James Merritt

Adenovirus-mediated p53 gene transfer in patients with advanced recurrent head and neck squamous cell carcinoma.

PURPOSE Standard therapies of head and neck squamous cell carcinoma (HNSCC) often cause profound morbidity and have not significantly improved survival over the last 30 years. Preclinical studies showed that adenoviral vector delivery of the wild-type p53 gene reduced tumor growth in mouse xenograft models. Our purpose was to ascertain the safety and therapeutic potential of adenoviral (Ad)-p53 in advanced HNSCC. PATIENTS AND METHODS Patients with incurable recurrent local or regionally metastatic HNSCC received multiple intratumoral injections of Ad-p53, either with or without tumor resection. Patients were monitored for adverse events and antiadenoviral antibodies, tumors were monitored for response and p53 expression, and body fluids were analyzed for Ad-p53. RESULTS Tumors of 33 patients were injected with doses of up to 1 x 10(11) plaque-forming units (pfu). No dose-limiting toxicity or serious adverse events were noted. p53 expression was detected in tumor biopsies despite antibody responses after Ad-p53 injections. Clinical efficacy could be evaluated in 17 patients with nonresectable tumors: two patients showed objective tumor regressions of greater than 50%, six patients showed stable disease for up to 3.5 months, and nine patients showed progressive disease. One resectable patient was considered a complete pathologic response. Ad-p53 was detected in blood and urine in a dose-dependent fashion, and in sputum. CONCLUSION Patients were safely injected intratumorally with Ad-p53. Objective antitumor activity was detected in several patients. The infectious Ad-p53 in body fluids was asymptomatic, and suggests that systemic or regional treatment may be tolerable. These results suggest the further investigation of Ad-p53 as a therapeutic agent for patients with HNSCC.

Adenovirus-Mediated p53 Gene Transfer in Sequence With Cisplatin to Tumors of Patients With Non–Small-Cell Lung Cancer

PURPOSE To determine the safety and tolerability of adenovirus-mediated p53 (Adp53) gene transfer in sequence with cisplatin when given by intratumor injection in patients with non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS Patients with advanced NSCLC and abnormal p53 function were enrolled onto cohorts receiving escalating dose levels of Adp53 (1 x 10(6) to 1 x 10(11) plaque-forming units [PFU]). Patients were administered intravenous cisplatin 80 mg/m(2) on day 1 and study vector on day 4 for a total of up to six courses (28 days per course). Apoptosis was determined by the terminal deoxynucleotidyl- transferase-dUTP nick-end labeling assay. Evidence of vector-specific sequences were determined using reverse-transcriptase polymerase chain reaction. Vector dissemination and biodistribution was monitored using a series of assays (cytopathic effects assay, Ad5 hexon enzyme-linked immunosorbent assay, vector-specific polymerase chain reaction assay, and antibody response assay). RESULTS Twenty-four patients (median age, 64 years) received a total of 83 intratumor injections with Adp53. The maximum dose administered was 1 x 10(11) PFU per dose. Transient fever related to Adp53 injection developed in eight of 24 patients. Seventeen patients achieved a best clinical response of stable disease, two patients achieved a partial response, four patients had progressive disease, and one patient was not assessable. A mean apoptotic index between baseline and follow-up measurements increased from 0.010 to 0.044 (P =.011). Intratumor transgene mRNA was identified in 43% of assessable patients. CONCLUSION Intratumoral injection with Adp53 in combination with cisplatin is well tolerated, and there is evidence of clinical activity.

Repeated intravesical instillations of an adenoviral vector in patients with locally advanced bladder cancer: A phase I study of p53 gene therapy

PURPOSE We investigated the feasibility, safety, and biologic activity of adenovirus-mediated p53 gene transfer in patients with locally advanced bladder cancer. PATIENTS AND METHODS Patients with measurable, locally advanced transitional-cell carcinoma of the bladder who were not candidates for cystectomy were eligible. On a 28-day cycle, intravesical instillations of INGN 201 (Ad5CMV-p53) were administered on days 1 and 4 at three dose levels (10(10) particles to 10(12) particles) or on either 4 or 8 consecutive days at a single dose level (10(12) particles). RESULTS Thirteen patients received a total of 22 courses without dose-limiting toxicity. Specific transgene expression was detected by reverse transcriptase polymerase chain reaction in bladder biopsy tissue from two of seven assessable patients. There were no changes in p53, p21waf1/cip1, or bax protein levels in bladder epithelium evident from immunohistochemical analysis of 11 assessable patients. Outpatient administration of multiple courses was feasible and well tolerated. A patient with advanced superficial bladder cancer showed evidence of tumor response. CONCLUSION Intravesical instillation of Ad5CMV-p53 is safe, feasible, and biologically active when administered in multiple doses to patients with bladder cancer. Observations from this study indicate that this treatment has an antitumor effect in superficial transitional-cell carcinoma. Improvements in the efficiency of gene transfer and the levels of gene expression are required to develop more effective gene therapy for bladder cancer.

Clinical evaluation of adenoviral-mediated p53 gene transfer: review of INGN 201 studies.

Gene therapy represents a new modality of cancer therapeutics based on novel mechanisms of action. Modified adenoviruses have important properties that lend themselves readily to commercial-scale manufacturing; with an excellent safety record, they have been used as the gene transfer vector in most clinical studies to date. They provide a potent means of delivering genes into cancer cells. Although multiple genes are involved in the development of malignancy, preclinical models (and now clinical studies) have proven that insertion of a single gene, p53, can arrest cancer cell growth and induce apoptosis and tumor regression in advanced cancers such as squamous cell cancer of the head and neck and non-small cell lung cancer. With equally strong rationale, clinical studies using combination approaches also have been initiated, with promising results in localized inoperable non-small cell lung cancer. In addition, the apparent safety, and especially the lack of adverse effects on normal tissues at injection sites following gene transfer, have stimulated an evaluation of intervention in the postoperative surgical adjuvant setting and in treating premalignancies. The goal of these studies has been to provide an antitumor locoregional effect. This may result in effective palliation in the advanced-disease setting, especially for diseases that do not metastasize widely. However, the most exciting near-term potential of p53 gene transfer using INGN 201 will be in up-front regimens in combination with surgery, radiation, and chemotherapy in the many clinical settings where local disease control remains suboptimal.

Cellular and humoral immune responses to adenovirus and p53 protein antigens in patients following intratumoral injection of an adenovirus vector expressing wild-type p53 (Ad-p53)

The immune responses of 10 patients with advanced non-small cell lung cancer receiving monthly intratumoral injections of a recombinant adenovirus containing human wild-type p53 (Ad-p53) to adenovirus and transgene antigens were studied. The predominate cellular and humoral immune responses as measured by lymphocyte proliferation and neutralizing antibody (Ab) formation were to adenovirus serotype 5 vector antigens, with increased responses in posttreatment samples. Consistent alterations in posttreatment cellular and humoral immune responses to p53 epitopes were not observed, and cytotoxic Abs to human lung cancer cells were not generated. Patients in this study had evidence of an antitumoral effect of this treatment with prolonged tumor stability or regression; however, neither Abs to p53 protein nor increased lymphocyte proliferative responses to wild-type or mutant p53 peptides have been consistently detected.

Evidence that transfer of functional p53 protein results in increased apoptosis in prostate cancer

Purpose: INGN 201 (Ad-p53) is a replication-defective adenoviral vector that encodes a wild-type p53 gene driven by the cytomegalovirus promoter. INGN 201 has been shown to have antitumoral activity against human prostate cancer cell lines. This study was undertaken to determine the safety of INGN 201 in patients with locally advanced prostate cancer, to assess transgene expression, and to evaluate antitumoral activity. Experimental Design: Our study included patients with clinical stage T3, T1c-T2a with Gleason score 8–10 disease, or T2a-T2b with Gleason score 7 disease and a prostate-specific antigen level >10 ng/ml. INGN 201 was administered by intraprostatic injection under ultrasonographic guidance. One course of INGN 201 was defined as three separate INGN 201 administrations 2 weeks apart. Biopsies at baseline and 24 h after the first administration were assessed for p53 protein by immunohistochemical staining and for apoptosis by terminal deoxynucleotidyl transferase-mediated nick end labeling assay. Results: A total of 38 courses of INGN 201 gene therapy were administered to 30 patients, of whom 26 underwent radical prostatectomy. There were no grade 3 or 4 adverse events related to INGN 201 administration. Of the 11 patients with negative baseline immunostaining for p53 protein, 10 had positive p53 immunostaining after the first administration of INGN 201, and 8 had an increase in apoptotic cells by terminal deoxynucleotidyl transferase-mediated nick end labeling staining. All 26 of the patients who underwent radical prostatectomy had significant residual viable prostate cancer, and 12 have experienced biochemical failure (median follow-up, 42 months). Conclusion: Intraprostatic INGN 201 gene therapy is safe and can reliably result in p53 protein production and apoptosis.

Phase I, Pharmacokinetic, and Pharmacodynamic Study of Intravenously Administered Ad5CMV-p53, an Adenoviral Vector Containing the Wild-Type p53 Gene, in Patients With Advanced Cancer

PURPOSE The purpose of this study was to assess the feasibility of administering Ad5CMV-p53, an adenoviral vector containing the wild-type p53 gene to patients with advanced malignancies, characterize the pertinent pharmacokinetic parameters, identify evidence of viral uptake in both normal and tumor tissue, and seek evidence of antitumor activity. METHODS Patients were treated with escalating doses of Ad5CMV-p53 intravenously over 30 minutes on days 1, 2, and 3, every 28 days. The clearance of circulating Ad5CMV-p53 (INGN 201) DNA was characterized in the plasma and paired tumor and skin biopsies were performed in patients treated at the two highest dose levels to assess vector uptake into tissues. RESULTS Seventeen patients received 36 courses of Ad5CMV-p53 at doses ranging from 3 x 10(10) to 3 x 10(12) virus particles (vp). Fatigue, nausea, vomiting, and fever were common, but rarely severe. Abnormalities of coagulation parameters, including decreases in fibrinogen and increases in fibrin degradation products at 3 x 10(12)vp, precluded additional dose escalation. Ad5CMV-p53 DNA could be detected in the plasma by polymerase chain reaction assay in the majority of patients at 14 days and 28 days at doses of 3 x 10(10) and higher. Six patients treated at 1 x 10(12)vp and 3 x 10(12)vp dose levels had Ad5CMV-p53 DNA detected within paired tumor tissue collected day 4. CONCLUSION Ad5CMV-p53 can be safely and repetitively administered up to 1 x 10(12)vp intravenously daily for 3 consecutive days. The absence of severe toxicities, the presence of circulating adenovirus 24 hours after administration, and detectable p53 transgene within tumor tissue distant from the site of administration demonstrates that systemic therapy with this adenoviral vector containing p53 is feasible.

Phase I trial of interferon-γ (IFN-γ) retroviral vector administered intratumorally to patients with metastatic melanoma

Background: Interferon-γ (IFN-γ) gene/retroviral vector cell vaccinations have generated protective responses from unmodified tumor cell challenges as well as a regression of established tumors in animal models. The purpose of this trial was to determine the feasibility and safety of a direct intratumoral injection of IFN-γ retroviral vector in advanced melanoma patients.Methods: This was a phase I study, in which 13 patients received a single daily injection of a retroviral vector with the IFN-γ gene for 5 consecutive days (1.5 × 108 colony-forming units total dose); patients subsequently underwent resection of the injected lesion to confirm DNA transduction in situ.Results: No toxicity related to the injected vector was observed. Replication competent retrovirus was not observed in any prepared samples (n = 65). IFN-γ expression was confirmed in 3 of 10 harvested tumor samples; one was equivocal, and DNA transduction was unable to be confirmed by enzyme-linked immunospot assay in six samples.Conclusions: An injection of IFN-γ gene/retroviral vector is well tolerated. DNA transduction was demonstrated in human subjects, confirming the feasibility of the direct injection approach for the gene therapy of solid tumors. Further trials to determine optimal schedule and potential efficacy are indicated.

Gene therapy approaches for the management of non-small cell lung cancer

Targeting the specific genetic lesions responsible for carcinogenesis and cancer progression is an attractive strategy for developing more effective anticancer therapies and reducing treatment-related toxicity. The restoration of defective tumor suppressor gene pathways by replacement of tumor suppressor genes in cancer cells has been studied in lung cancer. The most extensively studied agent is the wild-type p53 tumor suppressor gene delivered by an adenoviral vector. Clinical trials to date in non-small cell lung cancer and head and neck cancer have consistently shown evidence of gene transduction and expression, mediation of apoptosis, and clinical responses including pathologic complete responses. However, it also is clear that this approach can be improved. Promising avenues for investigation include improved gene delivery systems, induction of bystander effects, and adjuvant use of gene therapy with conventional chemotherapy, radiation therapy, and surgery. However, these strategies will need further refinement to succeed clinically. This review examines several important issues in cancer gene therapy in general and the most recent achievements in gene therapy for non-small cell lung cancer.

p53 Gene Replacement for Cancer&Interactions with DNA Damaging Agents

Clinical trials of p53 gene replacement have provided information that will be useful in the design of future gene therapy strategies. Direct intratumor injection has low toxicity and thus can be readily combined with existing treatments. Post-injection gene expression can be documented and occurs in the presence of an anti-adenovirus immune response. Importantly, this treatment can cause tumor regression or prolonged stabilization. Future research directions will include development of more efficient vectors, use of novel genes, and combined modality approaches. Unresectable tumors are a prominent problem in oncology, with proven therapies such as radiotherapy and chemotherapy controlling less than 20% of lung cancers. Based on the preclinical and clinical studies discussed, it now seems that these conventional therapies may provide renewed potential when used in conjunction with transfer of a functional p53 gene.Clinical trials of p53 gene replacement have provided information that will be useful in the design of future gene therapy strategies. Direct intratumor injection has low toxicity and thus can be readily combined with existing treatments. Post-injection gene expression can be documented and occurs in the presence of an anti-adenovirus immune response. Importantly, this treatment can cause tumor regression or prolonged stabilization. Future research directions will include development of more efficient vectors, use of novel genes, and combined modality approaches. Unresectable tumors are a prominent problem in oncology, with proven therapies such as radiotherapy and chemotherapy controlling less than 20% of lung cancers. Based on the preclinical and clinical studies discussed, it now seems that these conventional therapies may provide renewed potential when used in conjunction with transfer of a functional p53 gene.