review therapeutical practicepaşca maria dorina case report 29 severe outcome of influenza a h1n1...

1REVIEW

5 Chronic lymphocytic leukaemia. 1Chronic lymphocytic leukaemia. 1Malignant hemopathy with multiple 1Malignant hemopathy with multiple 1therapeutic options 1therapeutic options 1Ivănescu Ana-Maria, Oprea Mădălina, 1Ivănescu Ana-Maria, Oprea Mădălina, 1Coliță A., Turbatu A., Lupu Anca Roxana1Coliță A., Turbatu A., Lupu Anca Roxana1ORIGINAL PAPERS1ORIGINAL PAPERS112 Resistance to breast cancer treatment 1Resistance to breast cancer treatment 1induced by transport mechanisms1induced by transport mechanisms1Zob Daniela, Gruia Iuliana, Anghel Rodica1Zob Daniela, Gruia Iuliana, Anghel Rodica117 Risk factors for tuberculosis in HIV 1Risk factors for tuberculosis in HIV 1infected patients1infected patients1Stoian Andreea Cristina, Niculescu Irina, 1Stoian Andreea Cristina, Niculescu Irina, 1Cupșa A., Dumitrescu Florentina, Iocu 1Cupșa A., Dumitrescu Florentina, Iocu 1Cristina, Giubelan L., Marinescu Liliana1Cristina, Giubelan L., Marinescu Liliana122 Thrombo-prophylaxis in total hip 1Thrombo-prophylaxis in total hip 1arthroplasty using oral anti-thrombotics 1arthroplasty using oral anti-thrombotics 1anticoagulat – Rivaroxaban1anticoagulat – Rivaroxaban1Popescu Gh. I., Lupescu Olivera, Pătru 1Popescu Gh. I., Lupescu Olivera, Pătru 1Cristina, Nagea M., Vasilache Cornelia, 1Cristina, Nagea M., Vasilache Cornelia, 1Dimitriu Al., Macovei R. Al.1Dimitriu Al., Macovei R. Al.1THERAPEUTICAL PRACTICE1THERAPEUTICAL PRACTICE11271Communication strategies that improve 1Communication strategies that improve 1the relationship between pharmacist 1the relationship between pharmacist 1and patients with disabilities1and patients with disabilities1Paşca Maria Dorina1Paşca Maria Dorina1CASE REPORT1CASE REPORT11291Severe outcome of influenza a H1N1 1Severe outcome of influenza a H1N1 1rhombencephalitis in a 6-year-old 1rhombencephalitis in a 6-year-old 1child. Case report1child. Case report1Vișan Angelica, Drăgănescu Anca, 1Vișan Angelica, Drăgănescu Anca, 1Negulescu Cristina, Bilașco Anuţa, 1Negulescu Cristina, Bilașco Anuţa, 1Vasile Magdalena, Măntescu Ruxandra, 1Vasile Magdalena, Măntescu Ruxandra, 1Merișescu Mădălina, Slavu Diana, 1Merișescu Mădălina, Slavu Diana, 1Dogaru Cornelia, Marinescu Violeta, 1Dogaru Cornelia, Marinescu Violeta, 1Luminos Monica1Luminos Monica11331Cystic adrenal tumour - A case of 1Cystic adrenal tumour - A case of 1atypical abdominal pain1atypical abdominal pain1Săvulescu F., Nica R., Surdeanu D., 1Săvulescu F., Nica R., Surdeanu D., 1Cirlan C., Darmanescu M., Trifu V.1Cirlan C., Darmanescu M., Trifu V.1

Vol. XVIII Number 1 March 2014

National Institute of Infectious Diseases "Prof. Dr. Matei Balş"

Romanian Society of Pharmacology, Therapeuticsand Clinical Toxicology

Romanian Academy ofMedical Sciences

University of Medicine and Pharmacy "Carol Davila"

Journal published in cooperation with:

CNCSIS Category: B+ Code: 605

getCITEDIndexCopernicusEBSCOhost NLM (National Medical Library)

Academic Medical Database:

Founders

Emanoil Manolescu Mircea Angelescu Liviu Ioan Miclea

Editor-in-Chief TherapeuticsAdrian Streinu-Cercel (Professor, Member of Academy of Medical Science, Head of Department of Infectious Diseases, National Institute of Infectious Diseases "Prof. Dr. Matei Balş", University of Medicine and Pharmacy Carol Davila Bucharest)

Editor-in-Chief Clinical Pharmacology and ToxicologyVictor A. Voicu (Professor, Member of Romanian Academy, Head of Department of Pharmacology, Toxicology and Clinical Psychopharmacology, University of Medicine and Pharmacy Carol Davila Bucharest)

Associate EditorMonica Luminos (Associate Professor, National Institute of Infectious Diseases "Prof. Dr. Matei Balş", University of Medicine and Pharmacy Carol Davila Bucharest)

International Scientific Board

Laure Aurelian (Professor, Senior Associate, The Johns Hopkins School of Public Health), • Hege Christensen (Professor, School of Pharmacy, Uni-versity of Oslo, Norway), • Jaime Kapitulnik (Professor, The Hebrew University of Jerusalem, Israel), • Momir Mikov (Senior Lecturer, School of Pharmacy, University of Otago, New Zealand), • Stanislav Yanev (Professor, Head of Department Drug and Toxicology, Bulgarian Academy of Science, Bulgaria), • Olavi Pelkonen (Professor, Head of the Department of Pharmacology and Toxicology, University of Oulu, Finland), • Olivier Patey (Professor, Chef de service des maladies infectieuses et tropicales CHI, Villeneuve-Saint Georges, France), • George C. Rodgers (Professor of Pediatrics, Pharmacology and Toxicology, University of Louisville, Kentucky, USA), • Robert Smith (Professor, Brown Medical School, U.K), • Jean Paul Stahl (Professor, Rédacteur en chef de Médecine et Maladies Infectieuses, Elsevier Maison, Grenoble, France), • Michel Urbain (Chief of Research Department, Societe de Etude et de Research Biologique, Paris, France), • Andrei Iagăru (Assistant Professor, Department of Nuclear Medicine, Stanford University, USA) • Serafim Kastanakis (Professor, University of Crete, Greece)

Romanian Scientific BoardIoana Alina Anca (Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Eduard Apetrei (Professor, University of Medicine and Pharmacy Carol Davila Bucharest, Member of Romanian Academy of Medical Science), • Ştefan Sorin Aramă (Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Constantin Arion (Professor, University of Medicine and Pharmacy Carol Davila Bucharest, Member of Romanian Academy of Medical Science), • Anca Buzoianu (Professor, University of Medicine and Pharmacy Iuliu Haţieganu Cluj-Napoca) • Carmen Dorobăţ (Professor, University of Medicine and Pharmacy Gr.T. Popa, Iassy), • Constantin Dumitrache (Professor, Member of Romanian Academy, University of Medicine and Pharmacy Carol Davila Bucharest), • Leonida Gherasim (Professor, University of Medicine and Pharmacy Carol Davila Bucharest, Member of Romanian Academy of Medical Science), • Ioan Hăulică (Professor, Member of Romanian Academy, University of Medicine and Pharmacy Gr.T. Popa, Iassy), • Daniela Ion (Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Ion Fulga (Professor, Head of Department of Pharmacotherapy and Pharmacology, University of Medicine and Pharmacy Carol Davila Bucharest), • Sorin Leucuţa (Professor, University of Medicine and Pharmacy Oradea), • Radu Macovei (Professor, Head of Department of ICU-Toxicology, University of Medicine and Pharmacy Carol Davila Bucharest), • Alina Manolescu (researcher, ICSMCF), • Nicolae Miu (Professor, University of Medicine and Pharmacy Iuliu Haţieganu Cluj-Napoca), • Ostin C. Mungiu (Professor, Head of Department of Pharmacology and Clinical Toxicology, University of Medicine and Pharmacy Gr.T. Popa, Iassy), • Lucian Negruţiu (Professor, University of Medicine and Pharmacy Victor Babeş Timişoara), • Florian Popa (Professor, Chancellor of University of Medicine and Pharmacy Carol Davila Bucharest), • Irinel Popescu (Professor, Head of Department of Surgery and Liver Transplant, University of Medicine and Pharmacy Carol Davila Bucharest, Member of Academy of Medical Science), • Laurenţiu Mircea Popescu (Professor, Member of Romanian Academy, Head of Department of Celular Biology, University of Medicine and Pharmacy Carol Davila Bucharest), • Valeriu Popescu (Professor, University of Medicine and Pharmacy Carol Davila Bucharest, Member of Romanian Academy of Science), • Florica Stăniceanu (Professor, Univeristy of Medicine and Pharmacy Carol Davila Bucharest), • Dan Tulbure (Professor, Head of Department of ICU, University of Medicine and Pharmacy Carol Davila Bucharest), • Doina Ţăţulescu (Professor, University of Medicine and Pharmacy Iuliu Haţieganu Cluj-Napoca), • Coriolan Ulmeanu (Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Doina Velican (Researcher, Member of Romanian Academy of Medical Science), • Florin Căruntu (Associate Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Adrian Gabriel Popescu (Associate Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Anca Drăgănescu (Pediatrician, INBI Prof. Dr. Matei Bals, Bucharest) • Paraschiva Postolache (Associate Professor, University of Medicine and Pharmacy Gr.T. Popa, Iassy), • Alexandru Rafila (Associate Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Adriana Hristea (Associate Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Gabriela Leşanu (Lecture, University of Medicine and Pharmacy Carol Davila Bucharest) • Ion Lică (Lecture, University of Medicine and Pharmacy Carol Davila Bucharest), • Raluca Papacocea (Lecture, University of Medicine and Pharmacy Carol Davila Bucharest), • Maria Pasca (Associate Professor, University of Medicine and Pharmacy Targu Mures)• Voichiţa Lăzureanu (Assistant Professor, University of Medicine and Pharmacy Victor Babeş Timişoara), • Anca Macovei (Assistant Professor, University of Medicine and Pharmacy Carol Davila Bucharest), • Anca Streinu-Cercel (Lecture, University of Medicine and Pharmacy Carol Davila Bucharest), • Andrei Tica (Associate Professor, University of Medicine and Pharmacy Craiova) • Laura Topor (Assistant Professor, University of Medicine and Pharmacy Carol Davila Bucharest) • Mihail Tudosie (Assistant Professor, University of Medicine and Pharmacy Carol Davila Bucharest) • Ionel Alexandru Checheriță (Assistant Professor, University of Medicine and Pharmacy Carol Davila Bucharest) • Mihai Săndulescu (Assistant Professor, University of Medicine and Pharmacy Carol Davila Bucharest) • Toma Papacocea (Assistant Professor, University of Medicine and Pharmacy Carol Davila Bucharest)

General Registrar Of Editorial BoardVictoria Aramă (Professor, University of Medicine and Pharmacy Carol Davila Bucharest)

Issue EditorElisabeta Otilia Benea (Associate Professor, University of Medicine and Pharmacy Carol Davila Bucharest) Oana Streinu-Cercel Ana-Maria Tudor Alexandra Mărdărescu

Publishing EditorMihaela Cristina Negulescu

Editorial Office Institutul Naţional de Boli Infecţioase Prof. Dr. Matei Balş, Pavilionul IV, Etaj 41 Dr. Calistrat Grozovici Str., Sector 2, Bucureşti, C.P. 021105, O.P. 10

E-mail: [email protected] [email protected]

Published by SC Editura Rp. SRLCUI RO9954898, RC J40/7184/1997Address: 6 Codrii Neamțului Str., Bl. PM 26bis, Sc. A, Et. 8, Ap. 36, sector 3, Bucharest, Romania Tel/Fax: 031.80.40.513; 0724.356.578 E-mail: [email protected]: www.terapeutica.ro

ISSN 2066-0170

5XVII, Vol.18, Number 1/2014

Ana-Maria Ivanescu1 I.C.Bratianu Avenue, District 3, Bucharest, 030171e-mail: [email protected]

Summary. Chronic lymphocytic leukemia is a chronic lymphoproliferative disorder, indolent but het-erogeneous in evolution and difficult to predict. Beside cases whith long lasting evolution even without specific treatment, the clinical practice often faces cases with advanced stages, poor prognostic factors and high risk of disease progression in which choosing the best treatment tailored to each patients is a difficult task. Classical chemotherapy schemes have been effective but combining them with or shifting to new therapeutic agents, along with improved investigation techniques, led to a significant increase in the complete remission rates as well as improvement of survival, without risks of disease progression. During and after therapy, patients need to be thoroughly monitored, the detection of minimal residual disease increasing the chance to discover, early in the process, unresponsive cases or relapses. Complete clinical investigations and paraclinical tests, performed at the onset of disease have an important contribution for the selection of the best therapeutic option and moment of treatment initiation. Key words: chronic lymphocytic leukaemia, prognosis factors, chemotherapy, treatment options

1. Hematology Department - Coltea Clinical Hospital, Bucharest2. Carol Davila University of Medicine and Pharmacy, Bucharest

Ivănescu Ana-Maria1, Oprea Mădălina1, Coliță A.1,2, Turbatu A.1, Lupu Anca Roxana1,2

CHRONIC LYMPHOCYTIC LEUKAEMIA. MALIGNANT HEMOPATHY WITH MULTIPLE THERAPEUTIC OPTIONS

REVIEWTherapeutics, Pharmacology and Clinical ToxicologyVol XVIII, Number 1, March 2014Pages: 5 - 11 © Copyright reserved 2014

Chronic lymphocytic leukaemia (CLL) is a frequent malignant hemopathy, indolent

in terms of clinical course but with an extremely heterogeneous evolution. While several cases have been mentioned with survival rates spanning over decades without the need of specific treatment, many patients experience a rapid progression of their disease from the moment of diagnosis, with an unfavourable response to chemotherapy. An essential risk factor for CLL development is the patient’s medical history. Based on numerous CLL familial groups[1] were genetic analysis was possible, specialists noted that the age for CLL occurrence in the descendants of a known patient tends to decrease. According to a report released by National Cancer Institute Familial Registry, the average diagnostic age for familial cases was 58 years, 14 years below sporadic cases, this representing the only differentiation criterion[2]. Compared to non-family cases of CLL[3], in family cases the survival period was not significantly changed and the risk of aggressive transformation was

not increased either. In most sporadic cases the average age is 60 years at the time of diagnosis, a consequence of the decrement in the immune response given by the process of aging. The disease occurs more often in men than in women; 10% of cases are present in adults below 40, while CLL cases in children are quite scarce[4]. CLL is the only leukaemia not associated with exposure to ionised radiation. Chemical or alky-lating agents lack an aetiological impact on CLL and environment factors hardly have a major influence on the pathogenesis. [5,6]. Although there is no substantial evidence on viruses’ involvement in CLL’s aetiology, the infection with a Human T-cell lymphotropic virus- Human T-cell Leukaemia/Lymphoma Virus (HTLV-I) - triggers, however, some rare forms of leukaemia, namely adult T-cell Leukemia/Lymphoma.

At the onset, most patient are asymptomatic, the disease being caught at a routine check-up during which lymphocytosis is detected in the peripheral blood as well as lymphadenopathies and mild splenomegaly. In more advanced stages symptoms may be: asthenia, fatigue, fever, night sweats, weight loss over a short amount of time, anaemia [7].

These accessible clinical and paraclinical signs grounded the two staging systems currently used - Rai and Binet staging [8,9] (Figure 1).

6 Therapeutics, Pharmacology and Clinical Toxicology

The symptoms occur as a result of bone marrow and other tissues infiltration with malignant cells and the alteration of humoral and cellular immunity. CLL is

defined by immunodeficiency, many patients presenting autoimmune disorders, especially haemolytic anemia and autoimmune thrombocytopenia. Other signs are susceptibility to recurrent viral infections (e.g. Herpes Zoster), bacterial infections (e.g. Pneumoccocus), fungal (e.g. Candida, Cryptococcus), with persistent evolution, complications and resistance to conventional therapy.[7]

In terms of treatment, several options have become available with the main purpose to obtain a complete, long term remission, thus increasing the survival chances for CLL patients. Together with standard treatments whose efficiency has been proved on decades, present clinical trials perform an extensive search for innovative methods to improve the therapeutic response. A series of classical prognosis factors was identified: advanced stages of disease, lymphocyte doubling time <12 months, initial absolute lymphocyte count > 50.000/mmc, abnormal karyotype. New factors for unfavourable evolution have also been established, in close connection with modern investigation techniques: thymidine kinase positivity, high levels of Beta 2-microglobulin, positive CD38, positive ZAP-70, IgVH mutation status, cytogenetic abnormalities (p53 mutations, trisomy 12 or 17p, 13q, 11q deletion)[7,10,11,12].

Identifying the factors that influence the clinical evolution is essential and in close relation to the purpose of therapy, namely to provide the best life quality and to treat only when the patient becomes symptomatic. The treatment’s purpose is to obtain complete remission (CR), reduction of CLL cells from blood, bone marrow and lymph nodes, as much as possible. Remission is known in various forms: complete remission - undetectable CLL cells; minimal residual disease (MRD)- low CLL cell count, detectable only through highly sensitive tests (flow cytometry,

molecular biology), partial remission (PR)- detectable CLL, but the CLL cell count in blood and bone marrow is low and the lymph nodes diminish.

Considering each case’s particularities it is difficult to institute a standard first-line therapy, even less to agree on an optimal strategy. Hence, several strategies are in place: from “wait and watch” with a rigorous monitoring of symptoms, to the identification of high risk cases and early introduction of treatment - chemotherapy, immunotherapy, radiotherapy and stem cell transplant. Without bias for any of the abovementioned therapies, I will discuss briefly the specificity of the latter.

Monotherapy

Glucocorticoids are efficient as unique treatment for CLL in patients with haemolytic autoimmune anaemia, or autoimmune thrombocytopenia. In patients without autoimmune manifestations, glucocorticoids as single agents, can temporally control the diseases in about 10% of cases. They can function even for CLL patients with non-functional p53. Simultaneous therapy with H2 receptor antagonists and prophylactic antibiotherapy may decrease the risk of treatment complications[13, 14].

Alkylating agents - Chlorambucil: since it was first introduced in 1952, Chlorambucil has been the main alkylating agent used in CLL. Its oral administration renders it as a tolerable drug, without adverse events like: cystitis, alopecia or gastrointestinal disorders that can sometimes occur as a reaction to other alkylating agents. Myeloid and megakaryocytic series suffer little impact. In patients with advanced CLL, chlorambucil was administered for, at most, 6 months, in fixed dosage - 15mg/day until achievement of CR or stage 3 for toxicity. Some studies reported a higher complete or partial response rate (89,5%) compared to cyclophosphamide, doxorubicin, vincristine and prednisone treatment (CHOP) [15,16] but with the price of higher myelotoxicity related to long term use.

Bendamustine is another active agent for CLL. Trials on i.v. bendamustine administered for two consecutive days, every 4 weeks revealed a rate of overall response rates (ORR) varying from 56%-93% and CR rates from 7% to 29% in patients with relapsed or refractory CLL. [17]

Cyclophosphamide has a resembling activity as chlorambucil in CLL. Intermittent or simultaneous cyclophosphamide therapy predisposes at hemorrhagic cystitis, hence the recommendation is to administer it in a single dose, in the morning and the patients should be instructed to consume at least 2-3 liquids/day. [13,14,18].

Deoxyadenosine analogs. Fludarabine (2 fluoro-ara-AMP) is a fluorinated derivative of adenosine analogs, effective in CLL treatment. After i.v. or oral administration, the drug leads to complete or

Figure 1. Rai and Binet staging of chronic lymphocytic leukaemia


partial hematologic response in a large percentage of patients. Several trials reported ORR asociated with fludarabine phosphate injection, of approximately 45%, of which 10% presented complete remission, in patients previously treated with different agents. Furthermore, the ORR rate of about 70% - 38% with CR - has been observed in patients who received fludarabine as first line regimen. Fludarabine, alone, seems to be more effective in patients with CLL than other combined chemotherapy regimens, as COP[13, 19].

Cladiribine (2-chloro deoxyadenosine) is a deoxyadenosine analog which, as fludarabine, has proven active in CLL. Specialists used various schemes with different doses and ways of administration that turned out to be. Monthly courses of i.v. cladiribine led to 40-60% ORR in patients previously treated with alkylating agents and even higher percentages in naive patients. In naïve CLL patients who received oral cladiribine, the ORR reached 75%. However, cladiribine treatment failed to improve the survival rate, the mean survival being around 9 months, while unresponsive patients have an average survival period of 4 months. As with fludarabine, patients with no favourable clinical response after two cladiribine cycles should benefit from alternative therapies in order to decrease the level of toxicity. [20,21,22]

Polychemotherapy

Chlorambucil - Prednisone. As first line treatment in CLL, chlorambucil, in or without glucocorticoid associacion , has been used, at large scale, achieving a longer disease free survival, even in advanced stages. The main controversy regarding the ORR is still open, especially in what concerns choosing a conservative treatment for patients with early stages of disease versus polychemotherapy associated or not with immunotherapy. These options in relation with prognostic factors are under continuous assessment and the main objective in studies that have been designed to investigate the new therapeutic agents efficacy [21,23].

Cyclophosphamide - Vincristine - Prednisone .The cyclophosphamide-vincristine-prednisone (CVP) combination showed an outcome improvement both in naive patients and in some cases with refractory CLL. Almost a quarter of patients obtain complete remission in the aftermath of this therapy. No significant differences in terms of survival have, so far, been observed either in patients treated with CVP or those who received chlorambucil and prednisone. However, the latter may reach a response after CVP regimen administration. Apparently CVP treatment has no advantage over deoxyadenosine analogs, as fludarabine. [24]

Cyclophosphamide - Doxorubicine - Vincristine - Prednisone. Adding doxorubicine in CVP regimen has represented a highly studied decision in patients with advanced CLL. In those cases where doxorubicin

has been added, specialists noted that the survival rate expanded, on average, to over 4 years. Another interesting aspect highlighted by the studies was less significant role and benefit of vincristine within CHOP regimen, PR and ORR rates having been 64% and 75% in patients treated with CHOP, respectively 65% and 72% in those who received CVP [25].

Fludarabine - Cyclophosphamide (FC)Fludarabine-Cyclophosphamide combination may lead to clinical favourable outcome- CR after 4-6 administrations, for 30-35% of naive patients. This response rate is superior to the fludarabine regimen as a unique agent, the ORR rate and median disease-free survival period, were significantly better than patients only treated with fludarabine. [26]

Recent therapeutic methods

Alemtuzumab is a specific human monoclonal antibody- CD52, a glycosylated protein, linked to the cell membrane by a glycosylphosphatidylinositol (GPI) anchor, present on most lymphocytes. It can induce mediated lysis, antibody dependant cell-mediated cytotoxicity and represents a direct lymphocyte inhibitor, including malignant B lymphocytes. Intravenously or subcutaneously administered it has a significant activity for the treatment of patients with recurrent or relapsed CLL. Among its essential features, one can count the capacity to annihilate CLL lymphocytes with 17q13 deletions which are, in general, resistant to standard therapy [27,28]. The drug’s toxicity which occurs in most cases (~80%) consists of: shivers (90%), fever (85%), nausea (53%) and rash (33%). Except for the rash, the rest of these reactions usually decrease after the first administration. An important consequence of toxicity is immunosuppression, more than 50% of patients getting infections with opportunistic microorganisms (e.g. cytomegalovirus) during or after the treatment. [29,30]. As alemtuzumab seems to be the highest active drug in the process of blood and bone marrow leukemic cell destruction, which, usually are resistant to standard therapy, several studies have been performed in order to assess its use in consolidation therapy. [31,32,33].

Rituximab is a monoclonal CD20 antibody, initially used in follicular lymphoma. Although, compared to cells in the follicular lymphoma, CD20 is revealed at a lower level by B-cell lymphocytes in CLL, a series of clinical trials demonstrated that this monoclonal antibody is beneficial for CLL patients due to its role in inducing cellular apoptosis. [34,35,36]. When Rituximab is used as single agent it can induce PR in less than a third of patients. 1st or 2nd degree toxicity associated to the first administration appears in approximately 90% of cases, with frequent fever, shivers, nausea, vomiting, hypotension or dyspnoea. These manifestations are also subsequent to the increased white cell count, over 50x109/L at the time of


treatment initiation, when cytokine release syndrome may take place, as a result of TNF-α or interleukine-6 involment. Other adverse events are tumour lysis syndrome, drug induced neutropenia, disseminated intravascular coagulation (DIC). We can reduce difficulties related to treatment initiation by decreasing the rate of infusion and by first dose fraction, that is to say 100 mg Rituximab during the first day and the remaining of the dose, the following day. These reactions’ severity and risk scale down with the following infusions. Prior to the initiation of rituximab therapy, reactivation of an underlying infection with hepatitis B or C virus should be considered. In these situations the physician should reassess the liver function and a simultaneous antiviral therapy is recommended. [37,38, 38].

Fludarabine - Rituximab. This combination is well tolerated, its efficacy being higher than treatment with Fludarabine alone. Patients treated with the two drugs may obtain CR in almost half of the cases. Associated toxicity resembles toxicity noticed in Fludarabine, as unique agent. In addition, the median disease-free survival period turned out longer as a consequence of immunochemotherapies associacion.

Fludarabine-Cyclophosphamoide-Rituximab treatment appears to have an elevated efficacy, accounting for CR in a quarter of the cases and ORR for 73% of relapsed patients after first line treatment. A higher response rate could be observed in naive patients, in what concerns the median disease-free survival, it was improved compared to those treated with FC. [40,41].

New therapeutic agents

In recent years, many clinical studies focused on finding new therapy options for relapsed or refractory CLL[42]. The following table counts the newest agents and their action mechanism, achieved during clinical trials. (Table I)

Autologus stem cell transplantation. Several studies focused on the benefits of large dose chemotherapy, followed by stem cells transplantation, recovered from CLL patients. Considering the high probability that acquired stem cells could contain leukemic cells, autologus stem cell transplantation is a complex issue, even for treated patients with minimal residual disease. Although specialists approached various techniques to remove unwanted leukemic cells, a small number of cases succeeded in achieving a complete answer for CLL patients, i.e. autologus stem cell transplantation prolonged the period of survival, with no signs of progression.

Allogenic stem cell transplantation. For young patients with a poor prognosis, allogenic stem cell transplantation represents an option. Aggressive therapy may eliminate leukemic cells which are undetectable to molecular techniques for clonal immunoglobulin gene rearrangement. A five year survival period can be attained in about 75% of cases,

without additional therapy, while for approximately 25% of them, immunosuppressive therapy for graft versus host diseases is necessary. Results following allogenic transplant in patients with negative prognostic are encouraging, including for individuals with 17p deletion, but it is essential to undergo an initial thorough pre transplant assessment, with an estimation of the risk factors. [48,49,50].Complementary therapies

Spleen Radiotherapy (RT), in 3-8 Gy doses, 1,5 Gy per fraction every 2 days is highly indicated for symptomatic splenomegaly. [51]. Nodal, local or generaliezed RT can help in voluminous, obstructive splenomegalies. Splenectomy represents an option for splenomegaly generating abdominal discomfort, splenic sequestration or refractory to treatment-symptomatic autoimmune hemolytic anaemia. [52,53]. Leukopheresis is recommended when increased leukocyte count can trigger hyperviscosity syndrome. 400mg/kg I.V. immunoglobulines, every 3 weeks can be safely used for severe hypogammaglobulinemia, in order to prevent infection complications. [54] Each of the strategies described above has its own use within the clinical practice, providing many alternatives. However the therapeutic course of a CLL case can often be challenging. Therefore, specialists must perform an initial assessment of prognosis factors, rigorous and regulate monitoring of patients, as well as a thorough selection of the best therapy options, at the right moment, in order to obtain a qualitative remission and a long period of survival, free of any event connected to the disease.

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40. Wierda W, O’Brien S, Wen S, et al. Chemoimmunotherapy with fludarabine, cyclophosphamide, and rituximab for relapsed and refractory chronic lymphocytic leukemia. J Clin Oncol. 2005; 23:4070.[PMID: 15767647].41. Tam CS, O’Brien S, Wierda W, et al. Long-term results of the f ludarabine, cyclophosphamide, and rituximab regimen as initial therapy of chronic lymphocytic leukemia. Blood. 2008;112:975-980.42. Wu M,Akinleye A, Zhu X. Novel agents for chronic lymphocytic leukemia. J Hematol Oncol. 2013; 6:36.43. Jaglowski SM, Alan J, Flynn JM, et al. A phase Ib/II study evaluating activity and tolerability of BTK inhibitor PCI-32765 and ofatumumab in patients with chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) and related diseases. Journal of Clinical Oncology, ASCO Annual Meeting Proceedings. 2012; Post-Meeting Edition, Vol 30, No 15_suppl (May 20 Supplement),6508.44. Lynn Wang Y, Cheng S, Ma J, et al. BTK Inhibition Targets in Vivo CLL Proliferation Through Its Effects On B-Cell Receptor Signalling Activity. Blood. ASH Annual Meeting Abstracts, 2012; 120: 2903.45. Byrd JC, Blum KA, Burger JA, et al. Activity and tolerability of the Bruton’s tyrosine kinase (BTK) inhibitor PCI-32765 in patients with chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL): Interim results of a phase Ib/II study. Journal of Clinical Oncology, ASCO Annual Meeting Proceedings 2011 (Post-Meeting Edition);Vol 29, No 15_suppl (May 20 Supplement),: 6508.46. Byrd JC, Furman RR, Coutre S, et al. The Bruton’s Tyrosine Kinase (BTK) Inhibitor Ibrutinib (PCI-32765) Promotes High Response Rate, Durable Remissions, and Is Tolerable in Treatment Naïve (TN) and Relapsed or Refractory (RR) Chronic Lymphocytic Leukemia (CLL) or

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Ibrutinib

• Previously known as: PCI-32765• Potent and irreversible Bruton’s tyronise kinase (BTK) inhibitor; oral administration.• It inhibits: Chemokynes, cytokines, signalling pathways mediated by microenvironment as well as

ex vivo chemotaxis; • It reduces Ki67+ population and initiated apoptosis. [43,44] • It leads to several complete remissions and increase of survival period, even for cases of Richter’s

transformation or Bulky diseases [45,46,47].• In naive patients: results are encouraging. On a 26 patients lot, aged <65, the survival period,

without progression spanned to 15 months, in 96% of them. [47]. • Adverse events: nausea, fatigue, diarrhoea, rash, bleeding events- bruises. 3rd or 4th degree

reactions: diarrhoea, hematologic infections and toxicity (anaemia and/or thrombocytopenia), with an average 10-13% rate of incidence. [45,46].

Ofatumumab

• Human monoclonal antibody- CD20 that binds to a different CD20 epitope. • Higher complement dependant cytotoxicity and antibody dependant cell mediated cytotoxicity, in

pre clinical trials. • The antibody has been approved by FDA for treatment of refractory CLL patients to fludarabine

and alemtuzumab.

Lenalidomide • Immunomodulatory treatment promising in the area of salvage therapy for patients with relapsed/refractory CLL.

Idelalisib (GS-1101, CAL-101)

• Known as GS-1101 or CAL-101, a specific Phosphoinositide 3-kinase inhibitor (P13K), with apoptotic activity against CLL leukemic cells.

Veltuzumab

• A IInd generation, humanized monoclonal antibody, anti-CD20 which can bind selectively and irreversibly to CD20 molecule. The antigen binding site is similar with Rituximab, but with improved binding avidity. Prolonged subcutaneous administration produced encouraging results in naive patients, with relapsed/refractory CLL and acceptable tolerability (degree: 3-4).

XmAb5574

• CD19 monoclonal antibodies are currently being considered for clinical applications. XmAb5574 is a human monoclonal antibody, anti-CD19 with increased binding avidity. During pre clinical trials, XmAb5574 exhibited antitumor activity, including direct cytotoxicity, antibody dependant and cellular phagocytosis against leukemic cells. Unlike other anti-CLL, monoclonal antibodies, XmAb5574 didn’t display complement dependant cytotoxicity activity.

Navitoclax (ABT-263)

• Known as ABT-263. • It is an anti-apoptotic protein inhibitor, including Bcl-XL, Bcl-2 and Bcl-w with efficacy for

patients with relapsed or refractory CLL.

TRU-016• TRU-016 is a protein that targets CD37, present on normal and leukemic B cells. It has direct

toxicity and antibody-dependant cellular cytotxicity against CLL, superior to Rituximab. TRU-016 and TRU-016 and Bendamustine have both been used in the management of relapsed CLL.

Alvespimycin (17-DMAG, KOS-1022)

• Alvespimycin, a synthetic derivate of the antibiotic geldanamycin is a heat shock protein 90 inhibitor. It has a regulatory activity on genes and proteins involved in the proliferation and survival of CLL cells.

Table I. New therapy agents in CLL treatment [42]


Daniela Zob252 Soseaua Fundeni, 2-nd District, Buchareste-mail: [email protected]

Abstract. Breast cancer represents the most common form of cancer women develop. Although prognosis has significantly improved over the last years, due to increasing trends in early diagnosis and advanced treatment, the medical community still faces a series of severe problems. One of them is resistance to certain chemotherapy agents that constitutes the main cause of death in more than 90% of patients. Despite intense and thorough studies on resistance mechanisms, their relevance for the clinical practice continues to be unclear. The present paper aims to identify a possible resistance mechanism pattern to chemotherapy applied in breast cancers, which includes the attachment of cytostatics to albumin thiols, as a consequence of oxidized protein degradation. Given their chemical structure, these thiols may be responsible for the impossibility of cytostatics to bind to transport albumins, thus inducing therapeutic failures and implicitly resistance. The content of this article focuses on the measurement of albumin thiol levels in patients diagnosed with breast cancer that developed resistance after the first series of antitu-mor treatment. We also assessed the level of copper-carrying proteins, namely ceruloplasmin involved in oxidation-reduction reactions as well as the overall level of non-enzymatic endogenous antioxidants. The results reveal the occurrence of a significantly damaging oxidative stress that destroys the structure of transport proteins and, indirectly, can create resistance mechanisms. Key words: Breast cancer, resistance to chemotherapy, transport mechanisms, oxidative stress, albumin thiols, ceruloplasmin, antioxidants

Oncology Institute “Prof Al Trestioreanu”, Bucharest

Zob Daniela, Gruia Iuliana, Anghel Rodica

RESISTANCE TO BREAST CANCER TREATMENT INDUCED BY TRANSPORT MECHANISMS

ORIGINAL PAPERTherapeutics, Pharmacology and Clinical ToxicologyVol XVIII, Number 1, March 2014Pages: 12-16 © Copyright reserved 2014

Introduction

According to WHO estimates (International Agency for Research on Cancer- EUCAN)

for 2012, breast cancer is a major public health issue, known as the highest incidence rate neoplasia in women from Romania. Of all annually diagnosed forms of neoplasia - 25.22% (figure 1) represent breast cancer which also accounts for the largest mortality rate- 16.74% (figure 2).

The rough incidence for breast cancer in the European Union is 94.2/100.000 women (figura 3)while the mortality rate is 23.1/100.000 women (figura 4).

Despite the fact that prognosis for breast cancer has scaled up considerably during the last years, a high number of female patients still presents

an evolution in the disease. The increasing rate of incidence noticed until 2002, as a consequence of mammography introduction and uptake, tends to currently show a decreasing curve [1] also due to a lower use of postmenopausal oestrogen replacements. Even if the tendency in diagnosis increments in women over 50 (375/100.000) in comparison with women under 50 (42.5/100.000), approximately 23% of breast cancers are found in women under this age because this group of population corresponds to 73% of the overall feminine population. [2]

The treatment of the metastatic disease is a challenge for any clinician because many patients are susceptible to therapeutic failure and subsequently to death due to malignancy. In over 90% of patients with metastatic breast cancer resistance to various chemotherapy agents is the main cause of therapeutic failure. [3]

Even if resistance mechanisms have been identified and clarified in cell cultures they are less clear in the clinical practice (Cimoli et. Al 2004,


Uggla et. Al 2007). Resistance to chemotherapy may appear either prior to treatment administration (primary or innate resistance) or develop in time, after the exposure to a chemotherapeutic agent (acquired resistance). [4] Chemoresistance represents a major limitation for treatment and leaves few effective options [5]. Both innate and acquired resistance to taxane and anthracycline are common, without distinct mechanisms. [6] The most known in vitro mechanism that detects resistance to more than one class of chemically related agents (multidrug resistance) is the overexpression of efflux proteins. The most frequent drug efflux are members of the ATP binding cassette family (ABC): P-glycoprotein Pgp, also known as multidrug resistant protein – MDR or ABCB1, multidrug resistance associated protein 1- MRP-1 also named ABCG, while in breast cancer the resistance protein is BCRP or ABCG2. ABC cassette substrates transporter includes a diverse range of compounds, many of them being structurally independent. These proteins shield the cells and tissue by exporting toxins, including anticancer agents from cells in normal tissue as well as from cancer cells. [7]. Generally, ABCB1 transports large hydrophobic compounds, while ABCC1 and ABCG2 transport both hydrophobic drugs and large non ionic compounds. In relation to breast cancer, ABC proteins were previously involved in resistance to taxane and doxorubicin.

Remission failure and the development of drug-resistant cancer following antiangiogenic therapy were mostly linked to the unintentional induction of a hypoxic tumour microenvironment. [8][9] Estimates reveal that for 40-50% of breast cancer cases, the tumour site is in a hypoxic environment. [10] Hypoxia inducible factor (HIF1a), often detected within solid tumours in response to hypoxia leads to angiogenesis in order to form new blood vessels, necessary to tumour growth. [11][12] HIF1a may also be activated by nitric oxide, cytokines, growth factors, oncogene expression or by mutations in tumour suppressors. [13]

The present paper aims to identify a possible resistance mechanism pattern to chemotherapy applied in breast cancers, which includes the attachment of cytostatics to albumin thiols as a consequence of oxidized protein degradation. It is well known that tumour tissues produce excess endogenous antioxidants, especially metallothionein and sulphur proteins that once entered in circulation may interact with oxygen reactive species and suffer oxidation-reduction reactions (redox) which eventually can cause excess thiols production. Given their chemical structure, these thiols may be responsible for the impossibility of cytostatics to bind to transport albumins, thus inducing therapeutic failures and implicitly resistance. The content of this article focuses on the measurement of albumin thiols levels in patients diagnosed with

Figure 1. Cancer incidence for women in Romania

Figure 2. Cancer mortality for women in Romania

Figure 3. Cancer incidence for women in Europe

Figure 4. Cancer mortality for women in Europe


breast cancer that developed treatment resistance. We also assessed the level of copper-carrying proteins, namely ceruloplasmin involved in redox reactions as well as the overall level of non-enzymatic endogeneous antioxidants.

Material and methods

During January 2011 and January 2014 we observed 40 female patients, diagnosed with metastatic breast cancer who received cytostatic drugs for metastatic disease in the Oncology Institute “Prof Al Trestioreanu”, Bucharest - Medical Oncology Department I.

Inclusion criteria: 1. Age: >18 2. Histological confirmation of breast cancer 3. Presence of metastatic diseases 4. Immunohistochemical testing 5. Signed informed consent for the introduction

of their data in the study 6. The patients received adjuvant cytostatic

therapy or treatment for the metastatic disease and presented disease progression in less than six months from the last chemotherapy session or during chemotherapy.

Exclusion criteria: 1. Low IPECOG>3 performance status; serum

biochemical markers: bilirubin >1.5X normal values, AST, ALT >2.5X normal values except for hepatic metastases where a value <5X normal values is accepted, CBC when taken into records with ANC (absolute neutrophil count)<1500/mm3, T <100.000/mm3, Hb <8g/dl

2. Patients with no pathology confirmation of mammary neoplasia

3. Patients with no metastatic disease4. Patients with different forms of cancer

developed during the last 5 years 5. Patients whose disease didn’t progress in less

than 6 from the last adjuvant chemotherapy or therapy for metastatic disease.

The biological sample used in the study was the serum we collected from our patients subsequent to their signing the informed consent, also approved by the Ethical Committee in Oncology Institute “Prof Al Trestioreanu”, Bucharest. Based on the 5th exclusion criterion but following all the inclusion criteria we constituted a control group that had to be as homogeneous as possible, with no resistance mechanisms, from which we collected the same biological samples.

Measurement tests for plasma SH groups rely on the ability to develop a colour complex, measurable spectrophotometrically, with maximum absorbance at 412 nm to acid reaction 5, 5-dithiobis-2-nitrobenzoic acid (DTNCB), at room temperature.

Normal values range from 370-450 µmol/l. Increased values correlate with an excess production of thiols that represent the result of degradation of oxidized proteins. These thiol groups are highly reactive and through their electronic nature have the ability to attach easily to any circulatory compound whose structural core contains electronic metal groups (e.g. platinum).

Total antioxidants The used method relies on the serum’s ability

to reduce iron. At low pH, Felll complex- tripiridil-triazine (Felll –TPTZ) is reduced to the ferrous form with a new complex that develops an intense, measurable blue colour, with a peak of absorption of 539 nm. Any reaction with a potentially positive redox under the mentioned conditions may lead to Felll –TPTZ complex reduction. Using an excess Fe, the limiting factor of Felll –TPTZ complex and of colour formation represents the sample’s reduc-ing ability. The reaction measures the reduction of ferric ion complex-2,4,6-tri (2-piridil)-1,3,5- triazine (TPTZ) to a coloured product.

The reaction mixture is prepared on the spot and contains 25 ml of acetate buffer, 2, 5 ml of TPTZ solution and 2, 5 ml of FeCl3.

The reaction is monitored for 4 minutes, focusing on optical density to 593 nm. Results are obtained by multiplying the read absorbance value by a M40 spectrophotometer, with an index correction that takes into account the concentration of used solu-tions, the optical path, size of cuvettes, etc. Literature data [14] indicate as normal values for this serum type of test: 0,9-1,4 mmol/l. Elevated values may be due to the mobilization of antioxidant protection systems in response to oxidative stress, installed as a disease consequence or due to a diet rich in antioxidants, the patient chooses to adopt after the diagnosis disclosure.

CeruloplasminCeruloplasmin is a blue colour protein with

oxidase activity on polyamines, polyphenols and inorganic ferrous ions (Fe 2+). However, the bio-logic substratum is Fe2+, ceruloplasmin having the highest affinity for it. The catalytic oxidation of Fe2+ or complexes containing Fe2+ is called ferroxidase activity. There are several ways to determine fer-roxidase activity in ceruloplasmin, from which we opted for the Ravin method: p-phenylene diamine reaction in an acetate-acetic acid buffer. Ceruloplas-min carries a complex series of roles, among which: copper ion transport, oxidation of organic amines, Fe2+ oxidation to Fe3+ after its release from trans-ferrin and ferritin, antioxidant activity against lipid peroxidation, endogenous modulation of the inflam-matory response, stimulation of cell proliferation and angiogenesis [15]. Ceruloplasmin is an acute phase protein, with an intermediate amplitude response


to other proteins in acute phases, whose level au-guments 2-3 times in inflammations, pregnancy, traumas, and surgeries. CP seric concentration is a useful clinical indicator of Wilson’s disease. Each CP molecule contains 6 or 7 copper atoms, this being the state for 90-95% of serum copper [16].

The serum sample is incubated for 30 minutes with a known amount of ferric ion in a 0, 2 M environment to 5, 5 ph and a 37 ºC temperature. The reaction is stopped with a 0,5% sodium azide solution. The coloured product is quantitatively measured through spectrophotometry, by reading absorbance 540 nm from a control sample. Values are expressed in U.I. The enzyme quantity that converts 1 substrate μmol per minute is defined as 1 ceruloplasmin unit. Normal values vary between 80-120 U.I. ceruloplasmin.

Results The first phase was to establish the total values

of serum thiols in patients with constant evolution of the disease, during chemotherapy drug treatment compared to patients who developed no treatment resistance.

As Table no. 1 shows, the mean value increased

in both groups. Compared to the accepted maximum limit, the value increase in the control group is 2,2% while in the resistance group 21,5% (figure 5). This enables us to conclude that in the latter group resistance mechanisms can occur due to low transport capacity, where the albumins underwent a supplementary oxidative degradation and no longer have the means to attach and transport chemotherapy agents as effectively as in the non-resistance group.

The obtained values entitle us to determine redox reactions, measurable through the serum’s

ability to reduce iron. These reactions cause reactive degradation species, responsible for thiol production.

In table no. 2 the results reveal a 14% increase in the resistance group compared to the accepted maximum limit, which translates into intensified oxidation processes bred by the tumour mass and its reactivity, taking into account that the tumour tissue induces oxidative stress.

Control group Patients who developed resistance

460 μmol/l 547 μmol/l

Table I. Mean levels of thiols in investigated patients

(Normal values: 370-450 µmol/l)

Figure 5. Mean levels of thiols in investigated patients


1.53 mmol/l 1.60 mmol/l

Table II. Mean value of oxidation-reduction reactions- respon-sible for thiol production, assessed through FRAS technique

(Normal values: 0,9-4mmol/l)


114 U.I. 122 U.I. Table III. Values of ceruloplasmin activity in patients with

breast cancer (Normal values 80-120 U.I.)

Figure 7. Values of ceruloplasmin activity in patients with breast cancer

Figure 6. Mean value of oxidation-reduction reactions -respon-sible for thiol production, assessed through FRAS technique


In order to complete the redox panel we also determined the ferroxidase activity in serum ceruloplasmin from the same biological samples.

For the control group we observed values within normal limits while in the group with apparent resistance we noticed a light increase that can be the result of a minor intensification which we believe initiates resistance mechanisms within the transport of chemotherapy agents.

Conclusions Chemoresistance plays an important role in the

weak response and low rate of survival of patients with advanced or metastatic breast cancer. It is both a challenging and devastating phenomenon for oncologists and patients, since it entails various complex mechanisms. Understanding these mechanisms is crucial for the improvement of chemotherapy agents’ use, especially of taxanes and anthracyclines in breast cancer. Despite numerous clinical and experimental trials, many questions remain unanswered to date.

A hypothesis is that the excess of one of ABC transporter proteins cannot provide sufficient resistance to chemotherapy, with the possible involvement of other mechanisms that trigger resistance. [17] The clinical relevance of combination chemotherapy to diminish the induction of resistance to chemotherapy agents becomes clear and requires taking into consideration those mechanisms that rely on stress induction as main objective, especially hypoxia. Aggressive cancers may even use the agent in the process of angiogenesis transcription, which ultimately generates the growth of multiple tumour cells, resistant to treatment. Recent studies have demonstrated that the combination between angiogenesis and HIF1 inhibitors can produce better results than using each agent, alone. The occurrence of reactive to oxygen species in low concentration may induce a series of signalling events that create an angiogenic profile of the tumour tissue, ultimately its oxygenation leading to a devastating oxidative stress that destroys the structure of transporter proteins and indirectly, initiates resistance mechanisms.

A thorough understanding of resistance to chemotherapy agents, as well as its inherent processes, engendered either through innate mechanisms or as response to antitumor treatment will open the way to a rational design of targeted therapies, with a maximal effect on tumour cells and the limitation of side effects.

References1. L al S , Mahajan A , Chen WN, Chowbay B. Pharmacogenetics of target genes across doxorubicin disposition pathway: a review. Curr Drug Metab 2010, 11:115–128. 2. Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D. Global cancer statistics. CA Cancer J Clin 2011, 61:69–90.3. Murray S, Briasoulis E, Linardou H, Bafaloukos D, Papadimitriou C. Taxane resistance in breast cancer: mechanisms, predictive biomarkers and circumvention strategies. Cancer Treat Rev 2012, 38:890–903.4. Fojo T, Menefee M. Mechanisms of multidrug resistance: the potential role of microtubule-stabilizing agents. Ann Oncol 2007, 18(Suppl 5):v3–v8.5. Rivera E. Implications of anthracycline-resistant and taxane-resistant metastatic breast cancer and new therapeutic options. Breast J 2010, 16:252–263.6. McGrogan BT, Gilmartin B, Carney DN, McCann A. Taxanes, microtubules and chemoresistant breast cancer. Biochim Biophys Acta 2008, 1785:96–132.7. Zelnak A. Overcoming taxane and anthracycline resistance. Breast J 2010, :309–312.8. Rapisarda A, Melillo G. Overcoming disappointing results with antiangiogenic therapy by targeting hypoxia. Nat Rev Clin Oncol 2012, 9: 378–390. 42. 9. Bottsford-Miller JN, Coleman RL, Sood AK. Resistance and escape from antiangiogenesis therapy: clinical implications and future strategies. J Clin Oncol 2012, 30: 4026–4034.10. Ward C, Langdon SP, Mullen P, Harris AL, Harrison DJ, et al. New strategies for targeting the hypoxic tumour microenvironment in breast cancer. Cancer Treat Rev 2013, 39: 171–179.11. Semenza GL. Oxygen sensing, homeostasis, and disease. N Engl J Med 2011, 365: 537–547. 12. Semenza GL. Hypoxia-inducible factors: mediators of cancer progression and targets for cancer therapy. Trends Pharmacol Sci 2012, 33: 207–214. 13. Stroka DM, Burkhardt T, Desbaillets I, Wenger RH, Neil DA, et al. HIF- 1 is expressed in normoxic tissue and displays an organ-specific regulation under systemic hypoxia. FASEB J 2001, 15: 2445–2453.14. Reinisch N, Kiechl S, Mayr C, Schratzberger P, Dunzendorfer S, Kohler CM, Buratti T, Willeit J, Wiedermann CJ. Association of high plasma antioxidant capacity with new lesion formation in carotid atherosclerosis; a prospective study, Eur. J.Clin.Invest, 1998, 28,787-792.15. Ehrenwald E, Chisolm GM, Fox PL. Intact human ceruloplasmin oxidatively modifies low density lipoprotein. J Clin Investig; 1994, 93:1493-1501.16. Cousins RJ. Absorption, transport, and hepatic metabolism of copper and zinc: special reference to metallothionein and ceruloplasmin. Physiol Rev; 1985, 65:238-309.17. H Wang, T Vo, A. Hajar, S Li, XChen, A M Parissenti, D N Brindley Z Wang1*et al. BMC Cancer 2014, 14:37.


Andreea Stoian126 Calea Bucuresti, Craiovaemail: [email protected]

Abstract. Objectives: To identify risk factors for tuberculosis (TB) in patients with HIV/AIDS (PIH), through a retrospective study. Methods: A retrospective study compared the period 01/01/2006-31/12/2010 on two groups of PIH born between 1987-1990, horizontally infected with HIV in early childhood and ARV treatment: group A-16 PIH diagnosed with TB in 2010 and lot B-46 PIH without opportunistic infections at the time of inclusion in the lot (2010). Results: Demographics: group A-9M/7F; 7R/9U, median age = 20 years and group B-22M/24 F, 18 R/28U, median age = 21 years. Average BMI (kg/m²) in group A vs group B was: in T3-22.22 ± 4.18 vs 23.19 ± 3.99 (p=0.44) at T2-21.78±3.21 vs 22.43±3.78 (p=0,54) and T1-17,44±3,01 vs 22,11±4,18 (p=0.0001). Adherence of 95-100% (score 19-20) to ARV therapy was found in group A vs group B: the T3- 8 (53.33%) PIH vs 39 (84.78%) PIH (p=0.005; RR=3.29); the T2-7(43.75%) PIH vs 41 (89.13%) PIH ( p=0.0005; RR=5.18) and T1-9 (56.25%) PIH vs T3-44 (95.65%) PIH (p=0.001; RR=4.58). Average CD4 (cells/mm ³) in group A vs group B was: to the T3-331.09±182.54 vs 658.09±332.2 (p=0.0001), at T2-173±113.89 vs 616.3±377.16 (p=0.0004) and at T1-103.82±53.21 vs 682.5±388.22 (p=0.0001). Average HIV-VL (copies/ml) in group A vs group B, was: to the T3-7893.23±2839.63 vs 2123.2±623.83 (p=0.0001), at T2-24790.25±7653.87 vs 954.71±224.33 (p=0.0001) and at T1-99520.97±24745.52 vs 78.43 ± 29.21 (p=0.0001). AIDS related clinical events were found in group A, 5 PIH (31.25%) and in group B, 2 PIH (4.34%) (p=0.03; RR=3.57). Conclusions: Identified risk factors for TB in PIH were downward evolutionary trend of CD4 cell count and upward of HIV-VL, within the 5 years before detection of these infections and AIDS related clinical events (nonTB), within 3 years before TB diagnosis, with poor adherence at ARV therapy. Key words: HIV, TB, risk factors

1. University of Medicine and Pharmacy from Craiova2. “Victor Babes” Infectious Diseases Hospital from Craiova

Stoian Andreea Cristina1, Niculescu Irina1, Cupșa A.1, Dumitrescu Florentina1,Iocu Cristina2, Giubelan L.1, Marinescu Liliana2

RISK FACTORS FOR TUBERCULOSIS IN HIV INFECTED PATIENTS


Introduction

Tuberculosis (TB) is a major public health problem, causing high levels of morbidity

and mortality. The incidence, prevalence and mor-tality are high in PIH compared with the general population [1], and the diagnosis of TB is difficult. Approximately 13% of infections with M. tubercu-losis occur in PIH, representing the most common opportunistic infection worldwide [1]. Although TB in PIH causes high levels of viral replication and can occur at different levels of immunosuppression, higher risk of developing the disease correlates with progressively increased immunosuppression. In this context, it is important to forecast the diagnosis possibilities based on surrogate markers.

Objectives

To identify risk factors for TB in PIH through a retrospective clinical and biological study.

Methods

A retrospective study compared the period 01/01/2006-31/12/2010 on two groups of PIH born between 1987-1990, horizontally infected with HIV in early childhood and being treated with cARV: group A-16 PIH diagnosed with TB in 2010 and group B-46 PIH without opportunistic infections at the time of inclusion in batch (2010). In each group were analyzed: demographics, body mass index (BMI), blood count (CBC), CD4 cells count, HIV- viral load (VL) antiretroviral therapy adher-ence score and AIDS related events.

Patients were recruited between 01/01/2009-31/12/2010, monitored parameters were analyzed


retrospectively over a period of five years prior to diagnosis of TB, monitoring every two years.

The study was initiated in 2010 and mentioned parameters were retrospectively analyzed within 5 years before TB diagnosis, with monitoring every two years, as follows: the period between 01/01/2009-31/12/2010, in which they were di-agnosed TB, was noted T3, the period between 01/01/2007- 31/12/2008 was noted T2 and the period between 01/01/2005-31/12/2006 was noted T1. Were excluded from the study those PIH who had recurrent TB during the monitoring period.

Adherence assessment was based on completing „Periodic Assessment Questionnaire on ARV adher-ence”. Scores were quantified in fields as follows: adherence score 19-20 = very good adhesion (95% -100%), adherence score 17-18 = average adherence (85% - 90%), score <16 = non - adherence (<80%). Statistical analysis was performed using Data Analysis and Descriptive Statistics module of Mi-crosoft Excel XP. Quantitative characteristics were described by mean, standard deviation and median. Comparison test was performed by Student-t test, χ2 or Fischer. Differences were considered statisti-cally significant for a threshold p <0.05 and highly significant at p <0.001, at a confidence interval of 95% and a relative risk (RR)> 2.

Results

Demographics: A-9M/7F group; 7R/9U and group B-22M/24 F, 18 R/28U, without statistically signifi-cant differences (p = 0.77). PIH age at baseline in the study had a average value of 20.57 ± 7.33 years, respectively a median of 20 years in group A and an average of 20.73 ± 6.82 years and a median 21 years in group B, with no significant differences (p = 0.93).

Clinical and immunological staging of PIH at T1, was as follows: in group A (previously diagnosed

TB) 3 PIH in stage B2, 5 PIH in stage B3, 2 PIH in stage C2, 6 PIH in stage C3 and in group B: 2 PIH in stage B1, 8 PIH in stage B2, 7 PIH in stage B3, 3 PIH in stage in C1, 5 PIH in stage C2 and 21 PIH in stage C3. There were no statistically significant differences between PIH distribution of the two groups for clinical and immunological stages: B2 (p=1.00), B3 (p=0.26), C2 (p=1.00) and C3 (p=0.77).

Adherence of 95-100% (score 19-20) to ARV therapy was found in group A vs group B: the T3-8 (53.33%) vs PIH 39 (84.78%) PIH (p = 0.005; RR = 3.29) at T2-7 (43.75%) PIH vs. 41 (89.13%) PIH (p = 0.0005, RR = 5.18) and T1-9 (56,25% ) PIH vs. T3-44 (95.65%) PIH (p=0.001,RR=4.58).(table no I.)

Value of BMI at T1 was lower in group A (17.44 kg/ m²) compared to group B (22.11 kg/m²), the difference being statistically significant (p = 0.001). There were no statistically significant differences between average values of BMI at T2 (p = 0.541) and T3 (p=0.991) (table no II).

Nonspecific biological aspects at T1, found lower values for hemoglobin (Hb) and leukocytes (WBC) in group A compared to group B. The differences were statistically significant (p <0.05) (table no III) .

Evolution of biological parameters showed a progressive downward trend of Hb: in group A, the interval T2-T1. There were statistically significant differences in Hb values between the two groups at T1 (p = 0.03), but not at T2 (p = 0.59) or T3 (p = 0.45). (graphic no 1.)

Leucocytes evolution showed a progressive downward trend: in group A, the interval T3-T2-T1, with statistically significant differences versus group B at T1 (p = 0.033), at T2 (p = 0.011) and T3 (p = 0.0006) (graphic no 2)

No differences were found in platelet count (PLT) between group A and group B at T1 (199022.22 ± 113147.62 cells/mm³ versus 201009 ± 76056.98 cells/ mm³, p = 0.93), at T2 (218763.48 ± 9301.12 cells/

Table I. Adherence to antiretroviral therapy in group A vs B

Group A Group B Statistical significance

Number of cases (%)

adherence score19-20

Number of cases(%)


Number of cases (%)

adherence score<16

Number of cases (%)


Number of cases (%)


Number of cases (%)

adherence score<16

T1 9 (56.25) 4 (25) 3(18.75) 44 (95.65) 1 (2.18) 1 (2.18) p=0.001; RR=4.58

T2 7 (43.75) 4 (25) 5(31.25) 41 (89.13) 2 (4.35) 3 (6.53) p=0.0005; RR=5.18

T3 8 (53.33) 2 (12.5) 6 (37.5) 39 (84.78) 7(15.22) 0 p=0.005; RR=3.29


mm³ versus 222178.21 ± 8156.78 cells/mm³, p = 0.16) and T3 (208000.21 ± 9883.22 cells/mm³ versus 212000 ± 10,812, 13 cells/mm³, p = 0.19).

Patients distribution depending on CD4 cell count showed that most of PIH in group A had an advanced level of immunosuppression at T1 (11 PIH) and T2 (11 PIH) than in group B, most showing a slight immunosuppression at T1 (34 PIH), T2 (28 PIH) and T3 (28 PIH), the discrepancy is statistically significant (p<0.05) (Graphic no 3, graphic no 4).

Average value of CD4 cell count in group A

compared to group B was: at T3-331. 09 ± 182.54 cells/mm³ versus 658.09 ± 332.2 cells/mm³ (p = 0.0001) at T2-173 ± 113.89 cells/mm³ versus 616.3 ± 377.16 cells/mm³ (p = 0.0004) and T1-103, 82 ± 53.21 cells/mm³ versus 682.5 ± 388.22 cells/mm³ (p = 0.0001) (Graphic no 5).

Average value of ARN-HIV (copies/ml) in group A vs group B, was: to T3-7893.23 ± 2839.63 vs 2123.2 ± 623.83 (p = 0.0001), the T2-24790.25 ± 7653.87 vs 954.71 ± 224.33 (p = 0.0001) and T1-99520.97 ± 24745.52 vs 78.43 ± 29.21 (p = 0.0001) (Graphic no 6).

Group A1 Group A2 Statistical significance

AverageBMI (kg/m²) Std. Dev. Average BMI

(kg/m²) Std. Dev. p

T3 22.22 4.18 22.23 3.99 0.991T2 21.78 3.21 22.43 3.78 0.541T1 17.44 3.01 22.11 4.18 0.001

Table II. Body Mass Index value in group A vs B

Group Hb WBC NS(%) PMN (%) E (%) Ly (%) Mo (%) PLT

A

No 16 16 16 16 16 16 16 16

Average 10.43 5623.91 5.90 42.66 2.37 23.59 6.05 199022.22Std. Dev. 2.28 2149.78 5.89 29.59 2.76 12.81 3.61 113147.62Median 10 5100 7 57 2 20 6 191000

Minimum 7.2 1400 0 7 0 0 0 33400Maximum 15.1 20600 13 79 11 56 20 384000

B No 46 46 46 46 46 46 38 46Average 12.80 6888.22 1.92 60.34 2.87 30.42 3.45 201009Std. Dev. 4.11 1954.56 2.07 9.77 3.71 10.59 2.03 76056.98Median 12 6551 2 60 2 31 3 215000

Minimum 10.12 3100 0 32 0 5 0 153000Maximum 16.09 12800 7 81 22 58 10 497000

p 0.03 0.033 0.000 0.51 0.09 0.012 0.000 0.93

Table III. Biological issues in group A vs B, at T1

0

2

4

6

8

10

12

14

T1 T2 T3

Group A Group B

Graphic 1. Hb evolution in group A vs group B

0

1000

2000

3000

4000

5000

6000

7000

8000

T1 T2 T3

Group A Group B

Graphic 2. WBC Evolution in group A vs group B


AIDS-related clinical events (non TB) were found in group A, 5 PIH (31.25%) in T1-3 PIH herpes zoster and T2-1 PIH with cerebral toxo-plasmosis, 1 cryptococcocal meningitis PIH and in group B at T2-2 PIH (4.34%), herpes zoster; p = 0.03, RR = 3.57 (graphic no 7, graphic no 8).

The number evolution of AIDS-related events in group A showed an upward trend thereof, from T3 to T1.

Between T3-T1 changes required ARV: 12 PIH (75%) in group A (before TB diagnosis) versus 8 PIH (17.39%) in group B, p <0.0001. Reasons for changing ARV regimen were as follows: in group A, treatment failure at 12 PIH, of which, 2 PIH experienced adverse reactions to ARVs and 2 PIH showed resistance to ARVs, while in group B, 2 PIH

with treatment failure, 2 PIH with ARV adverse reactions and 4 ARV-resistant PIH.

Discussions

TB is more common in people underweight (BMI <18 kg/m²) weight loss being a predictor factor for the disease [2-4]; in my study, body mass index was lower in PIH with TB versus PIH without TB (17.44 kg/m² versus 22.11 kg/m²). The presence of weight loss, along with fever, cough and dyspnea, increase the sensitivity and specificity of the diagnosis of TB [5, 6].

Most patients with TB were non-adherent to an-tiretroviral therapy, at the time of TB diagnosis and during the five years before the diagnosis, the adher-ence decreased progressively during this period. By default, the immunosuppression and viral replication

0

2

4

6

8

10

12

T1 T2 T3

CD4<200 cells/mm? CD4=200- 499 cells/mm? CD4>500 cells/mm?

Graphic 3. PIH distribution of group A by their CD4 cell count

0

5

10

15

20

25

30

35

T1 T2 T3

CD4<200 cells/mm? CD4=200- 499 cells/mm? CD4>500 cells/mm?

Graphic 4. PIH distribution of group B by their CD4 cell count

331,09

173 103,82

658,09 616,3

682,5

T3 T2 T1

Group A Group B

Graphic 5. CD4 cell count evolution in group A versus group B

4,99 lg

3,89 lg 4,39 lg

1,89 lg

2,97 lg 3,32 lg

T3 T2 T1

Group A Group B

Graphic 6. ARN-HIV evolution in group A versus group B

No clinical events; 11 PIH (69%)

Cryptococcal meningitidis; 1 PIH

(6%)

Brain toxoplasmosis;

1 PIH (6%)

Herpes zoster; 3 PIH (19%)

Graphic 7. Non TB AIDS-related clinical events in group A

Graphic 8. Non TB AIDS-related clinical events in group A


increased, which contributed to the increase in the number of AIDS-related clinical events, during the period before TB screening.

The presence of anemia was correlated with the risk of TB in PIH [2, 5], similar data were obtained in our analysis: anemia and lymphopenia, monocytosis, were associated with the TB.

Immunodepression was more pronounced at PIH with TB compared with controls, both at the time of TB diagnosis (103.82 cells/mmc versus 682.55 cells/mmc) and at T3 (331.09 cells/mmc versus 658.09 cells/mmc) and T2 (173 cells/mmc versus 616.3 cells/mmc), data from the literature shows that tuberculosis can occur at different values of CD4 lymphocytes [7-9].

Viral replication level was highest for PIH with TB versus control group, before the diagnosis of TB (3.89 lg copies/ml vs 3.32 lg copies/ml in T3 and 4.39 lg copies/ml vs 2 , 97 lg copies/ml at T2) and also, at the time of disease diagnosis (4.99 lg copies/ml vs 1.89 lg copies/ml). Studies show that TB causes high levels of viral replication [7-10].

Study limitations: limited number of patients studied do not offer enough statistical power

Conclusions

The general context of developing TB infections in PIH is characterized by heightened degree of im-munosuppression, the rate of viral replication and the presence of AIDS-related clinical events, in conditions of poor adherence to antiretroviral therapy. At the time of diagnosis of TB found anemia and decreased BMI.

References1. World Health Organization –Data and statistic, http://www.who.int/tb/data, [accesed at 11/06/2012].2. Komati S., Shaw P.A., Stubbs N., et al. Tuberculosis risk factors and mortality for HIV-infected persons receiv-ing antiretroviral therapy in South Africa, AIDS, 2010; 24 (12): 1849-55.3. Lange C., Herzmann C., Miglori G.B., Gori A. Tu-berculosis in Hoffman C., Rockstroh J-HIV 2010. Medizin Fokus Verlag, Hamburg, 2010, 344-58.4. Sterling T.R., Pham P.A., Chaisson R.E. HIV in-fection-Related Tuberculosis: clinical manifestations and treatment, Clin Inf Dis, 2010; 50 (3): 223-30.5. Van Rie A., Westreich D., Sanne I. Tuberculosis in patients receiving antiretroviral treatment: incidence, risk factors and prevention strategies- JAIDS, 2011;56(4):349-556. Vidya C. Shankar. Clinical criteria can predict TB co-Infection among HIV patients, Int J Tuberc Lung Dis 2009; 13: 47-53.7. Bartlett J.G., Gallant J.E. Management of Infections, in Bartlett J.G., Gallant J.E: Medical Management of HIV Infection 2005-2006 Edition. John Hopkins Medicine Health Publishing Business Group, Baltimore, 2005; 329-84.8. Manas E., Pulido E., Pena J.M., et al. Impact of tuberculosis on the course of HIV-infected patients with a high initial CD4 lymphocyte count. Int J Tuberc Lung Dis 2008; 12: 417-23.9. Masur H. Management of Opportunistic Infectious As-sociated with Human Immunodeficiency Virus in: Mandell, Douglas and Bennett’s Principles and Practice of Infectious Diseases, 6th ed, 2005; 1679-1707.10. Goletti D., Weissman D. et al. Effect of Mycobac-terium tuberculosis on HIV replication. Role of immune activation, J Immunol 1996; 157: 1271-8.


Olivera Lupescu8 Calea Floreasca, Bucharest, Romaniae-mail: [email protected]

Abstract. Rivaroxaban, a novel anti-thrombotic drug, has been described as having a highly potent activity on the coagulation mechanism, due to its reversible inhibiting action on the central point of the coagulation cascade, Factor Xa. Published studies reveal some advantages of Rivaroxaban compared to previously used anti-thrombotics: it is orally administered and rapidly absorbed and it allows a fixed dose regimen as its pharmacokinetic and pharmacodynamic profile is predictable. Having similar cardio-vascular and hepatic side effects to other anti-thrombotic drugs and a low risk of bleeding, as shown by the published studies, Rivaroxaban can be considered well tolerated and effective in prevention of thrombembolic events. The authors present some aspects regarding their experience with this drug used in thrombo-prophylaxis following hip arthroplasty, a high thrombo-embolic risk procedure with increasing frequency in modern orthopedics. The results of this prospective study –post-operative blood loss, the incidence of thrombo-embolic events, side effects confirm the data presented in literature regarding the efficacy and safety of Rivaroxaban in major orthopedic surgery. Keywords: hip arthroplasty, thrombo-prophylaxis, oral anti-thrombotics, Rivaroxaban

1. Orthopedics and Trauma Clinic, Clinical Emergency Hospital Bucharest, Romania2. University of Medicine and Pharmacy Bucharest3. County Emergency Hospital Braila

Popescu Gh. I.1,2, Lupescu Olivera 1,2, Pătru Cristina 2, Nagea M.2, Vasilache Cornelia 3, Dimitriu Al.4, Macovei R. Al.1,2

THROMBO-PROPHYLAXIS IN TOTAL HIP ARTHROPLASTY USING ORAL ANTI-THROMBOTICS ANTICOAGULAT – RIVAROXABAN


Introduction

Thrombo-Embolic Disease (TED) including Deep Venous Thrombosis (DVT), Pulmo-

nary Embolism (PE) as well as their complications, Post-Thrombotic Syndrome (PTS) and Pulmonary Hypertension (PHT) represent a major cause of morbidity and mortality in modern medicine. Compared to the situation some decades ago, the interest for preventing Thrombo-Embolism (TE) significantly increased as studies revealed that it can occur after any type of pathology, regardless the organ involved.

Venous Thrombosis is based on an abnormal activation of the coagulation cascade, which has several steps; that is why research has been oriented towards finding drugs effective in different steps of this cascade to be used treatment and especially in thrombosis prophylaxis[1] .As major orthopedic

surgery is unanimously recognized as having a high thrombo-embolic risk, thrombo-prophylaxis is of particular interest for the physicians involved in this activity, surgeons and anesthetists, as well. After arthroplasty (hip, knee), the current standard is nowadays represented by Low Molecular Weight Heparins (LMWH), replacing the “classical”. Unfrac-tioned Heparine (UFH) nowadays used mainly in the treatment of TE, and less in prophylaxis, due to its numerous side effects (most of them balanced when LMWH are used).[2] Also indicated by the guides, Fondaparinux sodium, which indirectly af-fects Factor Xa, has been suggested to proved to have a high risk of bleeding; as for the Vitamin K Antagonists (VKA), they have some inconvenients :non-specific mechanism of action, slow onset and offset of the anticoagulating activity, and a narrow therapeutic window, frequent laboratory monitoring being required. [3]

Thus, the major demands for TE prophylaxis were drugs with higher specificity and predictable effect after a fixed unique oral dose with less bleed-ing risk. One group of drugs meeting this demand is represented by Direct Inhibitors, acting on Factor


IIa and Factor Xa. As the inhibition of one single molecule of Factor Xa (the activated form of Fac-tor X) produces the inhibition of 50 molecules of thrombin, it is considered to have the central role in the coagulation cascade[4]. Thus, Direct Inhibitors of Factor Xa were introduced in clini-cal practice; in orthopedics, their main indication was TE prophylaxis after elective surgery (hip and knee arthroplasty)

One of these is Rivaroxaban which directly, spe-cifically and competitively binds to human Factor Xa; its selectivity was evaluated to be more than 10 000-fold greater selectivity than for other serine proteases. It rapidly and reversibly inhibits factor Xa (both free and fibrin-bound), without needing without antithrombin as a cofactor, as well as the prothrombinase complex, but is has no direct ef-fect on thrombin activity or platelet aggregation.[5]

After oral administration, the absorbtion of Rivaroxaban is rapid, with Cmax reached within 2–4 hours after; it has a plasma protein binding of 92–95% and there are no major or active circulating metabolites as it is two-thirds metabolized, and one-third excreted as unchanged active substance in urine. From the two-thirds metabolized, half is eliminated renally and half is eliminated by faecal route. No dose adjustment is necessary in patients with mild or moderate renal impairment. Rivaroxa-ban is contraindicated in patients with clinically relevant bleeding risk, such as those having hepatic disease associated with coagulopathy and it may be used with caution in cirrhotic patients with moder-ate hepatic impairment (Child Pugh B) if it is not associated with coagulopathy.[6]

The efficiency and safety of Rivaroxaban were the objectives of the RECORD

(Regulation of Coagulation in major Orthopedic surgery reducing the Risk of DVT and PE ) I, II, III and IV studies which prospectively evaluated the efficacy and safety of a fixed daily dose of 10 mg of Rivaroxaban as thrombo-prophylaxis after hip and knee arthroplasty .[7-10]

Since hip and knee arthroplasty represent an important part of our activity, there has been a continuously growing interest in modern methods of thrombo-prophylaxis within our Clinic [11].This

study presents our experience regarding the preven-tion of thrombembolic events using RIVAROXA-BAN as XARELTO (10 mg Rivaroxaban per tablet)

Material and method

This prospective study evaluates 46 patients oper-ated between 01.01.2010 and 01.01.2013 in the Clini-cal Emergency Hospital Bucharest, Orthopedic and Trauma Clinic, who received Xarelto (Rivaroxaban 10mg) as thrombo-prophylactic agent. The study

group included 24 males and 22 females, with ages between 42 and 76 years (mean age 64 yrs). (Figure 1)

The operation performed was the same for all the patient- Total Hip Arthroplasty (THA)and the indication for surgery was represented by hip os-teoarthritis (OA), which was primary (30 cases-16 males, 14 females) and secondary (16 cases- 6 males, 10 females).

From the 30 patients with primary OA, there were involvement of the hip was unilaterally in 28 patients (18 males, 10 females) and bilaterally in 2 patients, all females, while in the “secondary OA group” the involvement was unilaterally in 13 cases (5 males, 8 females) and bilaterally in 3 cases (1 male, 3 females).(Table I)

Pre-operative all the patients followed the same standard protocol established together with the anesthetists.

I. Standard pre-operative evaluation, consisting of: a. anamnesis - in the study group , 32 patients

had significant medical history, with one or more of the following comorbidities: arterial hypertension

Figure 1. The group of patients – age and gender

Type of OA males Females TotalPrimary unilaterally 20 14 34Primary bilaterally 0 2 2

Secondary unilaterally 2 4 6Secondary bilaterally 2 2 4

Total 24 22 46

Table I. Etiology of the OA in the studied group


(20 patients), ischemic coronary disease (8 cases), atrial fibrillation (4 patients), allergic reaction to penicillin (1 patient), diabetes mellitus (4 patients), arterial occlusive disease (1 patient), gout (2 patients), ankylosing spondilitis (1 patient), rheumatoid arthritis (5 patients), leukemia (1 case), psoriasis (3 cases). There were 21 patients smoking in the study group (8 of them more than 20 cigarettes daily). None of the patients had had relevant clinical evidence/ medical history for DVT or PE, nor for liver or renal impairment. There were no patients within the study group receiving anti-coagulants in the last 6 months prior to surgery, but 8 patients received chronic treatment with Low-Dose–Aspirin 325mg (4 for atrial fibrillation and 4 for ischemic coronary disease), which was not considered as contra-indication for post-operative prophylaxis with Rivaroxaban

b. clinical examination-relevant for TED- 12 patients had varicose veins, 3 patients had clinical signs of chronic cardiac failure

c. paraclinical evaluation - ECG, pulmonary X-ray, lab tests for blood and urine; blood - hemoleucogram, serum glucose, urea, creatinine, ionogramme, liver function tests, proteins, albumin, CK (creatin-kinase), LDH(lactic-dehydrogenase/, alkaline phosphatase, ESR (erythrocyte sedimentation rate), fibrinogen, coagulation tests ); urine-biochemical,cellular and bacteriologic exam

d. pre-operative anesthetist evaluation, assessing the risk.e. no pre-operative pharmacological thrombo-

prophylaxis was performed; non-pharmacological anti-thrombotic measures were applied (proper hydration, elastic stocks for the patients with varicose veins).

Spinal anesthesia was used in 34 patients and general anesthesia in 12 cases; antibiotic treatment was administered to the patients at induction:• cephalosporin with aminoglycosides in 14 cases• cephalosporin alone in 6 cases• quinolone with aminoglycosides in 12 cases• quinolones alone in 8 cases• quinolones with glycopeptides in 6 cases

Lateral hip approach was performed in all the cases, except 3 cases, when the posterior approach was used because it had had been previously used for stabilizing the fracture of the posterior wall.

The type of the prosthesis was chosen depending the known criteria: age, bone stock (the quality and the quantity of bone) and the ability of the patient to respect the weight bearing recommendations; the correlation between age and the type of the prosthesis is represented in Figure 2.

Active suction was used in all the cases during surgery and Intraoperative Red Blood-Cell Salvage-Reinfusion System (so called “Cell Saver”) was used in 20 of the 46 cases; intra-operative blood loss was 500- 1050 ml (mean value 650 ml) (Figure 3)

Re-infusion was performed in two variants:

• reinfusion at the end of the operation (usually, after fascial suture), after collecting and processing all intra-operative blood – when intra-operative blood loss did not exceed 800 ml, or

• collecting 800 ml, processing and re-infusing the resulted product, then collect, process and reinfuse the rest of the blood at the end of surgery.Transfusion with total blood or RBC (red blood cells)

was intra-operatively indicated in 16 cases (12 cases operated without reinfusion system and 4 cases when reinfusion system was used, but there were more than 500 ml blood lost before finishing the acetabular preparation).

Peri-prosthetic and subfascial active drainages were installed; post-operative blood loss was monitored; the first outcome of this monitoring was to establish the moment of starting thrombo-prophylaxis, as the indication for that is the moment when hemostasis is proven to had been completed; the second outcome was to evaluate post-operative blood loss. Elastic bilateral stockings were used at the end of surgery in all the patients.

Rivaroxaban 10 mg daily (fixed dose) was given to the patients, starting after a mean time of 9.2 hrs post-operative, since in all the patients it was proven that haemostasis had been established before that. The moment of first administration was between 8-10 hours post-operative, depending on the post-operative drainage.

The patients were monitorised post-operative by: 1. clinical evaluation once daily and whenever

it was considered necessary, general and local (the aspect of the legs and wound surveillance with

Figure 3. Blood loss during surgery

Figure 2. Correlation between age and the type of prosthesis


drainage removal 48 hrs after surgery). 2. laboratory tests: the same complete pre-

operative tests were routinely performed in the evening of the operation day (4-6 hours after the end of the suture) and the next morning

3. Hemoleucograms and coagulation tests were perfumed for all the patients at discharge

4. Recommended control for suture removal, then 35-42 days after surgery

5. Compression ultrasound routinely performed for all the patients at discharge (which was after a mean time of 9.5 days in this group of patients, from 8-16 days) , then 35 - 42 days after surgery (when the treatment with Rivaroxaban was stopped); exceptionally, Compression Ultrasound was indicated whenever there was a suspicion of DVT

Results

The duration of Rivaroxaban treatment was 35- 42 days after surgery (mean time 39.2 days), no discontinuation of the treatment was described in the study group.

From the studied group, 6 patients developed persistent edema of the operated leg; all of them after more than 3 weeks from surgery (after 24, 25, 29, 30, 31 and 34 days correspondingly). Doppler compression ultrasound was performed and it detected thrombosis of the peroneal veins in one of the patients, and of the anterior tibial veins in 2 patients, so Unfractioned Heparine (UFH) treatment of DVT was started, following known protocols.

Doppler Compression Ultrasound was performed at 2,4, 8 and 12 weeks after Heparine treatment was started; the thrombi progressively reduced and disappeared 12 weeks after DVT onset; venography confirmed thrombus disappearance.

PE appeared in 1 patient(62 yrs, heavy smoker with varicose veins) 6 days after surgery.

Post-operative bleeding was within standards described in literature (600-1300 ml) with a mean value of 775 ml; post-operative transfusions were indicated (1- 4 units) depending on the hemograme. The variation of

the Hemoglobin in the study group is shown in Figure 4No post-operative abnormal bleeding, nor at the

surgical site, neither in different organs and systems, and no major bleeding (defined as life threatening bleeding or a bleeding which required reintervention) were described.

One of the patients developed an acute allergic reaction to Cephalosporin, which had no relation to Rivaroxaban and did not interfere with its

administration (Figure 5)Complete paraclinical evaluation (lab tests) were

performed corresponding to the local standard procedures (routine post-operative tests, then second day routine tests), only repeated if the patients had post-operative anemia and needed blood transfusions. No supplementary evaluation of coagulation was necessary, none of the patients developed thrombocytopenia during these standard tests.

Liver and renal function tests were modified at 16 patients in the post-operative set; four of these patients had these tests modified the second day; the values returned to normal for all the patients at discharge. Rivaroxaban treatment was continued, we concluded that no impairment of liver or renal function appeared, nor other side effects

Discussion

The significant impact that TED after major orthopedic surgery continues to have upon mortality and morbidity makes the problem of thrombo-prophylaxis still actual, since none of the available methods managed to completely erase the risk of these complications.

Rivaroxaban, a novel oral anticoagulant has been described as having two major advantages: the oral administration and the fixed dose, no matter weight or comorbidities. For the moment, in our country, the indications for thrombo-prophylaxis with Rivaroxaban are represented by elective surgery of the hip (total hip replacement) and knee.[12]

The prospective study in this paper confirm those resulted from the RECORD studies, which show that the benefit-risk ratio for Rivaroxaban is definitely inclined to the first one.

Figure 4. Post-operative blood loss

Figure 5. Clinical outcome - adverse events


There are certain aspects which must be discussed following presenting the results of our experience:

The compliance to long-term treatment is much better when the treatment is administered orally (such as Rivaroxaban) then injectably. This is very important especially when prolonged treatment is necessary, such as thrombo-prophylaxis after hip arthroplasty, since studies demonstrated that the risk of developing TED after this type of surgery persists for 6 weeks after surgery.[13] So it is very important for succesful thrombo-prophylaxis in hip arthroplasty including that with Rivaroxaban to respect the long-time prophylaxis rule, which, accord-ing to literature consistently reduces the incidence of thrombo-embolic events

The moment when thrombo-prophylaxis should be started (before or after surgery) has not been concluded, yet. Since none of the two methods (pre- or post-operative initiation of prophylaxis) proved to be clearly superior, the debate is still ac-tual, especially because theoretically the patient is moving (sometimes difficult, but still moving) before arthroplasty, so at least this pro-thrombotic factor is not active, yet. Still, if pre-operative prophylaxis is intended, LMWH are to be chosen, since the indica-tions of Rivaroxaban are restricted to post-operative thrombo-prophylaxis.

The precise moment of the first administration has to be established following the recommendations of the producer: there must be at least 8 hours after suture was finished, but in the same time hemostasis must be proven to have been achieved when Riva-roxaban is first administered, in order to allow the coagulation mechanism to achieve haemostasis. The surgeon must be sure that no abnormal bleeding oc-curs at the surgical site, otherwise no anti-thrombotic drug can be administered. As there might be dif-ferences between different anti-thrombotic drugs regarding the time between wound closure and first allowed administration, the rule of proven hemostasis is valid for all these drugs.[14]

Since there is a considerably difference between the designs of the studies and the timetables of the visits, the precise data referring to the incidence of different events within the study group are not to be compared to those obtained in the RECORD studies. Still, it must be underlined that within the study group there were no side events, no cases of liver and renal impairment, nor gastro-intestinal disturbances, thus suggesting that Rivaroxaban might reduce the risk of these unpleasant events .

Conclusions

Correct and complete identification of the TE risk by identifying all the thrombotic factors requires a thorough evaluation of the patient. Once TE risk

being established, thrombo-prophylaxis should be started , preferably with one of the new effective anti-coagulants which were continuously improved in order to obtain efficacy, safety, specificity , toler-ability and a convenient administration.

This analyses, even if performed on a small group of patients, demonstrated that conveniently orally administered Rivaroxaban demonstrated to be ef-fective, safe and well tolerated, since no side effects were described in the study group. Until its indica-tions will be enlarged, the properties of Rivaroxaban makes it an efficient thrombo-prophylactic agent to be used in prosthetic orthopedic surgery.

References1. Esmon C. Basic mechanisms and pathogenesis of venous thrombosis Blood Reviews, 2001, 23, 225-229.2. Samama C., Gigou F. Low-Molecular-Weight Heparin vs. Unfractionated Heparin in Femorodistal Reconstruc-tive Surgery: A Multicenter Open Randomized Study Annals of Vascular Surgery, 2002 , 9, 845-853.3. Mismetti P., Laporte S., Zufferey P. Prevention of venous thromboembolism in orthopedic surgery with vitamin K antago-nists: a meta-analysis; J Thomb Haemostasis 2002, .2, 1058-1070.4. Philippe C. Selective Factor Xa Inhibition Improves Ef-ficacy of Venous Thromboembolism Prophylaxis in Orthopedic Surgery;Thrombosis Research 2007, 3, 205-212.5. Duggan T., Scott J., Plosker L. Rivaroxaban: A Review of its Use for the Prevention of Venous Thromboembolism After Total Hip or Knee Replacement Surgery 2009, 69, 13, 1829-1851. 6. Rohde G. Determination of rivaroxaban – a novel, oral, direct Factor Xa inhibitor – in human plasma by high-per-formance liquid chromatography–tandem mass spectrometry

Drug Metabolism and Pharmacokinetics – Bioanalytics, Bayer HealthCare AG, Aprather Weg, 42096 Wuppertal, Germany.7. Erksson BI., Borris L., Friedman RJ.et al. Rivaroxaban versus Enoxaparin for Thromboprophylaxis after Hip Arthro-plasty, N Engj J Med 2008, 358:2765-2775.8. Kakkar AK, et al. RECORD 2 Investigators: Rivaroxaban versus Enoxaparin for Thromboprophylaxis after Hip Arthro-plasty, Lancet 2008; 372: 1-9.9. Lassen MR, Ageno W, Borris LC, et al. RECORD3 Investiga-tors. Rivaroxaban versus enoxaparin for thromboprophylaxis after total knee arthroplasty. N Engl J Med. 2008;358(26):2776-2786.10. Turpie AG, Lassen MR, Davidson BL, et al; RECORD4 Investigators. Rivaroxaban versus enoxaparin for thrombopro-phylaxis after total knee arthroplasty (RECORD4): a randomised trial. Lancet. 2009;373(9676):1673-1680. 11. Trusca T, Lupescu O, Nagea M, Patru C, Popescu Gh I. Early experiments in thrombo-prophylaxis with new oral anticoagulants - Rivaroxaban. Clujul Medical 2010;2:302-307.12. Eriksson BI, et al. J Thromb Haemost 2008;4:121–8.13. Eriksson BI, Kakkar AK, Turpie Ag. G.,et al. Oral riva-roxaban for the prevention of symptomatic venous thrombo-embolism after elective hip and knee replacement. J Bone Joint Surg Br 2009 91-B: 636-644. 14. Van Thiel D., Kalodiki E., Wahi R., Litinas E., Haque W., Rao G. Interpretation of Benefit-Risk of Enoxaparin as Comparator in the RECORD Program: Rivaroxaban Oral Tablets (10 milligrams) for Use in Prophylaxis in Deep Vein Thrombosis and Pulmonary Embolism in Patients Undergoing Hip or Knee Replacement Surgery. Clin Appl Thromb Hemost 2009; 15: 389-394.


Maria Dorina Pasca5 Dacia Traiana Str., Iernut, Jud. Murese-mail: [email protected]

Abstract. Patients with disabilities represent a particular category with whom the pharmacist has to know how to communicate and interrelate. Knowing the patient’s physical and psychological identity, the pharmacist will opt for an efficient communication and relationship, based on the idea that the person’s condition can comprise various disabilities: hearing, vision, intellect, motor skills. Given this context, the approach must be different and adapted to the specific needs of the patient. A succesfull communication approach leads to mutual respect, trust, acceptance and responsibility which, ultimately, engender new behaviors for any given context. Key words: pharmacist, person with disabilities, communication, acceptance, strategies

University of Medicine and Pharmacy Targu Mureş

Paşca Maria Dorina

COMMUNICATION STRATEGIES THAT IMPROVE THE RELATIONSHIP BETWEEN PHARMACIST AND PATIENTS WITH DISABILITIES

THERAPEUTICAL PRACTICETherapeutics, Pharmacology and Clinical ToxicologyVol XVIII, Number 1, March 2014Pages: 27-28 © Copyright reserved 2014

One of communication’s main roles is to send a message which, in turn, may or may not

be properly received. The two agents involved in this activity, the transmitter (T) and the receiver (R) must find an optimal way to receive and answer to the message content.

Once we become aware of the basic communication rules- to listen to the other, to observe, to control and to express ourselves (Paşca M. D. 2012) we will be able to reach our aimed target, bearing in mind what Stanton N (1995) considered as essential for the oral communication: to be perceived by the other (heard or read), to be understood, accepted and to stir a reaction (a change in behaviour or attitude). Hence, the communication between pharmacist and a patient with disabilities will be: efficient, constructive, decisive, appraising based on a ‘biunivocal’ interrelationship.

It is well known that in a drug store a person with disabilities can exhibit a particular behaviour that might trigger a successful dialogue or a failure in dialogue on behalf of the pharmacist. In order to prevent a communication clash between the pharmacist and the patient with disabilities, the former must know a minimal set of rules regarding the interrelationship with the latter. Subsequently, the

intrinsic and extrinsic bonding between cause (C) and effect (E) is established, leading to a predictable result, applicable in certain situations. This “communication formula” stands as a solution for various emotional states- uncertainty, anguish, rejection and ultimately it grounds the idea of patients’ rights and the rights of patients with disabilities.

From this viewpoint: safety, self esteem, respect and last but not least, the certainty that one is viewed as any other person breeds a series of features that the patient with disabilities will cultivate and manifest further on, through a positive attitude towards the pharmacist.

In practical terms, the pharmacist should opt for an attitude that inspires safety to the patient with disabilities. Consequently each patient should benefit from a specific interaction based on his/her condition: 1. Hearing loss: • Face to face interaction so that the patient

perceive the message visually; • Keep the face uncovered, especially the lips

creating a context for lip-reading; • Use a higher tone of voice and a slower pace of

speech should you notice the patient becomes slightly annoyed;

• Show him support by letting him know you understood what he said/transmitted- by nodding;

• If possible, present him the medicine, for safety reasons;


• Use a minimal set of mimics and gestures; • Display a respectful, understanding and

acceptance behaviour;• Smile and greed as you he leaves the pharmacy.

2. Visual impairment and blindness: Keep the tone of your voice calm so that the patient sense the feeling of trust;• If the patient is accompanied by a caregiver, you

should provide explanations for both of them; • Ask him about his preferences regarding the

medication (where this is possible); • If it is suitable and if he asks it, you can have

the patient feel, smell, weight the medicine and its packaging in order to consolidate the idea of credibility;

• Explain him as you write on the packaging how to use the medicine;

• If the patient initiates a discussion you can also engage, within reason;

• Offer to help him walk to the door, should you see he has difficulties moving and thank him for having accessed the services of your pharmacy.

3. Intellectual disability:• Avoid any signs of irritability towards the

patient’s disability;• Smile and encourage him to talk to you;• Keep your calm if he repeats a word, a phrase,

an information regarding a certain drug and help him relax ;

• Avoid any sign of irritability at his smile or permanent grin (without any reason)- consequences of his condition;

• Ignore any twitch, as he might not be aware of it; • Avoid any reply, despite any frustration you

might feel, to certain naming he may utter unaware of his actions;

• Refrain from responding to his way of perceiving the things you transmit, especially explanations regarding the medication and ways of taking it; bear in mind that in persons with intellectual disabilities the chronological age does not correspond to the psychological one.

4. Speech disorders:• Maintain a calm attitude if the person

stutters; • Ask the patient to slowly repeat what he

needs, if the pace of speech is too rapid;• If he addresses you in a whispery tone ask

him to speak a bit louder; • Put yourself in front of him in order to

follow his lips; • Give him the proper attention; • If the person show signs of aphasia help him

express verbally what he remembers about the topic concerned;

• If you notice signs of mutism ask him to show you the packaging of the medication or to hand you over his prescription. 5. Motor skills disorders:

• Put aside any prejudices regarding the patient’s global morphological disorder: growth, nutrition, attitude, skin, muscles, joints or behaviour;

• Avoid making any unjustified comments on a diet for persons with obesity or anorexia problems;

• Listen to his version on the prescribed medication;

• Consider him an equal partner of discussion;• Ask for his opinion, if this is necessary,

regarding certain situations or cases; • Show him proper respect; • Offer your support or the support he might

ask from you. The taxonomy of attitudes described above

is the very essence of the message: “Primum non nocere”- First do no harm!”

References1. Paşca M D. Comunicarea în relaţia medic- pacient, Ed. University Press, Tg. Mureş, 2012.2. Rusu A, Paşca MD, Hancu G. Ghidul farmacistului în comunicarea cu pacientul, Ed. University Press, Tg. Mureş, 2011.3. Stanton N. Comunicarea, Ed. Stiinţă şi Tehnică Bucureşti, 1995


Cristina Negulescu1 Dr. Calistrat Grozovici Str., Bucharest, Romaniaemail: [email protected]

Abstract. Introduction: Influenza infection is extremely common, but despite being usually self-limited, the frequent occurrence of epidemics entails a considerable economic burden. Influenza-associated en-cephalopathy/encephalitis is relatively rare, but carries significant morbidity and mortality. Case report: We report the case of a 6-year-old girl admitted to the intensive care unit of the National Institute of Infectious Diseases ”Prof. Dr. Matei Balş” with acute rhombencephalitis following influenza A H1N1 infection. The child survived but with severe neurological sequelae. Conclusions: Improving primary health care assistance from the point of view of prophylaxis through vaccination programs and improving patient management from the first signs of illness can lead to reducing the number of illnesses, but also to reducing the number of severe or complicated cases of influenza virus infection.Keywords: influenza, child, encephalitis, neurological sequelae, vaccination

1. National Institute of Infectious Diseases Prof. Dr. Matei Bals' Bucharest, Romania2. University of Medicine and Pharmacy 'Carol Davila', Bucharest, Romania

Vișan Angelica1,2, Drăgănescu Anca1, Negulescu Cristina1, Bilașco Anuţa1, Vasile Magdalena1, Măntescu Ruxandra1, Merișescu Mădălina2, Slavu Diana1, Dogaru Cornelia1,

Marinescu Violeta1, Luminos Monica1,2

SEVERE OUTCOME OF INFLUENZA A H1N1 RHOMBENCEPHALITIS IN A 6-YEAR-OLD CHILD CASE REPORT

CASE REPORTTherapeutics, Pharmacology and Clinical ToxicologyVol XVIII, Number 1, March 2014Pages: 29-32 © Copyright reserved 2014

Introduction

Influenza (flu) is a contagious acute respiratory illness caused by influenza viruses A and B,

manifested primarily with signs and symptoms of the upper and lower respiratory tract. Healthy immunocompetent children usually develop mild, self limited disease, but there are high-risk patients like children less than 1 year, patients with chronic illnesses, immunocompromised, or with poor social status that can develop severe forms of disease or life-threatening complications.[1,2]

Each year, about 20% of children worldwide develop symptomatic influenza A or B infections.[3]

The most frequent complications of influenza virus infection are acute otitis media, laryngotracheitis, bronchitis, bronchiolitis and interstitial pneumonia. Less often, influenza virus infection can lead to myositis, bacterial superinfection (most commonly Stafilococcus aureus and Streptococcus pneumoniae), neurologic complications, myocarditis, pericarditis, but also toxic shock syndrome associated to S. aureus infection.[1]

Neurological complications associated to influenza include aseptic meningitis, acute cerebellar ataxia, transverse myelitis, Guillain-Barre syndrome, acute necrotizing encephalitis, acute disseminated encephalomyelitis, encephalopathy, febrile seizures, acute mental status changes. [1].

In the last two decades, in Japan, there was a significant increase in the number of cases of encephalopathy and encephalitis associated to influenza infection, most predominantly seen in children less than 5 years of age.[4]. An important number of these cases were followed by death or severe neurological sequelae. [5]

The first cases of influenza A H1N1 pandemic virus infections associated with neurological complications were reported by CDC (Centers for Disease Control and Prevention) in 2009, 24th of July. The 4 patients were from Texas and between 7 and 17 years of age. Influenza A H1N1 virus was detected from the nasopharyngeal swab, but not from the corticospinal fluid (CSF). [5]

CDC recommends rapid antigen testing for evaluating clinical suspicion of influenza infection, especially in a respiratory illness outbreak. Most of the rapid tests have high specificity (90-95%), but sensitivity varies a lot from 10% to 80% (compared to viral culture or RT-PCR). [6]


http://www.cdc.gov/flu/professionals/diagnosis/clinician_guidance_ridt.htm)

The confirmation of an influenza infection can be done either on viral culture or RT-PCR.

Case report

We report the case of a 6-year-old-girl from a rural area, transferred from a county hospital to our intensive care unit on 12th of March 2014 with the clinical suspicion of acute encephalitis.

The onset of the disease was 4 days before the admission to our clinic with high fever, vertigo and vomiting (4-5 episodes/day). She was administered antipyretics by her family, without consulting their family physician. The day before admission to hospital she developed extreme agitation, visual hallucinations and then seizures.

Her personal and family medical histories were not significant, she was vaccinated according to our national schedule, but she was not vaccinated against influenza virus. In her family’s recent history all members presented respiratory symptoms without consulting their family physician.

On admission the child was comatose, with a GCS of 5-6/15, afebrile, pale, without cutaneous eruptions, with signs of dehydration, hypotrophy (weight=17 kg), with cold extremities, with a heart rate of 120 beats per minute, a blood pressure of 120/60 mmHg, a respiratory rate of 40 per minute. Oxygen saturation measured by pulse oximetry was 90% while breathing ambient air, with bilateral pulmonary rales. Her abdomen showed no signs of peritoneal irritation; she had acute urinary retention and diarrhea. The child was in a decorticate posture, with flexion of the upper limbs and hyperextension of the lower limbs.

After assessing her extremely severe condition by an ICU physician, the child was intubated and mechanically ventilated under sedation with midazolam.

A neurologic examination was obtained after intubation and sedation which showed: pupils on the midline, of 2 mm diameter with poor reactivity, deep tendon reflexes were absent, slight movement of right shoulder to painful stimulus.

Laboratory blood tests showed inflammatory syndrome (slightly elevated white blood cell count, a procalcitonin of 20.4 ng/ml), hemoconcentration (due to dehydration), Hemoglobin=16 mg/dl, Ht=48%, blood urea = 91,5 mg/dl, moderate elevated transaminase level (AST/TGP=80 U/l; ALT/TGO=130 U/l), hypopoteinemia (5,2g/dl) with hypoalbuminemia (2,93g/dl,), LDH = 1226 U/L; CK=577 U/l. Imunogramme: IgG=2,12g/l, (normal value for age: IgG=6-15g/l); IgA=8,69, (normal value for age: IgA=0,35-2,4g/l); IgM=17,7g/l (normal value

for age IgM=0,4-2,5g/l) Rapid antigen testing for respiratory viruses

using MariPOC test was positive for Influenza A virus from a nasopharyngeal sample. A confirmation test using RT-PCR from a nasopharyngeal sample was possible which tested positive for influenza A H1N1- 2009 virus.

A chest x- ray revealed bilateral bronchopneumonia. Emergency cerebral MRI was performed and

showed:• T2 and FLAIR signal hyperintensity patches,

with slightly T1 signal hypointensity, with infracentimetric restriction areas and progressive, intense, heterogeneous contrast enhancement areas localized in medulla, pons, midbrain, thalamus, posterior lenticular nuclei, supranuclear white matter; these lesions don’t have a homogenous T2 signal, containing almost fluid T2 signal in the anterior part of the medulla, retro pyramidal, posterior internal capsule.

• T2 and FLAIR signal hyperintensity spots and semi lunar area, without restriction of diffusion, without gadolinium contrast, disseminated in the white hemispheric matter, predominantly sub cortical, especially at the genu of corpus callosum.

• normal symmetric ventricular system• midline structures in normal position• angiography of the intracranial veins and dural

sinuses showed no abnormalities.

Brain MRI and clinical presentation made a clear diagnosis of Acute Rhombencephalitis with upper extension to the sub cortical areas.

Oseltamivir (10 days through nasogastic tube), IV Acyclovir (until laboratory confirmation of etiology), IV solu-Medrol pulse-therapy (3 days) , IV mannitol and IV human immunoglobulins therapy was initiated. Broad spectrum antibiotics were also administered due to bronchopneumonia (meropenem and linezolid), and solu-medrol

Figure 1. MRI cerebral showed contrast inhancement areas localized in medulla, pons, midbrain, thalamus, posterior

lenticular nuclei, supranuclear white matter


therapy was continued with IV dexametazone in progressively descendent doses, under gastric protection with IV H2 receptor inhibitor and proton pump inhibitor.

Due to clinical and biological evolution with severe thrombocytopenia (21.000/mm3), anemia (Hb 7,4g/dl) and hemorrhagiparous syndrome (blood through the nasogastric tube and macroscopic hematuria) administration of blood products was initiated (platelet concentrates, red blood cells, fresh frozen plasma)

The EEG showed beta rhythm, with symmetric hypo voltage. Hemoculture and nasal/pharyngeal cultures came back negative.

Serology testing was negative for HIV, Herpes simplex 1+2, Adenovirus, Coxsackie, Echovirus, Parainfluenzae viruses, Ebstein-Barr virus, Citomegalovirus. Serology for Mycoplasma p., Chlamydophila p., Borrelia burgdorferi, Yersinia spp were also performed and were negative.

After 14 days of mechanical ventilation (10 days in CPAP system) the patient was in a severe state, with hypotrophy, muscular atrophy, she had spontaneous respiration, cough reflex, pulmonary rales and rhonchi with periods of oxygen desaturation which necessitated oxygen therapy, a heart rate of 100 beats per minute, blood pressure of 100/70 mmHg, gastric tolerance through nasogastric tube, normal stool, urinary incontinence.

Neurologic examination revealed spontaneous opening of the eyes, lack of visual following or fixation, no execution of verbal commands, cranial nerve III paresis, pupils with anisocoria (left pupil>right pupil), mydriasis, diminished pupillary light reflex. Fundoscopic exam showed thin vessels with normal emergence. Spasticity was present with severe bilateral muscle contractures of pyramidal-extrapyramidal type, predominantly on the left side, brisk deep tendon reflexes, no plantar clonus, bilateral Babinski sign. The patient had suction automatism and no swallowing reflex. Nociceptive stimulation led to extension of the lower limbs.

The second brain MRI obtained after 20 days of hospitalization showed: • extension of the T2 and FLAIR hiperintensity

signal areas into the white matter of the hemispheres (semi-oval areas), in the parietal regions

• T2 and FLAIR hyperintensity signal bands located periventricular (lateral ventricles)

• moderate dilatation of ventricles V1, V2, V3,V4 • moderate enlargement of cortical sulci, especially

of the sylvian fissureThe overall conclusion of the second MRI

as compared to the first examination was the extension of the lesions into the white matter of the hemispheres, most probably due to a demyelinating

process, and the development of diffuse cortical atrophy.

The patient was transferred to a medical rehabilitation clinic for children with neuromotor disabilities, and after that to a centre for children with severe motor handicap because her family could not provide adequate care and support.

Discussion

In our case report, the possible risk factors that we could identify for such a severe neurological outcome were malnutrition, poor social status and lack of primary care assistance.

Although the child’s illness started 4-5 days before admission to hospital, her family approached medical assistance only after the child have had repeated seizures and became comatose.

Rapid antigen testing for respiratory viruses from a nasopharyngeal swab and RT-PCR confirmation from the same sample made an early (24 hours) and accurate diagnosis of infection with the pandemic strain of influenza A H1N1-2009 virus

Brain MRI evaluation was performed on admission and showed extensive, severe and early involvement of the neurologic system (medulla, pons, midbrain, thalamus, lenticular nuclei, supranuclear white matter) with influenza A H1N1 infection.

The treatment administered saved the child’s life with the cost of severe neurological sequelae. Neuromotor disability required institutionalization of the patient in a specialized centre.

Having in mind the high rate of morbidity and mortality of influenza infection in the poor social status population, it is necessary to develop a medical strategy to vaccinate this high risk population against influenza infection.

Figure 2. Extension of T2 and FLAIR hiperintensity signal areas into the white matter of the hemispheres in the parietal

regions. Moderate dilatation of ventricles V1, V2, V3, V4


Conclusions

Influenza infection is extremely common, but despite being usually benign and self-limited, the frequent occurrence of epidemics entails a considerable economic burden.[7]

The term „influenza-associated encephalopathy/encephalitis” (IAEE) is used because direct evidence of influenza virus in CSF is rarely demonstrated. In our case, the only noticeably finding was a positive PCR for influenza A from a nasopharyngeal swab;[8]

Acute encephalitis is one of the most severe influenza complications, responsible for high rates of mortality and morbidity with potentially severe neurological sequelae.[2]

Pr imar y medical ass istance can help diminish the economical burden of influenza infections through highlighting the importance of prevention methods like annual vaccination and by providing adequate medical care from the early stages of disease.

References1. Munoz FM, Mallory GB, Eduards MS. Clinical features and diagnosis of seasonal influenza in children, UpToDate, 2013.2. Pieter LA, Fraaij Heikkinen T. Seasonal influenza: The burden of diseases in children, Vaccine, August 2011.3. Pandemic influenza:a priority for the neurological community, The Lancet Neurology, 2009-10-01, vol 8, Issue 10, pages 869-869.4. Thorner AR, Hirsch MS, Baron EL. Clinical manifestations and diagnosis of pandemic H1N1 influenza (“swine influenza”), UpToDate, 2014.5. Henry J, Smeyne RJ, Jang H. Parkinsonism and neurological manifestation of influenza throughout the 20th and 21st centuries, Rev. Parkinsonism and Related Disorders, June 2010.6. Nicholson KG, Wood JM. Influenza, The Lancet, Nov. 2003, vol.362, Issue 9397,pag 1733-45.7. Fluss J, Ferey S. Mild influenza-associated encephalopathy/ encephalitis with a reversible splenial lesion in a Caucasian child with additional cerebellar features, Official Journal of the European Paediatric Neurology Society, 2010.8. Hussain MI, Ming CT, Yin LL. Neurologic manifestations and complications of pandemic influenza AH1N1 in Malaysian children: What have we learnt from the ordeal?; Brain and Development; March 2014.


Savulescu Alexandru Florin88 Mircea Vulcanescu Str., 010825 Bucharest, Romaniae-mail: [email protected]

Abstract. Introduction. Adrenal tumours are often discovered incidentally, while a patient is undergoing an imaging examination (abdominal ultrasound, computed tomography, magnetic resonance imaging), for a problem unrelated to the adrenal gland, usually having no signs or symptoms suggesting adrenal disorders. Case report. A female patient was presented for moderate abdominal pain which started 2 month prior to her admission, without having any other signs or symptoms. The usual blood tests came up normal. The abdominal ultrasound revealed a relatively large abdominal cystic mass, possibly belonging to the liver. The CT scan could not certify the origin of the tumour, but located it between the right hepatic lobe and the right kidney. Because of the patient’s growing pain, diagnostic laparoscopic surgery was performed. The surgery was completed with the excision of the abdominal mass. Results. The patient’s postoperative progress was positive, with complete pain remission. The histological findings revealed that the abdomi-nal mass was in fact a benign adrenal endothelial cyst. Conclusions. Unusual growing abdominal pain may benefit from diagnostic laparoscopy, especially when imaging procedures fail to offer accurate data. Minimal invasive surgery is the modern gold-standard for adrenal lesion excisions. Key words: adrenal tumour, abdominal pain, laparoscopy

1. Surgery Department, Central Clinical Emergency Military Hospital, Bucharest2. Dermatology Department, Central Clinical Emergency Military Hospital, Bucharest

Săvulescu F.1, Nica R.1, Surdeanu D.1, Cirlan C.1, Darmanescu M.2, Trifu V.2

CYSTIC ADRENAL TUMOUR - A CASE OF ATYPICAL ABDOMINAL PAIN

CASE REPORTTherapeutics, Pharmacology and Clinical ToxicologyVol XVIII, Number 1, March 2014Pages: 33-36 © Copyright reserved 2014

Introduction

Adrenal tumour formations are frequently discovered incidentally, being diagnosed during

a routine imaging examination (abdominal ultrasound, computed tomography, magnetic resonance imaging), or during an investigation for other medical conditions. These tumours are not suspected prior to imaging procedure, the patient usually having no signs or specific symptoms suggesting an adrenal pathology[1].

The widespread use of abdominal ultrasono-graphy and computed tomography led to an increase in intra-abdominal tumours discovered incidentally. Among these, adrenal formations are a relatively frequent discovery among the patients that undergo imagistic investigations. Both computed tomography and magnetic resonance imaging are very good methods for the visualisation and the characterization of adrenal tumors[2]. Approximately 1-2% of the patients going through an imaging investigation present adrenal lesions.

Asymptomatic patients, with cystic formations smaller than 3 cm do not require further investigation. In

contrast, for those with diameters larger than 6 cm surgery is required due to the risk of malignancy2, related to the increase in tumour size.

Minimally invasive surgery is the current gold standard in the adrenal gland surgery. The emergence of new technologies (Harmonic, Ligasure), which have become indispensable, helped so that the tumour size doesn’t stand as a contraindication to the laparoscopic approach, anymore. A new development was the advent of robotic surgery, the „Da Vinci“ surgical system which provides an exceptional 3D visualization and increases surgical accuracy.

Case report

A 19-year-old patient is hospitalized accusing mild pain located in the right lumbar region, flank and right upper quadrant, quasi-permanent nature. Symptoms started approximately two months prior to admission, initially as a painful discomfort, then increasing in intensity.

At an objective clinical examination, the devices and the systems did not reveal pathological changes. Paraclinical investigations commonly performed (blood count, serum biochemistry, ionograme, coagulation) were also normal.

The abdominal ultrasonography revealed a cystic tumour about 8 cm in diameter, possibly liver related,


raising the suspicion of hydatid cyst. Subsequently, blood samples were collected in order to assess the dosage of the anti-Echinococus antibodies (ELISA test), with negative results. The blood count revealed a normal leukocyte formula, without eosinophilia. From these results, the diagnosis of hydatid cyst became less plausible.

CT scan with contrast showed a cystic formation of 75/56/75 mm, with thin walls, located on the right side in the intrahepatic and renal areas, with homogeneous content, fluid densities, with mass effect on the liver, kidney, duodenum and inferior vena cava, but could not show precisely the organ affiliation. (Figure 1)

Due to increasing pain, laparoscopic surgery was needed (diagnostic laparoscopy to establish the organ belonging to the cystic formation). Intraoperatively, a cystic tumour 8-9 cm in diameter was revealed, located in

the intrahepatic and reanal areas but without belonging to the liver, therefore rising concerns of a benign cystic lesion of the right adrenal gland. It was decided for the surgery to continue with the ablation of the cystic tumour. (Figure 2).

The postoperative evolution was rapidly favourable, with full remission of pain related symptoms.

Postoperative histopathological examination established the diagnostic of adrenal endothelial cyst.

DiscussionsThe prevalence of adrenal tumours is not known

exactly, but they represent a rare pathology compared with other abdominal tumours. Most are benign tumour formations, discovered incidentally in asymptomatic stages. Biological laboratory samples are recommended only in the presence of one of the following pathology suggesting symptoms2: Cushing’s syndrome (24-hour urinary cortisol determination, dexamethasone suppression test), pheochromocytoma (24-hour dosing of the metanephrines, catecholamines and vanillic-mandelic acid within the urine for 24 hours), hyperaldosteronism (potassium levels determination). Laboratory tests for the screening of adrenal carcinomas

(the dosage of 17-ketosteroids within the urine for 24 hours) are not recommended due to lack of sensitivity and specificity. In addition, adrenal cancers are quite rare[2].

From the clinical point of view, the common feature of the three above-mentioned pathologies is the presence of high arterial blood pressure. In addition,

Figure 1. CT appearance of tumour formation

Figure 2. Intraoperative appearance of the cystic formation


Cushing’s syndrome is characterized by full moon facies, proximal muscle weakness, truncal obesity, vascular fragility with petechiae and ecchymosis, skin thinning. Pheochromocytoma is accompanied by paroxysmal hypertensive crisis episodes associated with headache, sweating and palpitations[4].

In our patient’s case, she presented only abdominal pain (initially in the form of discomfort), with no association to any of the symptoms or clinical signs listed above, indicating a hormonal change. Pain syndrome is explained by the compression of the cystic tumour on the surrounding structures, effect caused by the increased size of the lesion.

Due to the increasing pain, and to the impossibility to establish the organ affiliation of the abdominal tumour formation through body imaging, surgery was required initially for diagnosis. Once we established

the high probability of adrenal gland affiliation intraoperatively, also relying on the increased size of the tumour, we proceeded to ablation. According to literature, adrenal tumour formations over 6 cm in diameter have surgical indication, because of the increased possibility of malignancy. In contrast, tumour formations smaller than 3 cm showing no evidence of hormonal pathologies (without characteristic symptomatology , normal laboratory tests ) can be monitored by the family doctor[2]. Recommended tracking is done by computed CT scan or abdominal ultrasonography 3 months after diagnosis, then every 6 months for the next 2 years[5]. Patients presenting abnormal laboratory tests at the screening for endocrine pathologies associated with the presence of adrenal formations should be directed to the surgeon because all hormone -producing tumours should be surgically removed[2].

Regarding surgery for adrenal tumour masses, there is a controversy about the maximum size of tumours receiving laparoscopic resection. Some authors do not recommend the minimally invasive surgery for tumours over 6 cm in diameter[6]. Laparoscopic surgical

approach has two more main contraindications: tumour malignancy and invasion of vicinity[7]. None of these contraindications was present for the patient in question; therefore the intervention was done completely laparoscopically, conversion to laparotomy not being required. We believe that tumour size should not be a determining factor in the decision to perform laparoscopic surgery, especially if the imaging examinations display well defined formations.

In terms of aetiology and anatomical pathology, benign adrenal cystic formations can be divided in: hydatid cysts, essential cysts (epithelial, follicular, endothelial, lymphangiomal) and pseudocysts (TB, hemorrhagic cysts). In this case, the paraffin histopathology examination established the diagnosis of adrenal endothelial cyst. (Figure 3)

Conclusions

The prevalence of adrenal tumours increased proportionally with the development of imaging investigations, and their widespread use (ultra-sound, computed tomography, magnetic resonance imaging).

The risk of malignancy of an adrenal tumour formation discovered through imaging tests is consistent with the tumour size and, therefore, surgery for tumours with diameters larger than 6 cm becomes a priority.

Out of all the adrenal tumour formations, the cystic tumour accounts for the lowest percentage, being also mostly benign.

Laparoscopic surgery is useful for diagnostic purposes in patients whose imaging investigations cannot provide accurate data on the present pathology, being able to also provide therapeutic solutions during the same intervention.

Adrenal tumours may benefit from a minimally invasive surgical approach, laparoscopic or robotic - which is the current gold standard in adrenal surgery.

Figure 3. Microscopic appearance of the cyst


References1. Griffing G. Medscape – Adrenal incidentaloma. [Inter-active] Available at: http://emedicine.medscape.com/arti-cle/116587 [Accessed 9 march 2014]. 2013.2. Higgins JF J. Evaluation of incidental renal and adrenal mas-ses. Am Fam Physician, 2001. Volume 63(2), pages 288-295.3. Urology. Urology Care Foundation – Adrenal gland di-sorders. [Interactive] Available at: http://www.urologyhealth.org/urology[Accessed 9 march 2014]. 2011.4. Emedic.ro. emedic.ro – Ghiduri de practica medicala. [In-

teractive] Available at: http://www.emedic.ro/Ghiduri-de-prac-tica-medicala[Accessed 9 march 2014]. 2013.5. Herrera MF G. C. v. H. J. S. P. I. D. Incidentally disco-vered adrenal tumors: an institutional perspective. Surgery, Volume 110, pages 1014–21.1991. 6. MacGillivray DC, W. G. M. C. O. D. S. S. Laparoscopic resection of large adrenal tumors. Ann Surg Oncol, Volume 9(5), pages 480-5. 2002.7. Reincke M, S. M. End Text - Adrenal incidentalomas. [Interactive] Available at: http://www.endotext.org/chapter/adrenal-incidentalomas [Accessed 9 march 2014]. 2010.


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