Polymer nitric oxide donors potentiate the treatment of experimental solid tumours by increasing drug accumulation in the tumour tissue.

The delivery of nitric oxide (NO) specifically to solid tumours was explored in this study as a strategy to augment the passive accumulation of nanomedicines in tumours induced by the Enhanced Permeability and Retention (EPR) effect. An increase in accumulation was achieved by the binding of the chemical precursor of NO, based on an organic nitrate, to a water-soluble synthetic polymer drug carrier. Four structurally different N-(2-hydroxypropyl)methacrylamide (HPMA)-based polymer NO donors were synthesized. Depending on their chemical structure, two of these donors were hydrolytically stable, while two rapidly released the parent nitrate under acidic conditions, mimicking the intracellular environment. The polymer NO donors were shown to overcome the drawbacks related to low-molecular-weight NO releasing compounds, namely systemic toxicity, lack of site specificity, and fast blood clearance. The NO donors showed intracellular NO release upon incubation with tumour cells. In vivo, they potentiated the EPR effect, resulting in an increased accumulation of polymer-bound cytotoxic drug doxorubicin (Dox) in EL4 T-cell lymphoma inoculated in mice. This led to a better therapeutic outcome in the treatment of lymphoma with the high-molecular-weight polymer conjugates carrying Dox but not in the treatment with the free Dox. The localized augmentation of the EPR effect via the tumour-specific NO delivery system can be viewed as a promising strategy to potentiate polymer-based tumour therapy without increasing systemic toxicity.

[History of Immuno-therapy -  from Coley Toxins to Check-points of the Immune Reaction]. / Historie imunoterapie -  od Coley toxinu ke kontrolním bodum imunitní reakce.

Immunotherapy dates back to 1868 when German physicist Busch intentionally infected patients suffering from soft tissue sarcoma with erysipelas. Rapid tumor shrinkage was observed but response was only partial and tumor recurrence subsequently occurred. It was William B. Coley who in 1891 injected a patient with a soft tissue sarcoma with streptococcal cultures. Following a severe attack of erysipelas, the tumor underwent extensive necrosis and the patient remained diseasefree for eight years. The mixture of Streptococcus and other bacteria including Seratia marcescens, Staphylococcus and Escherichia coli was referred to as Coleys toxin and was used for the next 45 years. This first immunotherapy was replaced at the beginning of the 20th century by more exact radiotherapy and later on by first chemotherapy with yperit. However, immunotherapy is a treatment that uses patients own immune system to help fight cancer and as such has several advantages over other treatments. Thus, the next major milestones in immunotherapy came in the middle of the 80s as a) adoptive cell therapy relaying on patients tumor infiltrating lymphocytes, b) injection of recombinant cytokines such as rIL2, c) identification of the first tumorassociated antigens and d) development of tumor specific monoclonal antibodies. It was followed by dendritic cells vaccines. Tremendous progress has been made in the past two decades with regard to understanding the complex interactions between tumors and the immune system and developing innovative ways to manipulate the antitumor immune response. It is recently represented as blockage of immune checkpoint inhibitors.

[Escape Strategies of Tumors from Immune Surveillence]. / Únikové strategie nádoru pozornosti imunitního systému.

Immune system must be able to protect us from foreign dangerous pathogens, but on the other side, it must be able to recognize our own tissues and organs. Activity of the immune system is affected by many positive (stimulatory) and negative (inhibitory) signals. Some of these negative receptors protect us from damage of our tissues at a place of inflammation as it blocks too intensive or longlasting immune reaction. Thereby, they have a physiological protective function against strong inflammatory reaction and possible subsequent autoimmune pathology. However, some of these mechanisms are also utilized by tumors to avoid immune recognition and attention of the immune cells. Other tumor escape mechanisms involve increased production of cytokines and factors which are responsible for immunosuppressive tumor microenvironment where effective immune response is actively blocked. This review summarizes the most frequently used strategies, which are utilized by tumors to avoid immune recognition and/ or killing by the immune cells.

The comparison of in vivo properties of water-soluble HPMA-based polymer conjugates with doxorubicin prepared by controlled RAFT or free radical polymerization.

Two conjugates of anticancer drug doxorubicin (Dox) covalently bound by the hydrolytically degradable hydrazone bond to the polymer carrier based on water-soluble N-(2-hydroxypropyl) methacrylamide (HPMA) copolymers were synthesized and their properties were compared, namely their behavior in vivo. The polymer carriers differed in dispersity due to different methods of synthesis; the carrier with relatively high dispersity (HD) was prepared by free radical polymerization (Mw=29,900 g/mol, D=1.75) and the carrier with low dispersity (LD) by controlled radical polymerization (Mw=30,000 g/mol, D=1.13). Both polymer-Dox conjugates showed prolonged blood circulation and tumor accumulation of the drug in comparison with the free drug; e.g. the tumor-to-blood ratio for the polymer-bound Dox was 3-5 times higher. The LD polymer-Dox conjugate exhibited moderately higher tumor accumulation than the HD one at a dose of 1x15 mg Dox (eq.)/kg. Also, their anti-tumor activity did not differ when injected at this dose. However, the increase of the dose to 1x25 mg Dox (eq.)/kg resulted in the enhanced therapeutic activity of the conjugates, especially of the LD one with 100% of long-term survivals. The dispersity of polymer drug carriers influenced the tumor accumulation rate, which affected the overall anti-cancer activity of polymer-drug conjugates.

[The cost study of first- line treatment of metastatic colorectal carcinoma with bevacizumab- containing regimen in the Czech Republic]. / Analýza nákladu na 1. linii lécby metastatického kolorektálního karcinomu pri podání rezimu s bevacizumabem -  data z reálné klinické praxe v Ceské republice.

BACKGROUND: Bevacizumab, a humanized monoclonal IgG antibody against the vascular endothelial growth factor (VEGF), is reimbursed in combination with chemotherapy for the first and subsequent line treatment of patients with metastatic colorectal cancer (mCRC) in the Czech Republic. However, its high cost is a potentially limiting factor. We assessed the cost of bevacizumab in the treatment of mCRC in a comprehensive cancer center. PATIENTS AND METHODS: A total of 218 patients were included in our analysis. Cost data (examination, medication, hospitalization) were collected since the initiation of bevacizumab treatment to any tumor response (RECIST criteria: complete response -  CR, partial response -  PR, stable disease -  SD, progressive disease -  PD) and/ or to death. Minimal followup for all patients was 28 months. Costs were valued in Czech crowns (CZK) and converted to EUR (1€ = 25.14 CZK). RESULTS: PD was recorded in 194 patients (89% of patients). The mean cost of treatment to PD (median TTP 9.1 months) was 1,002,076.30 CZK (39,859.84 EUR). The majority of costs to PD was made by medication -  917,048.60 CZK (36,477.67 EUR) per patient. The mean cost to response PR, CR or SD was 1,105,823.10 CZK (43,986.60 EUR) after median 9.8 months of treatment (recorded for 21 patients), medication formed 1,023,827.70 CZK (40,725.05 EUR). During the study, 170 patients (78%) died. The mean of the total costs since initiation of treatment to death (median OS 18.8 months) was 1,338,874.20 CZK (53,256.70 EUR) -  out of that, medication was 1,184,251.10 CZK (47,106.25 EUR) per patient. CONCLUSION: Targeted bio-logical therapy is the largest part of the costs of mCRC therapy. Cost of bevacizumab made up to 69% of costs to PD -  687,608.20 CZK ( 27,351.20 EUR ) per patient. The majority of the total cost was formed by targeted drug therapy (bevacizumab in 1st line therapy, cetuximab and panitumumab in 2nd and 3rd line therapy); 58% of total costs since initiation of treatment to death -  778,233.80 CZK (30,956 EUR) per patient.

Synthesis of poly[N-(2-hydroxypropyl)methacrylamide] conjugates of inhibitors of the ABC transporter that overcome multidrug resistance in doxorubicin-resistant P388 cells in vitro.

The effects of novel polymeric therapeutics based on water-soluble N-(2-hydroxypropyl)methacrylamide copolymers (P(HPMA)) bearing the anticancer drug doxorubicin (Dox), an inhibitor of ABC transporters, or both, on the viability and the proliferation of the murine monocytic leukemia cell line P388 (parental cell line) and its doxorubicin-resistant subline P388/MDR were studied in vitro. The inhibitor derivatives 5-methyl-4-oxohexanoyl reversin 121 (MeOHe-R121) and 5-methyl-4-oxohexanoyl ritonavir ester (MeOHe-RIT), showing the highest inhibitory activities, were conjugated to the P(HPMA) via the biodegradable pH-sensitive hydrazone bond, and the ability of these conjugates to block the ATP driven P-glycoprotein (P-gp) efflux pump was tested. The P(HPMA) conjugate P-Ahx-NH-N═MeOHe-R121 showed a dose-dependent increase in the ability to sensitize the P388/MDR cells to Dox from 1.5 to 24 µM, and achieved an approximately 50-fold increase in sensitization at 24 µM. The P(HPMA) conjugate P-Ahx-NH-N═MeOHe-RIT showed moderate activity at 6 µM (∼10 times higher sensitization) and increased sensitization by 50-fold at 12 µM. The cytostatic activity of the P(HPMA) conjugate P-Ahx-NH-N═MeOHe-R121(Dox) containing Dox and the P-gp inhibitor MeOHe-R121, both bound via hydrazone bonds to the P(HPMA) carrier, was almost 30 times higher than that of the conjugate P-Ahx-NH-N═Dox toward the P388/MDR cells in vitro. A similar result was observed for P-Ahx-NH-N═MeOHe-RIT(Dox), which exhibited almost 10 times higher cytostatic activity than P-Ahx-NH-N═Dox.

High level of Tregs is a positive prognostic marker in patients with HPV-positive oral and oropharyngeal squamous cell carcinomas.

BACKGROUND: Human papillomaviruses (HPVs) have been proved as one of the etiological factors of oropharyngeal squamous cell carcinoma (OPSCC). Patients with tumors of viral etiology have a lower recurrence rate and better prognosis. OPSCC is linked to an alteration in the immune system. Only a limited number of studies have correlated both the immunological parameters and HPV status with patient prognosis. The aim of this study was to determine whether HPV infection and the immunological status influence patient prognosis individually or in concurrence. MATERIAL AND METHODS: Sixty patients with oral and oropharyngeal carcinomas were enrolled. They were divided into HPV-positive and HPV-negative groups based on the expression of HPV 16 E6 mRNA. Basic lymphocyte subpopulations were determined in the peripheral blood by means of flow cytometry. RESULTS: Significantly better disease-specific survival (DSS) was observed in patients with HPV-positive tumors. Nodal status, tumor grade, recurrence, and CD8+/Tregs ratio were identified as factors influencing DSS. A higher level of Tregs and a lower ratio of CD8/Tregs influenced overall survival (OS) independently of HPV status and age. Patients with HPV-positive tumors and high levels of Tregs survived significantly better than patients from the other groups. CONCLUSION: Better survival is associated with HPV positivity and elevated Tregs levels. Our data suggest that HPV infection and Tregs do not influence patient prognosis in concurrence.

HPMA copolymer-bound doxorubicin induces immunogenic tumor cell death.

Treatment of murine EL4 T cell lymphoma with N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer conjugates of doxorubicin (Dox) leads to complete tumor regression and to the development of therapy-dependent longlasting cancer resistance. This phenomenon occurs with two types of Dox conjugates tested, despite differences in the covalent linkage of Dox to the polymer carrier. Such a cancer resistance cannot fully express in conventional treatment with free Dox, due to substantial immunotoxicity of the treatment, which was not observed in the polymer conjugates. In this study, calreticulin (CRT) translocation and high mobility group box-1 protein (HMGB1) release was observed in EL4 cells treated with a conjugate releasing Dox by a pH-dependent manner. As a result, the treated tumor cells were engulfed by dendritic cells (DC) in vitro, and induced their expression of CD80, CD86, and MHC II maturation markers. Conjugates with Dox bound via an amide bond only increased translocation of HSPs to the membrane, which led to an elevated phagocytosis but was not sufficient to induce increase of the maturation markers on DCs in vitro. Both types of conjugates induced engulfment of the target tumor cells in vivo, that was more intense than that seen with free Dox. It means that the induction of anti-tumor immunity documented upon treatment of EL4 lymphoma with HPMA-bound Dox conjugates does not rely solely on CRT-mediated cell death, but involves multiple mechanisms.

[Cost of acute heart failure related readmissions]. / Náklady na rehospitalizaci pacientu s akutním srdecním selháním.

AIM OF STUDY: To assess direct in-patient cost and length of stay in the intensive care unit (ICU) and the standard cardiology unit in acute heart failure (AHF) readmissions. RESULTS: Out of 1 759 patients hospitalized with acute heart failure, 223 patients were readmitted to Faculty Hospital Brno-Bohunice (Czech Republic) during study period (61.4% male; mean age 71.2 years) with mean total cost CZK 85 120 (Euro 3 095) per length of stay 9.2 days and interventions. Comparing to the first hospitalization of study cohort (223 pts.) the decrease was recorded in mean room rate, length of stay and need of ICU stay (from 48% to 42% pts.), nevertheless ICU stay increased (from 3.7 days to 4.1 days). The growth of mean cost was recorded in both procedures in angiology (the decrease in number of coronary angiography which is cheaper was more remarkable than PCI decrease in readmitted patients) and arrhythmology (including device: pacemaker, ICD, CRT) which made 57.5% of total readmission costs. CONCLUSION: The difference in mean in-patient cost between the first and second hospitalization was 18%. The antiarrhytmic procedures had the most significant impact on total readmission cost and its variability, butwe assume that these procedures will reduce within next readmissions and their impact will weaken as in angiology procedures.

Novel star HPMA-based polymer conjugates for passive targeting to solid tumors.

Novel star polymer-doxorubicin conjugates designed for passive tumor targeting have been developed and their potential for treatment of cancer has been investigated. In the present study the synthesis, physico-chemical characterization, drug release, bio-distribution and preliminary data of in vivo efficacy of the conjugates are described. In the water-soluble conjugates the core of a molecule formed by poly(amido amine) (PAMAM) dendrimers was grafted with semitelechelic N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers bearing doxorubicin (Dox) attached by hydrazone bonds enabling intracellular pH-controlled hydrolytic drug release, or by GFLG sequence susceptible to enzymatic degradation. The controlled synthesis utilizing semitelechelic copolymer precursors facilitated preparation of polymer conjugates in a broad range of molecular weights (1.1-3.0·10(5) g/mol). In contrast to free drug or linear conjugates the star polymer-Dox conjugates exhibited prolonged blood circulation and enhanced tumor accumulation in tumor-bearing mice indicating important role of the EPR effect. The star polymer-Dox conjugates showed significantly higher anti-tumor activity in vivo than Dox?HCl or its linear or graft polymer conjugates, if treated with a single dose 15 or 5 mg Dox eq./kg. Method of tumor initialization (acute or chronic experimental tumor models) significantly influenced effectiveness of the treatment with much lower success in treatment of mice bearing chronic tumors.

Polymer conjugates of the highly potent cytostatic drug 2-pyrrolinodoxorubicin.

This paper describes the synthesis and biological evaluation of a conjugate of the highly cytotoxic drug 2-pyrrolinodoxorubicin (p-DOX) with an N-(2-hydroxypropyl)methacrylamide copolymer (PHPMA) as a water-soluble biocompatible polymer carrier, utilizing the advantageous concept of polymer-drug conjugates. The conjugate of p-DOX with HPMA copolymer (PHPMA/p-DOX) was prepared by reacting the PHPMA/DOX conjugate, where the DOX was bound via a hydrazone bond, with 4-iodobutyraldehyde. The hydrazone bond between the polymer and drug is susceptible to pH-controlled hydrolysis, enabling prolonged stability in circulation and fast p-DOX release under conditions mimicking the intracellular environment. The in vitro cytostatic activity of free p-DOX was in accordance with literature, whereas its PHPMA conjugate exhibited a 1.3- to 5-fold lower cytotoxicity, depending on the cancer cell line, when compared to the free p-DOX. This is in qualitative agreement with the data obtained for DOX and its HPMA copolymer conjugates. On mice bearing T-cell EL4 lymphoma, no tumor suppression was observed from the free p-DOX at a subtoxic dose of 0.1 mg/kg, whereas the PHPMA/p-DOX conjugate significantly inhibited the initial tumor growth at approximately equitoxic doses of 0.4 and 0.8 mg p-DOX eq/kg. However, moderately elevated doses of the p-DOX equivalent in the conjugate caused toxic effects, making accurate dosage setting essential.

Surface modification of polyimide sheets for regenerative medicine applications.

In the present work, two strategies were elaborated to surface-functionalize implantable polyimide sheets. In the first methodology, cross-linkable vinyl groups were introduced on the polyimide surface using aminopropylmethacrylamide. In the second approach, a reactive succinimidyl ester was introduced on the surface of PI. Using the former approach, the aim is to apply a vinyl functionalized biopolymer coating. In the latter approach, any amine containing biopolymer can be immobilized. The foils developed were characterized in depth using a variety of characterization techniques including atomic force microscopy, static contact angle measurements, and X-ray photoelectron spectroscopy. The results indicated that both modification strategies were successful. The subcutaneous implantation in mice indicated that both modification strategies resulted in biocompatible materials, inducing only limited cellular infiltration to the surrounding tissue.

Synergistic action of doxorubicin bound to the polymeric carrier based on N-(2-hydroxypropyl)methacrylamide copolymers through an amide or hydrazone bond.

The cytostatic effects of polymeric conjugates based on N-(2-hydroxypropyl)methacrylamide copolymers (PHPMA) and containing doxorubicin bound through amide and hydrazone bonds (mixed conjugates) were compared with the cytostatic effects of monoconjugates containing drug bound through an amide or hydrazone bond. One group of mixed conjugates was formed from two comonomers containing doxorubicin bound to the methacryloyl group through a spacer and an amide (DOX(AM)) or hydrazone (DOX(HYD)) bond via copolymerization with HPMA. A second group of mixed conjugates was formed from two different interconnected HPMA copolymers, one containing DOX(AM) and the other DOX(HYD), forming a high-molecular-weight branched structure. The third mixed polymeric system was a simple mixture of monoconjugates DOX(AM)-PHPMA and DOX(HYD)-PHPMA. Simultaneous treatment with all mixed forms of the polymeric derivatives of doxorubicin significantly increased antitumor efficacy after application of monoconjugates, suggesting a synergizing effect that could be used in designing new doxorubicin-containing therapeutic systems.

Doxorubicin release is not a prerequisite for the in vitro cytotoxicity of HPMA-based pharmaceuticals: in vitro effect of extra drug-free GlyPheLeuGly sequences.

A systematic study was designed to elucidate differences in cytostatic activity in vitro between HPMA-based doxorubicin conjugates synthesized using different polymerization techniques and differing in peptidyl side chain. A polymer-drug conjugate containing doxorubicin (DOX) bound to HPMA copolymer backbone through the enzymaticaly non-cleavable sequence GlyGly shows low but significant cytotoxicity in vitro in seven cancer cell lines of mouse (EL4, 38C13, 3T3, BCL1) and human (SW620, Raji, Jurkat) origin. The low cytotoxicity can be considerably increased by the presence of additional drug-free GlyPheLeuGly side chains. P1 conjugate, i.e. non-targeted HPMA copolymer bearing doxorubicin bound via a biodegradable GlyPheLeuGly sequence, synthesized by direct copolymerization of HPMA with monomeric doxorubicin and thus without additional drug-free GlyPheLeuGly sequences is less effective compared to PK1 synthesized by polymer analogous reaction and thus containing extra drug-free GlyPheLeuGly sequences. Significant activity-enhancing effect was not seen with other amino acid/oligopeptide sequences (e.g., Gly or GlyGly). The activity-enhancing effect of GlyPheLeuGly sequences is more obvious in the conjugate containing doxorubicin bound to HPMA through GlyGly sequence. Derivatization of the terminal carboxyl group of the extra GlyPheLeuGly side chains (amide, N-substituted amide, free carboxyl) does not significantly influence the cytotoxicity of the conjugates. The presence of the GlyPheLeuGly sequence in the conjugate structure increases its rate of intracellular accumulation. Normal cells (Balb/c splenocytes) accumulate less polymer-doxorubicin conjugate compared to cancer cells (T cell lymphoma EL4, B cell lymphoma Raji and T cell leukemia JURKAT).

New HPMA copolymer-based drug carriers with covalently bound hydrophobic substituents for solid tumour targeting.

Various conjugates of anticancer drug doxorubicin (Dox) covalently bound by the hydrolytically degradable hydrazone bond to the drug carrier based on N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers were synthesised. Structure of the conjugates differed in the type and the content of hydrophobic substituent (dodecyl, oleic acid and cholesterol moieties) introduced into the polymer structure. In aqueous solutions the conjugates self-assembled into high-molecular-weight supramolecular structures, such as polymeric micelles or stable hydrophilic nanoparticles 13-37 nm in diameter, depending on the type and the content of hydrophobic substituents. Treatment of mice bearing EL-4 T cell lymphoma with the conjugates in the therapeutic regime of drug administration (i.v.) resulted in significant tumour regression with up to 100% of long-term survivors, depending on the dose and the detailed structure of the carrier. The nanoparticles formed by the conjugate bearing cholesterol moiety exhibited prolonged blood circulation and enhanced tumour accumulation indicating an important role of the EPR effect in excellent anticancer activity of the conjugate.

Induction of systemic antitumour resistance with targeted polymers.

Conjugates based on N-(2-hydroxypropyl)methacrylamide (HPMA) represent a new generation of antibody-targeted polymeric anticancer drugs with both cytotoxic and immunoprotecting/immunomobilizing activity. 20-90% of mice that are cured of EL4 mouse T-cell lymphoma, BCL1 mouse B-cell leukaemia and 38C13 mouse B-cell lymphoma by injection of doxorubicin-HPMA conjugate develop a long-lasting memory and systemic antitumour resistance. It is suggested that the main activity of the polymeric drug, directly after application is - due to the high level of the drug - of cytotoxic and cytostatic nature. Thereafter, long-term conjugates persist at low concentration in the circulation, which are capable of mobilizing the defence mechanisms of the host. Until now, seven patients with generalized carcinoma were treated with doxorubicin-HPMA-human-Ig conjugate. Disease stabilization, lasting from 6 to more than 18 months, was recorded.

Antibody-targeted polymer-doxorubicin conjugates with pH-controlled activation.

The paper is dealing with the synthesis and properties of new non-targeted or antibody-targeted polymer drug conjugates, bearing doxorubicin (DOX) attached via a spacer susceptible to pH-controlled hydrolysis (hydrazone conjugates), designed as anticancer drugs facilitating site-specific therapy. These conjugates are stable in a pH 7.4 buffer, modeling conditions during transport in the body, but release DOX and activate it inside target cells as a result of pH changes when going from outside to inside the cells. Conjugates containing an antibody directed against T lymphocytes bind effectively and specifically T cell lymphoma EL 4 cells. Cytotoxicity of the hydrazone conjugates is higher than that of classic conjugates, depending on the detailed structure of the polymer, the spacer between the drug and polymer carrier and method of antibody conjugation. Cytotoxicity of some of the conjugates is comparable even with that of the free drug. In both protective and therapeutic regimes of drug administration, the in vivo anti-tumor activity of the conjugates containing DOX was enhanced with long-term survivors (T-cell lymphoma EL 4, C57BL/6 mice) in comparison with much less effective free DOX or a classic P(N-(2-hydroxypropyl)methacrylamide)HPMA-DOX conjugate (already clinically tested).

HPMA copolymers containing doxorubicin bound by a proteolytically or hydrolytically cleavable bond: comparison of biological properties in vitro.

N-(2-Hydroxypropyl)methacrylamide (HPMA) copolymer carrier containing the anticancer drug doxorubicin bound either by a proteolytically degradable bond (non-targeted PK1 or targeted with alpha-CD71 mAb) or by a hydrolytically degradable bond were synthesised and tested in vivo for various biological properties. Mouse 38C13 B-cell lympoma was used as a well established and defined cell line for this study. 38C13 cells are sensitive to free doxorubicin and IC50 was very low, about 0.014 microM. PK1 showed a strongly decreased cytostatic effect, IC50 being 12.6 microM. alpha-CD71 targeted conjugate, which can be considered as an antibody-targeted form of PK1, had IC50 0.358 microM. HPMA copolymer with doxorubicin bound via a hydrolytically sensitive bond (HYD conjugate) showed a high cytostatic effect with IC50 about 0.052 microM. We demonstrated that HYD conjugate inhibited DNA synthesis and induced p21(Waf1/Cip1) protein expression (p21(Waf1/Cip1) is cyclin-dependent kinase inhibitor which blocks cell cycle progression) as quickly as free doxorubicin, whereas PK1 acted much more slowly. Similarly, apoptosis induction measured by Annexin V binding and Caspase 3 activity was detected later after incubation of cells with PK1 or alpha-CD71 targeted conjugate. Apoptosis was manifested by elevation of bax and bad mRNA levels, which was much more rapid and intense in the case of free doxorubicin and HYD conjugate. Expression of antiapoptotic genes as well as cyclin-dependent kinases was surprisingly not affected.

Polymeric anticancer drugs with pH-controlled activation.

The paper is dealing with the synthesis and properties of new, nontargeted or antibody-targeted pH-sensitive polymer-doxorubicin (DOX) conjugates designed as anticancer drugs facilitating site-specific therapy. These conjugates are stable and inactive during transport in the body but activate inside target cells as a result of pH changes outside and inside the cells. Cytotoxicity of the conjugates depends on the detailed structure of the polymer and of the spacer between the drug and polymer carrier. In both protective and therapeutic regimes of drug administration, the in vivo antitumor activity of the pH-sensitive conjugates containing DOX was significantly enhanced (T-cell lymphoma EL 4, C57BL/16 mice) in comparison with the free DOX or classic PK1, the PHPMA-DOX conjugate clinically tested at present.

Clinical implications of N-(2-hydroxypropyl)methacrylamide copolymers.

Different anticancer drugs, farmorubicin, doxorubicin, paclitaxel and cis-platin have been conjugated through a Gly-Phe-Leu-Gly tetrapeptide side chain to a water-soluble synthetic polymeric carrier based on N-(2-hydroxypropyl)methacryalmide (HPMA) non-targeted or targeted with galactosamine and/or human IVIg and used in Phase I clinical trials. Conjugation of the drugs to the polymeric carrier that is non-toxic and non-immunogenic in man significantly decreased their non-specific organ toxicities and increased maximum tolerated dose up to 5 times. Macromolecular therapeutics based on HPMA have radically different pharmacokinetics. Drugs are not released from their polymeric carrier and remain in the peripheral blood and urine of patients mostly in their polymer-bound form. A clinical response against some refractory cancers was recorded in Phase I clinical trials. It was also demonstrated that doxorubicin-HPMA copolymer conjugates containing an immunoglobulin moiety have both cytostatic and immunomobilizing activity.