. This increased to 97% (HEp-2) and 49% (FaDu) at an MOI of 10,000. For HEp-2, maximum expression occurred during the first 48 hours after exposure (52% at 24 hours, 48% at 48 hours; MOI 500), then declined by 40% per day. This rapid decline may be caused by dilution of the gene through cell proliferation, because normalizing for the increase in total protein shows that the total number of cells expressing lacZ is stable from days 1 to 4. FaDu and HEp-2 were then transduced by AD.RSVtk and exposed to 20 microM ganciclovir for 24 hours. Significant tumor cell killing, as measured by a colony forming assay, occurred at an MOI of 2 for HEp-2 and 20 for FaDu. At an MOI of 200, 100% of HEp-2 and 97% of FaDu cells were killed. Next, subcutaneous tumor nodules derived from FaDu and HEp-2 were established in the flanks of SCID mice. Direct intratumoral injection of Ad.RSVtk followed by 7 days of ganciclovir therapy resulted in an adenovirus dose dependent reduction of tumor growth, and an actual size reduction of established tumor nodules at the highest does (10(10) plaque forming units). In conclusion, an adenovirus vector can efficiently transduce HNSCC cell lines in vitro. Maximum marker gene expression occurred during the first 48 hours after transduction. Transduction by Ad.RSVtk followed by exposure to ganciclovir resulted in tumor cell killing in vitro and in vivo.
基因转移为头颈部鳞状细胞癌（HNSCC）提供了新的治疗可能性。为此，我们证明了一种复制缺陷型腺病毒载体（Ad.RSVlacZ）能够在体外有效地将外源基因转导至人HNSCC细胞系，并且腺病毒介导的单纯疱疹病毒胸苷激酶（Ad.RSVtk）转移，随后暴露于更昔洛韦，可在体外和体内导致肿瘤细胞死亡。暴露于Ad.RSVlacZ后，细胞系HEp-2和FaDu分别在感染复数（MOI）为10和100时出现lacZ表达。在MOI为10,000时，HEp-2的lacZ表达增加到97%，FaDu增加到49%。对于HEp-2，在暴露后的前48小时内出现最大表达（24小时时为52%，48小时时为48%；MOI 500），然后每天下降40%。这种快速下降可能是由于基因通过细胞增殖而稀释，因为对总蛋白增加进行归一化显示，从第1天到第4天，表达lacZ的细胞总数是稳定的。然后用AD.RSVtk转导FaDu和HEp-2，并将其暴露于20 microM更昔洛韦24小时。通过集落形成试验测量，在HEp-2的MOI为2和FaDu的MOI为20时出现显著的肿瘤细胞杀伤。在MOI为200时，100%的HEp-2细胞和97%的FaDu细胞被杀死。接下来，在SCID小鼠的侧腹建立源自FaDu和HEp-2的皮下肿瘤结节。直接瘤内注射Ad.RSVtk，随后进行7天的更昔洛韦治疗，导致肿瘤生长呈腺病毒剂量依赖性减少，并且在最高剂量（10(10) 噬斑形成单位）时，已建立的肿瘤结节实际尺寸减小。总之，腺病毒载体能够在体外有效地转导HNSCC细胞系。最大标记基因表达在转导后的前48小时内出现。Ad.RSVtk转导后暴露于更昔洛韦可在体外和体内导致肿瘤细胞死亡。
