Cattedra di Oncologia Medica, Dipartimento di Endocrinologia e Oncologia Molecolare e Clinica, Università degli studi di Napoli "Federico II", Naples, Italy (E-mail: fortunatociardiello@yahoo.com)
Emerging novel strategies of cancer treatment are based on the selective down-regulation of specific molecular targets involved in the process of neoplastic development and progression [1]. Antisense nucleic acids that bind to specific mRNAs have been shown, in cell cultures and in preclinical models, to be effective agents for interfering with the endogenous expression of several mammalian genes [2]. Increasing experimental evidence has been provided to support the view that antisense-induced reduction in the expression of genes directly involved in cancer development and progression could inhibit cancer cell growth [34]. Antisense oligonucleotides are short synthetic sequences of DNA or RNA capable of hybridising specifically to the mRNA of a chosen target gene [3]. Antisense oligonucleotides bind to the target mRNA by WatsonCrick base pairing, resulting in the inhibition of mRNA processing or translation into protein by different mechanisms, such as prevention of mRNA transport, of mRNA splicing or translation blockade [3]. Cleavage of the target mRNA by RNase H is presumably the most relevant mechanism of antisense action [5]. RNase H is a ubiquitous endonuclease usually involved in DNA replication. RNase H cleaves the RNA strand of a DNARNA heteroduplex. Since antisense oligonucleotides with a natural phosphodiester backbone are subject to rapid degradation by cellular endogenous nucleases, a variety of sugar, base and/or backbone modifications have been made for the potential clinical development of antisense therapeutics [6]. Phosphorothioate (PS) analogues represent the most widely used class of antisense compounds currently being tested in clinical trials in cancer [78]. Although phosphorothioate oligonucleotides are not the ideal antisense drugs since they have a series of potentially toxic non-sequence specific side effects, such as complement activation and thrombocytopenia, some of them have shown encouraging results in early clinical trials [78]. A number of antisense oligonucleotides against different genes that code for important cellular protein involved in cancer cell signaling, proliferation and survival, including protein kinase A, protein kinase C, c-raf, c-Ha-ras, have been tested in phase I clinical trials [4, 78]. In this respect, one of the most promising antisense oligonucleotides in clinical development is an 18-mer PS-oligonucleotide targeting the human bcl-2 mRNA (G3139, GenasenseTM).
Bcl-2 is the prominent member of a family of proteins that are responsible for dysregulation of apoptosis and prevention of death in cancer cells [910]. Antiapoptotic bcl-2 family members, including bcl-xL, and proapoptotic proteins, such as BAD and BAX, interplay with each other to control the pathways leading to the release of cytochrome c from the mitochondrial membrane, the activation of caspase cascade and, finally, to the execution of apoptosis [910]. Bcl-2 overexpression and/or activation has also been correlated with resistance to chemotherapy, to radiotherapy and to development of hormone-resistant tumours [1113]. Moreover, it has been suggested that Bcl-2 overexpression results in the up-regulation of VEGF expression with increased neoangiogenesis in human cancer xenografts [14]. Therefore, Bcl-2 appears to be a relevant target for cancer therapy.
In this issue of Annals of Oncology, Rudin et al. report the results of an elegant and well conducted pilot trial on the combination of anti-Bcl-2 G3139 antisense oligonucleotide and paclitaxel in a group of 12 patients with heavily pretreated advanced small-cell lung cancer [15]. G3139 was administered as a continous i.v. infusion for 7 days (3 mg/kg/day) and paclitaxel was given as a 3 h infusion (150 mg/m2) on day 6. Therapy was given every 3 weeks. This combination was feasible and well tolerated. Although no objective responses were observed in this chemorefractory group of patients, disease stabilisation was observed in two patients, in one of which it lasted for 30 weeks. Furthermore, the authors were able to show preliminary evidence of Bcl-2 protein down-regulation by antisense treatment in the patient with stable disease.
G3139 has been evaluated in a number of phase I/II stud- ies in different malignancies, including melanoma, non-Hodgkins lymphoma (NHL) and prostate cancer, in which antitumour activity has been demonstrated [1619]. The first results of a phase I dose-escalation clinical trial evaluating subcutaneous administration of the bcl-2 antisense oligonucleotide G3139 were published as a preliminary report in 1997 [16] and as a complete study in 2000 in 21 patients with non-Hodgkins lymphoma [17]. Local inflammation at the infusion site was the most common side effect observed, the maximum-tolerated dosage was 147.2 mg/m2/day and the dose-limiting toxicity was thrombocytopenia. One complete response and two minor responses were reported. Nine patients had disease stabilisation and nine patients showed disease progression. However, reduction in Bcl-2 protein by antisense treatment was documented in only half of the evaluable patients. More recently, based on their preclinical studies showing that G3139 administration decreases Bcl-2 protein, enhanced tumour cell apoptosis, and in combination with systemically administered dacarbazine leads to major tumour responses in models of human melanoma xenografts in mice [11], Jansen et al. have conducted a phase I/II clinical trial on the combination of G3139 and chemotherapy in 14 chemotherapy-resistant metastatic melanoma patients [18]. G3139 could be safely administered by continuous i.v. infusion in combination with full-dose dacarbazine (DTIC). This trial also demonstrated that G3139 treatment caused a down-regulation of Bcl-2 protein in the majority of the tumour biopsies and that this biological activity was associated with major clinical responses in melanoma patients. The overall survival of this group of patients exceeded 1 year. Transient thrombocytopenia at 12 mg/kg/day was dose-limiting in patients who also received full-dose DTIC treatment. On this basis, a large international multicentre phase III randomised trial is currently in progress in patients with advanced melanoma using a 5-day pretreatment regimen of G3139 administered by continuous i.v. infusion at a dose of 7 mg/kg/day, followed by DTIC at 1000 mg/m2.
Collectively, the results of the early clinical trials with the Bcl-2 antisense oligonucleotide G3139, including the study of Rudin et al. in this issue of Annals of Oncology, are an excellent example of the possibility of translating the knowledge on molecular mechanisms underlying cancer development and progression into the production of selective anticancer drugs that could be useful in a clinical setting. In this respect, the preclinical results and the early clinical data on the potentiation of the antitumour activity of conventional cyotoxic agents even in heavily pretreated and largely chemoresistant advanced cancer patients are particularly promising. In fact, the enhancement of the antitumour efficacy of conventional cytoxic treatments by interfering with Bcl-2 activation may have relevant clinical implications. Treatment with conventional doses of cytotoxic drugs in combination with inhibitors of key signals involved in the control of cell proliferation and apoptosis has been proposed as an effective and rational anticancer treatment that is less toxic and more tolerable than other clinical approaches for increasing the activity of cytotoxic drugs, such as high-dose chemotherapy [20]. Further carefully designed early clinical trials and large phase III studies, such as the ongoing trial on DTIC in combination with G3139 in advanced melanoma patients, will allow researchers to define in a reasonable period of time the role of anti-Bcl-2 therapy in the medical treatment of cancer.
F. Ciardiello & G. Tortora
Cattedra di Oncologia Medica, Dipartimento di Endocrinologia e Oncologia Molecolare e Clinica, Università degli studi di Napoli "Federico II", Naples, Italy (E-mail: fortunatociardiello@yahoo.com)
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