Parvum Administration Research Articles

Innate Immunity Stimulation during the COVID-19 Pandemic: Challenge by Parvulan.

Aim We report an open spontaneous anecdotical retrospective survey of Corynebacterium parvum administration to 4000 fragile immune-depressed and multimorbid patients treated with a killed C. parvum strain to enhance innate immunity, integrating the adaptative immune response for long-standing antinfectious resistance. Methods A total of 4000 patients (1900 men and 2100 women) with mild, moderate, or chronic disease, appealing to our Second Opinion Medical Consultation Network, signed an informed consent form and were injected subcutaneously with C. parvum. The treatment was followed up to 6 months, completing the short form of the medical outcome health survey questionnaire (SF-36) directly by the patients or their parents and monitoring their health status regularly via telemedicine (Skype, WhatsApp, mail, etc.) or outpatients visits. Results The main efficacy endpoints, as assessed by the SF-36 questionnaire are: significant improvements in the mental and physical role functioning score (p < 0.02), better general health; social role performance (p < 0.02), vitality (p < 0.03), and a significant pain reduction (p < 0.03). A quick (48-72 hours) symptoms improvement and/or complete regression of the herpetic eruptions was observed in 1000 affected patients with disappearance or relieve of herpetic neuralgia (reduced in 80% of cases); also full recovery or frequency reduction (30%) of recurrent cystitis and prostatitis in 120 affected patients. Last but not least, a life quality improvement in 100 oncologic patients of overall 200 cases. A significant increase in the lymphocyte count (p < 0.01), mainly helper and killer lymphocytes, was noted 6 months after Parvulan injection vs. the baseline. The asymptomatic SARS-CoV-2 patients who were incidentally enrolled in our survey were tested at the sixth month for antibodies against SARS-CoV-2, and 14 patients had high levels of SARS-CoV-2 antibodies. The incubating COVID infections of the Parvulan-injected patients even if frail and multimorbid recovered in short term (48-96 hours) with a benign clinical course, without need of drugs administration except for the variants, such as Delta and Omicron, whose infections lasted on average one week and required some antipyretics and low-dose steroids for a few days. Conclusions Our results confirm that C. parvum is quite safe and effective in supporting immune-compromised patients when epidemic or pandemic events increase the life risk and any kind of infection and complication rate. Further double-blind placebo evidence-based studies are urgently required, and our numerically substantial not sponsored spontaneous observation is exclusively intended to promote further evidence-based double-blind institutional studies.

Electron Transport Chain Activity in Normal and Activated Rat Macrophages

The pivotal role played by the macrophage in specific and nonspecific immunity suggests that the physiological status of the macrophage may effect the overall regulation of the host defense system. Many studies have evaluated macrophages as effector cells by examining expression of surface markers, cytokine release, or tumor killing in the presence of challenge to host defenses. In this report, the physiological parameter of mitochondrial respiration in freshly isolated rat macrophages is shown to be regulated upon activation in vivo. Assay conditions for the reduction of MTT [3(4,5-dimethylthiazol-2)-2,5-diphenyltetrazolium bromide] in rat macrophages were optimized and used to quantitate electron transport chain activity as a measure of mitochondrial respiration. Corynebacterium parvum administration significantly increased the activity of the mitochondrial electron transport chain in both peritoneal (120% increase, 0.18 ± .01 vs 0.40 ± .03, P < 0.01) and liver macrophages (143% increase, 0.12 ± .02 vs 0.30 ± .06, P ± 0.01) as detected by augmented MTT reduction. It is demonstrated further that MTT reduction is distinct from the respiratory burst activity of macrophages and supports the mitochondrial localization of intracellular MTT reduction in this cell type. These results demonstrate that electron transport chain activity is a physiological indicator of macrophage activation.

Use of prostaglandin I2 analog in treatment of massive hepatic necrosis associated with endothelial cell injury and diffuse sinusoidal fibrin deposition.

Endothelial cell damage causes massive hepatic necrosis as a result of fibrin deposition in the hepatic sinusoids. When a stable analog of prostaglandin I2, beraprost sodium, was administered to rats given either dimethylnitrosamine, carbon tetrachloride, or endotoxin following Corynebacterium parvum administration, the hepatic necrosis produced in each was attenuated, but to a greater extent in the dimethylnitrosamine and endotoxin/Corynebacterium parvum models, where fibrin deposition in the hepatic sinusoids occurs, as compared to the carbon tetrachloride model, where such fibrin deposition does not occur. Beraprost sodium reduced the expected increase of portal venous pressure in the endotoxin/Corynebacterium parvum model without affecting plasma thrombin-antithrombin III complex levels. Beraprost sodium also significantly reduced cell killing of both isolated rat hepatocytes and hepatic sinusoidal endothelial cells exposed to tert-butyl hydroperoxide when compared to controls. Beraprost sodium could prove to be a therapeutic candidate for the treatment of hepatic necrosis, particularly in cases associated with fibrin deposition in the hepatic sinusoids because of its fibrin clot-clearing action.

Nitric Oxide Synthase Expression Is Induced in Hepatocytes in Vivo during Hepatic Inflammation

Nitric oxide (NO ·) is a short-lived biologic mediator produced by the enzyme NO · synthase (NOS) which exists in constitutive and inducible isoforms. Previously, we have shown that hepatocytes express an inducible NOS in vitro following exposure to the combination of lipopolysaccharide and inflammatory cytokines. The purpose of the present study is to characterize the induction of NOS in vivo in rat hepatocytes during chronic hepatic inflammation triggered by Corynebacterium parvum injection and to correlate NO · synthesis with the timing of liver injury. Using Northern blot hybridization, hepatocyte-inducible NOS mRNA was detected 3 days after C. parvum administration and was not found in normal hepatocytes. Hepatocyte NOS activity was significantly increased 3 to 7 days after C. parvum. Plasma concentrations of nitrite and nitrate (NO - 2 + NO - 3), the stable end products of NO · oxidation, increased from a basal concentration of 21.0 ± 2.5 to 2439.6 ± 364.2 μ M 3 days after injection. Urinary excretion of NO - 2 + NO - 3 also increased in a parallel manner. Plasma liver injury enzymes were elevated three to sixfold in vivo at 3 to 5 days following C. parvum and coincided with the period of maximal NO production. The results show that NO · is produced directly by hepatocytes in vivo during hepatic inflammation and suggest a role for NO · in mediating the hepatic response to inflammatory stimuli.

Alteration in Natural Defense Activity Against NKSusceptible B16 Melanoma Cells after Treatment with Corynebacterium parvum

We have investigated the effects of administration of C. parvum on host anti-metastatic activity against B16 melanoma H-2L, a natural killer (NK) sensitive clone with a low expression of H-2 b. The anti-metastatic activity was estimated by monitoring the following two points. One was the survival ratio at an early stage after an intravenous (iv) inoculation of radiolabeled B16 H-2L cells, the other was the formation of pulmonary metastases after iv injection with the tumor cells. Administration of C. parvum showed a biphasic change in the NK activity of the spleen cells and the peritoneal exudate cells (PEC) in mice. At an early phase (2–3 days) after administration of C. parvum, the NK activity of the spleen cells and PEC was significantly augmented. On the other hand, at a later phase (14 days) after C. parvum administration, the NK activity was deeply depressed. In correlation with NK activity of the mice treated with C. parvum, the anti-metastatic activity of the hosts was augmented in the early phase, whereas a depressed level of anti-metastatic activity was observed in the late phase after administration of C. parvum. These results suggest that the modification of NK activity is a possible basis for modulation of anti-metastatic activity by C. parvum.

Toxicity of IL-2 and Corynebacterium parvum following direct intracranial injection.

The toxicity of IL-2 and/or C. Parvum when injected directly into the mouse brain was examined by survival and histopathology using different numbers of injections, different doses of IL-2, different solvents for the IL-2, and different doses and routes of administration of C. Parvum. Two injections was found significantly to increase mortality (19%) over a single injection (4%). Mortality from two injections of 30,000 U (23%) or 60,000 U (20%) was higher than from two injections of 15,000 U (12%). The mortality from two injections with normal saline as solvent was much higher (29%) than from two injections with sterile water (19%) or D5W (9%). Two injections of IL-2 given simultaneously with C. Parvum showed a much higher mortality (26%) than other doses and routes of C. Parvum administration. Mice dying acutely (6-24 days) of toxicity showed an extensive mononuclear infiltrate at the site of injection. The brains of surviving mice (sacrificed at 30 days) showed a mild residual mononuclear cell infiltrate with the exception of mice which had received IL-2 and C. Parvum simultaneously. Brains from this latter group had an extensive residual mononuclear cell infiltrate.

Splenocytes from tumor-bearing Corynebacterium parvum treated mice maintain interleukin-2 and -3 activity.

Systemic and local administration of the bacterium Corynebacterium parvum (more accurately known as Propionibacterium acnes) is reported to exert antitumor action via activated macrophages or short-lived cytotoxic T lymphocytes (CTL), respectively. This study examined the effect of C. parvum treatment on resulting in vitro interleukin levels, which are components in the sequence of events leading to the development of effective CTL. C. parvum administration prevented palpable fibrosarcoma development. This was concomitant with restoration and maintenance of normal interleukin-2 (IL-2) and interleukin-3 (IL-3) levels and prevention of suppressor cell development in mice injected with both tumor cells and vaccine. Our finding of C. parvum-induced maintenance of IL-2 and IL-3 levels and apparent lack of suppressor cell formation lends support to the idea of local C. parvum antitumor action possibly being mediated by CTL arising via the interleukin cascade.

The quantity and function of pulmonary alveolar macrophages after splenectomy and Corynebacterium parvum.

We have previously shown that Corynebacterium parvum (C. parvum), a nonspecific immunomodulator, partially protects splenectomized and nonsplenectomized mice when challenged with aerosolized pneumococci. We report here the effects of both splenectomy and C. parvum on the phagocytic function of the lavageable pulmonary alveolar macrophage (PAM). Groups of young adult male Sprague Dawley rats underwent splenectomy or sham operation 3 weeks before injection of C. parvum 1.5 mg IP (or saline) per animal. One week postinjection PAM's were harvested. The in vitro phagocytic indices (PI) for PAM incubated with tritiated thymidine-labeled S. aureus or Streptococcus pneumoniae, type 14, opsonized with normal rat serum, were determined. Splenectomy had no effect on lung weights, PAM yield, or PAM phagocytic activity. C. parvum administration significantly increased spleen weight in sham-operated animals, but had no effect on lung weights, PAM yield, or phagocytic activity of either control or splenectomized animals. Splenectomy in the adult rat did not induce a phagocytic defect in the PAM and thus the lavageable PAM cannot be considered a significant site of the postsplenectomy defect. Since C. parvum protects animals from respiratory challenge with Streptococcus pneumoniae but does not alter the number or activity of lavageable alveolar macrophages, we hypothesize that C. parvum protection is more likely related to our previous finding of an increased clearance of blood-borne bacteria by the expanded and enhanced reticuloendothelial system.

Intraperitoneal administration of Corynebacterium parvum in patients with ascitic ovarian tumors resistant to chemotherapy: effects on cytotoxicity of tumor-associated macrophages and NK cells.

AbstractEight ovarian cancer patients with ascites resistant to various chemotherapy protocols were treated i.p. with C. parvum in an effort to obtain a palliative reduction of peritoneal effusions. C. parvum(7–14 mg) was given i.p. on days 0, 7, 28 and then at monthly intervals. Three patients showed no clinical evidence of therapeutic benefit from C. parvum administration. Three subjects had a complete disappearance of ascites, and two additional patients showed a marked reduction of their effusions. Solid tumor masses were not affected by i.p. C. parvum. C. parvum injection caused a rapid decrease of the percentage of tumor cells in the ascites as early as 7–15 days after the first treatment. Concomitantly, the percentage of polymorphs and, to a lesser extent, of macrophages in the ascitic tumor increased, whereas tumor‐associated lymphoid cells (TAL) were not altered in a consistent pattern. The tumoricidal activity of peripheral blood monocytes and natural killer (NK) cells was not consistently modified after C. parvum treatment. Pretreatment NK activity of TAL was usually very low and was not appreciably modified by i.p. C. parvum. In one patient, whose TAL had NK activity in the range of normal PBL, C. parvum administration was followed by a profound depression of intratumor NK cytotoxicity. The tumoricidal activity of tumor‐associated macrophages (TAM) was not enhanced following C. parvum administration; 14–21 days after the first C. parvum inoculation, a marked decrease of the cytolytic capacity of TAM was observed, cytotoxicity returning to pretreatment values thereafter. These observations do not elucidate the mechanism of the limited, but significant, anti‐tumor activity of i.p. C. parvum in human ovarian carcinomatous ascites. The lack of enhancement of macrophage cytotoxicity after i.p. C. parvum, predicted on the basis of data in rodents, cautions against the empirical use in humans of agents assumed to affect human neoplasia through activation of host defense mechanisms.

Inhibition of cell-mediated cytotoxicity by Corynebacterium parvum

We investigated the effect of altering dose and route of Corynebacterium parvum (C. parvum) administration on the adjuvant's inhibition of cell-mediated cytotoxicity (CMC). Primary in vivo and secondary in vitro CMC of C57B1/6 mice alloimmunized to P815 were depressed if C. parvum was administered systemically (IV or IP) but not when it was given SC. Similarly, only systemic C. parvum generated cells capable of suppressing in vitro CMC. Primary and secondary CMC in spleen was equally inhibited by 700 and 70 μg, whereas suppressor cell activity was marked with 700 μg and minimal with 70 μg. Administration of C. parvum SC admixed with alloantigen resulted in early enhancement and late depression of primary CMC. Secondary CMC was depressed but suppressor activity was absent. Dissociation of CMC depression from suppressor cell generation indicates that these phenomena can be separated under certain conditions.

Dissociation of biological activities of Corynebacterium parvum by chemical fractionation

Fractionation of Corynebacterium parvum (C. parvum) by phenol-water extraction resulted in components which differentially affected the cell-mediated cytotoxic response to tumor alloantigens in mice. The residue of the extraction (Fraction C) and its light component (Fraction D) retained all of the activities of the whole micro-organism, i.e. inhibition of primary spleen cell cytotoxicity, inhibition of in vitro memory cell-mediated cytotoxicity (CMC) and generation of macrophage-like suppressor cells. Administration of the phenol phase extracted material (Fraction A) inhibited both the primary and memory cytotoxic responses, but did not elicit suppressor macrophages. Destruction of carbohydrate components of Fraction D significantly reduced its ability to inhibit the memory response and to elicit suppressor macrophages. In contrast, its ability to inhibit the primary cytotoxic reponse was unaffected. This study clearly demonstrates that the several effects of C. parvum administration on the cell-mediated cytotoxic response to tumor alloantigens can be associated with different components of the organism. Furthermore, the dissociation of the effects indicate that the generation of primary cytotoxic lymphocytes, the priming of memory cytotoxic lymphocytes, and the induction of regulatory macrophages are not necessarily linked during the generation of a normal cytotoxic response.

Inhibition of memory cell-mediated cytotoxic response by systemic administration of Corynebacterium parvum

Intravenous administration of Corynebacterium parvum to mice during a developing immune response to alloantigens resulted in the marked inhibition of the generation and expression of memory cell-mediated cytotoxic response in the spleen. The inhibition was observed following rechallenge in vivo or by in vitro culturing with the same alloantigen. The impairment in vitro was due, in part, to the generation of regulatory cells which were non-T phagocytic cells, probably macrophages activated by C. parvum administration. These suppressor macrophages appear to act by inhibiting proliferation and clonal expansion of memory cytotoxic cells.

Role of mononuclear phagocytes in decreased hepatic drug metabolism following administration of Corynebacterium parvum

Administration of BCG, Corynebacterium parvum and a number of chemical immunostimulants is known to cause marked decreases in hepatic microsomal drug metabolizing enzyme activity. In this study we found that the per cent inhibition of drug metabolism 7 days following C. parvum vaccine (350 μg per mouse i.p.) was related directly to the extent of splenomegaly. Prior splenectomy, or whole body irradiation, performed concurrently or 2 h after administration of C. parvum prevented the decrease in microsomal enzyme activity. Silica and carrageenan, which are known to lyse macrophages, also blocked the C. parvum effect on drug metabolism, but only if injected 2 h prior to C. parvum administration. A strain of C. parvum with little antitumor activity produced slight splenomegaly and elevation of serum LDH, but drug metabolizing enzyme activity and cytochrome P-450 content remained normal. Characteristic focal intrahepatic inflammatory lesions, consisting principally of macrophages and lymphocytes with adjacent necrotic hepatocytes, were found in splenectomized mice with normal levels of microsomal enzyme activity and cytochrome P-450. However the number and severity of inflammatory lesions generally were related to these parameters and to serum transaminase levels. These results define a role for the reticuloendothelial system in the depression of drug metabolism found after administration of C. parvum.

An evaluation of corynebacterium parvum during remission maintenance therapy in pediatric patients with acute lymphoblastic leukemia

The biologic impact and clinical toxicity of Corynebacterium parvum administered at a dose of 5 mg/M2 by intravenous or subcutaneous routes were evaluated in 18 children receiving combination chemotherapy for maintenance of acute lymphoblastic leukemia (ALL) in remission. Several nonspecific immunologic and hematologic parameters were evaluated. Patients were also monitored for changes in cutaneous sensitivity to histamine. No changes in any parameter were observed in patients after only one course of C parvum injection. However, after 6-10 courses, glass-adherent peripheral blood leukocytes of C parvum-treated patients augmented the response of PHA-stimulated autologous lymphocytes. In all nine patients studied who received C parvum injection subcutaneously for at least six months, there were significant increases in the mean bone marrow myelocyte-erythrocyte (ME) volumes compared to pretreatment values. These results suggest that periodic evaluations are desirable in patients receiving repeated administration of C parvum, since changes in immunologic and hematologic responses may be demonstrable only after several injections. In contrast to the reported experience in adults, subcutaneous C parvum administration children was not well tolerated, whereas intravenous infusion was generally well tolerated.

Toxicity manifestations following intravenous Corynebacterium parvum administration to patients with ovarian and cervical carcinoma

Manifestations of clinical toxicity were evaluated following 341 courses of intravenous Corynebacterium parvum adjuvant immunotherapy in patients with ovarian and cervical carcinoma. Most patients exhibited symptoms of minor toxicity, which decreased in intensity as subsequent courses of therapy were administered. Temperature elevations to 38.5° C. were the most objectively measured signs of toxicity but temperature elevations greater than 38.5° C. occurred following only 20.5 per cent of the infusions. Blood pressure alterations were not a serious problem and no serious central nervous system or renal toxicity was noted. The minor side effects should not preclude the use of C. parvum as an immunopotentiating agent if it is shown to be beneficial in the treatment of human malignant disease.

Corynebacterium parvum-induced radiosensitivity and cycling changes of hematopoietic spleen colony-forming units.

Ten days after total-body irradiation with 550 rads of b0Co, spleen colonies were observed in adult C57BL mice. A change in radiosensitivity induced by Corynebacterium parvum, as measured by increased numbers of colony-forming units that survived the 550 rads, began shortly after C. parvum stimulation and extended for at least 7 days before irradiation. C. parvum given 4-24 hours before, followed by high specific activity [3H]thymidine (HSATT) 1 hour before total-body irradiation greatly reduced survival of the stem cells that formed spleen colonies (CFUs) and CFUs radiosensitivity to control levels. The HSATT sensitivity by "suicide" assay in vivo and the time-response change in radiosensitivity corresponded with the decrease in radiosensitivity, which showed that CFUs were stimulated by C. parvum administration and entered the S-phase shortly after stimulation. The data indicated a resting population close to the S-phase. After stimulation, this population entered S-phase. Syngeneic mouse lymphoma cells injected iv 24 hours earlier did not elicit any effect as a stimulus to CFUs radiosensitivity change.

In Vitro Antitumor Effect of Lymphoid Cells From Corynebacterium parvum-Treated Mice: Effect of Route of C. parvum Administration2

Corynebacterium parvum administration in CBA mice resulted in the stimulation in the peritoneal exudate and other lymphoid organs of cells which nonspecifically inhibited the tumor growth in vitro. The distribution of the antitumor activity in different organs was dependent on the route of C. parvum injection. Both ip and iv treatments stimulated antitumor activity in the peritoneal exudate, whereas the sc route was ineffective. Only iv treatment stimulated the antitumor activity in the blood, and only the sc route produced the antitumor activity in the lymph node. The antitumor activity following the sc treatment was confined mainly to nodes draining the site of C. parvum injection. All routes of treatment provoked some antitumor activity in the spleen, although the degree of the activity varied with the route in the order of ip, iv, and sc. The antitumor activity in all organs peaked between 4 and 14 days after C. parvum injection. This activity in all organs, except the spleen, disappeared by day 42 post treatment. The results are discussed in the light of the in vivo observations on the antitumor effects of C. parvum.

Observations followingCorynebacterium parvum administration to patients with advanced malignancy.A phase I study

There has been increasing interest regarding the use of Corynebacterium parvum (CP) with other modalities in the management of primary cancer. Due to the paucity of specific information available relative to CP toxicity, a Phase I study was carried out in patients with advanced disease. The purpose of the investigation was not to evaluate the effect of CP on tumor growth. from 273 injections of CP in 40 patients it was observed that following intravenous (i.v.) infusion of CP: a) a febrile response and chills of considerable severity occured in almost all patients and did not appreciably diminish in intensity following repetitive administrations; b) nausea, vomiting, headache, and confusion were not infrequent; c) a "flu-like" syndrome lasting 24 to 48 hours occurred following almost all courses of CP; d) blood pressure elevations occurred on occasion and were related to the severity of other-side-effects; hyper- or hypo- tension was not a problem; e) ther were no anaphalactic reactions. Pretreatment with a single administration of 100 mg of hydrocortisone prior to CP infusion markedly and in some instances dramatically diminished the toxicity and made acceptable the use of i.v. CP on an outpatient basis. The use of i.v. CP in patients with cerebral metasteses may be hazardous. Subcutaneously administered CP resulted in a significant number of undesirable local reactions. Evaluation of delayed cutaneous hypersensitivity response, immunoglobulins, complement, and E- and EAC-rosette-forming cells during CP administration failed to demonstrate significant change from injection values. Results were similar whether hydrocortisone pretreatment was or was not employed. From the standpoint of toxicity it now seems appropriate to use i.v. CP, particularly following pretreatment with hydrocortisone, in a controlled clinical trial to evaluate its therapeutic effectiveness in the management of primary cancer.

Antitumor antibodies and immunoglobulin class and subclass levels in Corynebacterium parvum-treated mice.

Changes in immunoglobulin class and subclass levels and the development of antitumor antibodies were assessed in normal and tumor-bearing mice challenged with Corynebacterium parvum. C. parvum administration resulted in a marked increase in certain immunoglobulin levels, especially Ig G2b, and in the development of antibodies reacting with syngeneic and allogeneic tumor cells. The serologic changes induced by C. parvum were dependent on the dose and route of administration; preliminary studies suggested that they may have been largly independent of T-cell function. These changes were suppressed by the administration of gold salts, which also inhibited the antitumor effect of C. parvum.

The effect of Corynebacterium parvum therapy on immunoglobulin class and IgG subclass levels in cancer patients.

Detailed serological studies have been undertaken in a small group of cancer patients receiving nonspecific immunotherapy with Corynebacterium parvum (C. parvum). These patients included 4 cases of recurrent malignant melanoma, 2 of stomach cancer and 2 of recurrent breast cancer. They all received an initial i.v. infusion of 20 mg of a formol killed suspension of C. parvum followed by 2 mg (i.m.) at weekly intervals for 10-11 weeks. This protocol consistently resulted in an increase in the circulating IgG levels of all patients but had a variable effect on their IgA, IgM and IgE levels. Increases in the concentration of all 4 IgG subclasses contributed to the overall increase in IgG levels and these changes ranked IgG2 greater than IgG1 greater than IgG3 = IgG4. It also had an inconsistent effect upon the levels of alpha-macroglobulin in pregnancy but the levels of normal serum alpha2-macroglobulin were virtually unchanged. Pre-existing antibodies to C. parvum were noted in all the patients. Titres rose appreciably following C. parvum administration and remained at high, though fluctuating levels, throughout the 100-day period of observation. Absorption studies suggested that the development of antibodies to C. parvum accounted in part for the increased IgG levels noted following this form of therapy. The significance of these changes in relation to the possible anti-tumour effect of C. parvum is discussed.