The development of malignancies in renal allograft recipients with special emphasis on Kaposi's sarcoma

Moosa, M. R. (2002-03)

Thesis (PhD)--Stellenbosch University, 2002.


ENGLISH ABSTRACT: Renal transplantation is undoubtedly the best treatment for patients with irreversible renal failure. As a prelude to establishing the nature of malignancies in renal transplant patients we sought to determine factors influencing the outcome of renal transplantation. The survival of renal allografts and of recipients is influenced by a number of demographic, clinical and therapeutic factors. Some of these factors have been better studied than others, and we sought to establish the influence of particular factors on our own patients and allografts. The total number and nature of malignancies developing in these patients subsequent to transplantation was also established. All patients transplanted in our unit between April 1, 1976 and March 31, 1999 were included in the study. In the study period, 542 patients received 623 renal allografts. Demographic details were analysed. Patient and graft outcomes were assessed using Kaplan-Meier survival analysis. The survival curves were compared using univariate analysis; results that were significant were subjected to multivariate analysis. The influence of a number of factors on graft and patient survival were assessed and compared. The impact of a variety of variables on the number and behaviour of malignancies was also established. Patient and graft survival were superior in recipients who were aged less than 40 years; cyclosporine improved graft survival but not patient survival. Early graft loss was associated with a high patient mortality rate. Contrary to the experience elsewhere, black and white patients had similar outcomes after renal transplantation. Of the 542 recipients 41(8.1%) developed malignancies with Kaposi's sarcoma occurring, in 21 patients and skin cancers in 13 patients. The relative risk for the Kaposi's sarcoma development was 235. Kaposi's sarcoma was the most common tumour in non-white patients (accounting for 79% of malignancies in this group) and occurred less than 2 years after transplantation. Kaposi's sarcoma was equally common in male and female recipients. Under cyclosporine the latent period to malignancies was reduced but the frequency remained unaffected. Kaposi's sarcoma skin lesions were present in all the affected patients, with the lower limbs the most common site of involvement. Kaposi's sarcoma responded to reduction of immunosuppression without the need for complete discontinuation, and with preservation of renal function. Extracutaneous involvement occurred in over one quarter of the patients and invariably proved fatal in all patients with visceral organ involvement. The histopathology of posttransplant Kaposi's sarcoma was the same as that described in the other epidemiological forms of the disease. White male recipients were at the greatest risk of developing skin cancers after renal transplantation. Squamous cell carcinomas were relatively more common and were found in sun-exposed areas. The lesions were treated only by local excision and none metastasized. Malignant lymphoma, breast cancer and lung cancer occurred in individual patients but the relative risk of all these lesions were close to unity. Patients with preexisting cancers did not develop recurrences following transplantation. SECTION 2 Both immunosuppression and immunostimulation are thought to play a role in the development of Kaposi's sarcoma after renal transplantation. We investigated the quantitative and qualitative aspects of the immune system of patients who had developed Kaposi's sarcoma. The lymphocyte phenotypes were established using flow cytometry while transformation studies were performed using mitogens. Pokeweed was used as the B-cell mitogen, and concanavalin A and phytahaemagglutinin were the T-cell mitogens. Cell mediated immunity was also tested using delayed type hypersensitivity skin tests and the serum immunoglobulin levels were estimated. Firstly, with regard to humoral immunity, 2/3 of the patients had normal serum immunoglobulin levels, although the B-cell count was reduced in all the patients on immunosuppression. B-cell transformation tests with pokeweed mitogen revealed that B-cell function was not impaired in patients with Kaposi's sarcoma. The patients with decreased immunoglobulin levels also appeared to be malnourished as evidenced by low albumin levels. Secondly, CD3 and CD4, but not CD8, cell counts were reduced in patients with Kaposi's sarcoma. The transformation analyses revealed significant differences compared to controls, with reduced responses in patients with Kaposi's sarcoma. Thirdly, natural killer (NK) cell numbers were also reduced in patients with Kaposi's sarcoma. There were no significant differences in delayed type hypersensitivity skin reactions that could not be accounted for by racial differences. Cellular immunity is impaired in patients with Kaposi's sarcoma with a reduction in the number of NK cells. Both of these components of the immune system are important in protection against malignant transformation. SECTION 3 Kaposi's sarcoma is an important complication of renal transplantation. If the human herpesvirus 8 (HHV-8) causes Kaposi's sarcoma, the virus should be present in all Kaposi's sarcoma lesions and be drastically reduced or cleared from involved tissue on remission of the Kaposi's sarcoma. Fourteen renal transplant patients with cutaneous Kaposi's sarcoma, including autopsy material from two cases, were investigated for the presence of HHV-8. A second skin biopsy was taken from 11 survivors, after remission of Kaposi's sarcoma, from normal skin in the same anatomical region as the first biopsy. Remission was induced by reduction or cessation of immunosuppression. A peripheral blood sample was collected simultaneously with the repeat biopsy. A nested polymerase chain reaction assay was used to detect HHV-8 DNA in the biopsy tissue and peripheral blood mononuclear cells followed by direct sequencing of polymerase chain reaction product to detect any nucleotide changes. HHV-8 DNA was detected in all the cutaneous Kaposi's sarcoma and all the visceral Kaposi's sarcoma samples, as well as a number of Kaposi's sarcoma-free organs including the thyroid, salivary gland, and myocardium that have not been described before. Mutations in the viral DNA could be demonstrated in all patients. The mutations found were related more to that seen in AIDS-Kaposi's sarcoma cases than that found in African endemic Kaposi's sarcoma cases. HHV-8 sequences could be detected in follow-up frozen skin biopsies of five patients but were negative in the equivalent formalin-fixed specimens. Viral DNA was also detected in 2 of 11 peripheral blood mononuclear cell samples collected at the time of the follow-up skin biopsies. Reduction or withdrawal of immunosuppression allows the immune system to recover sufficiently to reduce viral replication with subsequent viral persistence and low-grade viral replication that coincides with clinical remission of the Kaposi's sarcoma lesions. This provides further evidence for the important etiological role played by HHV-8 in the pathogenesis of posttransplant Kaposi's sarcoma. SECTION 4 The recently discovered HHV-8 is an important factor in the aetiopathogenesis of Kaposi’s sarcoma. The reason for the exceptionally high prevalence of Kaposi's sarcoma in our area, as well as that of other developing countries, remains unexplained. We investigated the seroprevalence of the virus in the different healthy subjects as well as organ donor-recipient pairs. All recipients were tested at the time of transplantation, as were the paired donors. Control subjects tested were healthy blood donors, Renal Unit staff, and household contacts of patients with Kaposi's sarcoma. An enzyme-linked immunoassay (ELISA) to the whole virus was used for screening and all positives were confirmed using ELISA to the latent ORF 73 antigen. The prevalence of HHV-8 was similar in all groups and averaged less than 6%. After transplantation the seroprevalence increased to almost 20% but neither the transplanted kidney nor blood transfused perioperatively could account for the increase. Kaposi's sarcoma developed in 3 of the 116 patients transplanted. All patients with Kaposi's sarcoma were proven to be HHV-8 seropositive before the development of the disease. Two of the patients who developed Kaposi's sarcoma were seropositive before transplantation. No patient who received a graft from a seropositive donor developed Kaposi's sarcoma. We refute the notion that a high prevalence of HHV-8 in the general population is responsible for the high prevalence of Kaposi's sarcoma in our population or that the donor organ is a major source of infection in renal transplant recipients. Reactivation, rather than primary infection appears to be the source of the virus after renal transplantation.

AFRIKAANSE OPSOMMING: Nieroorplanting is ongetwyfeld die beste behandeling vir pasiente met onomkeerbare nierversaking. As ‘n aanloop om die aard van maligniteite in nierooorplantingspas'fente vas te stel het ons gepoog om die faktore wat die uitkoms van nieroorplantings bemvloed te bepaal. Die oorlewing van oorgeplante niere en van nierontvangers word deur ‘n aantal demografiese, kliniese en terapeutiese faktore bemvloed. Sommige van hierdie faktore is beter ondersoek as ander and ons het gepoog om die invloed van sekere faktore op ons eie pasiente en oorgeplante niere te bepaal. Die getal en aard van maligniteite wat ontwikkel het in hierdie pasiente na nieroorplanting is ook gedokumenteer. Alle pasiente in ons eenheid in wie ‘n nier tussen 1 April 1976 en 31 Maart 1999 oorgeplant was, is in die studie ingesluit. Tydens die studieperiode het 542 pasiente 623 niere ontvang. Demografiese detail is ontleed. Pasient- en nieroorplantings uitkomste is beraam deur gebruik te maak van Kaplan-Meier oorlewing analiese. Die oorlewingskurwes is vergelyk deur gebruik te maak van enkelveranderlike ontledings; noemenswardige resultate is onderwerp aan meerveranderlike ontledings. Die invloed van ‘n aantal faktore op oorgeplante nier- en pasientoorlewing is ondersoek en vergelyk. Die impak van ‘n verskeidenheid veranderlikes op die getal en gedrag van maligniteite is ook ondersoek. Pasient oorlewing asook oorlewing van oorgeplante niere was beter in ontvangers onder die ouderdom van veertig jaar. Vroee verlies van ‘n oorgeplante nier het verband gehou met ‘n hoe pasientmortaliteit. Siklosporien het die oorlewing van oorgeplante niere verbeter, maar nie die van pasiente nie. In teenstelling met die ervaring elders, het swart en wit pasiente soortgelyke uitkomste uitkoms gehad na ‘n nieroorplanting. Van die 542 ontvangers, het 41 (8.1%) maligniteite ontwikkel; Kaposi se sarkoom het in 21 pasiente voorgekom en velkanker in 13 pasiente. Die relatiewe risiko (“relative risk") vir die ontwikkeling van Kaposi se sarkoom was 235. Kaposi se sarkoom was die algemeenste tumor in swart en gekleurde pasiente (verantwoordelik vir 79% van maligniteite in die groep) en het binne twee jaar voorgekom. Kaposi se sarkoom was ewe algemeen in manlike en vroulike ontvangers. Met behandeling deur middel van siklosporien het die latente periode totdat maligniteite ontwikkel het verkort, maar die insidensie daarvan het onveranderd gebly. Velletsels geassosieer met Kaposi se sarkoom was teenwoordig in alle pasiente met die vel van die onderste ledemate die mees algemeen betrokke ligging. Die sarkoom het gereageer op vermindering van immuunonderdrukking, sonder die nodigheid vir volkome onttrekking, en met die bewaring van nierfunksie. Ekstrakutane betrokkenheid het in meer as ‘n kwart van die pasiente voorgekom en was altyd noodlottig in pasiente met viserale aantasting. Die histopatologie van postoorplanting Kaposi se sarkoom was dieselfde as die wat beskryf is vir die ander epidemiologiese vorms van die siekte. Wit mans het die hoogste risiko vir die ontwikkeling van velkankers na nieroorplanting gehad. Plaveiselsel karsinoom was betreklik meer algemeen en het in son-blootgestelde areas voorgekom. Die letsels was uitsluitlik met lokale eksisie behandel en geen pasiente het metastases ontwikkel nie. Maligne limfoom, borskanker, en longkanker het in enkele pasiente voorgekom maar die relatiewe risiko van al die letsels was om en by een gewees. Nie een van die pasiente met vorige maligniteite het herhaling van die tumore na oorplanting ontwikkel nie. AFDELING 2 Die vermoede is dat beide immuunonderdrukking en immuunstimulasie ‘n rol speel in die ontwikkeling van Kaposi se sarkoom na ‘n nieroorplanting. Ons het die kwantitiewe en kwalitatiewe aspekte van die immuunsisteem van pasiente wat Kaposi se sarkoom ontwikkel het na ‘n nieroorplanting, ondersoek . Limfosiet fenotipes is met behulp van vloeisitometrie bepaal, terwyl transformasiestudies uitgevoer is deur gebruik te maak van mitogene. “Pokeweed” is gebruik as die B-sel mitogeen, en konkanavalien A en fitaheemagglutinien was die T-sel mitogene. Sel-gemedieerde immuniteit was ook getoets deur die gebruik van vertraagde tipe hipersensitiwiteit veltoetse. Die serum immunoglobulien vlakke was ook bepaal. Eerstens, met betrekking tot humorale immuniteit, het 2/3 van die pasiente normale serum immunoglobulienvlakke gehad, alhoewel die B-seltelling verminder was in al die pasiente op immuunonderdrukking. B-seltransformasie-toetse met “pokeweed” mitogeen het getoon dat B-sel funksie nie ingekort was in pasiente met Kaposi se sarkoom nie. Die pasiente met verminderde serum immunoglobulinvlakke het ook wangevoed voorgekom soos die verlaagde serum albumienvlakke uitgewys het. Tweedens was CD3 en CD4 seltellings, maar nie CD8 nie, verlaag in pasiente met Kaposi se sarkoom. Betekenisvolle verskille is ook aangetoon met T-sel transformasietoetse in vergelyking met kontroles, met verminderde response in Kaposi se sarkoom pasiente. Derdens was natuurlike dodersel (NK) getalle ook minder in pasiente met Kaposi se sarkoom. Daar was geen noemenswaardige verskille in vetraagde tipe hipersensitiwiteit velreaksies wat nie deur rasseverskille kon verklaar word nie. Sellulere immuniteit is ingekort in pasiente met Kaposi se sarkoom met ‘n verlaging in die aantal NK selle. Beide die komponente van die immuunstelsel is belangrik vir beskerming teen maligne transformasie. AFDELING 3 Kaposi se sarkoom is ‘n belangrike komplikasie van nieroorplanting. As die menslike herpesvirus-8 (HHV-8) Kaposi se sarkoom veroorsaak, behoort die virus teenwoordig te wees in alle letsels en as Kaposi se sarkoom remissie ondergaan behoort dit drasties te verminder of te verdwyn in weefsel waarin dit voorkom. Veertien nieroorplantingspasiente met Kaposi se sarkoom van die vel, insluitend outopsiemateriaal van twee gevalle, is ondersoek vir die teenwoordigheid van HHV- 8. ‘n Tweede velbiopsie van dieselfde anatomiese area as die eerste is uitgevoer op 11 oorlewende pasiente na remissie van die sarkoom. Remissie was deur die vermindering of onttrekking van immuunonderdrukking bewerkstellig. ‘n Perifere bloedmonster is by dieselfde geleentheid as die tweede biopsie geneem. ‘n Geneste polimerase kettingreaksietoets (“nested polymerase chain reaction”) is gebruik om die teenwoordigheid van HHV-8 DNA in die biopsieweefsel en perifere bloed mononukluere selle te bepaal, gevolg deur direkte volgordebepaling (“sequencing”) van die polimerase kettingreaksieproduk om enige nukleotiedveranderings te dokumenteer. HHV-8 DNA is waargeneem in al die kutane Kaposi se sarkoom en al die viserale Kaposi se sarkoom monsters, sowel as in weefsel waar die Kaposi se sarkoom nie voorgekom het nie en waar die teenwoordigheid van die virus nie tevore beskryf is nie, soos die skilklier, speekselklier, en hartspier. Mutasies in die virale DNA kon in alle pasiente aangetoon word. Die mutasies wat gevind is, was nader verwant aan die wat in VIGS-Kaposi se sarkoom beskryf is as die wat in endemiese Kaposi se sarkoom in Afrika gevind word. HHV-8 volgordes kon waargeneem word in bevrore opvolg-velbiopsies van vyf pasiente, maar was afwesig in die ekwivalente formaliengefikseerde monsters. Virus DNA is ook waargeneem in 2 van 11 perifere bloed mononukluere selmonsters wat versamel is tydens die opvolg velbiopsies. Vermindering of onttrekking van immuunonderdrukking laat die immuunsisteem toe om genoegsaam te herstel om virale replikasie te verminder met daaropvolgende teenwoordigheid en laegraadse virale replikasie wat ooreenstem met kliniese remissie van letsels van Kaposi se sarkoom. Dit verskaf verdere bewyse van die belangrike oorsaaklike rol wat deur HHV-8 gespeel word in die patogenese van postoorplanting Kaposi se sarkoom. AFDELING 4 Die onlangs-ontdekte HHV-8 is ‘n belangrike faktor in die etiopatogenese van Kaposi se sarkoom. Die rede vir die buitengewone hoe prevalensie in ons gebied, sowel as die van ander ontwikkelende lande, is nie voor die hand liggend nie. Ons het die seroprevalensie van die virus in verskillende gesonde persone, sowel as orgaan donor-ontvanger pare ondersoek. Alle nierontvangers en hul gepaarde donors is getoets ten tye van die nieroorplanting. Getoetste kontrole persone was gesonde bloedskenkers, niereenheid personeel en huishoudlike kontakte van pasiente met vorige Kaposi se sarkoom. ‘n Ensiemgekoppelde immuuntoets (enzyme-linked immnoassay; ELISA) vir die volledige virus was gebruik vir sifting en bevestiging is verkry vir alle positiewe toetse deur gebruik te maak van van ‘n ELISA vir die latente ORF 73 antigeen. Die prevalensie van HHV-8 was vergelykbaar in alle groepe en was gemiddeld minder as 6%. Na oorplanting het die prevalensie gestyg tot byna 20%, maar nog die oorgeplante nier nog perioperatiewe bloedoortappings kom die styging verklaar. Kaposi se sarkoom het in 3 van 116 van die pasiente wat ‘n oorplanting ondergaan het, ontwikkel. Al die pasiente met Kaposi se sarkoom was HHV-8 seropositief voor die ontwikkeling van die Kaposi se sarkoom. Twee van die pasiente wat Kaposi se sarkoom ontwikkel het was seropositief voor die oorplanting. Geen pasient wat ‘n nier van ‘n seropositiewe donor ontvang het, het Kaposi se sarkoom ontwikkel nie. Ons weerle die stelling dat ‘n hoe prevalensie van HHV-8 in die algemene bevolking verantwoordelik is vir die hoe prevalensie van Kaposi se sarkoom onder ons nieroorplantingspasiente of dat die donornier ’n belangrike bron van infeksie is. Dit wil voorkom of heraktivering, eerder as primere infeksie, ‘n bron van die virus is na nieroorplanting.

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