soblidotin and Pancreatic-Neoplasms

Pancreatic cancer has the worst prognosis of all cancers with a dismal 5-year survival rate. Hence, there is a tremendous need for development of new and effective therapy for this tumor. In an earlier study we reported a potent antitumor activity of Auristatin PE (AuriPE) against pancreatic tumor. In addition, we have also reported that bryostatin 1 (bryo1) induces differentiation of leukemia cells, but the effect of bryo1 has not been investigated in pancreatic tumors. This is the first report where we demonstrate that bryo1 induces differentiation and potentiates the antitumor effect of AuriPE in a human pancreatic tumor (PANC-1) xenograft model. A xenograft model was established by injecting the PANC-1 cells s.c. in severe combined immune deficient (SCID) mice. After development of the s.c. tumors, tumors were dissected and small fragments were transplanted in vivo to new SCID mice, with a success rate of 100% and a doubling time of 4.8 days. The SCID mouse xenograft model was used to test the in vivo differentiation effect of bryo1 and its efficacy when given alone or in combination with AuriPE. Sections from paraffin-embedded tumors excised from untreated (control) SCID mice revealed typical poorly differentiated adenocarcinoma of the pancreas. Interestingly, sections of s.c. tumors taken from bryo1-treated mice revealed carcinomas that were much lower grade and less aggressive, and displayed prominent squamous and glandular differentiation. In this study, the tumor growth inhibition (T/C), activity score and cure rate for bryo1, AuriPE and bryo1+AuriPE were 80%, (+) and 0/4; 0.0%, (++++) and 3/5; and 0.0%, (++++) and 3/4, respectively. Mice treated with either AuriPE or bryo1+AuriPE were free of tumors for more than 150 days and were considered cured. The use of bryo1 as a novel differentiating agent and its combination with AuriPE should be further explored for the treatment of adenocarcinoma of the pancreas.

Topics: Animals; Antineoplastic Agents; Bryostatins; Drug Synergism; Female; Humans; Lactones; Macrolides; Mice; Mice, SCID; Oligopeptides; Pancreatic Neoplasms; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Pancreatic cancer is one of the most incurable and lethal human cancers in the United States. To facilitate development of novel therapeutic agents, we previously established an orthotopic pancreatic tumor model that closely mimics the natural biological behavior of human pancreatic cancer. In this study, magnetic resonance imaging (MRI) techniques were developed to detect tumor formation noninvasively and monitor serially tumor growth kinetics in this orthotopic model used for experimental drug testing. By using an optimized T2-weighted imaging method, we were able to distinguish human pancreas cancer from normal mouse pancreas. Orthotopic tumor formation was detected as early as day 1 after tumor cell implantation with a tumor volume as small as 12 mm3. Mice with evidence of tumor were separated into four treatment groups: control, auristatin-PE, gemcitabine, and their combination. After treatment, the mice were imaged at least three times before termination of the experiment. Comparison between MRI tumor volume measurements and tumor weights made at biopsy resulted in a correlation coefficient of 0.98. The tumor growth curves constructed from serial magnetic resonance imaging (MRI) measurements clearly showed tumor growth inhibition in treated mice compared with the control group. As expected, the group treated with the combination had the highest response rate compared with either auristatin-PE or gemcitabine alone, and the data were statistically highly significant (p < 0.004). From these results, we conclude that noninvasive MRI can be used to monitor serially therapeutic response in this orthotopic human pancreatic tumor model and can be used in the future to evaluate novel antitumor agents before human studies.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Female; Humans; Magnetic Resonance Imaging; Mice; Mice, SCID; Oligopeptides; Pancreas; Pancreatic Neoplasms; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Pancreatic adenocarcinoma is the fifth leading cause of cancer related deaths in the United States. Treatment for this disease has largely been unsuccessful, which may partly be due to insufficient data regarding the molecular mechanisms of chemotherapeutic drugs currently being used as single agents or in combined modality regimens. In this study, we investigated the molecular mechanisms by which auristatin-PE, a newly developed experimental agent, and gemcitabine, a commercially available anti-cancer agent, exert their inhibitory effects on pancreatic cancer cell lines containing wild-type p53 (HPAC) and mutant p53 (PANC-1). Our results showed that auristatin-PE and gemcitabine inhibited cell growth and induced cell cycle arrest in G2/M and S phase, respectively. Auristatin-PE also induced apoptosis in both cell lines. Western blot analysis showed that auristatin-PE up-regulated the expression of wt-p53, p21WAF1 and Bax, and down-regulated Bcl-2 and cyclin B in HPAC cells, while only up-regulation of p21WAF1 and Bax was observed in PANC-1 cells. These results suggest that auristatin-PE may induce apoptosis and p21WAF1 expression through p53-dependent or independent pathways, and that up-regulation of p21WAF1 and Bax and down-regulation of Bcl-2 may be the molecular mechanism through which auristatin-PE inhibits cell growth and induces apoptosis. Furthermore, the up-regulation of p21WAF1 and down-regulation of cyclin B may contribute to the G2/M cell cycle arrest. Combination of auristatin-PE and gemcitabine showed significantly greater inhibition of cell growth and up-regulated expression of p21WAF1 and Bax. From these results, we conclude that the selection of therapeutic agents based on their molecular mechanism may improve therapeutic outcome, and that auristatin-PE may be more effective in the treatment of pancreatic cancer when given in combination with gemcitabine, rather than as a single agent.

Topics: Adenocarcinoma; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Cell Cycle; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Deoxycytidine; Flow Cytometry; Gemcitabine; Gene Expression; Humans; Oligopeptides; Pancreatic Neoplasms; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Tumor Cells, Cultured; Tumor Suppressor Protein p53

Adenocarcinoma of the pancreas generally remains an incurable disease by available treatment modalities, demanding the development of a suitable cell-culture/animal model and the discovery and evaluation of novel therapeutic agents. We report the clonal preservation of a human pancreatic cell line (KCI-MOH1) established from a 74-year-old African-American man diagnosed with pancreatic cancer. Initially the human primary tumor was grown as a xenograft in SCID mice and, subsequently, a cell line was established from tumors grown as a xenograft as reported in our earlier publication. The molecular characterization of the primary tumor, the tumors grown as xenograft, and the cell line all revealed similar genotypic properties. By using an automated DNA sequencer, a K-ras mutation (codon 12, GGT to CGT, Gly to Arg) was detected in the pancreatic tumor tissue taken from the patient, whereas no p53 mutation was detected. The same K-ras mutation and unaltered p53 was also found in the xenograft tumor and in the KCI-MOH1 cell line. Chromosome analysis of the cultured cells revealed: 42,XY,add(3)(p11.2),der(7)t(7;12) (p22;q12),-10,-12,add (14)(p11),-18,add (20)(q13),-22/84, idemx2, which is the same chromosome complement found in xenograft tumors. The KCI-MOH1 cell line grows well in tissue culture and forms tumors in the SCID mice when implanted subcutaneously, as well as in orthotopic sites. The KCI-MOH1 cell line-derived SCID mouse xenograft model was used for efficacy evaluation of bryostatin 1, auristatin-PE, spongistatin 1, and gemcitabine alone and in combination. Tumor growth inhibition (T/C expressed as percentage), tumor growth delay (T - C), and log 10 kill for these agents were 38%, 22 days, and 0.53; 15%, 30 days, and 0.80; 24%, 25 days, and 0.66; and 10%, 33 days, and 0.90, respectively. When given in combination, two of seven gemcitabine + auristatin-PE-treated animals were free of tumors for 150 days and were considered cured. Animals treated with a combination of bryostatin 1 and gemcitabine and a combination of spongistatin and gemcitabine produced remissions in only one of seven mice. From these results, we conclude that (a) this is the first study illustrating that clonal characteristics of primary pancreatic tumors remained unchanged when implanted in mice and as a permanent cell line grown in vitro; and (b) there is a synergistic effect between gemcitabine and selected marine products tested in this study, which is more apparent in the gemcita

Topics: Adenocarcinoma; Aged; Animals; Antineoplastic Agents; Bryostatins; Deoxycytidine; DNA Mutational Analysis; Ethers, Cyclic; Gemcitabine; Genes, p53; Genes, ras; Humans; Karyotyping; Lactones; Macrolides; Male; Mice; Mice, SCID; Neoplasm Transplantation; Oligopeptides; Pancreatic Neoplasms; Tumor Cells, Cultured

Pancreatic adenocarcinoma is one of the most incurable and least understood of all human cancers. It is the fourth leading cause of cancer-related mortality in males (after lung, prostate, and colon) and in females (after lung, breast, and colon) in the United States with <2-3% of patients surviving >5 years. In an attempt to search for more effective therapies for this disease, we report here, for the first time, an effective treatment, the combination of gemcitabine and auristatin-phenethylamine (PE), against an orthotopic implantation of a human pancreatic adenocarcinoma cell line (HPAC) in severe combined immunodeficient (SCID) mice. Tumor implantation was performed by injecting 100 microl of the HPAC cell suspension (1 x 10(6) cells) directly into the pancreas of 5-week-old SCID mice. After implantation, tumor formation was checked twice a week. All palpable tumors were detected within 21 days (100% take rate), and tumors were confirmed histologically to be pancreatic adenocarcinoma. For the subsequent efficacy trial, tumor-bearing SCID mice were randomized into four groups with five mice in each group. One served as a control, the second received gemcitabine alone (2.5 mg/kg/injection i.p.), the third received auristatin-PE alone (2.0 mg/kg/injection i.v.), and the fourth group received the combination of gemcitabine (i.p.) and auristatin-PE (1.5 mg/kg/injection i.v.). All animals were euthanized 7 days after the completion of their treatments, and the pancreases were resected. Histological examination revealed the tumors to be adenocarcinoma. The tumors were composed of diffuse sheets of cells interrupted by glandular spaces containing secretory material. Cytologically, the tumor cells were large, pleomorphic, and hyperchromatic. Many cells contained intracellular lumina containing mucin. Immunohistochemical studies showed strong p21WAF1 (p21) expression but no immunoreactivity with p53 and Her-2/neu antibodies. The mean pancreatic weight in the gemcitabine/auristatin-PE combination group was significantly (P = 0.014) lower (0.84 +/- 0.639 g) when compared with those of the control (2.91 +/- 1.19 g) and gemcitabine alone (1.84 +/- 0.796 g; P = 0.064) groups. In addition, the mean weight in the combination group approached statistical significance when compared with the auristatin-PE group alone (1.16 +/- 0.635 g; P = 0.028). We conclude that the combination of gemcitabine and auristatin-PE is an effective treatment against HPAC tumors in this xenog

Topics: Adenocarcinoma; Animals; Antineoplastic Combined Chemotherapy Protocols; Deoxycytidine; Disease Models, Animal; Female; Gemcitabine; Mice; Mice, Inbred ICR; Mice, SCID; Oligopeptides; Pancreatic Neoplasms; Transplantation, Heterologous