Significant Delay of Lethal Outcome in Cancer Patients Due to Peroral Administration of Bacillus oligonitrophilus KU-1

Treatment of cancer patients remains a serious medical problem and the development of alternative treatment strategies is therefore of great importance. In this connection, we developed a new bacterial-based, anticancer method. Ten cancer patients (three males, seven females) were involved in this study. Bacterial suspension of stationary phase Bacillus oligonitrophilus KU-1 was used as a remedy for peroral administration. In five patients, side effects (sicchasia, slight blood, and intracranial pressure gain) were detected, but all patients showed significant delay of lethal outcome without serious side effects. In conclusion, the suggested method was, in our opinion, a good alternative to conventional chemo- and radiotherapy techniques. In order to evaluate its efficiency for various tumors, a double-blind, placebo-controlled, multicenter study is needed.


INTRODUCTION
Oncological diseases remain a significant health problem despite enormous scientific efforts and investments during the last decades. Numerous protocols for chemo-and radiotherapeutic treatment of various cancers have been developed [1,2,3,4,5,6]. However, a very low efficiency, especially in advanced cancer patients, as well as serious side effects [7,8] evoke a necessity to work up some alternative treatment strategies.
Using some theoretical backgrounds suggested by Tokin [9] and Voronkov et al. [10], a new bacterialbased, anticancer treatment strategy has been suggested. As our experience showed [11], safety and relative efficiency were the main hallmarks of the developed method.

METHODS
Ten cancer patients (three males, seven females, age 42-76 years) were involved in this study. Although type of cancer varied from patient to patient, all of them were characterized by the presence of distant metastasis. Bacterial suspension of Bacillus oligonitrophilus KU-1 strain was used as remedy. The bacterial strain was isolated from the soil of Kazan city in Russia, according to published techniques [12]. After identification, B. oligonitrophilus KU-1 was cultivated in liquid medium of Alexandrov as described previously [11]. It has been shown that B. oligonitrophilus KU-1 does not have any toxicity [11]. Stationary phase B. oligonitrophilus KU-1 culture (0.5-1.0 × 10 9 cells/ml) was used for peroral administration according to previously reported scheme [11]. It is important to note here that freshly frozen bacterial culture can be used for administration. For this purpose, a small aliquot of liquid bacterial culture (e.g., 100 ml) can be frozen in a plastic container and then unfrozen before administration. B. oligonitrophilus KU-1 culture can be stored in frozen form for a month without deprivation of anticancer activity. To compare difference in overall survival, we used paired Student t test. A p value of <0.01 was considered to indicate significance.

RESULTS
Case 1 -Female, born in 1929. In 1996, sigmoid colon tumor was diagnosed. Operation on primary tumor ablation was made at Tatarstan Republican Hospital (Kazan, Russia). Two years later, tumor relapse with metastases into left ureter (left kidney was nonfunctional) and loops of thin bowels was found. Concomitant diseases were hypertension, diabetes, adiposity, and postoperative ventral hernia. In December 1998, enterostomy, ileotransversostomy, and herniotomy were performed. At bifurcation of aorta, a tumor (15 × 9 cm) was observed. Metastases into loops of small intestine and ileum were the reason for intestinal obstruction. There were carcinomatous nodes at transversal colon and small intestine. The patient was declared inoperable. The patient refused chemotherapy. Since August 1999, minimal doses of B. oligonitrophilus KU-1 culture were received (10 ml/day). Then, administration of bacteria increased up to 50 ml/day, but in September 2000, administration of bacteria stopped due to intolerance. In November 2000, the patient died due to stroke. Case 2 -Female, born in 1930. In early 1997, breast adenocarcinoma with vertebral column metastases, metastases in ribs and upper extremities determined (Т 3 N 1 М 1 ). The associated illnesses were diabetes, allergy, and renal calcinosis. Operation on primary tumor ablation was made at All-Russian Oncological Center (Moscow, Russia). In 1997, chemotherapy and radiotherapy were performed. Since March of 1997, the patient received B. oligonitrophilus KU-1 culture (400 ml/day). In July 1997, bandilization of metastases was observed. After that time, bacteria administration was stopped. In autumn of 1997, bandle state was watched. However, the administration of stimulator of bone stock growth ("Bonephos") resulted in tumor relapse and death in November 1998. Case 3 -Female, born in 1957. In late 1999, ovary tumor with metastases into liver and abdominal cavity as well as undifferentiated rectum tumor (T 4 N x M 1 ). In February 2000, hysterectomy and resection of greater omentum were made at Government State Service "Oncology" (Kazan, Russia). Sigmostoma and rectum were not ablated. There was about 3 l of ascitic fluid in the abdominal cavity. Right ovary was in the form of thick-walled cyst (20 × 18 × 15 cm). In the liver, there were three roundish formations (up to 3 cm in diameter). In the rectosigmoid section, there was Schnitzler's metastasis (2 × 3 × 1 cm). Urine analysis (May 5, 2000): weight, 1.010; pH 6.0; glucose and protein were absent. Ultrasonography (March 31, 2000): liver was heterogeneous parenchyma near the edge of costal margin. There were hyperechoic formations (in SVI, 55 mm in diameter; in SVII, 41 mm in diameter). There were foci of disintegration in the center of each hyperechoic formation. In the left lobe of liver, there was hyperechoic formation (31 mm in diameter). Gall bladder was increased. Ultrasonography (May 6, 2000): liver was with variety of hyperechoic metastatic formations of 69 mm in diameter (some of them were with fluid inclusions). Kidneys, spleen, heart, lungs, and pancreas were without pathology according to X-ray examination. In May 2000, administration of B. oligonitrophilus KU-1 culture was started. The initial level of bacteria administration was 200 ml/day; since August and October, daily administration was increased up to 400 and 600 ml, respectively. Simultaneously, topotecan was received. Ultrasonography of liver (February 2, 2001): numerous metastases (47-123 mm in diameter) were detected. During all this time, the patient received daily 500-700 ml of B. oligonitrophilus KU-1 culture. Due to liver failure, the patient died in July 2001.    (Fig. 1). The patient rejected chemo-and radiotherapy. Urine analysis (the same date): protein and glucose were absent; specific gravity, 1.002; reaction was acid. Since April 2001, the patient began to receive B. oligonitrophilus KU-1 culture (200 ml/day). Since December 2001, daily administration decreased up to 100 ml. In August 2002, total destruction of D11 and D12 bodies and significant abnormalities in D8-D10 were revealed in roentgenograph. Diagnosis: metastases in D11-D12. In July-August 2004, ultrasonography of liver, kidneys, pancreas, and spleen without pathology. In February 2004, the patient stopped administration of B. oligonitrophilus KU-1 culture considering him cured. In July 2004, computed X-ray imaging was made and cancer bandilization was observed (Fig. 2)  vertebral body with wedge-shaped deformity and slight consequent kyphosis are detected. There are sclerotic bony fragments, but the presence of lytic process is also evident; this is most conspicuous in the vertebral arches. Surrounding soft tissues are somewhat widened. There is another lytic area (approximately 1.5 × 2.5 cm) on the left anterior aspect of Th.X. vertebral body adjacent to Th.X. intervertebral space, affecting the cortical bone as well. A third lytic area is demonstrated in Th.IX. Vertebral body on the right side with a size of approximately 1 × 2.5 cm. It has lobulated contours with sclerotic margins. There are moderate sclerotic degenerative appositions at the Th.XI. facet joints. There is no significant spinal canal stenosis at this point. Lytic areas in Th.XI. vertebra, involving the arches: white arrows; lytic lesion in Th.IX. vertebral body with sclerotic margin: short yellow arrow; lesion in Th.X. vertebral body at its lower rim: red arrow.

FIGURE 2.
MRIs of patient with backbone metastases (after treatment with bacteria). There is progressive spinal deformity: collapse of the anterior part of Th.XI. vertebral body is complete with more prominent wedge-shaped deformity. Posterior sclerotic bony elements shifting towards the canal cause significant stenosis (at least 50% in AP direction). Remarkable osteophytic appositions have developed on the right lateral aspect of Th.X.-XI. intervertebral spaces. There is marked progression of sclerotic degenerative changes in the facet joints as well. There is sclerosis in the vertebral arches instead of the formerly observed lytic areas. No evidence of soft-tissue mass. The lytic area affecting the left anterior aspect of Th.X. vertebral body became demarcated by irregular slerotic margin. The lesion in the Th.IX. vertebral body is unchanged. Compression fracture of Th.XI. vertebral body was probably caused by lytic bone pathology, which seemed to affect Th.X. vertebral body as well. From the available data, the nature and dignity of this process cannot be determined with confidence; both malignant neoplasia (metastasis) and benign tumors/tumor-like lesions may be taken into consideration. The follow-up, however, reveals progression only in the secondary deformity and the accompanying degenerative changes with consolidation and demarcation of the lytic components. This could be explained by an effect of successful antineoplastic, bacterial-based treatment of a malignant tumor. The probability of malignancy would set lower. The lesion in the Th.IX vertebral body had slightly different imaging characteristics that have not changed in the follow-up period, hence it may represent benign pathology different from the one affecting Th.X-XI. segments. The most striking finding of the follow-up scan is the evolution of significant bony spinal canal stenosis. Sclerosis in place of former lytic areas in Th.XI. vertebra: white arrows; unchanged lytic lesion in Th.IX. vertebral body with sclerotic margin: short yellow arrow; lesion in Th.X. vertebral body at its lower rim with sclerotic margins: red arrow. and 5 × 7 cm) with a wide distribution into peritoneum was determined (T 4 N x M 1 ). In addition, multiple metastases into greater omentum were observed. Cystatin was received before operation. In that period, the patient was very ill. In August 2002, hysterectomy and ablation of greater omentum were made at Samara Oncology Center (Samara, Russia). Since September 2002 until February 2003, the patient received B. oligonitrophilus KU-1 (300 ml/day). Increase in intracranial pressure was observed while blood pressure was normal. After CA125 normalization, bacteria were received with prophylactic aim (50 ml/day, a week of administration was changed by a week of interruption). In May 2004, ultrasonography revealed metastases into cellular tissue of pelvis minor. Chemotherapeutic course of treatment (cystatin, 110 mg) was perceived very badly (there were retching and asthenovegetative syndrome). Since June 2004, the patient increased administration of B. oligonitrophilus KU-1 (300 ml/day). According to ultrasonography data, size of metaplastic cancer in cellular tissue of pelvis minor decreased up to 30%. Then, the patient continued to receive B. oligonitrophilus KU-1 (50 ml/day, a week of administration changes by a week of interruption). In January 2005, the patient was hospitalized due to relapse of thrombophlebitis. Clot was operated. The patient received antithrombin drugs.  in the right lobe: 27 × 25 mm at S 6 -S 7 ) with fluid substances (10 mm in diameter), which are characteristic feature of undifferentiated tumor. Pancreas, spleen, and gall bladder were without pathology. In that period, the patient received 5-fluorouracil with leucovorin. Since April 2004, chemotherapy was absent; administration of B. oligonitrophilus KU-1 was increased up to 500 ml/day. Ultrasonography data (August 2004): liver metastases increased up to 7 cm in diameter, metastases in epicolon (until 15 mm in diameter), and retroperitoneal lymphatic glands were also revealed. In August 2004, the patient made cruise along Volga River. In the end of August, the patient worked at vegetable garden (he transported pails with potatoes). In September 2004, there was further decrease in state of health. Since September 2004, the supporting therapy (vicasol, prednisolone) was received. Since the end of September 2004, there was a drastic decrease in health state, quick development of jaundice and cachexy. Also, there was no appetite; insomnia and oedemata of legs were presented. The patient died in October 2004.
For comfort of data comparison, some key information is presented in Table 1. At present, there are three patients alive with rectal adenocarcinoma, thyroid adenocarcinoma, and in situ cancer. Their current survival periods are 7, 3, and 9 years, respectively. The patient with rectal adenocarcimoma (female, born in 1937) receives B. oligonitrophilus KU-1 regularly twice a year with prophylactic aim, while the two others have no restrain.

DISCUSSION
The ability of some probiotic microorganisms to reduce the risk of cancer is a well-established phenomenon. It was shown that probiotics could reduce risk of cancer in the gastrointestinal tract [13,14] as well as at other sites of the human organism [15,16]; therefore, it is reasonable to suggest that probiotics have a systemic effect rather than a local one [17]. Although our results confirmed cancer-preventive properties of B. oligonitrophilus KU-1, the main finding is that peroral administration of B.
oligonitrophilus KU-1 may be a promising treatment approach. Assuredly, our research had a significant limitation because it was not double-blind or placebo-controlled. Also, computed tomography and magnetic resonance tomography data were absent for the majority of the studied patients. Four patients received conventional treatments in parallel. In all patients, however, deterioration in health state was detected after administration of cytostatic agents and radiotherapy. In this connection, it is interesting to note that parallel administration of B. oligonitrophilus KU-1 allowed us to downplay side effects of chemo-and radiotherapy, since hematological status was normalized owing to applied bacterium. Fig. 3 demonstrates survival periods for the studied patients. It is clear from the figure that all patients demonstrated increased survival period in comparison with the standard prognosis given by the physician and presented in the literature [18]. Table 2 shows that the observed differences were significant. Concerning effectiveness of our method, the following remarks should be made. Of course, prognosis depends on various factors like type of cancer, stage of disease, age, and environmental factors. However, we consider that some approximation can be made. Namely, life prolongation is possible (approximately 12-18 months over standard prognosis) in patients with undifferentiated tumors. In patients older than 40 and younger than 60 with moderately differentiated tumors, elevation of survival over standard prognosis can reach 2 years. In patients with high-grade differentiated tumors and an absence of metastases into the vitally important organs (i.e., liver or lungs), life prolongation can reach 2-3 years over standard prognosis. Finally, we consider that total elimination of revealed metastases is possible in patients older than 60 (especially in case of breast cancer). Unfortunately, our method may be ineffective in children, especially with undifferentiated tumors. We also consider some recommendations important for a favorable outcome: a diet without preservatives, emulsifiers, synthetic dyes, smoked food, roast, marinades, pickles, or spirituous liquors; sour-milk diet, boiled veal meat and fish, boiled buckwheat, porridge, millet gruel, boiled rice, vegetables, and green tea are the most recommended eatables, while supercooling, superheating, isolation, psychic anxieties, and stress are contraindicated.
Finally, therapeutic efficiency, safety, and low price allow us to conclude on the principal utility of the suggested method. We hope that our results will be confirmed in double-blind, placebo-controlled clinical trials.