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Synchrotron nanoprobe X-ray absorption (XAS) studies of a potent organo-osmium arene anticancer complex in ovarian cancer cells at subcellular resolution allow detection and quantification of both OsII and OsIII species, which are distributed heterogeneously in different areas of the cells.

Four chiral Os(II) arene anticancer complexes have been isolated by fractional crystallization. The two iodido complexes, (S(Os),S(C))-[Os(η(6)-p-cym)(ImpyMe)I]PF6 (complex 2, (S)-ImpyMe: N-(2-pyridylmethylene)-(S)-1-phenylethylamine) and (R(Os),R(C))-[Os(η(6)-p-cym)(ImpyMe)I]PF6 (complex 4, (R)-ImpyMe: N-(2-pyridylmethylene)-(R)-1-phenylethylamine), showed higher anticancer activity (lower IC50 values) towards A2780 human ovarian cancer cells than cisplatin and were more active than the two chlorido derivatives, (S(Os),S(C))-[Os(η(6)-p-cym)(ImpyMe)Cl]PF6, 1, and (R(Os),R(C))-[Os(η(6)-p-cym)(ImpyMe)Cl]PF6, 3. The two iodido complexes were evaluated in the National Cancer Institute 60-cell-line screen, by using the COMPARE algorithm. This showed that the two potent iodido complexes, 2 (NSC: D-758116/1) and 4 (NSC: D-758118/1), share surprisingly similar cancer cell selectivity patterns with the anti-microtubule drug, vinblastine sulfate. However, no direct effect on tubulin polymerization was found for 2 and 4, an observation that appears to indicate a novel mechanism of action. In addition, complexes 2 and 4 demonstrated potential as transfer-hydrogenation catalysts for imine reduction., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We have compared the organometallic arene complexes [(eta(6)-biphenyl)M(ethylenediamine)Cl](+) RM175 (M=Ru(II)) and its isostructural osmium(II) analogue AFAP51 (M=Os(II)) for their ability to induce cell detachment resistance from fibronectin, collagen IV and poly-l-lysine, and cell re-adhesion after treatment, their effects on cell migration and cell viability, on matrix metalloproteinases production, and on primary tumour growth of MCa mammary carcinoma, the effect of human serum albumin on their cytotoxicity. There are differences between ruthenium and osmium. The Os complex is up to 6x more potent than RM175 towards highly-invasive breast MDA-MB-231, human breast MCF-7 and human epithelial HBL-100 cancer cells, but whereas RM175 was active against MCa mammary carcinoma in vivo and caused metastasis reduction, AFAP51 was not. Intriguingly the presence of human serum albumin in the growth medium enhanced the cytotoxicity of both compounds. RM175 increased the resistance of MDA-MB-231 cells to detachment from substrates and both compounds inhibited the production of MMP-2. These data confirm the key role of ruthenium itself in anti-metastatic activity. It will be interesting to explore the activity of osmium arene complexes in other tumour models and the possibility of changing the non-arene ligands to tune the anticancer activity of osmium in vivo.

The possibility of using electron emitters to cure a cancer with metastatic spread depends on the energy of the emitted electrons. Electrons with high energy will give a high, absorbed dose to large tumours, but the absorbed dose to small tumours or single tumour cells will be low, because the range of the electrons is too long. The fraction of energy absorbed within the tumour decreases with increasing electron energy and decreasing tumour size. For tumours smaller than 1 g, the tumour-to-normal-tissue mean absorbed dose-rate ratio, TND, will be low, e.g. for 131I and 90Y, because of the high energy of the emitted electrons. For radiotherapy of small tumours, radionuclides emitting charged particles with short ranges (a few microm) are required. A mathematical model was constructed to evaluate the relation between TND and electron energy, photon-to-electron energy ratio, p/e, and tumour size. Criteria for the selection of suitable radionuclides for the treatment of small tumours were defined based on the results of the TND model. In addition, the possibility of producing such radionuclides and their physical and chemical properties were evaluated. Based on the mathematical model, the energy of the emitted electrons should be < or = 40 keV for small tumours (< 1000 cells), and the photon-to-electron energy ratio, p/e, should be < or = 2 to achieve a high TND. Using the selection criteria defined, five low-energy electron emitters were found to be suitable: 58Co, 103mRh, 119Sb, 161Ho, and 189mOs. All of these nuclides decay by internal transition or electron capture, which yields conversion and Auger electrons, and it should be possible to produce most of them in therapeutic amounts. The five low-energy electron-emitting radionuclides identified may be relevant in the radiation treatment of small tumours, especially if bound to internalizing radiopharmaceuticals.

New organometallic complexes have been synthesized by association of an active organic molecule with a metallic element such as Pt, Rh, Ir, Pd, Os. Their trypanocidal activity was studied in vitro and in vivo against T. b. brucei. The more active compounds were pentamidine derivatives. The Ir- COD-pentamidine complex, and Iridium (I) cationic and organometallic complex showed and in vitro activity at 60 micrograms/l. Moreover, all infected mice were cured by this compound subcutaneously administered in a single dose at 0.5 mg/kg (0.317 mumol/kg). In the same conditions, pentamidine cured all the mice at 5 mumol/kg. Ir-COD-pentamidine (or P1995) was 16 fold more efficient than pentamidine. Since the chemotherapeutic index of this molecule was 7.5 fold higher than those of pentamidine, P1995 can be considered as a potential trypanocidal drug of the future.

Antigen induced arthritis (AIA) was elicited in 28 adult rabbits using ovalbumin. All animals developed an intensive monarthritis manifested by local heat and swelling. Two, 7 and 21 days later, respectively, 0.3 ml of 1 per cent OsO4 was injected into the arthritic knees of three groups of 9 rabbits. The rabbits were killed 1, 3 and 6 months later. No modifying influence on the severe to complete cartilage destruction was evident. One group of 8 rabbits received only OsO4 and the cartilage of these animals remained intact after 6 months.

Two hundred and one patients (305 knees) with rheumatoid arthritis received intra-articular osmic acid in one or both knees. Assessment was based on pain relief, warmth, tenderness, size and presence of effusion, degree of synovial thickening and range of pain-free movement. Satisfactory results were obtained in 61% at one year gradually reducing to 22% over a five-year period. If only those knees with none or minimal joint damage were considered, then a satisfactory result was obtained in a higher percentage, 74% at one year and 38% at five years.

In the U.S., the chief source of new osmium is copper refining, where this metal is produced as a byproduct. Probably less than 10% of the osmium in the original copper ore is recovered, and 1000-3000 oz troy of osmium is lost each year to the environment as the toxic, volatile tetroxide from copper smelters. In 1971, about 2000 oz troy of osmium was domestically refined, most of which was from secondary sources. An additional 4169 oz troy of osmium was toll-refined. Major uses for osmium tetroxide identified are for catalysis, especially in steroid synthesis, and for tissue staining. Minor uses of osmium metal are for electrical contacts and for imparting hardness to alloys for mechanical pivots, etc. Unreclaimed osmium tetroxide that reaches wastewater streams is probably rapidly reduced by organic matter to nontoxic osmium dioxide or osmium metal, which would settle out in the sediment of the water course. Waste osmium metal, itself innocuous and chemically resistant, would be oxidized to the toxic tetroxide if incinerated. Because of the small amounts used and their wide dispersal, the amounts of osmium tetroxide in wastewater and air should pose no hazard to man or the environment. The chief acute toxic effects of osmium tetroxide are well known and include eye and respiratory-tract damage. Few data are available that provide information on possible effects of nonacute exposure resulting from environmental contamination by osmium. However, workers continually exposed to osmium tetroxide vapors (refiners and histologists) and rheumatoid arthritis patients who have received intra-articular injections of osmic acid solutions have shown no apparent damage from exposure to low levels of osmium.

Osmic acid synoviorthesis is an adjuvant form of local treatment for rheumatoid arthritis, the value of which has been demonstrated by a large number of studies over some 30 years. However, it is widely considered that the stability of the results of this type of synoviorthesis is less satisfactory than that obtained with isotope synoviorthesis and that osmic acid may damage the articular cartilage. The authors analyzed the results of a retrospective study of 90 rheumatoid knees treated by this method with a follow-up of more than 5 years (mean period 9 years). With such a follow-up, a satisfactory clinical result was seen in 44.5 p. cent of cases and radiological evaluation of the knees treated showed no worsening in initial signs in an identical percentage of cases. Best results were obtained in joints where there was little radiological damage and in cases of inflammatory or only moderately progressive arthritis, good results may also be hoped for in more severe or advanced cases. The good overall quality of the results obtained with a mean follow-up of 9 years would be in favour of the value of osmic acid synoviorthesis in the local treatment of rheumatoid arthritis.

A case of general yersinosis in a male physician is described. Most probably the infection was acquired nosocomially in a paediatric ward. The infection affected several organs including the liver, but osmium tetroxide was nevertheless used to perform chemical synovectomy in a knee joint suffering from active arthritis. The arthritis was cured, and the already existing liver affection did not progress. The only complaint from the patient, months after the osmic acid injection, was a strong metallic taste on the tongue. More than a year after the intra-articular injection, the metallic taste disappeared. The amount of osmium in the patient's blood was then calculated to be less than 100 molecules of osmium per ml blood.

Synoviorthesis of the knee joint with OsO4 is a simple procedure with few short or long term side effects. The substance used is inexpensive and easy to handle. All studies published so far have confirmed its suitability in the treatment of synovitis of the knee and various other joints. The initial good results deteriorate steadily, primarily in the first year following injection. However, intra-articular administration of OsO4 may be repeated without risk, and achieves as good results as the first synoviorthesis. The less advanced the lesions and the less active clinically and biologically the systemic manifestations, the better the results. Osmic acid does not appear to damage the joint either radiologically or functionally. The effect of OsO4, although probably shorter in duration, is comparable to that of yttrium-90, with the additional advantage that it can be used in young patients. 100 mg (5 ml of 2% solution) can be considered the optimal dose. Synoviorthesis with osmic acid can be performed before going on to surgical synovectomy. Synoviorthesis with osmic acid is ineffective in the treatment of gonarthrosis.

The use of radioactive yttrium (90Y) in the treatment of rheumatoid knee joints is described. Clinical results in 48 knees after a mean observation period of more than 3 years are reported: 60 percent excellent or good at one year, 46 percent at two years, 33 percent at three years and 17 percent at four years. The number of failures or relapses exceeded 50 percent between 30 and 36 months. The average rate of relapse in successful cases was 12.8 percent per year. The most important factors associated with unsatisfactory result were advanced articular destruction and progression of the systemic disease.

One hundred mg of osmic acid (with corticosteroid and lidocaine) was injected for therapeutic purposes into the knee in 10 patients suffering from various types of recalcitrant synovitis (mostly rheumatoid arthritis). The effects of this injection were followed by examination of synovial fluid, blood, and urine of these patients, collected at various intervals after intra-articular injection. As a control, a group of 3 patients (5 knees) received corticosteroid and lidocaine intra-articularly, and another group (6 patients, 7 knees) was injected with lidocaine only, both with the same total volume as the first group. Blood and urine examinations revealed essentially no effects of osmic acid, whereas in synovial fluid a strong inflammatory reaction was observed up to the second day after the injection, though the effusion soon disappeared. In a group treated with lidocaine + corticosteroid, a mild, early inflammatory reaction was noted, while lidocaine alone produced only a dilution effect, and the exudate remained relatively longer. Thus, the early local inflammatory reaction due to osmic acid is an expression of the necessary drastic effect of this beneficial therapeutic agent.

Radiosynoviorthesis plays an important role in local treatment of rheumatoid arthritis. Depending on the size of the joint, radionuclides are used at different depths of penetration. Data from the literature and our own results with 396 treated joints indicate that in 2/3 of cases good therapeutic effects are achieved, e.g. the effect on the local inflammatory process is comparable with surgical synovectomy. This is in agreement with the results of our longterm follow-up of 141 joints after radiosynoviorthesis. As with surgical synovectomy, early synoviorthesis was most successful; however, longterm observations revealed a slight increase in local recurrences. These and primary non-responder cases show a good response to secondary injection of radionuclides. The effect of intraarticular injection of isotopes is possibly due to irradiative action on the inflamed cells, with subsequent fibrosis of the synovium. Side effects of radiosynoviorthesis are rare and generally avoidable. Since late complications cannot be excluded, careful consideration should be given to the indication for radiosynoviorthesis. Radiosynoviorthesis represents an alternative treatment to surgical synovectomy in patients over 40 years of age.

Ninety-nine adult RA patients with knee joint synovitis were randomized into two groups. The knee joints of the first group (52 patients) were treated with osmic acid and those of the second group (47 patients) with a placebo. After 6 months, the incidence of hydrops and pain was statistically less significant in joints treated with osmic acid. the result was better in joints without advanced radiological destruction. It is concluded that osmic acid is still of benefit in the local treatment of rheumatoid knee joint synovitis at an early stage.

Punch biopsy specimens from the cartilage of the lateral condyle of the femur in 29 patients suffering from rheumatoid arthritis were taken during surgical synovectomy of knee joints and investigated by light or electron microscopy. Eleven patients had never received osmic acid injections in this particular joint, whereas 18 had had such injections 1-24 months prior to surgery. By light microscopy only minor differences could be seen between controls and cartilage treated with osmic acid, whereas with electron microscopy, increased amounts of dark-staining cell debris were visible in samples taken after osmic acid treatment. The perilacunar matrix appeared normal. There was no irregularity of collagen fibres at the surface of the cartilage. It is concluded that osmic acid causes limited superficial damage to the cartilage. The question whether this is of any clinical consequence in the form of subsequent degenerative joint disease, must be solved by clinical follow-up studies.

The effectiveness of osmic acid and yttrium-90 in the treatment of synovitis of the knee in rheumatoid arthritis is compared in 126 patients followed-up for 3 years. Ninety-one knees were injected with osmic acid and eighty-four knees with yttrium-90. Osmic acid appeared to be more effective than yttrium-90 throughout the period of the follow-up but the difference only reached statistical significance (p less than 0.05) at 3 years. Both therapies were well tolerated by patients and should be considered as an alternative to operative synovectomy.

Abnormal radiological opacities are sometimes observed after intra-articular injection of osmic acid. These opacities are radio-opaque because osmium is a heavy metal (atomic number = 76). They are usually found near the suprapatellar pouch which is the usual injection site. A parasynovial injection (or back flow from the joint cavity) of some of the osmic solution, followed by concentration and fixation of the osmic deposits at this level, seems to be the cause of these radiological opacities. This could be the reason for the poor clinical results encountered in some cases of osmic acid therapy followed by such deposits.

An anatomical and radiologic study of a synovial autopsy specimen 32 months after an intra-articular injection of osmic acid for the local treatment of a rheumatoid synovitis. Osmium deposits are seen within peripheral cells of a fat lobule and also in a few histiocytes of the scar tissue which replaced the rheumatoid synovitis. These osmium deposits in the fat cells are present in a sufficient quantity to give a radiologic opacity.

A clinical and experimental study was carried out on knee joints of rabbits and in humans. In one knee joint of each rabbit 0.3 ml 1% osmic acid or 0.3 ml 5% Varicocid was injected, the remaining uninjected joints serving as controls. The animals were killed after 1 and 24 hours, 15, 45 and 90 days. Study with the electron microscope after osmic acid injection revealed necrosis of the chondrocytes in the superficial layer and upper middle zone. The articulations injected with Varicocid exhibited only slight lesions of the chondrocytes in the upper layer, consisting of dilated rough endoplasmatic reticulum, mitochondria lacking matrix and christae; the electron density of the layer seemed to be diminished. Biopsy of the non-weight-bearing cartilage of the lateral femoral condyle was performed in 4 patients with Rheumatoid Arthritis and arthritis of the knee in the onset stage. Two of these patients received one intra-articular injection of 10 ml 1% osmic acid, 5 ml 2% xylocaine and 50 mg hydrocortisone and the other 2 patients 6 ml 5% Varicocid. Biopsy of the cartilage in the areas adjacent to the site of the first biopsies 1 and 3 1/2 months after treatment with osmic acid and 2 and 3 1/2 months after treatment with Varicocid showed in the first two cases necrosis of the chondrocytes in the superficial layer and upper middle zone of the articular cartilage and depletion of the mucopolysaccharides in the ground substance, revealed by PAS and alcian blue staining. No lesion of the knee joint cartilage was observed in the other two cases. In view of the results obtained, chemical synovectomy with Varicocid can be considered as having received further support.

A study of 113 knees of 72 patients treated with intra-articular injection of osmic acid for chronic exudative synovitis caused by various rheumatic diseases is reported. Certain cases presented abnormal radiological opacities, statistically more frequent after the injection of 200 mg than after 100 mg of osmic acid (33.9% and 15.7%). These images appear early after the injection and can remain for years. They should not be confused with intra-articular calcium deposits, or with synovial ossifications or various periosteal osteogenic processes. Their appearance does not seem to be related to the rheumatic disease treated or the therapeutic result obtained. They reflect the fixation of osmium in the synovial fatty tissue, which causes a fibrous reaction at this level. Their longterm action in the cicatrization of the synovia, or their part in a relapse of an inflammatory episode should be clarified.

Thirty-seven New Zealand rabbits were observed for up to 1 year after their knee joints were instilled with 1 ml of 1% osmic acid. A selective destruction of synovial lining cells occurred with a concurrent synovitis, but lining cell regenerated by the eighth week. Nonprogressive abnormalities were also present in the cartilage. When an immune synovitis was induced in both knees of 16 rabbits, the osmic acid treated joint demonstrated a significantly greater synovitis and progressive aberrations compared to the untreated knee.

Arthroscopy of the knee, including synovial biopsy under direct vision, was performed in 22 patients (21 cases of arthritis, 1 of osteoarthrosis), treated 15 days to 12 months previously by osmic or isotopic synoviorthesis. The aspects seen after the 16 failures were compared with those seen after good results (4 cases) and after recurrences (2 cases). In arthritis, failure was due either to persistent synovitis, to synovial necrosis or to a combination of the two. Arthroscopy is useful since the discovery of inflammatory lessions justifies repeat synoviorthesis, whilst necrosis indicates the need for articular lavage.

43 rheumatoid knees with persistent effusion were treated by intra-articular injections of triamcinolone hexacetonide (40 mg) or combined triamcinolone - osmic acid. Both treatments produced long-lasting improvements in the treated knees. Osmic acid seemed to potentiate the effect of the steroid. This demonstrated by measurement of joint circumference and local 99mTc uptake. In the general clinical evaluation the difference between the treatments was not noticeable. The percentage of 'excellent' and 'good' improvements was 36 at 12 months in the steroid-group and 44 in the steroid-osmic acid group.

The authors report their experience of 61 injections of osmic acid into the knee during the course of rheumatoid polyarthritis. The results stated to be very good and good, are classified by the disappearance of pain, swelling and stiffness, or a improvement in any one of these criteria. As observed in 67% of cases at the third month the result following changed with time. While these favourable results persisted into the fifth month, there was a remarkable maintenance of them to the 30th month in those patients who it was able to follow so long. The most distinct and lasting results were seen in those joints which showed the least radiological change. Tolerance of the injection of osmic acid was satisfactory apart from several painful reactions which improved in the 24 hours following injection. Two cases of thrombo-phlebitis were seen and one case of rupture of the synovium. If the results obtained with osmic acid appear less impressive than those obtained with isotopes, the simplicity of the method attracts attention as it needs no complicated equipment and is applicable to young invalids.

Ten IgM polyclonal rheumatoid factor (RF) preparations isolated from sera from various patients with rheumatoid arthritis (RA) were investigated with respect to their variable heavy chain (VH) subgroups. They were tested in a haemagglutination inhibition system using red cells sensitized with myeloma proteins with known chemical VH subgroups and anti-VH subgroup specific antisera. Most of the preparations showed a considerable degree of restriction to one VH subgroup. Seven of the IgM-RF preparations were restricted to the VHIII subgroup, two to the VHI subgroup and one to the VHII subgroup. However, a weak reaction in other VH subgroup systems was seen in several instances. Two normal IgM fractions from healthy persons showed no VH subgroup restriction, and showed a rather similar degree of reaction in all the three subgroup systems.

In rabbits with immune arthritis the numbers of lymphoid aggregates and pyroninophilic and non-pyroninophilic cells per unit area of synovium were calculated from sections taken at various intervals after intra-articular drug injections. A model of immune arthritis based on the results is presented. In the active phase of the arthritis, plasma cells, plasmablasts and lymphoid aggregates proliferate, while the number of lymphocytes decreases. In the inactive phase after treatment, the cell pattern is the reverse. The ratio of pyroninophilic to non-pyroninophilic cells proved to be a good measure of the activity of the arthritis. A favourable therapeutic effect was achieved with triamcinolone hexacetonide and combined steroid-osmic acid, but not with osmic acid alone.

This study presents the macroscopic and histologic results of 35 knee arthroscopies performed on patients with rheumatoid arthritis, some months after an yttrium or osmic acid intraarticular injection. The procedure was most often performed after a failure of the injection or a relapse of synovitis. Arthroscopy provides an understanding of the cause of synoviorthesis failure--insufficient action of the product on the synovitis or its poor diffusion, fibri-nonecrotic deposits, or cartilaginous lesions--and may be used both diagnostically and therapeutically.

Synovial fluid total, dialysable and non-dialysable hydroxyproline were determined in patients with Rheumatoid Arthritis before intra-articular osmic acid injection and on days 2 and 4 after this local treatment. On day 2 the increase in dialysable hydroxyproline paralleled the increase in non-dialysable hydroxyproline. Patients with the highest pre-treatment dialysable hydroxyproline levels also had the highest levels of this fraction on day 2 suggesting that articular damage proceeds after osmic acid injection. However, on day 4 dialysable hydroxyproline levels were consistently lower than before the treatment; the same pattern was observed with non-dialysable hydroxyproline, except for all the joints but one, where effusion recurred 6-12 months after osmic acid treatment. This study suggests that the determination of synovial fluid non-dialysable levels 4 days after osmic acid injection may provide a clue to the prediction of recurrent joint effusion and possibly justify a complementary injection either with osmic acid or with a beta-emitting isotope.

Unusual radiological opacities have been observed in 29 cases after intraarticular injection of osmic acid in 113 knees of 72 patients under treatment for a chronic exudative synovitis in different types of rheumatic diseases. They are statistically more frequent after the injection of 200 mg than after 100 mg (33.9 and 15.7% respectively). Most of the time, they are located in the suprapatellar pouch. The opacities appear soon after the injection and can last for several years. They do not seem to have an unfavorable influence on the clinical course. They appear to be caused by a deposition of osmium in synovial fat tissue. From here the osmium might stimulate fibrosis. The radiological image has to be differentiated from intra-articular calcifications, synovial ossifications and various periosteal osteogenic processes.

Antigen-induced arthritis (AIA) was induced in adult rabbit knee joints, using ovalbumin as antigen. After 4 weeks, intra-articular injections of 0.3 ml 1% osmium tetroxide or 100 muCi 198Au were given. The same injections were also given to normal animals. The animals were observed for up to 12 weeks. In the normal animals, no affection of articular cartilage could be seen by light microscopy. In the arthritic animals, however, severe destruction of articular cartilage was seen. The degree of destruction in the osmium tetroxide-treated animals was similar to or even worse than that of the controls, whereas the 198Au-treated animals showed a lesser degree of destruction than both controls and osmium tetroxide-treated animals. Autoradiograms, using 3H-thymidine, showed affection articular cartilage in all groups, i.e. both normal and arthritic.

47 patients with rheumatoid arthritis and synovitis were treated with intra-articular injection of hydrocortisone and 52 patients with both hydrocortisone acetate and osmic acid. In another series of 24 patients with synovitis in both knees were injected with osmic acid to the more seriously affected knee and with methylprednisoloneacetate to the other knee. Osmic acid gave better improvement than corticosteroid alone, although the difference between th groups of the latter series was not statistically significant. On the basis of previous investigations and this study osmic acid is one alternative to radiation or surgical synovectomy in rheumatoid synovitis. The therapeutic result is not always satisfactory but facility of treatment and minor side effects are the advantages of osmic acid therapy.