Streptococcus pyogenes in tumor treatment: the past, present and future

2019/2020 Joana Oliveira e Sousa Ferraz Brandão Streptococcus pyogenes in tumor treatment: the past, present and future Fevereiro, 2020 Mestrado Integrado em Medicina Área: Doenças Infecciosas Tipologia: Monografia Trabalho efetuado sob a Orientação de: Doutor António Carlos Megre Eugénio Sarmento Trabalho organizado de acordo com as normas da revista: Nature reviews journals Joana Oliveira e Sousa Ferraz Brandão Streptococcus pyogenes in tumor treatment: the past, present and future Fevereiro, 2020 UC Dissertação/Projeto (6º Ano) - DECLARAÇÃO DE INTEGRIDADE Eu, Joana Oliveira e Sousa Ferraz Brandão, abaixo assinado, nº mecanográfico 201307053, estudante do 6º ano do Ciclo de Estudos Integrado em Medicina, na Faculdade de Medicina da Universidade do Porto, declaro ter atuado com absoluta integridade na elaboração deste projeto de opção. Neste sentido, confirmo que NÃO incorri em plágio (ato pelo qual um indivíduo, mesmo por omissão, assume a autoria de um determinado trabalho intelectual, ou partes dele). Mais declaro que todas as frases que retirei de trabalhos anteriores pertencentes a outros autores, foram referenciadas, ou redigidas com novas palavras, tendo colocado, neste caso, a citação da fonte bibliográfica. Faculdade de Medicina da Universidade do Porto, 24/02/2020 Assinatura conforme cartão de identificação: ________________________________________________ Joana O.s FerrazBrandão UC Dissertação/Projeto (6º Ano) – DECLARAÇÃO DE REPRODUÇÃO NOME Joana Oliveira e Sousa Ferraz Brandão NÚMERO DE ESTUDANTE E-MAIL 201307053 [email protected] DESIGNAÇÃO DA ÁREA DO PROJECTO Medicina Clínica TÍTULO DISSERTAÇÃO/MONOGRAFIA (riscar o que não interessa) Streptococcus pyogenes in tumor treatment: the past, present and future ORIENTADOR Doutor António Carlos Megre Eugénio Sarmento COORIENTADOR (se aplicável) ASSINALE APENAS UMA DAS OPÇÕES: É AUTORIZADA A REPRODUÇÃO INTEGRAL DESTE TRABALHO APENAS PARA EFEITOS DE INVESTIGAÇÃO, MEDIANTE DECLARAÇÃO ESCRITA DO INTERESSADO, QUE A TAL SE COMPROMETE. É AUTORIZADA A REPRODUÇÃO PARCIAL DESTE TRABALHO (INDICAR, CASO TAL SEJA NECESSÁRIO, Nº MÁXIMO DE PÁGINAS, ILUSTRAÇÕES, GRÁFICOS, ETC.) APENAS PARA EFEITOS DE INVESTIGAÇÃO, MEDIANTE DECLARAÇÃO ESCRITA DO INTERESSADO, QUE A TAL SE COMPROMETE. DE ACORDO COM A LEGISLAÇÃO EM VIGOR, (INDICAR, CASO TAL SEJA NECESSÁRIO, Nº MÁXIMO DE PÁGINAS, ILUSTRAÇÕES, GRÁFICOS, ETC.) NÃO É PERMITIDA A REPRODUÇÃO DE QUALQUER PARTE DESTE TRABALHO. Faculdade de Medicina da Universidade do Porto, 24/02/2020 Assinatura conforme cartão de identificação: ______________________________________________ X Joana 0.5 Ferraz Brandão À minha Família e ao meu desporto, a Ginástica Acrobática, que me ensinaram a aceitar todas as oportunidades que a vida propõe com empenho e dedicação. Aos meus Amigos, que me orgulho de ver crescer e tornarem-se todos os dias pessoas mais completas, sem nunca perderem o seu singular sentido de humor e vontade de entrega ao outro. À Tuna Feminina de Medicina do Porto e à Família que construí na mui nobre Faculdade de Medicina por me terem ensinado a importância da abstração das convicções pessoais pelo bem de uma causa maior e que, independentemente dos percalços que surgirem, no final tudo se concretizará. STREPTOCOCCUS PYOGENES IN TUMOR TREATMENT: THE PAST, PRESENT AND FUTURE Joana Brandão1 Doutor António Sarmento2 1 Estudante de Mestrado Integrado em Medicina, Faculdade de Medicina da Universidade do Porto 2 Director do Serviço de Doenças Infecciosas, Centro Hospitalar Universitário de São João Professor Catedrático Convidado da Faculdade de Medicina da Universidade do Porto Correspondence: Joana Oliveira e Sousa Ferraz Brandão Faculdade de Medicina da Universidade do Porto Alameda Professor Hernâni Monteiro 4200-319 Porto E-mail: [email protected] Serviço de Doenças Infecciosas Director de Serviço: Doutor António Sarmento Alameda Professor Hernâni Monteiro 4200-319! Abstract In light of the recent emergence of immunotherapy as a possible treatment in Oncology, and considering the concept of spontaneous tumor regression following an infectious episode, the present work aimed to understand the role of infectious diseases in tumor treatment, specifically concerning the pathogen Streptococcus pyogenes. The pertinence of this bacteria in cancer treatment has been investigated throughout the last century and has achieved promising results, but it still isn’t a reliable tool. This work reviews the most relevant aspects of Streptococcus pyogenes usage in cancer treatment, including: Streptococcus pyogenes-based formulations, administration protocols used, possible mechanisms of action for the evidenced tumor regression, main side effects observed and possibilities for clinical assessment of responders and non-responders, as well as the most relevant differences in the outcomes observed in different clinical studies. The current evidence suggests that Streptococcus pyogenes has established itself as a reliable inducer of the immune system, which can positively contribute to cancer treatment. However, further clinical evaluations are still needed to validate the present findings and allow for a conclusion to be drawn. Key words: streptococcus pyogenes; OK432; immunotherapy; bacterial infection; tumoral regression! 2 Resumo Aliando a recente expansão da Imunoterapia, uma opção promissora no tratamento de tumores e cancro, ao conceito antigo de regressão espontânea de massas tumorais pós-infeção, surge a pertinência de avaliar o papel das doenças infecciosas no tratamento tumoral. No âmbito da presente revisão, abordou-se com especial enfoque o Streptococcus pyogenes. A relevância desta bactéria advém de ainda não se ter consagrado como uma ferramenta de tratamento, apesar dos resultados promissores que têm vindo a ser obtidos no último século. Assim, nesta revisão estão reunidos os aspetos mais relevantes do uso desta bactéria no tratamento de massas tumorais, nomeadamente: preparações obtidas com base no Streptococcus pyogenes, protocolos de administração, possíveis mecanismos de ação para a regressão tumoral evidenciada, principais efeitos colaterais observados e possibilidades de monitorização clínica, assim como as diferenças mais relevantes nos resultados observados em diferentes estudos clínicos. As evidências atuais sugerem que o Streptococcus pyogenes é um indutor do sistema imunológico, o que poderá ser uma importante contribuição para a abordagem dos doentes com tumores e cancro. No entanto, para se ter uma noção concreta do seu papel, constatamos que são ainda necessários mais estudos clínicos. Palavras-chave: streptococcus pyogenes tumor treatment; OK432; imunoterapia; infeção bacteriana; regressão tumoral 3 cells. First, an adenovirus prototype was used as vector; afterwards a spliceosome-mediated RNA- transplicing technology was attempted.33,34 Both experiments were conducted in vitro and showed successful outcomes. In both cases, it was reported that tumoral cells produced streptolysin and this led to membrane disruption and tumor cell death. This matter introduced Streptococcus pyogenes as a promising new agent in suicide gene cancer therapy.33,34 Before the use of streptolysin, the use of streptococcal gene Em55 was also attempted, given that it encodes M protein, a surface virulence factor. This M protein binds to glycosaminoglycans (GAGs) and enhances antiphagocytic and antiopsonization activity of Streptococcus pyogenes that further increased the immune system’s response.35,36 Then it was attempted to insert a DNA plasmid containing Em55 gene in tumoral cells, that were subsequently used to create a vaccine. 37 Although it seemed promising, no further studies were found regarding this matter. III. Clinical studies of Coley Vaccine and OK432 From the reports we reviewed concerning Coley’s Vaccine and OK432’s in vivo application, we were able to assemble the most important and consensual features. III-a) Route of administration The following methods were explored: intramuscularly, locally intra- or peritumoral and intravenously.5,14 Although in the early experiments, much concern about first applications as intratumoral injection was given, thinking that it could precipitate severe complications such as tumor lysis syndrome or circulatory collapse14, in the later studies intratumoral injection was indicated as the most effective method52-68. In fact, in cases of malignant ascites intraperitoneal was the best choice22,69-71. In the most recent experiments, the systemic approach didn’t seem as powerful in reducing tumor burden specially concerning pancreatic cancer.15 III-b) Administration protocol Both single and multiple administrations of the streptococcal formulas were attempted. Single administration seems to have the fewest effects58, however it had a significant positive result in intrapleural administration in intrapleural malignancies.72-77 10 Multiple exposure, with a 24 to 48 hours interval between injections, showed the greatest efficacy14, having results ranging from lower recurrence rate, enhancement of disease stability and better survival rate, to complete tumor regression.6,43,58,78-87 Concerning the dosage of streptococcal content in the vaccines, it has been customized through different studies, and no standardization was found in the articles reviewed. However the first experiments mention a gradual increase of the vaccine’s bacterial content until a fever of >39ºC was present.12,14 III-c) Combination therapy Regarding the reports on the usage of streptococcal derivates to treat cancer patients, several reported that this procedure was conducted after the patient had undergone other treatments or was used as a complement to other therapies. More specifically, it was used as a complement to pleurodesis72-77, in combination with chemotherapy, after mastectomy with axillary dissection in breast cancer patients51,88, as an adjuvant agent to chemo- or/and radiotherapy and as an immune maturation agent 16,89-92. These promising reports indicate that streptococcal formulations could enhance tumor treatment of other antineoplastic drugs7,15, allowing the use of lower radiation doses, even below those proposed as tumoricidal.93 Finally, as indicated before, its application also allowed for the decrease of analgesic dosage.2 III-d) Clinical assessment of response to application of Streptococcus pyogenes derivates in cancer patients From the reports that regarded a clinical approach to the use of streptococcal derivates to treat cancer patients, especially through OK432 and mixed bacterial vaccines, the most relevant observations were gathered concerning the changes in physical examination and possible laboratorial criteria to assess a positive response. Good outcomes were often reported, including diminished growth tumor, significantly reduced healing time51,88, reduction of recurrences, controlled dissemination52-54,56,57,63 and prolonged survival time.16,22,26,43,55,59-62,64-71,89-92,94,95 Concerning the assessment of the clinical signs and symptoms, a shift in the characteristics of the tumoral mass was observed macroscopically — first by becoming gradually less hard and immobile, and afterwards paler, softer and more mobile, until it discharged a caseous secretion or 11 simply regressed.5 One important aspect reported was that no apparent effect on healthy tissues was observed, not even when it was used in combination with radiation therapy.93 Moreover, in closed cavities, the sclerosing effect of the injection with OK432 was able to enhance adhesion and obliterate the newly created space, with the most successful cases being those where the injection was applied in macrocystic lesions (>1cm), with concomitant aspiration of the cavity’s content before the injection of OK432.12,75 Nonetheless, there were reports indicating that streptococcal formulas were unable to produce a significant impact if curative surgery had already been performed.56,95 Concerning the laboratorial complementary assessment of response to the injection of Streptococcus pyogenes components, both blood and cutaneous tests were reported to have promising results. Hanaue et al. observed that the presence of a normal or higher peripheral leucocyte count and the levels of lymphocyte and the percentage of T cells correlated significantly with survival and anticipated a better outcome.96 Later, Chirigos et al. reported that the T cell role in survival was more specifically due to an increment on NK cytotoxicity observed with just 0,001µg/ mL of OK432, and with peak activity at 1µg/mL.97 Moreover, Aoyagi et al. noted the reduction of CEA to normal values within 6 months of OK432 injection.54 Regarding the effects of Streptococcus pyogenes exposure on skin tests, Hananue et al. reported a significant correlation with survival concerning phytohemagglutinin skin test (PHA skin test) and serum titer glutinine. The first test, PHA skin test, consists of phytohemagglutinin application in the skin, that nonspecifically stimulates normal lymphocytes, thus indicating the efficiency of a healthy cell-mediated immunity system. In the second test, a significant correlation with life-span rate (p<0,01) was observed when the serum antibody titer rose 64 times higher, after stimulation with Streptococcus pyogenes components.96 The most widely used skin tests were PPD and SU-PS skin test, and both showed good correlation with life span rate.98 The PPD is a test that uses a purified protein derivate of tuberculin and was also linked to a better survival rate96, specially when it was observed the conversion from negative to positive 2 to 6 weeks after OK432 peritumoral injection.16,98 Nonetheless, there was one study which reported no change in intratumoral injection 89 and another describing no changes when the injection was systemic.16 Concerning the SU-PS skin test, in healthy patients the reaction is usually positive and changes to negative as the cancer progresses.94 Its mechanism of action is still not well understood98, but there are several reports arguing that in most cancer patients it re-shifted from 12 negative to positive 2 to 6 weeks after the OK432 injection.89,94,98 Further analysis showed that whilst SU-PS changed reactivity, it was also detected an increase in lymphocytes, T-helper cells and leucocytes (due to neutrophils enhancement).98 This confirmed that a delayed hypersensitive reaction is in the basis of the response to Streptococcus pyogenes anti-tumor activity, and therefore is able to recall an antigen and anticipate a response to OK432 injection.94 Although, the mechanisms behind Streptococcus pyogenes formulas usage in cancer treatment have been studied extensively, there is a lack of solid clinical evidence approaching its effects on patients with tumors. Nonetheless, we gathered some informations concerning the tumors in which OK432 had been used. In prospective studies, the biggest flaw was the small number of subjects, but they still reported the possibility of a reliable efficacy of OK432 in the treatment of recurrent cervical cysts and lymphoceles99-101, metastatic and/or recurrent head and neck tumor102, sialoceles post-parotidectomy103, juvenile nasopharyngeal angiofibroma104, malignant brain tumors16,92, oral melanoma105, gastric carcinoma52,57, hepatocelular carcinoma65,68, colorectal carcinoma72, malignant ascites68 and malignant pleural effusions.75 Accordingly, the observations collected retrospectively concerning OK432 in treatment of refractory/recurrent ovarian cancer86,87 and malignant pleural effusions73,76,77 were also promising, having reported an increase in disease stability and better outcomes with survival rate free of disease being significant after OK432 administration. III-e) Randomized controlled trials of OK432 in tumor treatment Two multi-center randomized trials conducted in Japan reported that intratumoral injection of OK432 before curative surgery in gastric cancer had a positive impact in the 5- and 10-years survival rate in stage III cancers, as well as it was effective in reducing the regional lymph node metastasis. Nonetheless, both reported that overall there were no differences in outcomes between experimental and control groups.53,56 This matter was later clarified by Oba et al. in a meta-analysis with 8009 patients that further concluded that adjuvant OK432 injection followed by curative surgery improved survival rate.63 More recently, in an individual patient meta-analysis, this improvement was considered as borderline significant in stage III and IV gastric cancers, leaving the most significant response in earlier cancer stages.95 Recently, in the last 10 years (2010-2019) only three randomized controlled trials were found to have been conducted with streptococcal lysates, namely OK432. The three randomized controlled trials found concerned approaches towards seroma formation after axillary 13 lymphadenectomy in breast cancer. In all there was a significant correlation between OK432 use and reduced incidence of seroma after axillary lymphadenectomy and better outcomes.51,88,106 Thus, it was reported significantly less volume and duration of drainage after surgery, fewer and smaller seroma occurrences and shorter duration of healing time.51,84,88 Nevertheless, the impact of its use on survival time wasn’t obtained due to inadequate follow up time.106 This vouches for OK432 as a potent, effective and safe sclerosing agent that can complement axillary lymphadenectomy surgical protocols.84,88 III-f) Main side effects of streptococcal components use Most of the adverse reactions were described as mild, ranging from constitutional symptoms like fatigue38 or anorexia38,53, to more localized manifestations on the site of injection, or local inflammation, hyperemia50,75, chest pain17,75,107 and abdominal pain.53 The most frequently reported event was fever, often controllable with antipyretic therapy, onl y a few studies repo r t e d t o ha v e st o p e d t r e a tme n t as soon as fever developed.17,38,50,52,55,75,98,102,107,108 Intraorbital or intracranial injections require close monitoring, since rapid elevations of intracranial pressure can happen, and preventive measures must be taken.16,109 Six patients were reported to have fatal adverse events due to embolism, acute nephritis, hemorrhage and inadequately dosed first injections.5 14 Discussion Contrary to older misconceptions, some authors concluded that infection or fever doesn’t always have to be an unwanted and frightening circumstance in cancer patients and it doesn’t necessarily represent the organism’s failure to defend itself. We acknowledge the existence of a dichotomy between infection and cancer since it has been established that a chronic infection enhances the malignancy’s risk.14 However, there are some evidences that an infection can also stimulate the immune system and trigger a reaction that will lead to tumor regression. Additionally, current methods for cancer treatment (chemotherapy, radiation and often invasive surgeries) have many disadvantages, such as high morbidity rates.2,7,9 Thus, the adequate therapy for tumors should have a good anticancer activity with nearly no toxicity, the ability to recognize and selectively act in neoplastic cells without damaging healthy tissues, and the capability to overcome neoplastic cells constant alterations.2 Fehleisen, Busch, Coley and Coley-Nauts believed that infection could treat tumors. Their work can contribute to a change of paradigm in cancer treatment.13,14,110,111 In our opinion, their work is promising and also alerts for new conceptions that can contribute for scientific advances.1,12,14 Thus, from this review, we think that immunotherapy with streptococcal formulas can be a valuable contribution for the treatment of cancer. However, the lack of further research in this area is a possible explanation for its limited use.14,15,98 In the future, we propose that further investigation should be carried out regarding some more unclear subjects such as optimal administration condition, which tumors will be the most susceptible, the best timing to initiate treatment, vaccine dosage, interval time between re-injections, injection site, indicators of good responders, and unwanted events and how to avoid them.6,10,15,62,112,113 One of the limitations of this paper, was the inability to review some papers as we did not master the language. The lack of double blinded randomized studies is also a reality. Therefore, it is the authors belief that this matter is worth further investigation and investment. We acknowledge that in the recent years many breakthroughs were presented in the immunotherapy field, but there still is a vast amount of solutions and strategies that are yet to be uncovered, as well as older methods that might have been forgotten, but may still prove revolutionary.! 15 References 1. Hobohm, U. Fever and cancer in perspective. Cancer Immunol Immunother 50, 391-396 (2001). 2. Kucerova, P. & Cervinkova, M. Spontaneous regression of tumour and the role of microbial infection--possibilities for cancer treatment. Anticancer Drugs 27, 269-277 (2016). 3. Negishi, Y. et al. Studies of natural killer activity and augmentation by OK-432 in patients with gynecological malignancies. Gynecol. obstet. Invest 21, 40-46 (1986). 4. Linnebacher, M., Maletzki, C., Emmrich, J. & Kreikemeyer, B. 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