Inhibition of breast cancer cell growth in vitro by a tyrosine kinase inhibitor.

Human breast cancer cell proliferation is regulated by growth factors that bind to receptors with intrinsic tyrosine kinase (TK) activity, including the epidermal growth factor (EGF) receptor. To determine whether inhibition of receptor TK activity inhibits tumor growth, we studied the effects of a tyrosine kinase inhibitor, RG-13022, on cultured human breast cancer cells. RG-13022 represents a class of compounds which have been shown to inhibit preferentially the TK activity of the EGF receptor in a cell-free system and also to inhibit EGF-stimulated growth of cultured cells. RG-13022 significantly inhibited EGF-stimulated autophosphorylation of its receptor in two breast cancer cell lines that have abundant, although not amplified, EGF receptor content (MDA-231 and T47D). RG-13022 also inhibited EGF-stimulated DNA synthesis and proliferation of T47D and MCF-7 breast cancer cells in a reversible and dose-dependent manner. Inhibition was observed at 0.1 microM, and it was maximal at 10 microM. The effect was rapid (within 3 h), persisted for 18 h, and was partially reversed by 24 h at 1 microM. At 5 microM, inhibition persisted for more than 50 h. Inhibitory effects were also observed in a panel of estrogen receptor-positive and estrogen receptor-negative breast cancer cell lines. RG-13022 inhibited not only EGF-induced growth but also growth stimulated by insulin, insulin-like growth factor I, insulin-like growth factor II, or transforming growth factor alpha. RG-13022 also totally blocked estrogen-stimulated phosphorylation of the EGF receptor, as well as estrogen-induced cell proliferation, suggesting that functioning TK pathways are required for estrogen action. The TK inhibitor RG-13022 is a potent inhibitor of hormonally regulated growth of human breast cancer. Tyrosine kinase inhibitors have the potential of providing a new strategy for the "endocrine therapy" of breast cancer.

Water soluble 20(S)-glycinate esters of 10,11-methylenedioxycamptothecins are highly active against human breast cancer xenografts.

Water-soluble 20(S)-glycinate esters of two highly potent 10,11-methylenedioxy analogues of camptothecin (CPT) have been synthesized and evaluated for their ability to eradicate human breast cancer tumor xenografts. The glycinate ester moiety increases the water solubility of the 10,11-methylenedioxy analogues 4-16-fold. However, in contrast to CPT-11, a water-soluble CPT analogue that was recently approved for second line treatment of colorectal cancer, the 20(S)-glycinate esters do not require carboxylesterase for conversion to their active forms. The glycinate esters are hydrolyzed to their parent, free 20(S)-hydroxyl active analogues in phosphate buffer (pH 7.5) and in mouse and human plasma. The glycinate esters are also 20-40-fold less potent than CPT-11 in inhibiting human acetylcholinesterase. In vivo, we examined 20(S)-glycinate-10,11-methylenedioxycamptothecin, 20(S)-glycinate-7-chloromethyl-10,11-methylenedioxycamptothecin, and CPT-11. We found that the two 10,11-methylenedioxy analogues had antitumor activity against breast cancer xenografts that was comparable to that of CPT-11. Our results indicate that water-soluble 20(S)-glycinate esters of highly potent CPT analogues provide compounds that maintain biological activity, do not require interactions with carboxylesterases, and do not inhibit human acetylcholinesterase.

Preclinical antitumor activity of 6-hydroxymethylacylfulvene, a semisynthetic derivative of the mushroom toxin illudin S.

6-Hydroxymethylacylfulvene (HMAF; MGI 114) is a novel semisynthetic antitumor agent derived from the sesquiterpene mushroom toxin illudin S. In vitro cytotoxicity determinations produced IC50 concentrations (concentrations required for 50% inhibition of growth) ranging from 160 nM in sensitive MCF-7 human mammary carcinoma cells to 17 microM in relatively insensitive murine B16 melanoma cells. In vivo antitumor activity was consistent with in vitro sensitivity. HMAF was very effective in human tumor xenograft models, including MX-1 breast carcinoma, MV522 lung adenocarcinoma, and HT-29 colon carcinoma, but not murine B16 melanoma or P388 leukemia. Excellent responses were observed in animals bearing MX-1 tumors administered i.v. or i.p. doses of 3-7.5 mg/kg daily for 5 days, with complete regression recorded in 29 of 30 animals administered i.v. HMAF. Extensive tumor shrinkage was also observed with MV522, and significant tumor growth inhibition was obtained with HT-29 when animals received 5 daily i.p. doses ranging from 3.75 to 7.5 mg/kg. Complete regressions were also observed in individual animals with MV522 and HT-29. The excellent activity of HMAF in several human solid tumor xenografts, including the more refractory MV522 and HT-29 models, warrants the further investigation of this novel agent in clinical trials.

Enhanced antitumor activity of 6-hydroxymethylacylfulvene in combination with irinotecan and 5-fluorouracil in the HT29 human colon tumor xenograft model.

6-Hydroxymethylacylfulvene (MGI-114) is a semisynthetic analogue of the toxin illudin S, a product of the Omphalotus mushroom. MGI-114 induces cytotoxicity in a variety of solid tumors in vivo, including the refractory HT29 human colon cancer xenograft. In this study, the potential application of MGI-114 in the treatment of colon cancer was further explored by evaluating the activity of MGI-114 in combination with irinotecan (CPT-11) and 5-fluorouracil (5FU). Groups of 9 nude mice bearing HT29 xenografts were treated with either single agent MGI-114, CPT-11, or 5FU, or MGI-114 in combination with CPT-11 or 5FU. MGI-114 was administered at doses of 3.5 and 7 mg/kg i.p. daily on days 1 through 5, and CPT-11 and 5FU were administered at doses of 50 and 100 mg/kg i.p. on days 1, 12, and 19. In the single agent studies, MGI-114, CPT-11, and 5FU all resulted in decreased final tumor weights compared with vehicle-treated controls (P<0.05), but only MGI-114 at 7 mg/kg produced partial responses. When MGI-114 at 3.5 mg/kg was combined with CPT-11, significant decrements in final tumor weights occurred compared with monotherapy with the same doses of MGI-114 and CPT-11 (P< or =0.001). Also, administration of the low-dose combination (MGI-114 at 35 mg/kg and CPT-11 at 50 mg/kg) resulted in final tumor weights similar to those achieved after administration of high-dose MGI-114 as a single agent. Moreover, the combination of MGI-114 and CPT-11 produced partial responses in nearly all of the animals, with some animals achieving complete responses. The outcome with the combination of MGI-114 and 5FU was less striking, with fewer partial responses and no complete responses. These results suggest enhanced activity when MGI-114 is combined with CPT-11, and clinical trials to further evaluate this combination regimen are planned.

Tumor efficacy and bone marrow-sparing properties of TER286, a cytotoxin activated by glutathione S-transferase.

TER286 is a latent drug activated by human glutathione S-transferase (GST) isoforms P1-1 and A1-1 to produce a nitrogen mustard alkylating agent. M7609 human colon carcinoma, selected for resistance to doxorubicin, and MCF-7 human breast carcinoma, selected for resistance to cyclophosphamide, both showed increased sensitivity to TER286 over their parental lines in parallel with increased expression of GST P1-1. In primary human tumor clonogenic assays, the spectrum of cytotoxic activity observed for TER286 was both broad and unusual when compared to a variety of current drugs. In murine xenografts of M7609 engineered to have high, medium, or low GST P1-1, responses to TER286 were positively correlated with the level of P1-1. Cytotoxicity was also observed in several other cell culture and xenograft models. In xenografts of the MX-1 human breast carcinoma, tumor growth inhibition or regression was observed in nearly all of the animals treated with an aggressive regimen of five daily doses. This schedule resulted in a 24-h posttreatment decline in bone marrow progenitors to 60% of control and was no worse than for a single dose of TER286. These studies have motivated election of TER286 as a clinical candidate.

Enhanced antitumour activity of 6-hydroxymethylacylfulvene in combination with topotecan or paclitaxel in the MV522 lung carcinoma xenograft model.

6-Hydroxymethylacylfulvene (HMAF; MGI 114; Irofulven) is a semisynthetic analogue of the toxin illudin S, which is a product of the Omphalotus mushroom. MGI 114 induces cytotoxicity against a broad range of solid tumours in vivo, including the drug-refractory MV522 human lung cancer xenograft. In this study, the potential application of MGI 114 in the treatment of lung cancer was explored by evaluating the activity of MGI 114 in combination with either topotecan (TPT) or paclitaxel. Groups of eight nude mice bearing MV522 xenografts were treated with MGI 114, TPT or paclitaxel as single agents and with MGI 114 in combination with TPT or paclitaxel. MGI 114 was administered at doses of 2.5 and 5.0 mg/kg intraperitoneally (i.p.) daily on days 1-5, while TPT and paclitaxel were administered at doses of 0.5 or 1.0 mg/kg and 20 mg/kg, respectively, i.p. on days 1-5. In the single-agent studies, MGI 114, TPT and paclitaxel all resulted in decreased final tumour weights compared with vehicle-treated controls. As single agents, TPT, at the 0.5 mg/kg dose level, and paclitaxel, at the 20 mg/kg dose level, produced partial shrinkages (PSs). All combinations of MGI 114, and either TPT or paclitaxel, produced decrements in final tumour weights compared with monotherapy with the same doses of MGI 114, TPT and paclitaxel. Although all animals treated with the combination of MGI 114 and paclitaxel experienced PSs or complete shrinkages (CSs) (or died), analysis of the time to tumour doubling revealed that the combination of MGI 114 and TPT at 2.5 and 0.5 mg/kg, respectively, was synergistic. These results suggest that cytotoxic activity is enhanced when MGI 114 is combined with either TPT or paclitaxel, and clinical trials to further evaluate these combination regimens are warranted.

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.

Design of new topoisomerase II inhibitors based upon a quinobenzoxazine self-assembly model.

A new class of pyridobenzophenoxazine compounds has been developed as topoisomerase II inhibitors for anticancer chemotherapy. These compounds were designed based on a proposed model of a quinobenzoxazine self-assembly complex on DNA. They showed excellent inhibitory effects on several tumor cell lines with nanomolar IC50 values. Their cytotoxic potency correlates with their ability to unwind DNA and inhibit topoisomerase II.

Transgenic mice with pigmented intraocular tumors: tissue of origin and treatment.

PURPOSE: To describe the cell of origin, tumor progression, light and electron microscopic appearance, immunohistochemical properties, and response to frequently used anticancer therapies in two transgenic models of intraocular melanoma. METHODS: Two lines of transgenic mice that develop pigmented intraocular tumors were produced with the SV40 T and t antigens under the control of the mouse tyrosinase gene. Tumors were sequentially studied and characterized by light microscopy, electron microscopy, and immunohistochemistry stains. Tumor response to two cycles of dacarbazine was assessed on the basis of tumor size in one group of animals. Response to external beam irradiation was measured by survival time in other animals. RESULTS: Two lines of transgenic mice developed bilateral intraocular tumors with complete penetrance and without primary cutaneous melanomas. Tumors developed first in the retinal pigment epithelial layer, with subsequent retinal and choroidal invasion, extraocular extension, and metastasis. Tumors stained positive for S-100, HMB-45, and Fas-ligand. Electron microscopy revealed polarization of tumor cells with basement membrane formation, microvilli, immature melanosomes, and abundant endoplasmic reticulum. Dacarbazine significantly reduced tumor size in these mice, and a trend toward dose-dependent decrease in survival was found with external beam irradiation. CONCLUSIONS: Tumors developed from the retinal pigment epithelium. Their histology and growth, however, closely resembled that of human choroidal melanoma. This model may be a useful tool for future studies of endogenous primary pigmented tumors limited to the eye. Response to standard therapies suggests it can serve as a model with which to evaluate therapeutic modalities.

An investigation of the antitumour activity and biodistribution of polymeric micellar paclitaxel.

PURPOSE: To evaluate in vitro cytotoxicity, in vivo antitumour activity and biodistribution of a novel polymeric (poly(DL-lactide)-block-methoxy polyethylene glycol) micellar paclitaxel. METHODS: Hs578T breast, SKMES non-small-cell lung, and HT-29 colon human tumour cells were exposed, either for 1 h or continuously, to conventionally formulated paclitaxel (Cremophor paclitaxel) or polymeric micellar paclitaxel. After a period of incubation, cytotoxicity was measured using a radiometric system. In the in vivo antitumour study, B6D2F1 mice, bearing P388 leukaemia tumour intraperitoneally (i.p.), were treated with polymeric micellar paclitaxel or Cremophor paclitaxel by i.p. injection. The number of deaths and body weights were recorded. In the biodistribution study, CD-1 mice were given micellar paclitaxel i.p. at a dose of 100 mg/kg. The mice were sacrificed after a given time and the organs were harvested. Paclitaxel in the organs was extracted by acetonitrile and analysed using HPLC. RESULTS: The polymeric micellar paclitaxel showed similar in vitro cytotoxicity to Cremophor paclitaxel against the tumour cell lines. The polymeric micellar formulation of paclitaxel produced a fivefold increase in the maximum tolerated dose (MTD) as compared with Cremophor paclitaxel when administered i.p. In addition, micellar paclitaxel was more efficacious in vivo when tested in the murine P388 leukaemia model of malignancy than Cremophor paclitaxel when both were administered i.p. at their MTDs. Micellar paclitaxel-treated animals had an increased survival time and, importantly, long-term survivors (20% of those tested) were obtained only in the polymeric paclitaxel formulation group. Biodistribution studies indicated that a significant amount of paclitaxel could be detected in blood, liver, kidney, spleen, lung and heart of mice after i.p. dosing of the polymeric micellar paclitaxel formulation. CONCLUSION: These preliminary results indicate that polymeric micellar paclitaxel could be a clinically useful chemotherapeutic formulation.

Evidence of enhanced in vivo activity using tirapazamine with paclitaxel and paraplatin regimens against the MV-522 human lung cancer xenograft.

PURPOSE: Tirapazamine (3-amino-1,2,4-benzotriazine 1,4-dioxide; SR 4233) is a bioreductive agent that exhibits relatively selective cytotoxicity towards cells under hypoxic conditions and can enhance the antitumor activity of many standard oncolytics. In the present study we examined the interaction between tirapazamine in vivo with paclitaxel and paraplatin in two- and three-way combination studies using the MV-522 human lung carcinoma xenograft model. METHODS: Agents were administered as a single i.p. bolus, with tirapazamine being given 3 h prior to paclitaxel, paraplatin, or their combination. Tumor growth inhibition (TGI), final tumor weights, partial and complete responses, and time to tumor doubling were determined after drug administration. RESULTS: Tirapazamine as a single agent was ineffective against this human lung tumor model. A substantial increase in TGI was seen in animals treated with the triple-agent regimen (tirapazamine-paclitaxel-paraplatin) compared to animals treated with double-agent regimens that did not include tirapazamine. The addition of tirapazamine to paclitaxel-paraplatin therapy resulted in a 50% complete response rate; there were no complete responses seen when only the paclitaxel-paraplatin combination was administered. Time to tumor doubling was also significantly improved with the addition of tirapazamine to the paclitaxel and paraplatin combinations. Tirapazamine did not increase the toxicity of paclitaxel, paraplatin, or their combinations as judged by its minimal impact on body weight and the fact that no toxic deaths were observed with tirapazamine-containing regimens. CONCLUSIONS: These results are important since recent studies have suggested that the combination of paclitaxel and paraplatin may be particularly active in patients with advanced stage non-small-cell lung cancer. Since tirapazamine can significantly improve efficacy, but does not appear to enhance the toxicity of paclitaxel and paraplatin, its evaluation in future clinical trials in combination with paclitaxel-paraplatin-based therapy appears warranted.

[Accessory spleen in the pancreatic tail -- a neglected entity? A contribution to embryology, topography and pathology of ectopic splenic tissue]. / Die Nebenmilz im Pankreasschwanz-eine vernachlässigte Entität? Ein Beitrag zur Embryologie, Topographie und Pathologie der Ektopien des Milzgewebes.

According to autoptic studies, accessory spleens may be found in 10% to 15% of the population, in 1% to 2% they are located in the pancreatic tail. They thus have to be taken into account in the differential diagnosis of intra- and peripancreatic tumorous lesions. After splenorenal fusion, they can be found pararenally and retroperitoneally, and after splenogonadal fusion they can descend into the pelvis or scrotum. Since they usually are asymptomatic, they are mostly discovered accidentally. The diagnosis is ascertained by a scintigram with technetium-99-marked, heat-damaged red blood cells. If resection is necessary, the organ adherent to the encapsulated accessory spleen should be preserved. By means of three case reports (intrapancreatic accessory spleen, retroperitoneal accessory spleen and torsion-infarcted wandering spleen), the surgical relevance of ectopic splenic tissue is discussed.

[Variations in the course of the inferior laryngeal nerve. Surgical anatomy, classification, diagnosis]. / Verlaufsvarietäten des Nervus laryngeus inferior. Chirurgische Anatomie, Klassifikation, Diagnostik.

Because of multiple variations in course, the inferior laryngeal nerve shows a great variety of topographic relations to adjacent cervical structures. It may recur in the tracheoesophageal groove or anteriorly or posteriorly to it. It can pass under, over, or through the ramifications of the inferior thyroid artery. If Zuckerkandl's tubercle is enlarged, the nerve may be luxated. It is firmly fixed to the ligament of Berry by tight adhesions. Before entering the larynx, the nerve may show multiple ramifications. It may also recur around the inferior thyroid or vertebral artery. On the right, a nonrecurrent nerve is found in 0.6-0.8% of individuals, always in coincidence with a "lusorian" artery. Three course variations can be distinguished: descending (type I), horizontal (II), and ascending (III). A nonrecurrent nerve on the left is extremely rare, as it can only be found as a combination anomaly of a right-sided lusorian artery with situs inversus viscerum. The divided inferior laryngeal nerve shows recurrent and nonrecurrent ramifications. A nonrecurrent inferior laryngeal nerve can be indirectly ruled out preoperatively by demonstration of a normally developed brachiocephalic trunk via colour-coded duplex ultrasound.

[Bacterial infections of the abdominal aorta]. / Bakterielle Infektionen der Aorta abdominalis.

There is a high mortality of bacterial aortitis particularly as it is usually not recognized before the stage of rupture. Therefore the disease should early be considered in obscure febrile conditions. Problems of diagnosis and therapy are discussed on two own cases. A 63-year old man had a rupture of a small infrarenal aortic aneurysm in the course of salmonella sepsis, a 79-year old woman had a pyogenic osteomyelitis of a lumbar vertebra that spread to the aorta and caused its rupture. In both cases surgical treatment consisted of bleeding control followed by in-situ reconstruction.

[Fibrovascular esophageal polyp--diagnosis and therapy]. / Der fibrovasculäre Oesophaguspolyp--Diagnostik und Therapie.

Fibrovascular polyps are extremely rare benign neoplasms of the esophagus. The most prominent clinical symptom is enoral tumor regurgitation, which may lead to asphyxiation by pharyngeal impaction. Usually fibrovascular polyps cause dysphagia and progressive weight loss. Diagnosis by endoscopy and barium swallow may be unexpectedly difficult. The most frequent incorrect diagnoses are achalasia or an intramural or mediastinal tumorous mass compressing the esophagus. CT scan and MR imaging are of little help. Small polyps may be resected endoscopically by means of electrocautery or Nd:YAG laser ablation. In most cases, however, surgical resection is required. Since the basis of the polyp is usually located subcricoidally, tumor exposure and resection are achieved by esophagotomy via a left cervical approach. Thoracotomy is seldom required.

[Traumatic arteriovenous fistula between superior thyroid artery and vein]. / Traumatische arteriovenöse Fistel zwischen Arteria und Vena thyreoidea superior.

Arteriovenous fistula is a rare complication following vascular trauma. We report a case of an arteriovenous fistula of the neck after a penetrating injury. The only presenting symptoms were a thrill and a machinery murmur. There were no signs of cardiac insufficiency. Duplex scan and arteriography detected an arteriovenous fistula between the superior thyroid artery and the superior thyroid vein. Surgical therapy was straight-forward. Treatment should be carried out as soon as possible, and is then usually simple and successful.

[Single injuries to distal leg arteries]. / Singuläre Unterschenkelarterienverletzungen.

The appropriate treatment of an injury to a single arterial vessel of the calf is still a matter of discussion. Isolated injury of one of the calf arteries is generally not considered to cause severe ischemia of the leg. Other factors such as the degree of concomitant trauma to bones, nerves, veins and soft tissue, which may impair collateral circulation, seem to represent the real threat for the survival of the extremity. On the other hand, high numbers of amputations were reported after the Second World War following ligation of an injured single vessel of the calf. Concomitant injuries are poorly documented in these reports. Consequently a good physical examination as well as arteriography, duplex ultrasound scan and a high index of suspicion are mandatory to evaluate the impaired circulation of the calf and to prevent hasty ligation of a single vessel.

[Massive gastrointestinal hemorrhage following internal drainage of pancreatic pseudocysts]. / Massive Gastrointestinalblutung nach innerer Drainage von Pankreaspseudocysten

In operative treatment of pancreatic pseudocysts by inner drainage there is a risk of massive gastrointestinal bleeding particularly following an anastomosis to the stomach. In 10 patients in whom cystogastrostomy or cystoduodenostomy had been performed elsewhere a second laparotomy was necessary because of acute bleeding. In one patient a cystadenoma of the pancreas had been anastomosed to the duodenum by mistake at the previous operation. The leak of obliteration of the cyst is suggested to be the most important factor in the pathophysiology of bleeding. Inner pseudocyst drainage in a disconnected small bowel loop therefore principally should be performed at the lowest point of the cyst. The indication for an inner cyst drainage, however, must be closely examined since simultaneous pathologic changes in the pancreas often justify a resection to remove the origin of the cyst and, further, averting the complications of an inadequate inner pancreatic cyst drainage.

[Delayed surgery in acute pancreatitis]. / Verzögerte Operation bei akuter Pankreatitis.

In 32 patients with acute pancreatitis, delayed operation was performed between 13 and 44 days after onset of the illness. The indications for the operation were development of a palpable mass together with clinical deterioation and other complications. In all patients we found a necrotizing pancreatitis and/or abscesses of the pancreas. The surgical procedure consisted of digital removal of necrotic tissue (sequestrotomy) and/or abscess incision in 20 patients, of left-sided resection in 11 patients and partial duodenopancreatectomy in 1 patient. Twenty-three patients survived, 9 died.

[Ruptured aneurysm of the pancreaticoduodenal artery. A rare etiology of acute abdomen]. / Das rupturierte Aneurysma der Arteria pancreaticoduodenalis. Eine seltene Ursache des akuten Abdomens.

Even among the uncommon aneurysms of the visceral arteries the aneurysm of the pancreaticoduodenalis artery is considered a rarity. Etiologically, numerous factors must be taken into account, the most significant one being arteriosclerosis. The clinical presentation is unspecific and ambiguous. CT and, above all, intra-arterial DSA allow for a diagnosis. A generous consideration of indicating operative intervention, even in asymptomatic patients, is especially justified because of the imminent risk of rupture. The preferable therapy consists of elimination of the aneurysm either conventionally by proximal and distal ligature of the pancreaticoduodenalis artery or endovascularly by embolization. In the future a treatment with coated stents (TPEG) would also seem possible. Special attention must be paid to concomitant occlusive disease in other visceral arteries since measures for vessel reconstruction may be required because of intraoperative impairment of the collateral circulation. We report on the rupture of an aneurysm of the pancreaticoduodenal inferior artery in association with celiac axis occlusion.