Partial Tumor Regression Research Articles

Therapeutic Targeting of the Endothelin-A Receptor in Human Ovarian Carcinoma: Efficacy of Cytotoxic Agents is Markedly Enhanced by Co-administration with Atrasentan

The endothelin-1/endothelin-A receptor autocrine pathway is overexpressed in ovarian carcinoma. We explored the efficacy of atrasentan (ABT-627), a small orally active endothelin- A receptor antagonist, in monotherapy and combination therapy on HEY ovarian carcinoma xenografts. Atrasentan (2 mg/kg per 24 hours i.p. for 21 days) induced similar inhibition of tumor growth as paclitaxel (20 mg/kg i.v. three times a day every 4 days) with a reduction of 65% compared to control. The co-administration of atrasentan enhanced the efficacy of cytotoxic agents, such as taxanes or platinum compounds. Administration of atrasentan in combination with paclitaxel caused a strong antitumor effect. Remarkably, four of ten mice bearing HEY xenografts had no histological evidence of tumors. Tumor growth inhibition was accompanied by a significant decrease of molecular effectors involved in angiogenesis and invasion and by enhanced tumor cell apoptosis. Moreover, although cisplatinum as a single agent (5 mg/kg i.p. on day 1) markedly inhibited HEY tumors, atrasentan was very effective in potentiating this effect, with partial or complete tumor regression. The antitumor, anti-angiogenic, and apoptotic activities obtained with atrasentan and the enhanced efficacy of cytotoxic agents provide a rationale for its clinical evaluation in ovarian carcinoma.

Telomerase as a tumor-associated antigen for cancer immunotherapy

Telomerase reverse transcriptase hTERT is an attractive target for cancer immunotherapy given its broad expression in human tumors and its demonstrated immunogenicity. Human and murine model systems demonstrate that CD8(+) cytotoxic T-lymphocytes (CTL) and CD4(+) helper T-lymphocytes can recognize dominant epitopes derived from TERT. CTL kill TERT-positive tumor cells of multiple histologies, although there is some disagreement regarding the level of processing and presentation of certain TERT peptides within the context of MHC class I molecules. CTL recognizing modified, low-affinity cryptic TERT epitopes have also been generated that protect against tumor challenge in a murine model. Several phase I clinical trials testing hTERT as a cancer vaccine target have shown the induction of T-cell immune responses but minimal toxicities, including bone marrow toxicity, in patients with multiple types of cancer. Several studies report some patients experiencing clinical benefit, including partial tumor regression, providing further encouragement for hTERT as broadly applicable target for cancer immunotherapy.

21. Induction of CEA in Pulmonary Vasculature to Study Targeting Strategies for Cancer

Truly systemic gene transfer in vivo has been limited by the ability of vectors to localize to the tumor as well as immune inactivation, non-specific adhesion and loss of particles in the circulation. Toxicity can also result from an immune response to the systemic virus and transduction of other cells. We have previously shown that retroviral vectors can be carried to human xenografts by tumor targeted T cells. In immunodeficient mice we observed regression of disseminated tumors without evidence of toxicity. To address a more clinically relevant model, we have now used tumor specific T cells to deliver retroviral vectors to tumors in immunocompetent mice. We used T cells from OT-1 TCR transgenic mice, whose T cell receptor recognizes the SIINFEKL peptide epitope from chicken ovalbumin, ova, presented in the context of H2Db by B16 cells that stably express ova (B16ova). We have previously shown that the adoptive transfer of OT-1 T cells in vivo results in the partial regression of B16ova tumors. We have developed two methods to engineer OT-1 T cells to deliver retroviral vectors. In the first method, OT-1 T cells were generated to produce retrovirus by transducing them with an HSV-derived amplicon vector encoding the gag, pol and env genes, and a retroviral vector encoding the beta-galactosidase gene. In the second method, OT-1 T cells were incubated with retroviruses that can adsorb to the cell surface whilst entering the cell at very low efficiency. We have demonstrated that OT-1 T cells that have retrovirus adsorbed to their surface will hand this virus off to target cells in culture. For example, 5 × 105 OT-1 T cells loaded with retroviral stocks at an MOI of about 100, washed and then co-cultured with B16 cells resulted in transduction of target B16 cells at a level of 2.7 × 103 c.f.u. T cell hand off-mediated target cell infection is strictly dependent upon the presence of a functional envelope and is not the result of carry over of virus in the media. Handoff is more efficient when the viral envelope utilizes a receptor that is poorly, or not, expressed on the T cell itself. OT-1 T cells coated with retrovirus can both kill and transfer virus into B16ova cells. This transfer does not entirely depend upon the presence of an intact envelope on the viral particle surface.

Potentiation of oncolytic adenoviral vector efficacy with gutless vectors encoding GMCSF or TRAIL.

Oncolytic adenoviral vectors selectively replicate in and lyse human tumor cells, providing a promising means for targeted tumor destruction. However, oncolytic vectors have limited capacity for incorporation of additional genetic material that could encode therapeutic transgenes and/or transcriptional regulatory control elements to augment the efficacy and/or safety of the vector. Therefore, we hypothesized that coadministration of an oncolytic vector with a replication-defective, gutless adenoviral vector encoding a therapeutic transgene would result in replication of both vectors within a tumor and potentiate antitumor efficacy relative to the use of either vector alone. We constructed gutless vectors encoding the murine granulocyte-macrophage colony-stimulating factor (AGVmGMF) or human tumor necrosis factor alpha-related apoptosis-inducing ligand (AGVhTRAIL) gene and tested the ability of these vectors to augment the efficacy of an oncolytic vector (Ar6pAE2fE3F) in a potentiating vector strategy. In Hep3B cells in vitro, cotreatment with Ar6pAE2fE3F increased transgene expression from AGVhTRAIL and permitted replication of AGVhTRAIL, suggesting that an oncolytic vector can propagate gutless vector spread in vivo. In pre-established Hep3B xenograft tumors, neither gutless vector alone inhibited tumor growth; however, coadministration of AGVmGMF or AGVhTRAIL with Ar6pAE2fE3F significantly reduced tumor growth relative to Ar6pAE2fE3F alone. Additionally, use of AGVhTRAIL with Ar6pAE2fE3F increased the number of complete or partial tumor regressions observed at study end. These data provide evidence that coadministration of an oncolytic vector with a gutless vector holds promise for potentiating tumor ablation efficacy.

90 Hepatocyte GP130/STAT activation results in liver protection in immune medi-ated hepatitis

tal HCCs (Hepa 1-6 cells in C57L/J mice). However, tumor vaccination strategies might be limited by low levels of tumor infiltration by effector Tlymphocytes. In the present study we therefore analyzed, if intatumoral cytokine/chemokine expression may enhance the efficiency of mAFP-specific DNA-vaccination in a highly aggressive primary murine HCC model. Materials and Methods: Subcutaneous HCC tumors expressing AFP (BW7756) were generated in C57L/J mice by transplantation of tumor pieces. DNA-vaccination was performed at days 4 and 18 by intramuscular injection of expression vectors encoding mAFP, mIL-12 and mGM-CSF. 9 days after DNA-vaccination tumors were injected with adenoviral vectors expressing mIL-12 (Ad-IL-12) and IP-10 (Ad IP-10) or b-galactosidase (Ad-b-gal) as control. Results and Discussion: DNA-vaccination alone, tumor injections with Ad-b-gal and the combination of DNA-vaccination with Ad-b-gal did not show therapeutic efficiency in mice bearing aggressive BW7756 HCCs. Intratumoral adenoviral mIL-12+IP-10 expression resulted in partial tumor regressions, but did not significantly improve animal survival. In contrast, combination of AFP-specific DNA-vaccination with intratumoral Ad-IL12 and Ad IP-10 injection resulted in tumor regressions in all treated animals and prolonged animal survival. These results demonstrate for the first time, that combination of DNA-vaccination with intratumoral gene therapeutic expression of cytokines and chemokines results in improved therapeutic efficiency. This synergistic effect might be caused by enhanced infiltration of the tumor tissue with effector lymphocytes generated by the DNA-vaccination. 90 HEPATOCYTE GP130/STAT ACTIVATION RESULTS IN LIVER PROTECTION IN IMMUNE MEDI-ATED HEPATITIS

Partial regression of primary cutaneous melanoma: is there an association with sub-clinical sentinel lymph node metastasis?

Whether partial regression of a primary melanoma has an adverse impact on prognosis is controversial. As an indirect mechanism of addressing this question we drew a correlation between the histopathological characteristics of 107 cutaneous melanomas and the presence of sub-clinical metastasis in corresponding sentinel lymph nodes. Partial regression of the primary tumor, defined as focal replacement of the lesion by a scar, unrelated to a previous biopsy, was observed in 20 (19%) cases in the group as a whole. Excluding cases in which an accurate Breslow thickness of the primary melanoma could not be established and/or the presence of a capsular nevus was detected in the sentinel node, a total of 97 remained. Seventeen cases (Breslow thickness 0.63-9.7; mean 2.4 mm) showed partial regression and 80 (Breslow thickness 0.25-7.00; mean 1.8 mm) were devoid of regression. Of the 17 cases with regression 5 (29%) had nodal metastasis (by histopathology and/or molecular analysis) and of the 80 cases without regression 23 (29%) had nodal metastasis (by one or both evaluations). Our data reveals no association between partial regression of the primary melanoma and sentinel node involvement by the disease. The Breslow thickness proved to be the only significant independent variable related to nodal metastasis. Of interest, ulceration of the primary lesion was significantly associated with nodal disease on univariate, but not on multivariate, analysis. While acknowledging that the cohort size may lack the statistical power to demonstrate subtle associations, our data supports the known relevance of tumor thickness and ulceration to regional lymph node metastasis and thereby, to outcome of melanoma in its early stages, but fails to support a similar role for partial regression.

Predictive factors in locally advanced rectal cancer treated with preoperative hyperfractionated and accelerated radiotherapy

This study examines the prognostic significance of pathologic factors in patients with primary locally advanced rectal cancer treated prospectively with preoperative radiotherapy. From 1992 to 1998, 104 patients with rectal cancer of grades T3 or T4 and any N underwent preoperative radiotherapy followed by surgical resection. Survival curves were estimated according to the Kaplan-Meier method. Correlation of outcome with clinicopathologic variables (pathologic tumor and lymph node staging, histology, radial resection margin [RRM], clearance, vessel involvement, and tumor regression grade [TRG], quantitated in 5 grades) was evaluated using the Cox proportional hazards model. None of the patients achieved a histologically confirmed complete pathologic response, but 79% of the patients showed partial tumor regression (TRG2–4) and 21% did not show any tumor regression (TRG5). Among the tumors, 22% were of a mucinous type. The RRM was free of tumor in 76% of the surgical specimens. The median clearance was 2 mm. Vascular invasion was present in 37 cases (36%). In the univariate analysis, lymph node metastases, absence of tumor regression, positive RRM, and vascular invasion were correlated with adverse overall survival and disease-free survival; absence of tumor regression, positive RRM, and clearance <2 mm were correlated with local recurrences; and advanced pT stage was correlated only with disease-free survival. However, in the multivariate analysis, only lymph node metastases and RRM were independent prognostic factors for overall survival and disease-free survival, and clearance <2 mm was an independent prognostic factor for local control. Pathologic parameters remain strong determinants of local recurrence and survival in locally advanced rectal cancer, treated preoperatively with hyperfractionated and accelerated radiotherapy. We show that patients with advanced pT, positive lymph nodes, vascular invasion, positive RRM, clearance <2 mm, or absence of tumor regression are known to have poor clinical outcome.

Endoscopy in the diagnosis and therapy of inoperable esophageal carcinoma after radiotherapy and combined radiochemotherapy

Treatment of inoperable (locally advanced) esophagus cancer, even today, has been a subject of numerous clinical researches. Combination of several clinical modalities is needed, of radiotherapy and chemotherapy the most frequently. This combination applied as neoadjuvant or concomitant brought to improvement of treatment results of these patients. Local control is still a big problem, since majority of these patients die of local treatment "failure". Because of that, estimation of the local control is very important, i.e. assessment of the tumor response the treatment. For those reasons, a study has been conducted on 52 patients with locally advanced esophagus tumor, who in the first group of 25 patients, were treated only by radiotherapy, and in the second group of 27 patients, by combined radio-chemotherapy. Pretherapy evaluation, evaluation of esophagus stenosis, as well as the tumor regression to conducted treatment, were done in all patients by radiography and endoscopy and the findings were compared: there was no statistically significant difference between investigated methods (p > 0.05). Although bigger number of the tumor complete regressions was verified by radiography (6 in the first group and 5 in the second group of patients) in relation to endoscopic finding (3 in the first group and 2 in the second group of patients), which had no microscopical evidence of remaining disease. Partial tumor regression made greater differences between investigated methods (10:4 in the first group and 6:3 in the second group of patients).

Synergistic antitumor effect of bispecific CD19 x CD3 and CD19 x CD16 diabodies in a preclinical model of non-Hodgkin's lymphoma.

To target NK cells against non-Hodgkin's lymphoma, we constructed a bispecific diabody (BsDb) with reactivity against both human CD19 and FcgammaRIII (CD16). Bacterially produced CD19 x CD16 BsDb specifically interacted with both CD19(+) and CD16(+) cells and exhibited significantly higher apparent affinity and slower dissociation from the tumor cells than from effector cells. It was able to induce specific lysis of tumor cells in the presence of isolated human NK cells or nonfractionated PBLs. The combination of the CD19 x CD16 BsDb with a previously described CD19 x CD3 BsDb and CD28 costimulation significantly increased the lytic potential of human PBLs. Treatment of SCID mice bearing an established Burkitt's lymphoma (5 mm in diameter) with human PBLs, CD19 x CD16 BsDb, CD19 x CD3 BsDb, and anti-CD28 mAb resulted in the complete elimination of tumors in 80% of animals. In contrast, mice receiving human PBLs in combination with either diabody alone showed only partial tumor regression. These data clearly demonstrate the synergistic effect of small recombinant bispecific molecules recruiting different populations of human effector cells to the same tumor target.

Cancer immunotherapy with peptide-based vaccines: what have we achieved? Where are we going?

Many human tumor-associated antigens (TAAs) have recently been identified and molecularly characterized. When bound to major histocompatibility complex molecules, TAA peptides are recognized by T cells. Clinical studies have therefore been initiated to assess the therapeutic potential of active immunization or vaccination with TAA peptides in patients with metastatic cancer. So far, only a limited number of TAA peptides, mostly those recognized by CD8(+) T cells in melanoma patients, have been clinically tested. In some clinical trials, partial or complete tumor regression was observed in approximately 10%-30% of patients. No serious side effects have been reported. The clinical responses, however, were often not associated with a detectable T-cell-specific antitumor immune response when patients' T cells were evaluated in ex vivo assays. In this review, we analyze the available human TAA peptides, the potential immunogenicity (i.e., the ability to trigger a tumor-specific T-cell response) of TAA peptides in vitro and ex vivo, and the potential to construct slightly modified forms of TAA peptides that have increased T-cell stimulatory activity. We discuss the available data from clinical trials of TAA peptide-based vaccination (including those that used dendritic cells to present TAA peptides), identify possible reasons for the limited clinical efficacy of these vaccines, and suggest ways to improve the clinical outcome of TAA peptide-based vaccination for cancer patients.

Importance of tumor regression assessment in predicting the outcome in patients with locally advanced rectal carcinoma who are treated with preoperative radiotherapy

BACKGROUND Locally advanced rectal carcinoma has a poor prognosis. However, since the introduction of preoperative radiotherapy, the outcome of patients with rectal carcinoma has been reported to have improved. Nevertheless, to the authors' knowledge few data are available regarding the histopathologic response to radiotherapy as assessed on surgical specimens as a potential predictive factor for outcome. METHODS To estimate the effect of radiotherapy on rectal carcinoma, the authors retrospectively reviewed the surgical specimens of 102 patients with T3-4, N0 or ≥ N1 rectal carcinoma and 1 patient with T2 but N1 rectal carcinoma. All patients were treated preoperatively with a hyperfractionated accelerated radiotherapy schedule in a prospective protocol (Trial 93-01). Using a standardized approach, tumor regression was graded using a system that varies from Grade 1 (tumor regression Grade [TRG] 1) when complete tumor regression is observed to Grade 5 (TRG5) when no tumor regression is observed. RESULTS Radiotherapy resulted in tumor downstaging in 43% of the patients. There were 2 pT1 tumors (2%), 21 pT2 tumors (20%), 66 pT3 tumors (64%), and 14 pT4 tumors (14%) after treatment. Regional lymph nodes were involved in 55 patients (53%). None of the patients demonstrated a complete tumor regression after radiotherapy, but in 79% of the specimens a partial tumor regression was observed (TRG1: 0%; TRG2: 20%; TRG3: 39%; TRG4: 20%; and TRG5: 21%). The median actuarial overall survival (OS) and disease-free survival (DFS) were 52 months. Actuarial local recurrence rates at 2 years and 5 years were 6.4% and 7.6%, respectively. Univariate analysis showed the actuarial DFS to be significantly lower in patients with lymph node metastases (P = 0.0004) and advanced pT stages (pT3-4) (P = 0.03). A favorable outcome for OS, DFS, and local control was observed in patients with TRG2-4 (i.e., responders) compared with patients with TRG5 (i.e., nonresponders), but also in patients with low residual tumor cell density (TRG2, 3, and 4). On multivariate analysis, TRG remained an independent prognostic indicator for local tumor control. CONCLUSIONS Tumor regression as well as residual tumor cell density were found to be predictive factors of survival in rectal carcinoma patients after preoperative radiotherapy. Even after preoperative radiotherapy, the pathologic stage of the surgical specimen remained a prognostic factor. The use of a standardized approach for pathologic evaluation must be implemented to allow comparison between the results of various treatment approaches.

Importance of tumor regression assessment in predicting the outcome in patients with locally advanced rectal carcinoma who are treated with preoperative radiotherapy

Locally advanced rectal carcinoma has a poor prognosis. However, since the introduction of preoperative radiotherapy, the outcome of patients with rectal carcinoma has been reported to have improved. Nevertheless, to the authors' knowledge few data are available regarding the histopathologic response to radiotherapy as assessed on surgical specimens as a potential predictive factor for outcome. To estimate the effect of radiotherapy on rectal carcinoma, the authors retrospectively reviewed the surgical specimens of 102 patients with T3-4, N0 or > or = N1 rectal carcinoma and 1 patient with T2 but N1 rectal carcinoma. All patients were treated preoperatively with a hyperfractionated accelerated radiotherapy schedule in a prospective protocol (Trial 93-01). Using a standardized approach, tumor regression was graded using a system that varies from Grade 1 (tumor regression Grade [TRG] 1) when complete tumor regression is observed to Grade 5 (TRG5) when no tumor regression is observed. Radiotherapy resulted in tumor downstaging in 43% of the patients. There were 2 pT1 tumors (2%), 21 pT2 tumors (20%), 66 pT3 tumors (64%), and 14 pT4 tumors (14%) after treatment. Regional lymph nodes were involved in 55 patients (53%). None of the patients demonstrated a complete tumor regression after radiotherapy, but in 79% of the specimens a partial tumor regression was observed (TRG1: 0%; TRG2: 20%; TRG3: 39%; TRG4: 20%; and TRG5: 21%). The median actuarial overall survival (OS) and disease-free survival (DFS) were 52 months. Actuarial local recurrence rates at 2 years and 5 years were 6.4% and 7.6%, respectively. Univariate analysis showed the actuarial DFS to be significantly lower in patients with lymph node metastases (P = 0.0004) and advanced pT stages (pT3-4) (P = 0.03). A favorable outcome for OS, DFS, and local control was observed in patients with TRG2-4 (i.e., responders) compared with patients with TRG5 (i.e., nonresponders), but also in patients with low residual tumor cell density (TRG2, 3, and 4). On multivariate analysis, TRG remained an independent prognostic indicator for local tumor control. Tumor regression as well as residual tumor cell density were found to be predictive factors of survival in rectal carcinoma patients after preoperative radiotherapy. Even after preoperative radiotherapy, the pathologic stage of the surgical specimen remained a prognostic factor. The use of a standardized approach for pathologic evaluation must be implemented to allow comparison between the results of various treatment approaches.

Effects of interleukin 2 therapy on lymphocyte magnesium levels

Interleukin 2 (IL-2) can cause partial or complete tumor regression in approximately 20% of patients with renal cell carcinoma. Among the many physiologic effects of IL-2, decreased serum levels of the divalent cations magnesium (Mg) and calcium have been demonstrated, with corresponding decreases in their urinary excretion. We investigated the effect of IL-2 on lymphocyte Mg levels among patients receiving three different dosing regimens. Twenty-eight patients with metastatic renal cell carcinoma were treated with high-dose intravenous, low-dose intravenous, or subcutaneous IL-2 therapy. Serum ionized Mg, urinary Mg, and peripheral blood mononuclear cell Mg levels were measured in samples from patients during treatment and compared with pretreatment levels. Serum Mg and ionized Mg levels decreased for all patients within 12 hours of treatment (P <.005) and remained low for the duration of therapy. Urinary Mg decreased in parallel with serum levels in all patients (P <.005). The peripheral blood mononuclear cell Mg content per cell increased within 24 hours of treatment (P <.005). The magnitude of these changes was similar during the first week of treatment for patients receiving intravenous or subcutaneous administration of IL-2. During IL-2 therapy, lymphocyte Mg increases coincident with serum Mg depletion. Mg availability may have functional implications for lymphocyte proliferation and integrin function. (J Lab Clin Med 2002;139:5-12)

Pre-clinical evaluation of a methotrexate-albumin conjugate (MTX-HSA) in human tumor xenografts in vivo.

Methotrexate covalently bound to human serum albumin in a 1:1 molar ratio (MTX-HSA) is a new macromolecular drug which is currently being studied in phase I clinical trials by the German Association for Medical Oncology (AIO) Phase I/II study group. Previous studies have shown that MTX-HSA differs favorably from unbound MTX in terms of plasma half-life time, tumor accumulation of albumin and uptake mechanisms into cancer cells. To achieve optimal drug efficacy, repeated treatment cycles were necessary. To evaluate the anti-tumor activity of MTX-HSA and MTX in pre-clinical in vivo models, we selected 7 solid human tumor xenografts growing s.c. in nude mice and administered drug either i.p. or i.v. weekly for 3 weeks. The maximal tolerated dose (MTD) of MTX-HSA in nude mice was 12.5 mg/kg given i.p. on days 1, 8 and 15, whereas the MTD for free MTX was 100 mg/kg given i.v. MTX-HSA was significantly more active (p > 0.01) than MTX in 3 models. In the soft tissue sarcoma SXF 1301, MTX-HSA effected complete remission/cure after a single injection, whereas free MTX resulted in short-lasting, partial tumor regression. In the prostate-cancer model PRXF PC3M, MTX-HSA produced growth inhibition of 92.8% of control or an optimal test/control (T/C) of 7.2% compared to a T/C of 20.8% for MTX (p = 0.05). In the osteosarcoma model SXF 1410, optimal T/C values were 10.2% and 14.5%, respectively (p = 0.025). In lung cancers LXFE 409 and LXFL 529, bladder cancer BXF 1258 and breast cancer MAXF 449, both compounds were inactive. The improved therapeutic effects seen in 3 xenograft models under MTX-HSA treatment are promising and might be due to specific accumulation of the compound in solid tumors owing to their enhanced permeability and retention effect. Thus, clinical development of MTX-HSA will continue and sarcomas as well as prostate cancers will be included as potential target tumors for upcoming clinical phase II trials.

Overexpressed Cell Surface Interleukin-4 Receptor Molecules Can Be Successfully Targeted for Antitumor Cytotoxin Therapy

A variety of human solid cancer cell lines and primary cell cultures has been reported to overexpress high-affinity receptors (R) for interleukin-4 (IL-4), a pleiotropic immunoregulatory cytokine. The significance of IL-4R expression is not known; however, IL-4 is able to upregulate adhesion molecules, inhibit cell proliferation, and mediate signal transduction in tumor cell lines. To target IL-4R, we produced a chimeric protein composed of a circular permuted IL-4 and a mutated form of Pseudomonas exotoxin [termed IL4(38-37)-PE38KDEL or cpIL4-PE]. The recombinant cpIL4-PE was highly cytotoxic to cancer cells, but not toxic to normal B cells, T cells, monocytes, and CD34+, even though these cells express detectable numbers of IL-4R. The cytotoxicity was specific because excess of recombinant IL-4 neutralized the cpIL4-PE effect. To further develop this molecule, in vivo antitumor activity was tested in animal models of human cancer. This agent showed remarkable antitumor activity in AIDS-Kaposi's sarcoma, glioblastoma multiforme, and breast cancer models in immunodeficient animals. cpIL4-PE caused partial or complete regression of established human tumors. Preclinical efficacy and toxicity studies provided a therapeutic window in which this cancer-targeted agent could be used. On the basis of these studies, we initiated a Phase I clinical trial for the treatment of recurrent glioblastoma multiforme. Our preliminary clinical results suggest that cpIL4-PE has antitumor activity against the deadliest form of brain tumors, without detectable toxicity to normal brain tissues. Thus, IL-4 receptors represent novel targets for cancer cytotoxin therapy.

Antitumor activity and toxicity of novel nitroheterocyclic phosphoramidates.

A series of novel nitroheterocyclic phosphoramidates has been evaluated for antitumor activity in murine and xenograft tumor models and for toxicity in mice. Significant increases in lifespan and long-term survivors were noted in L1210 leukemia and B16 melanoma models, and both complete and partial tumor regressions were observed in the MX-1 breast cancer xenograft model. All compounds exhibited some degree of toxicity to granulocyte/macrophage progenitors in the bone marrow of mice. Two drugs were selected for further toxicologic, histopathologic, and pharmacokinetic evaluations. Toxicity of potential clinical significance was observed only in the bone marrow at the highest drug dose; otherwise no significant abnormalities in blood chemistries or organ histopathology were noted. The bone marrow lesions consisted of reduced numbers of progenitor cells in the myeloid and erythroid series; platelets were not affected. The compounds were eliminated rapidly by first-order kinetics, with half-lives in the 4-12 min range. The best of these compounds exhibits excellent antitumor activity and minimal toxicity at therapeutically effective doses in mice.

The expression of CD70 and CD80 by gene-modified tumor cells induces an antitumor response depending on the MHC status.

The expression of costimulatory molecules such as CD70 or CD80 by gene-modified tumor cells has been shown to enhance the antitumor immune response based mainly on T lymphocytes. However, most human tumors show defects of major histocompatibility complex (MHC) expression, preventing them from being recognized by MHC-restricted T cells. To investigate if coexpression of CD70 and CD80 costimulatory molecules induces comparable antitumor responses in low and high MHC-expressing tumor cells, we used two low immunogenic murine tumor models, the B16.F10 melanoma and the TS/A mammary adenocarcinoma cell lines expressing, respectively, low and high levels of MHC class I molecules. Transfection of both CD70 and CD80 genes resulted in an increased capacity of gene-modified tumor cells to costimulate in vitro the proliferation and cytokine production of optimally activated lymphoid cells. Coexpression of CD70 and CD80 by the two tumor cell lines, TS/A and B16.F10, resulted in both cases in partial regression of subcutaneous tumors. Immunochemical analysis and studies in nude mice showed that, even in the B16.F10 model, T cells had a significant role in the antitumor response induced by combining CD70 and CD80. However, rejection of the CD70/CD80-transfected tumor cells appeared more effective in the MHC class I high TS/A model, leading to a protection against parental tumor cells. B16.F10 and TS/A transfectants were then tested with fibroblasts genetically modified to secrete interleukin-12 (IL-12) as a therapeutic vaccine in mice bearing parental tumors. In the two models tested, the injections of irradiated IL-12 and CD70/CD80 gene-modified cells generated an antitumor response to established tumors leading to the slowing down of the tumor growth rate. Although the mechanisms remain to be defined, these findings suggest that the combination of several immuno-modulatory molecules could provide additional strategies for cancer immuno-gene therapy, even for MHC expression-deficient tumors.

Conventional and novel strategies in the treatment of adrenocortical cancer.

Adrenocortical carcinoma is a highly malignant neoplasm with an incidence of two per million people per year. Several treatment strategies have resulted in temporary or partial tumor regression but very few cases have attained long survival. Surgical resection of the primary tumor and metastases is most effective. Several chemotherapeutic protocols have been employed with variable success. Mitotane (o,p'-DDD) is an adrenalytic drug effective in inducing a tumor response in 33% of patients treated. Mitotane requires metabolic transformation for therapeutic action. Tumors may vary in their ability to metabolize mitotane and the ability of tumors to transform mitotane may predict the clinical response to the drug. Preliminary data show a possible correlation between metabolic activity of neoplastic adrenocortical tissue and response to mitotane. We have attempted to develop mitotane analogs with enhanced adrenalytic effect. Compared to mitotane, a di-chloro compound, the bromo-chloro and di-bromo analogs appear to have a greater effect. Future approaches to the treatment of adrenocortical carcinoma are likely to be based on blocking or reversing the biological mechanisms of tumorigenesis. Angiogenic and chemotactic mechanisms may play a role in adrenal tumor growth and inhibition of these mechanisms may result in inhibition of tumor growth. New mitotane analogs with greater adrenalytic potential could be a promising approach to developing more effective and selective therapies for adrenal cancer. Alternative approaches should attempt to suppress tumor growth by means of compounds with anti-angiogenic and anti-chemotactic activity.

Mouse α-fetoprotein-specific DNA-based immunotherapy of hepatocellular carcinoma leads to tumor regression in mice

alpha-Fetoprotein (AFP) is a tumor-associated protein that is frequently expressed at high levels in hepatocellular carcinoma (HCC). The aim of the study was to characterize self-reactive cytotoxic T lymphocytes (CTLs) directed against murine AFP (mAFP) after DNA-based immunization in mice. To study CTL responses, mAFP-expressing recombinant vaccinia viruses were generated. An HCC tumor model was established in C57L/J mice by injection of syngeneic endogenously mAFP-expressing Hepa1-6 cells. Gene gun and intramuscular coimmunizations of DNA expression vectors encoding mAFP with plasmids encoding murine interleukin (IL)-12, granulocyte-macrophage colony-stimulating factor, or IL-18 induced weak CTL activity against mAFP in different mouse strains. Some mice developed anti-mAFP antibody responses, suggesting breaking of immunologic ignorance. No hepatocyte damage was detectable despite low-level endogenous hepatic mAFP expression. Therapeutic immunizations of mice bearing mAFP-expressing murine HCCs induced partial regression of tumors. A significant survival benefit was observed in mice immunized with mAFP expression vector DNA but not in untreated mice or in mice immunized with mock/cytokine plasmid DNA. The data show that AFP may be used as a potential self tumor antigen to induce CTL and CD4(+) T cell-mediated regression of AFP-expressing HCC by DNA-based immunization.

Metastatic Breast Tumour Regression Following Treatment by a Gene-Modified Vaccinia Virus Expressing MUC1 and IL-2.

MUC1 is expressed by glandular epithelial cells. It is overexpressed in the majority of breast tumours, making it a potential target for immune therapy. The objectives of the present study were to evaluate the anti-tumour activity and tolerance of repeated administration of TG1031 (an attenuated recombinant vaccinia virus containing sequences coding for human MUC1 and the immune stimulatory cytokine IL-2) in patients with MUC1-positive metastatic breast cancer. This was an open-label, randomised study comparing two dose levels, 5 × 10E6 and 5 × 10E7 pfu, with 14 patients in each arm. The treatment was administered intramuscularly every 3 weeks for the first 4 doses and every 6 weeks thereafter, until progression. Two patients had a partial tumour regression (> 50%), and 15 patients had stable disease as their best overall response until at least the 5th injection. Partial regression lasted for 11 months in one patient and for 12 months in the second patient who then underwent surgical resection of her hepatic metastases. The most frequent adverse events included inflammation at injection site: 7 patients, itching or pain at injection site: 5 patients, and moderate fever: 6 patients. One responding patient developed antinuclear, anti-DNA, and increased anti-TPO antibodies after the fifth injection, and which resolved at the end of treatment. The treatment regimes were well tolerated with a low toxicity profile. Although clinical efficacy remains limited, this study demonstrates the potential use of MUC1-based immune therapy in breast cancer.