Immunotherapy of osteogenic sarcoma with transfer factor

Summary Fifteen patients with osteogenic sarcoma were treated with transfer factor derived from leukocytes of their household contacts. Eight of the fifteen patients were tumor-bearing, and transfer factor therapy was correlated with increased cell-mediated immunity in peripheral blood lymphocytes and with lymphocytic infiltrates into the tumors. There was no marked increased in survival time as compared with historical controls, but this therapy did not accelerate the disease, and there were no untoward side effects.Seven of the fifteen patients were disease-free when transfer therapy was initiated shortly after surgical removal of the primary tumor (five patients) or solitary pulmonary metastases (two patients). These patients received transfer factor injections every 2 weeks for 1–2 years. Six of the seven patients are disease-free 62–82 months after surgery. Compared with probabilities of 5-year survival computed from historical controls, this is significant at P<0.008. Abbreviations used in this paper are: CI, cytotoxicity index; GCT, giant cell tumor of bone; HHC, household contacts; HHC_os, household contacts of patients with osteogenic sarcoma; MIC, mononuclear inflammatory cell; TFCI, transfer factor cytotoxicity index; TSTF, tumor-specific transfer factor.     

Cellular immunity in patients with malignant melanoma and their household contacts

Summary Immune reactivity was measured in control subjects and in 86 patients with malignant melanoma using four tests of cellular immunity. In addition, cellular immune reactivity to melanoma extract was studied in 13 household contacts of patients with melanoma. Thirty percent (4/13) of the household contacts showed reactivity to melanoma extract as determined by lymphocyte stimulation as compared to 20% of patients and 5% of controls. Seventy-two percent (8/11) household contacts showed reactivity as measured by the indirect MIF test, compared to 38% of patients and 23% of normal controls. As determined by the leukocyte migration inhibition test, 71% (5/7) of household contacts showed reactivity to melanoma extract versus 20% of patients and 22% of controls. The number of household contacts studied was low; however, it was observed that these subjects showed responses to melanoma extract with considerably greater frequency than did normal control subjects, and the frequency of positive responses in patients with melanoma was intermediate between that of the household contacts and the normal controls.This work was supported in part by grants AI-10495, AI CA-10686, and CA 13671 from the U.S. Public Health Service. Dr. Spitler is supported by a National Institutes of Health Career Development Award (AI-43012).     

Case report: Metaplastic carcinoma presenting as a breast abscess

Metaplastic breast carcinoma (MBC) is a rare neoplasm containing a mixture of epithelial and mesenchymal elements. The epithelial component is usually ductal carcinoma but may include other variants of breast carcinomas including squamous carcinoma and osteogenic sarcoma. There is a relative paucity of data regarding such tumours. Metaplastic carcinoma carries a prognosis not dissimilar to that of comparable ductal carcinoma. This is the case of a 57 year old patient with MBC presenting with a breast abscess. A thorough literature search has not revealed any previous reports of MBC presenting as a breast abscess.     

Immunospecificity of non-histone chromosomal proteins in 89Sr-induced osteogenic sarcoma (mouse).

Antibodies against non-histone chromosomal proteins for 89Sr-induced osteogenic sarcoma (mouse) were prepared by immunization of rabbits. The immunoreactivity of this antigen was then compared with those of non-histone chromosomal proteins from Ehrlich ascites tumor, normal mouse liver, and calf thymus by the method of quantitative microcomplement fixation. The non-histone chromosomal proteins of 98Sr-induced osteogenic sarcoma, fractionated by hydroxylapatite chromatography, exhibited significant affinity for the antibodies. Similar proteins from Ehrlich ascites tumor, normal mouse liver, or calf thymus were virtually inactive, indicating the tissue-specificity of 89Sr-induced osteogenic sarcoma proteins.     

Development of cellular anti-tumor immunity in chickens bearing tumors induced by Rous sarcoma virus.

The technique of peripheral lymphocyte stimulation in response to antigen was used to demonstrate the existence of cell-mediated anti-tumor immunity in chickens bearing tumors induced by avian sarcoma viruses. The expression of this anti-tumor response against virus-containing transformed cell culture supernatant fluids and cell extracts varied among animals in terms of time after inoculation with the oncogenic agent. Animals whose tumors had completely regressed rapidly lost the ability to mount such continuing immunity. In addition, we found evidence that some normal animals express endogenous levels of natural immunity against neoplasms induced by these viruses and/or against the viruses themselves.     

Inhibition of normal allogeneic lymphocyte mitogenesis by a factor released from human tumor cells in culture

Summary Human tumor and normal cell lines in culture were examined for the release of factors capable of inhibiting lymphocyte blastogenesis. Supernatants from tumor cell cultures of melanoma, carcinoma (lung, colon, breast), sarcoma, and normal fibroblasts inhibited normal lymphocyte response to PHA. Only supernatants from the tumor cell lines C1 (colon carcinoma), 734B (breast carcinoma), and 231 (breast carcinoma) were found to inhibit both PHA and ConA responses significantly. The two breast carcinoma cell lines, 734B and 231, which also were capable of inhibiting lymphocyte responses to PPD and alloantigens, were investigated further. The inhibition of lymphocyte blastogenesis caused by the supernatant of these two cell lines could not be overcome by the addition of added mitogen. Further experiments showed that the inhibition was not due to nutrient deficiencies and the supernatants were not directly toxic to the lymphocyte cultures as judged by trypan blue exclusion.     

Synchronous primary mammary osteosarcoma and invasive breast cancer. A case report – Pathohistological and immunohistochemical analysis

Primary osteogenic sarcoma of the breast is a rare neoplasm, diagnosed mainly by pathohistological and immunohistochemical analysis.We hereby present a case of primary osteogenic sarcoma in the right breast of a 62-year-old woman with synchronous appearance of an invasive ductal carcinoma. Clinical findings are manifested with two separate painless formations 2.5 cm/2 cm and 1.5 cm/1 cm in size, located on the border of the upper and lower lateral quadrant of the right breast. No axillary lymphadenopathy was diagnosed. The pathohistological and immunohistochemistry findings of both tumors revealed a synchronous manifestation of two distinct neoplasms – epithelial and non-epithelial. Multimodality treatment consisted of Patey''s radical mastectomy; 3 cycles of adjuvant chemotherapy; postoperative 50 Gy radiotherapy to the chest wall followed by additional 3 cycles of chemotherapy and anti-estrogen hormonotherapy.Due to the rarity of osteogenic mammary sarcoma, even more so in a combination with epithelial breast tumors, its clinical features are unclear and optimal treatment remains controversial. Considering the poor prognosis of the combination of both malignomas, we discuss a number of diagnostic and therapeutic issues.     

Evidence of Hepatitis A Virus Person-to-Person Transmission in Household Outbreaks

The person-to-person transmission of the hepatitis A virus primarily occurs in enclosed spaces, particularly in the presence of inadequate hygiene conditions and a high proportion of susceptible individuals. Thus, intimate family contact stands out as a risk factor for HAV infection dissemination. The present study aimed to evaluate the occurrence of household HAV transmission. Blood samples were collected from patients with hepatitis A (index cases) and their family members (contacts) that were referred to an ambulatory care clinic specializing in viral hepatitis. A total of 97 samples were collected from 30 families with a confirmed hepatitis A case (index case). Serological and molecular techniques for the diagnosis of hepatitis A were conducted on all samples. HAV infection (anti-HAV IgM + and/or HAV RNA +) was detected in 34.3% (23/67) of the contacts; 34.3% (23/67) of the contacts were immune to HAV, and 31.4% (21/67) were susceptible. In the household contacts, HAV immunity was significantly associated with older age; susceptibility to infection and HAV infection were associated with younger age. Household outbreaks were detected in 16/30 families studied. Co-circulation of subgenotypes IA and IB was found in the household outbreaks, and person-to-person transmission was evidenced in six of the household outbreaks, with 100% homology between the index case and contact strains. The results demonstrated the relevance of HAV household transmission, reaffirming the need for hepatitis A vaccine administration in susceptible contacts and effective infection control procedures to prevent the extension of household outbreaks.     

The effect of anti-VEGF therapy on immature myeloid cell and dendritic cells in cancer patients

Impairment of dendritic cells (DC), the most effective activators of anticancer immune responses, is one mechanism for defective antitumor immunity, but the causes of DC impairment are incompletely understood. We evaluated the association of impaired DC differentiation with angiogenesis-associated molecules D-dimer, vascular endothelial growth factor (VEGF), urokinase plasminogen activator (uPA), and plasminogen activator inhibitor (PAI-1) in peripheral blood from 41 patients with lung, breast, and colorectal carcinoma. Subsequently, we studied the effect of administration of the anti-VEGF antibody (bevacizumab) on DC maturation and function in vivo. Compared with healthy volunteers, cancer patients had a bias toward the immunoregulatory DC2, had deficits in DC maturation after overnight in vitro culture, and had a significant increase in immature myeloid cell progenitors of DC (0.50 +/- 0.31% vs. 0.32 +/- 0.16% of peripheral blood mononuclear cells, respectively, P = 0.011). A positive correlation was found between the percentage of DC2 and PAI-1 (R = 0.50) and between immature myeloid cells and VEGF (R = 0.52). Bevacizumab administration to cancer patients was associated with a decrease in the accumulation of immature progenitor cells (0.39 +/- 0.30% vs. 0.27 +/- 0.24%, P = 0.012) and induced a modest increase in the DC population in peripheral blood (0.47 +/- 0.23% vs. 0.53 +/- 0.30%). Moreover, anti-VEGF antibody treatment enhanced allo-stimulatory capacity of DC and T cell proliferation against recall antigens. These data suggest that DC differentiation is negatively associated with VEGF levels and may be one explanation for impaired anticancer immunity, especially in patients with advanced malignancies.     

Interaction of TaqI polymorphism at exon 9 of the vitamin D receptor gene with the negative lepromin response may favor the occurrence of leprosy

Controversies over the vitamin D receptor (VDR) acting as a susceptibility factor in Mycobacterium sp. infections may be the result of incorrect population stratification. The risk of leprosy occurrence conditioned by VDR polymorphism was investigated by stratifying the population of a highly endemic Brazilian region into negative and positive Mitsuda responses. Leprosy patients (102) and a group of healthy nonconsanguineous household contacts (68) were genotyped for the VDR TaqI polymorphism (T/t). TT and Tt genotypes were not considered to be risk factors as their odds ratios (OR) were not different from those presented by the negative Mitsuda response individuals. The combination of the tt genotype and the negative Mitsuda test provided an occurrence rate 13 times higher in leprosy patients than in controls with positive Mitsuda responses. This suggests that there is a higher risk of leprosy development when individuals carry this unfavorable combination, and demonstrates a possible synergistic role of these two variables in leprosy susceptibility via effects on cellular immunity.     

Postoperative Depression of Tumour-directed Cell-mediated Immunity in Patients with Malignant Disease

Leucocytes from 46 melanoma patients, 45 breast carcinoma patients, and 95 control donors were tested by the leucocyte migration test against the supernatants of homogenates of malignant melanomas, breast carcinomas, simple breast tumours, and breasts showing simple cystic disease. By comparison with controls inhibition of migration occurred significantly more frequently when tumour patients'' leucocytes were exposed to extracts of histogenetically similar tumours.Cell-mediated immunity to tumour-associated antigens was measured in 12 patients with breast carcinoma and 12 with malignant melanoma immediately before surgical operation and in the postoperative period. All patients tested before operation showed significant inhibition of migration on contact with extracts of histogenetically similar tumours. Postoperatively the degree of leucocyte migration inhibition was reduced in all patients with melanoma and breast carcinoma. Significant inhibition of leucocyte migration returned in most patients 6-22 days after operation.     

Monoclonal antibodies that distinguish non-small cell from small cell lung cancer

Two murine IgG2Ak monoclonal antibodies (703D4, 704A 1) were produced and characterized after immunization with a human large cell lung cancer line (NCI-H 157). These antibodies detect different epitopes on 31 kilodalton [35S]methionine incorporating protein(s). Radiobinding and immunohistochemical studies show these antibodies bind to most (11/13) human non-small cell lung cancer (adenocarcinoma, epidermoid, and large cell), but not to small cell lung cancer (0/11) tumors tested. The epitopes these antibodies recognized are also expressed on human melanomas (7/8), two other tumors (osteogenic sarcoma, renal cell carcinoma), but not on many other human tumors (breast, colon, neuroblastoma, lymphoid), and not on a panel of normal adult human tissues. Because the antigen(s) are preserved after fixation and because of their ability to distinguish lung cancer types from each other and normal tissues, they should be of clinical, as well as of biologic interest.     

Positive regulation of normal and tumoral mammary epithelial cell proliferation by fibroblasts in coculture

Summary In the mammary gland, mesenchymal-epithelial interactions are of paramount importance during normal and tumoral developments. We have studied the paracrine growth regulation of a variety of breast epithelial cells in coculture with normal or pathological breast fibroblasts. Two models of coculture were used in which the two cell types were seeded and grown, either together in microchamber slides or separated by a microporous membrane. Under these two conditions, all fibroblasts were shown to stimulate the proliferation of the hormono-responsive breast carcinoma MCF-7 cell line, suggesting that cell contacts were not indispensable for the paracrine stimulation of MCF-7 cell growth by fibroblasts. Moreover, in the Transwell coculture system, the proliferation of a variety of other breast carcinoma cells (MDA-MB231, T47D, and BT-20) was also stimulated by fibroblasts. However, the amplitude of the proliferative response seemed to be dependent on the carcinoma cell line considered. Moreover, the proliferative response of normal mammary epithelial cells to the presence of fibroblasts was shown to be significantly higher than the tumor cell response. The nature of the tissue of fibroblast origin, normal or pathological, did not influence the growth response of the epithelial cells. In this study, we thus demonstrate that fibroblasts are able to stimulate the proliferation of normal and carcinoma cells through paracrine exchange mechanisms. We also conclude that the target epithelial cell phenotype will essentially determine the extent of the proliferative response.     

Human tumor and normal tissue reactivity of the anti-(breast cancer) monoclonal antibody BA-Br-3 and its similarity to the anti-(epithelial membrane antigen) monoclonal antibody E29

Summary A mouse monoclonal antibody (BA-Br-3) raised against the breast carcinoma cell line CAMA-1 was previously shown to react with a 300-kDa globule-like glycoprotein from human milk fat also expressed in the cytoplasm and on the surface of human carcinoma cells of different histological types. In this report the reactivity of this mAb with a large number of normal and malignant human tissues was analyzed using immunoperoxidase techniques. When tested on sections of both fresh-frozen tissues and formalin-fixed, paraffin-embedded tissues, BA-Br-3 reacted with a formalin-resistant antigenic determinant expressed by normal and malignant epithelial cells. Preferential reactivity was observed at the apical portion of ductal epithelial cells in normal breast and in glandular epithelia distributed in several other organs. Reactivity with mucin-like secretions in the lumina of ducts was also found. BA-Br-3 reacted mostly in heterogenous staining patterns with 88% of 49 breast carcinoma specimens tested, regardless of their histological type or whether they were primary or secondary neoplasms. Testing of epithelial malignant tumors other than breast carcinomas with this antibody showed that 127 of 151 (84%) were also reactive. mAb BA-Br-3 and E29 (a commercially available anti-(epithelial membrane antigen) shared very similar staining patterns and distributions of reactivity with breast and other epithelial tumors. However, BA-Br-3 showed a significantly higher percentage of reactivity with melanoma (33% versus 6%,P = 0.003) and a trend toward a higher percentage of reactivity with sarcoma (55% versus 27%,P >0.05). This antibody, therefore, defines a molecule that is a member of the mucin-like epithelial membrane antigen family. Further studies are warranted to determine its usefulness in antibody-directed cancer diagnosis, prognosis, and immunotherapy.     

Which contacts of patients with meningococcal disease carry the pathogenic strain of Neisseria meningitidis? A population based study

Objectives: To determine the prevalence of the pathogenic strain of Neisseria meningitidis in contacts of patients with meningococcal disease, and to determine which contact groups are likely to be carriers and warrant chemoprophylaxis. Design: Population based study. Setting: Norwegian county of Telemark. Subjects: 1535 primary contacts of 48 patients with meningococcal disease, and 78 secondary contacts. Interventions: Carriers of the pathogenic strain were treated with rifampicin. All household members and kissing contacts under 15 years of age were treated with oral penicillin. Contacts were taught to recognise the symptoms of meningococcal disease. Results: In 27 of 48 cases investigated, contacts carrying the pathogenic strain of N meningitidis were found. A total of 42 such contacts were identified. Contacts were stratified into three classes according to the assumed closeness of contact with patients. In class 1 (household members and kissing contacts) the prevalence of the pathogenic strain was 12.4% (95% confidence interval 5.5% to 19.3%). In classes 2 and 3 the prevalence was 1.9% (0.9% to 3.4%) and 1.6% (0.14% to 3.1%). Conclusions: There is a high rate of carriage of the pathogenic strain of N meningitidis in patients’ household members and kissing contacts, and this supports the practice of giving chemoprophylaxis to these contacts. The prevalence of carriage among other contacts is 2-3 times that found in the general population (0.7%); the benefits of chemoprophylaxis to these contacts may be marginal.

Key messages

• Contacts of patients with meningococcal disease have a 12.4% (95% confidence interval 5.5% to 19.3%) risk of carrying the pathogenic meningococcus if they are kissing contacts or household members
• The risk of carriage of the pathogenic strain for two groups of contacts less close than household members or kissing contacts is 1.9% (0.9% to 3.4%) and 1.6% (0.14% to 3.1%)

     

Immunohistochemical phenotyping of human solid tumors with monoclonal antibodies in devising biotherapeutic strategies

Summary A panel of 14 monoclonal antibodies (MoAbs) (4 raised against breast cancer, 6 against colon cancer and 4 against melanoma) were used to phenotype frozen sections of tumor biopsies obtained from 110 patients, by avidin-biotin-peroxidase complex techniques. We observed heterogeneity of antigen expression among the multiple metastatic lesions of single patients, as well as among tumor lesions from different patients with similar tumor histotypes. A wide range of cross-reactivity of anti-(breast-carcinoma) and anti-(colon-carcinoma) MoAbs with other carcinoma histotypes and limited reactivity with melanoma and sarcoma was detected. Some of our anti-melanoma MoAbs were also found to cross-react with selected carcinomas. Nine of the 14 MoAbs most reactive with carcinomas of diverse histotypes have been identified. A mixture or cocktail of different MoAbs could be selected for each individual patient in order to achieve binding of MoAbs with most, if not 100% of tumor cells. This study illustrates the approach that we have taken to individualize the cocktail of MoAbs for the development of patient-specific therapeutic immunoconjugates.     

Androgen metabolism in male and female breast tissue

Incubation studies have been carried out using normal breast tissue and breast tissue from patients with gynecomastia, mammary dysplasia and breast carcinoma to determine the pattern of androstenedione metabolism. All tissues formed estrone (E₁) and testosterone (T) in all incubations. Estradiol (E₂) was isolated in incubations of tissue from 1 to 6 patients with mammary dysplasia, 5 of 6 patients with gynecomastia and in all incubations with normal and carcinoma tissue. Estrone formation was lowest in mammary dysplasia and gynecomastia, and higher in apparently normal breast tissue. The greatest E₁ formation was found in incubations with breast carcinoma tissue, although there was considerable variation within this tissue group. Estradiol formation was low in all tissues, with the highest conversion rates in carcinoma tissue. Testosterone formation in carcinoma tissue was greater than in mammary dysplasia or gynecomastia, but similar to apparently normal tissue. These results indicate that breast tissue from different pathological states varies in its capacity to aromatize androstenedione (A) to estrogenic products and to convert it to other androgens. They have also shown that the pattern of metabolism is distinctive for the nature of the pathological abnormality.     

Characterization of the antitumor activities of human tumor necrosis factor-alpha and the comparison with other cytokines: induction of tumor-specific immunity

We have investigated the in vitro and in vivo antitumor activities of recombinant human tumor necrosis factor-alpha (rHuTNF-alpha) against Meth A sarcoma. Meth A sarcoma cells were found to a) be relatively insensitive in vitro to rHuTNF-alpha, and b) express low numbers of TNF-alpha receptors. Intraperitoneally implanted Meth A sarcoma was insensitive to the antitumor effects of rHuTNF-alpha. In contrast, rHuTNF-alpha was highly efficacious against subcutaneously implanted Meth A sarcoma. Biodistribution studies with 125I- or 3H-labeled rHuTNF-alpha demonstrated that, after intravenous administration, the majority of the labeled rHuTNF-alpha localized in the kidney, lungs, and liver. Only low levels of radiolabel were found in subcutaneous Meth A implants. These results support the in vitro data on the low number of TNF-alpha receptors on Meth A sarcoma cells. The ability of rHuTNF-alpha to induce regression of established (7 days) subcutaneous Meth A implants, positively correlated with the degree of both macroscopic and microscopic tumor necrosis. In addition, recombinant human tumor necrosis factor-beta (lymphotoxin) and recombinant murine tumor necrosis factor-alpha induced similar levels of necrosis. Other lymphokines with known antitumor activities, recombinant human interferon-gamma, murine interferon-gamma, and human interleukin 1 alpha, failed to induce detectable necrosis of Meth A sarcoma. Mice which had rejected subcutaneous Meth A sarcoma implants after rHuTNF-alpha treatment and which were later challenged subcutaneously with Meth A sarcoma or other noncross-reacting chemically induced sarcomas were found to be specifically immune to Meth A sarcoma. In addition, low levels of cytotoxic antibodies reactive to Meth A sarcoma were detected in the sera of 21 of 30 Meth A immune mice. Histological evaluation of the hemorrhagic tumor necrosis induced by rHuTNF-alpha suggests that the primary lesion is vascular, possibly directly on the endothelial cells. The mechanisms involved in the generation of specific cell-mediated antitumor immunity in this model are at present unknown.     

Use of carminomycin on children and adolescents with osteogenic sarcoma]

The therapeutic effect of carminomycin was studied in clinic at different treatment schemes with respect to 14 children and juvenile patients with osteogenic sarcoma. Pronounced local effect evident from disappearance of the pain and in some cases decrease of the metastatic tumor were noted in the patients with metastases of the osteogenic sarcoma to the bones or relapses of the primary tumor. Subjective improvement and objective effect were observed respectively in 90 and 53 per cent of the patients with metastases into the lungs and pronounced lung symptomatology.     

Haemophilus influenzae type B meningitis: a contagious disease of children.

The families of 126 consecutive patients with Haemophilus influenzae type B meningitis were surveyed for secondary invasive H influenzae disease among household contacts. A total of 120 of the families were contacted. In six cases no contact was possible and the medical record was reviewed. Some 555 household contacts were found; 31% (171) were under 5 years of age. A secondary case was defined as a household contact with H influenzae type B isolated from blood or cerebrospinal fluid more than 24 hours, but less than 30 days, after admission to hospital of the index case. Four secondary cases were identified, all in children aged under 5 years. The secondary attack rate in children under 5 years or less in the month after exposure to an index case was thus 2.3%, 800 times the endemic attack rate for H influenzae meningitis. This is a conservative estimate since five additional contact cases were documented, but not included in the secondary attack rate. Young contacts of a child with H influenzae meningitis are thus at significant risk of life-threatening secondary disease.