The first clinical use of a live-attenuated Listeria monocytogenes vaccine: A Phase I safety study of Lm-LLO-E7 in patients with advanced carcinoma of the cervix
Introduction
Invasive carcinoma of the cervix (ICC) is the second most common cancer in women worldwide with over 450,000 new cases and 230,000 deaths annually, most of them occurring in developing countries [1]. Persistent infection with high-oncogenic risk human papillomavirus (HR-HPV) types is recognized as a necessary, but not sufficient, cause of cervical cancer [2], [3], [4]. HPVs 16 and 18 are the most prevalent types in malignant lesions, accounting for over 70% of ICC and over 50% of high-grade precursor lesions [5]. Although the advent of prophylactic HPV vaccines may have a significant impact on the incidence of ICC in the future, the worldwide implementation of such vaccines remains a challenge, particularly in developing countries [6]. Additionally, these vaccines are not intended to treat pre-existing HPV infections and associated malignancies, which require therapeutic vaccines, mostly targeting the E6 and E7 HPV oncoproteins [7].
Listeria monocytogenes (Lm) is a food-borne gram-positive bacterium that can occasionally cause disease in humans. Listeriosis is an uncommon infection, primarily affecting elderly individuals, newborns, pregnant women and immunocompromised individuals [8]. In addition to strongly activating innate immunity and inducing a cytokine response that enhances antigen-presenting cell (APC) function, Lm has the ability to replicate in the cytosol of APCs after escaping from the phagolysosome, which requires the virulence factor listeriolysin O (LLO) protein [9], [10]. This unique intracellular life cycle allows antigens secreted by Lm to be processed and presented in the context of both MHC class I and II molecules, resulting in potent cytotoxic CD8+ and Th1 CD4+ T-cell-mediated immune responses [10], [11].
Lm has been extensively investigated as a vector for cancer immunotherapy in pre-clinical models [12], [13], [14], [15], [16]. Lm-LLO-E7 is a recombinant live-attenuated Lm that secretes the antigen HPV-16 E7 fused to a non-hemolytic LLO [16]. Previous studies have shown that genetically fusing an antigen to LLO enhances the immunogenicity of tumor-associated antigens, resulting in a better therapeutic efficacy against established tumors [15], [16], [17].
Immunization of mice with Lm-LLO-E7 induces regression of established tumors expressing E7 and confers long-term protection [16]. Moreover, it is able to overcome immunological tolerance and impair the development and severity of autochthonous tumors in an E7 transgenic mouse model [18]. The therapeutic efficacy of Lm-LLO-E7 correlates with its ability to induce E7-specific tumor-infiltrating CTLs, mature dendritic cells, reduce the number of intratumoral regulatory CD4+ CD25+ T cells and inhibit tumor angiogenesis [19].
Lm has also a number of inherent advantages as a vaccine vector. The bacterium grows very efficiently in vitro without special requirements and it lacks LPS, which is a major toxicity factor in gram-negative bacteria, such as Salmonella[20]. Genetically attenuated Lm vectors also offer additional safety as they can be readily eliminated with antibiotics, in case of serious adverse effects and unlike some viral vectors, no integration of genetic material into the host genome occurs. Although the potential of Lm as a vaccine vector has been proven in pre-clinical studies, the feasibility of using live-attenuated Lm vectors as a cancer immunotherapeutic in humans has not been demonstrated yet. In a previous study, healthy volunteers were given orally escalating doses of an actA/plcB-deleted Lm strain without serious adverse effects or long-term sequelae [21]. However, no previous studies with Lm vectors have been done in cancer patients, who are the target population for Lm-LLO-E7 clinical use. Moreover, the highly attenuated Lm-LLO-E7 vector is given intravenously. We report herein the first clinical use and safety of a live-attenuated Lm vector to treat patients with advanced ICC, who had failed prior chemotherapy, radiotherapy, and/or surgery.
Section snippets
Construction of the Lm-LLO-E7 vaccine
Construction of Lm-LLO-E7 was previously described [16]. Briefly, the prfA-defective Lm strain XFL-7, which is derived from the streptomycin-resistant wild-type Lm 10403S strain [22], was transformed with the multicopy plasmid pGG55. This plasmid contains an expression cassette with the HPV-16 E7 gene fused to a truncated hly gene that encodes the first 441 residues of LLO. Additionally, pGG55 contains a mutated copy of the prfA gene that partially restores the virulence of XFL-7 and it is
Pre-clinical toxicity evaluation of Lm-LLO-E7
The MTD for IV administration of Lm-LLO-E7 in BALB/c female mice was 2.8 × 107 CFU. In the group that received 2.8 × 109 CFU, splenomegaly was observed in 2 out of 5 mice on day 7 after injection and it was defined as an AE. No deaths occurred at the doses tested. A dose of 2.8 × 107 CFU corresponds to a human dose of 7.8 × 1010 by body mass or 7.5 × 109 by surface area, a range that is consistent with the clinical doses used in this clinical trial. Importantly, an IV dose of 2.8 × 107 CFU of Lm-LLO-E7 was
Discussion
To our knowledge, this is the first clinical report demonstrating safety and feasibility on using a live-attenuated Lm vaccine in patients with cancer. Intravenous administration of Lm-LLO-E7 was consistently associated with a flu-like syndrome in all patients, characterized by pyrexia, chills, headache, tachycardia, hypotension, nausea and vomiting. These AE were acute and transient in most patients, resolving in the first 12 h after infusion. While well tolerated in the lower two dosage
Acknowledgments
The authors wish to acknowledge Dr. Yvonne Paterson from the University of Pennsylvania for her insight and guidance in the use of live Listeria monocytogenes vaccines, and Drs. Benigno Figueroa and Jorge Herrera-Rodriguez for their clinical work in Mexico. Dr. Pavle Vukojevc and Ms. Jelena Tasic for their essential work in the oversight of this clinical trial. This study was funded by Advaxis, Inc.
References (41)
Global cancer statistics in the year 2000
Lancet Oncol
(2001)- et al.
Listeriolysin O: a phagosome-specific lysin
Microbes Infect/Institut Pasteur
(2007) - et al.
Cisplatin nephrotoxicity: mechanisms and renoprotective strategies
Kidney Int
(2008) - et al.
Carboplatin versus cisplatin in solid tumors: an analysis of the literature
Ann Oncol
(1998) - et al.
Topotecan in squamous cell carcinoma of the cervix: a phase II study of the gynecologic oncology group
Gynecol Oncol
(2000) - et al.
HPV16/18 E7-pulsed dendritic cell vaccination in cervical cancer patients with recurrent disease refractory to standard treatment modalities
Gynecol Oncol
(2006) - et al.(1992)
- et al.
Human papillomavirus is a necessary cause of invasive cervical cancer worldwide
J Pathol
(1999) - et al.
Chapter 1: HPV in the etiology of human cancer
Vaccine
(2006) - et al.
Chapter 3: HPV type-distribution in women with and without cervical neoplastic diseases
Vaccine
(2006)
Initial lessons learned in HPV vaccination
Gynecol Oncol
Therapeutic human papillomavirus vaccines: current clinical trials and future directions
Expert Opin Biol Ther
Foodborne listeriosis
Clin Infect Dis
Immune responses to Listeria monocytogenes
Nature Rev Immunol
Delivery of a viral antigen to the class I processing and presentation pathway by Listeria monocytogenes
J Exp Med
Cancer immunotherapy targeting the high molecular weight melanoma-associated antigen protein results in a broad antitumor response and reduction of pericytes in the tumor vasculature
Cancer Res
Development of a Listeria monocytogenes based vaccine against prostate cancer
Cancer Immunol Immunother
Listeria-based vaccines for cancer treatment
Curr Opin Mol Ther
Fusion to Listeriolysin O and delivery by Listeria monocytogenes enhances the immunogenicity of HER-2/neu and reveals subdominant epitopes in the FVB/N mouse
J Immunol
Two Listeria monocytogenes vaccine vectors that express different molecular forms of human papilloma virus-16 (HPV-16) E7 induce qualitatively different T cell immunity that correlates with their ability to induce regression of established tumors immortalized by HPV-16
J Immunol
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