A novel retinoic/butyric hyaluronan ester for the treatment of acute promyelocytic leukemia: preliminary preclinical results.

All-trans retinoic acid (ATRA) represents the therapy of choice for patients with acute promyelocytic leukemia (APL). However, patients often relapse due to ATRA-resistance. The molecular basis of APL alterations indicates that addition of a histone deacetylase inhibitor to ATRA may restore the sensitivity to retinoids. We explored the in vitro and in vivo effects of a novel retinoic/butyric hyaluronan ester (HBR) on a retinoic acid (RA)-sensitive human myeloid cell line, NB4, and on its RA-resistant subclone, NB4.007/6. In vitro, HBR induced growth arrest and terminal differentiation in RA-sensitive NB4 cells (as confirmed by an increased expression of CD11 family members and nitroblue tetrazolium assay), whereas it inhibited the growth of RA-resistant cells by apoptosis, paralleled by an increase in the levels of caspase 3 and 7. In vivo, HBR treatment of NB4-inoculated severe combined immunodeficient mice resulted in a statistically significant increase in survival time (P<0.0001), comparable to that induced by a maximum tolerated dose of RA alone. Also on P388-inoculated mice, HBR was active in contrast to RA that was completely ineffective. Present findings suggest that, owing to the simultaneous presence of RA and an histone deacetylases inhibitor, HBR might be useful in controlling the proliferation of RA-resistant cells and the differentiation of RA-sensitive cells.

Pharmaceutical development of a parenteral lyophilized formulation of the antimetastatic ruthenium complex NAMI-A.

This paper describes the development of a stable pharmaceutical dosage form for NAMI-A, a novel antimetastatic ruthenium complex, for Phase I testing. NAMI-A drug substance was characterized using several spectrometric and chromatographic techniques. In preformulation studies, it was found that NAMI-A in aqueous solution was not stable enough to allow sterilization by moist heat. The effect of several excipients on the stability of the formulation solution was investigated. None of them provided sufficient stability to allow long-term storage of an aqueous solution of NAMI-A. Therefore, a lyophilized product was developed. Five different formulations were prepared and subjected to thermogravimetric (TG) analysis and stability studies at various conditions for 1 year. Minimal degradation during the production process is achieved with a formulation solution of pH 3-4. Of the acids tested, only hydrochloric acid (HCl 0.1 mM) both stabilized the formulation solution and was compatible with the lyophilized product. This product was stable for at least 1 year when stored at -20 degrees C, 25 degrees C/60% relative humidity (RH) and 40 degrees C/75% RH, and was also photostable.

Reduction of lung metastasis by ImH[trans-RuCl4(DMSO)Im]: mechanism of the selective action investigated on mouse tumors.

NAMI-A (imidazolium trans-imidazoledimethylsulfoxidetetrachlororuthenate, ImH[trans-RuCl4(DMSO)Im]) is a new ruthenium compound active against lung metastasis of solid metastasizing tumors. We have tested this compound in mice with Lewis lung carcinoma or MCa mammary carcinoma in order to compare the effects on primary tumor and lung metastases with possible alterations of cell cycle distribution of tumor cells. We have also investigated whether there were unequal tissue accumulations of the compound itself at different dose levels ranging from 17.5 to 70 mg/kg/day given for six consecutive days. NAMI-A caused a reduction of metastasis weight larger than that of metastasis number; we explain this finding as the capacity of NAMI-A to selectively interfere with the growth of metastases already settled in the lungs. However, this specificity is not simply related to a larger concentration of NAMI-A in the lungs than in other tissues. Following i.p. treatment, NAMI-A rapidly disappeared from the peritoneal cavity; its low blood concentration may be caused by rapid renal clearance. These data provide further evidence for a selective anti-metastasis effect of the ruthenium complex NAMI-A. The reduction of lung metastasis is followed by a significant prolongation of the host's life-time expectancy, indicating a therapeutic benefit of NAMI-A on lung metastases from solid tumors.

Hyaluronic acid as drug delivery for sodium butyrate: improvement of the anti-proliferative activity on a breast-cancer cell line.

The potential clinical utility of sodium butyrate, a natural compound known to inhibit tumor-cell growth, is hampered by the difficulty of achieving effective in-vivo concentrations. The short half-life (about 5 minutes) of sodium butyrate results in rapid metabolism and excretion. To increase the availability of sodium butyrate over a longer period of time, we co-valently linked it to hyaluronic acid (a component of the extracellular matrix). Its major advantages as a drug carrier consist in its high biocompatibility and its ability to bind CD44, a specific membrane receptor frequently over-expressed on the tumor-cell surface. The degree of substitution of hyaluronic acid with butyrate residues ranged from d.s.=0.10 to d.s.=2.24 (1.8-28.4% w/w). The biological activity of hyaluronic-acid-butyric-ester derivatives was evaluated in terms of the inhibition of the growth of the MCF7 cell line and compared with that of sodium butyrate. After 6 days of treatment, we observed a progressive improvement of the anti-proliferative activity up to d.s.=0.20; thereafter, the anti-proliferative effect of the ester derivatives decreased. Fluorescence microscopy showed that after 2 hr of treatment fluorescein-labelled compounds appeared to be almost completely internalized into MCF7 cells, expressing CD44 standard and variant isoforms. These findings indicate that hyaluronic acid could offer an important advantage in drug delivery, in addition to its biocompatibility: the ability to bind to CD44, which are known to be frequently over-expressed on the tumor-cell surface.

Aging of the human synovium: an in vivo and ex vivo morphological study.

Age-associated changes of the human synovium have been investigated by microarthroscopy, optical and electron microscopy, immunohistochemistry, and cytochemistry. The knee joints of nineteen 15- to 56-year-old subjects, classified as normal by inspection, were carefully examined by microarthroscopy; small synovial tissue biopsy specimens from both the suprapatellar pouch and the medial tibiofemoral gutter were taken. Microarthroscopy showed that the villi were more numerous and the vascular network and cell distribution and profiles less regular in aged individuals. These data were confirmed by scanning electron microscopy, which also showed large areas of the synovial surface devoid of cells and collagen bundles in contact with the joint cavity in aged subjects. Light and transmission electron microscopy confirmed these data and allowed evaluation of the number, distribution, shape, and internal organization of cells as well as the distribution of vessels and the organization of the extracellular matrix in the full thickness of the synovium (down to 2 mm). Particular attention was paid to synovial lining cells, among which three main phenotypes could be recognized: synthetic type (present at all ages and hypertrophied in aged subjects), macrophagelike (increasing with age), and fibroblastlike. Collagen increased with age. Further studies are needed for comprehensive understanding of age-associated changes in the human synovium.

Comparative ototoxic potential of hyaluronic acid and methylcellulose.

In experimental animal studies, exogenous hyaluronan (HA) has been shown to exert beneficial effects on the healing of tympanic membrane perforation. As any other exogenous substance, HA may prove potentially toxic, by filling the middle ear cavity, to the sensory cells of the organ of Corti. Electrophysiological (ABR) and morphological studies were carried out in the rat to examine the auditory function and the structure of the sensory epithelium. Rats received either HA or hydroxy-propyl-methyl-cellulose by trans-tympanic injection (middle ear cavity was completely filled up) and were compared to untreated, age- and weight-matched rats. In both treated groups ABR revealed transitory, mild conduction hearing loss, in particular for high frequencies, until day 7 postinjection. This loss recovered completely within the 15th day. Morphologically, no significant degenerative/necrotic lesions were observed in the organ of Corti, from both treated groups.

Intra-articular treatment of knee osteoarthritis. A comparative study between hyaluronic acid and 6-methyl prednisolone acetate.

The efficacy and tolerability of 20 mg of hyaluronic acid were compared in an open, randomized trial with that of 40 mg of 6-methylprednisolone acetate, administering them both by an intra-articular route once a week for 3 weeks to patients suffering from inflammatory knee osteoarthritis. The results of the study showed that for up to one week after the end of treatment hyaluronic acid's analgesic activity was comparable to that of the steroid, while at the end of the follow-up (45 days after the end of treatment) all the pain monitoring parameters presented significant differences in favour of the HA-treated group. Both treatments were well tolerated, since no local or systemic adverse reactions were observed.

[Intraluminal ultrasound echotomography: a new method in the study of arterial angioplasty. Evaluation of a case]. / Ecotomografia intraluminale ad ultrasuoni: un nuovo metodo per lo studio dell'angioplastica arteriosa. Valutazione di un caso.

Intravascular ultrasound angioscopy allows to better define the structure of arterial wall and the effect of angioplasty. We report the case of a patient previously treated by coronary angioplasty, who underwent peripheral angioplasty for a stenosis of the left iliac artery. Intravascular ultrasound imaging performed before and after balloon dilatation clearly identified the plain of dissection and the partial resolution of the stenosis in a better way compared to digital angiography.

[Effectiveness of regional anesthesia for loco-regional carotid surgery. Retrospective review of 147 interventions]. / Validità dell'anestesia loco-regionale nella chirurgia della carotide. Studio retrospettivo su 147 interventi.

The Authors present a retrospective review that compares general anesthesia with regional anesthesia performed during carotid endarterectomy surgery. The study includes 147 procedures, performed on 144 patients: 76 had general anesthesia and 71 had regional anesthesia. After emphasizing the importance of conscience preservation during regional anesthesia to allow a simple and secure monitoring of the patient's neurologic functions, the Authors point out the decrease of peri- and postoperative accidents, whether neurologic or not, obtained with regional anesthesia. Another essential advantage of this technique is the possibility to insert the transitory shunt only when objectively necessary, ulteriorly reducing neurologic complications. Cervical plexus block is primarily indicated for high risk patients with instable neurologic simptomatology, severe stenosis of the contralateral carotid, coronary heart disease. The technique has proved to be of easy execution, safe and well accepted by the patient and the surgeon.

[Spontaneous reparation of post-traumatic tympanic perforation: an experimental study in rats]. / Riparazione spontanea della perforazione timpanica post-traumatica: studio sperimentale nel ratto.

In most patients tympanic membrane perforation spontaneously repairs itself. Nonetheless, in several cases the perforation persists due either to traumatic or phlogistic damage. Although the membrane perforation healing process has been under study for over a century there are still two contrasting theories. One of these theories asserts that repair takes place through the growth of granulation tissue while the other believes that epithelial migration is at the basis of healing. In the present study an experimental animal model (rat) was used in order to assess the staging of the natural evolution of post-traumatic membrane perforation and to characterize, from a morphological and sub-microscopic point of view, the cellular population and microstructural aspects of the extracellular matrix. Transmission electron microscopy (TEM) and scanning electron microscopy (SEM) were employed to evaluate the progress in tympanic membrane healing and to make a comparison with the most recent theories. Eighteen Sprague-Dawley rats weighing 230-260 gr of the same age (10 weeks) were used in the study. The animals were anesthetized by intraperitoneal injections of Na-pentobarbital (NembutalR) (45 mg/Kg) and, under operating microscope, bilateral perforation of the upper rear quadrant of the pars tensa was performed with a myringotomy lancet. The animals were subjected to periodic follow-ups over next 30 days. Three animals were sacrificed during each control and the tympanic membrane was removed for TEM and SEM study. On the basis of the present study the following conclusions can be drawn: a) primary healing of the lesion is through granulation tissue; b) the healed T.M. is composed of three normal layers as is the normal T.M.; c) the presence of fibroblasts in the intermediate neoformed fibrous layer leads one to conclude that its extracellular matrix is produced and organized "in situ"; d) the neoformed lamina propria has a disorganized, fibrous structure.

Arterial revascularization by laser angioplasty: first Italian experiences.

Eighty-seven laser angioplasty procedures were performed on 85 patients (73 men and 12 women). In the first phase (54 patients) a broad selection process included long occlusions (23.9 cm avg) and vessels with heavy calcification; in the second phase (31 patients) the authors treated vessels with shorter occlusions (10 cm avg) and less calcific. The primary success rate, based on number of procedures (73.5% overall) was 69% in the first phase but reached 81.3% in the second phase. There were 14 perforations (16%), 8 dissections (9.4%), and 4 peripheral emboli (4.7%). Six-month follow-up shows that the Winsor index maintained its initial improvement in 76.5% of all patients, or 62% in just the long occlusions. These data and a review of the cases suggest that no more of 10-20% of patients with claudication in lower extremities can be treated by laser angioplasty; the authors consider ideal candidates those patients who noncalcific lesions in the common iliac or superficial femoral arteries at the III medium level with obstructions shorter than 8 cm and good runoff.

[Evaluation of complications during laser angioplasty with a hot tip system in the treatment of peripheral arteriopathy]. / Valutazione delle complicanze durante laser angioplastica con sistema a punta calda nel trattamento delle arteriopatie periferiche.

"Hot tip" laser angioplasty has been used in the treatment of peripheral artery disease. The purpose of our study (the largest in Italy) is to evaluate the complications during the recanalization of stenotic and occluded iliac and femoro-popliteal arteries, using a hot tip laser system coupled to an Argon or Nd Yag laser. One hundred twenty-three patients have been treated. The complications were: arterial spasm in 2 patients; distal embolizations in 6 patients; arterial dissections in 35 patients; vessel wall perforations in 20 patients. Vessel wall perforation is the more severe complication and did never allow to complete the procedure. The presence of intravascular calcified deposits increases the risk of perforation, due to the loss of the coaxial position of the probe. Only the development of a guided laser system could allow a safer delivery of laser radiation, reducing the risk of arterial wall perforation.

Evolution of carotid atheromatous lesions in endarterectomized patients.

A noninvasive follow-up using duplex ultrasonography was conducted in 128 patients who had undergone carotid endarterectomy from January 1987 to December 1988. Repeated scans of the operated area revealed a distinct increase in thickened ultrasonographic features (32% vs 8%), and a stenosing lesion was detected in 8 patients. There was also a parallel increase in the number of subjects with thicknesses of the intima adjacent to the endarterectomy area exceeding 2.5 mm. In 7% of cases, dilatation was detected in the operated area and in 27% the margin of the area was raised. The study also dealt with the contralateral carotid artery, where a progression of atheromatous involvement was observed with an increased number of cases of hemodynamically significant stenosis and 3 cases of occlusion. Periodic duplex ultrasonography in endarterectomized patients proves useful for the early detection of hemodynamically significant stenoses or of structural features potentially capable of generating emboli.

Controlled comparison of ketanserin and nifedipine in Raynaud's phenomenon.

Twenty-eight patients suffering from either primary or secondary Raynaud's phenomenon were treated with nifedipine and ketanserin. Each patient was treated with one of the two drugs administered after an adequate washout period. Furthermore each patient was submitted before and after treatment with each drug to computerized digital thermometry to evaluate the therapeutic response. The data obtained during the intake of the two drugs at zero, five, and twenty-three minutes were compared with thermometry-relevant baseline data at the same periods. Ketanserin proved to be useful in the treatment of Raynaud's phenomenon and statistically significantly superior (alpha less than 0.05) with respect to nifedipine in the thermometric controls and also in the subjective evaluation of the patients (p less than 0.02). In this study nifedipine did not show particular efficacy. Furthermore only 2 patients had to discontinue treatment with ketanserin, whereas 8 had to discontinue treatment with nifedipine (p less than 0.001).

[Prognostic factors in surgery of ruptured aneurysms of the abdominal aorta]. / Fattori prognostici nella chirurgia degli aneurismi dell'aorta addominale rotti (nostra esperienza in 83 pazienti).

From 1978 to 1986 83 patients underwent surgery for ruptured abdominal aortic aneurysm. The intraoperative and post-operative mortality rates was 34%. This multivariant analysis has been performed with intention of identifying the specific factors that affected the surgical outcome of ruptured abdominal aortic aneurysms. Increased intraoperative mortality rates were associated with heart disease, while postoperative mortality rates were associated with preoperative hypotension, BUN, levels higher than 40 mg/dl and other factors like duration of operation, blood less, blood transfusion and pre-operative hematocrit.

[Aneurysms of the gluteal artery (description of a clinical case and review of the literature)]. / Gli aneurismi dell'arteria glutea (Descrizione di un caso clinico e revisione della letteratura).

The authors show a case of superior gluteal artery aneurysm they operated successfully. Starting from this case the Authors review the literature about the etiology, the clinical characteristics, the diagnosis and the various methods of treatment of this lesion.

Intra-articular injections of hyaluronic acid in the treatment of painful shoulder.

Twenty-nine patients (mean age, 61 years) received three intra-articular injections of 10 mg of hyaluronic acid in 1 ml of phosphate buffer; the injections were given at three-day intervals. Six of the patients had osteoarthritis, six had Duplay's disease, and 17 had both osteoarthritis and Duplay's disease. The patients were evaluated after each injection and 11 days after the last injection. On all measures of treatment effectiveness, including joint mobility, pain, and analgesic consumption, the patients showed rapid and significant improvement. No side effects were reported.

[Subclavian and innominate steal syndrome (our experience with 29 patients)]. / Il furto della succlavia e dell'anonima (nostra esperienza in 29 pazienti).

Subclavian steal syndrome and innominate steal syndrome are well-known reasons of cerebral vascular insufficiency. Today this pathology can be corrected by extra-anatomic by-passes. In our experience in recent years (1980-1986) these by-pass (in our patients carotid-subclavian and subclavian-subclavian by-pass) gave very good outcomes. These are simple and poor-risk operations. Their patent after many years is very good.