Voreloxin Is an Anticancer Quinolone Derivative that Intercalates DNA and Poisons Topoisomerase II

Topoisomerase II is critical for DNA replication, transcription and chromosome segregation and is a well validated target of anti-neoplastic drugs including the anthracyclines and epipodophyllotoxins. However, these drugs are limited by common tumor resistance mechanisms and side-effect profiles. Novel topoisomerase II-targeting agents may benefit patients who prove resistant to currently available topoisomerase II-targeting drugs or encounter unacceptable toxicities. Voreloxin is an anticancer quinolone derivative, a chemical scaffold not used previously for cancer treatment. Voreloxin is completing Phase 2 clinical trials in acute myeloid leukemia and platinum-resistant ovarian cancer. This study defined voreloxin's anticancer mechanism of action as a critical component of rational clinical development informed by translational research.Biochemical and cell-based studies established that voreloxin intercalates DNA and poisons topoisomerase II, causing DNA double-strand breaks, G2 arrest, and apoptosis. Voreloxin is differentiated both structurally and mechanistically from other topoisomerase II poisons currently in use as chemotherapeutics. In cell-based studies, voreloxin poisoned topoisomerase II and caused dose-dependent, site-selective DNA fragmentation analogous to that of quinolone antibacterials in prokaryotes; in contrast etoposide, the nonintercalating epipodophyllotoxin topoisomerase II poison, caused extensive DNA fragmentation. Etoposide's activity was highly dependent on topoisomerase II while voreloxin and the intercalating anthracycline topoisomerase II poison, doxorubicin, had comparable dependence on this enzyme for inducing G2 arrest. Mechanistic interrogation with voreloxin analogs revealed that intercalation is required for voreloxin's activity; a nonintercalating analog did not inhibit proliferation or induce G2 arrest, while an analog with enhanced intercalation was 9.5-fold more potent.As a first-in-class anticancer quinolone derivative, voreloxin is a toposiomerase II-targeting agent with a unique mechanistic signature. A detailed understanding of voreloxin's molecular mechanism, in combination with its evolving clinical profile, may advance our understanding of structure-activity relationships to develop safer and more effective topoisomerase II-targeted therapies for the treatment of cancer

Applications of stochastic modeling in air traffic management:Methods, challenges and opportunities for solving air traffic problems under uncertainty

In this paper we provide a wide-ranging review of the literature on stochastic modeling applications within aviation, with a particular focus on problems involving demand and capacity management and the mitigation of air traffic congestion. From an operations research perspective, the main techniques of interest include analytical queueing theory, stochastic optimal control, robust optimization and stochastic integer programming. Applications of these techniques include the prediction of operational delays at airports, pre-tactical control of aircraft departure times, dynamic control and allocation of scarce airport resources and various others. We provide a critical review of recent developments in the literature and identify promising research opportunities for stochastic modelers within air traffic management

Assessment of Erwinia amylovora diversity and virulence: from strain molecular typing to immuno-flow cytometry of infected Pyrus fruits

Tese de mestrado, Microbiologia Aplicada, Universidade de Lisboa, Faculdade de Ciências, 2020Erwinia amylovora is a phytopathogenic bacterium and the causative agent of fire blight, a destructive disease that affects several members of the Rosaceae family. Pear and apple are particularly susceptible hosts of this bacterium, representing a major concern due to their socioeconomic value. The devasting nature of this disease has led to the classification of E. amylovora as a quarantine organism. Although this bacterium is widespread throughout the world, its emergence in Portugal happened relatively recently when compared to other European countries. Since its first report in 2006 in Fundão region, the number of outbreaks has increased, leading Portugal to loss its statute of Integral Protected Area within the European Union, in 2019. Factors such as the absence of a cure, false-negatives results and the need to understand unknown aspects about the life cycle of E. amylovora, lead to the interest in conducting characterization studies and developing alternative laboratory methods as an attempt to make preventive measures more effective. A set of Portuguese and foreign E. amylovora isolates was characterized by molecular, pathogenicity and virulence studies. Molecular characterization used CRISPR-PCR and genomic fingerprintings, whereas pathogenicity and virulence were assessed by biological tests on immature fruitlets of Pyrus communis cv. “Rocha”. In addition, a flow cytometry and an immuno-flow cytometry protocols were developed using pure and mixed bacterial cultures for the detection and cell viability assessment of E. amylovora in planta. A total of two CRISPR genotypes, A and D, was observed revealing a low genetic diversity among the E. amylovora isolates tested. Genomic fingerprinting reinforced the high homogeneity of this species. In contrast, a wide diversity in virulence was displayed. Flow cytometry and immuno-flow cytometry protocols were validated, revealing the capacity to detect and distinguish different viability states of E. amylovora artificially inoculated in pear fruitlets. In the future, the established protocols may help shed some light over previously undiscovered aspects of E. amylovora life cycle.Erwinia amylovora é uma bactéria fitopatogénica Gram-negativa responsável pela doença comummente denominada por fogo bacteriano. A designação desta doença está relacionada com o desenvolvimento de necroses de cor castanha a preta no hospedeiro durante o processo de infeção, cuja aparência se assemelha a uma queima. Outros sintomas típicos incluem o surgimento de lesões, exsudado bacteriano, cancros nos ramos e tronco, bem como de mumificação dos frutos. Originalmente nativa da América do Norte, E. amylovora foi detetada pela primeira vez no final do século 18. Atualmente, por consequência da ação humana, a doença está amplamente disseminada pelo globo. Devido à elevada capacidade de propagação e ao efeito potencialmente devastador para os seus hospedeiros, E. amylovora foi classificada como um organismo de quarentena. Os hospedeiros desta bactéria pertencem à família Rosaceae, na qual estão inseridos géneros de plantas agrícolas importantes a nível socioeconómico, tais como Malus e Pyrus, Em Portugal, a presença de fogo bacteriano foi reportada pela primeira vez em 2006 no Fundão, data relativamente tardia aquando comparação do surgimento da doença nos restantes países da Europa. Desde então, a ocorrência de situações reportadas em várias regiões do país tem vindo a aumentar, fazendo com que o país perdesse o estatuto de Zona Integral Protegida na União Europeia em 2019. Alguns fatores que tornam esta doença preocupante são a inexistência de cura e o desconhecimento de alguns aspetos do ciclo de vida de E. amylovora. Assim, são aplicadas estratégias de controlo que incluem abordagens complementares, tais como medidas preventivas, profiláticas e de erradicação. Outro fator preocupante é a ocorrência de resultados falso-negativos aquando aplicação dos métodos utilizados para o diagnóstico desta bactéria. Estes podem dever-se a uma concentração bacteriana insuficiente ou à indução do estado viável não cultivável em E. amylovora, o qual pode estar associado, por exemplo, a condições climáticas adversas e à utilização de compostos cúpricos como medida profilática. Um diagnóstico errado pode conduzir à falta de aplicação de medidas de controlo e, consequentemente, acarretar consequências catastróficas. O presente trabalho teve dois objetivos. O primeiro consistiu na caracterização de um conjunto de isolados de Erwinia amylovora da Coleção Portuguesa de Bactérias Fitopatogénicas através de estudos genómicos, de patogenicidade e de virulência. O segundo compreendeu o desenvolvimento e validação de protocolos de citometria de fluxo e de imuno-citometria de fluxo, para servirem como método de diagnóstico alternativo na deteção e avaliação da viabilidade celular de populações de E. amylovora presentes em material infectado. Um total de 48 isolados de E. amylovora foram caracterizados genotipicamente a partir da utilização CRISPR-PCR e de fingerprintings genómicos, nomeadamente rep- e MSP-PCR. A patogenicidade e virulência destes isolados foram caracterizadas a partir da utilização de frutos imaturos de Pyrus communis cv. “Rocha”, os quais foram artificialmente infetados por um subconjunto dos 48 isolados. Após 6 e 12 dias de infeção, foram registados o tamanho da lesão necrótica e a presença/ausência de exsudado bacteriano, respetivamente. Para avaliação da viabilidade celular de E. amylovora por citometria de fluxo, foram testados três fluoróforos diferentes, nomeadamente Syto9, PI e DIBAC4(3). Estes foram aplicados em células tratadas por calor e em células não tratadas. A aplicação do protocolo de imuno-citometria de fluxo compreendeu o uso do anticorpo monoclonal Ea7A IVIA e de um anticorpo secundário conjugado com FITC. Ambos os protocolos foram testados primeiramente numa cultura pura de E. amylovora e posteriormente em culturas mistas. Por fim, os dois protocolos foram implementados na deteção e avaliação de viabilidade celular de E. amylovora presente em peras imaturas cv. “Rocha” infetadas artificialmente. A técnica CRISPR-PCR permitiu diferenciar os isolados em dois grupos consoante a presença ou ausência da duplicação do spacer 1029, nomeadamente em genótipo A e genótipo D, respetivamente. A análise dos isolados Portugueses permitiu verificar que ambos os genótipos estão presentes no país desde 2010 e que existe uma ligeira predominância do genótipo D, o qual foi registado em 17 dos 31 isolados. Em termos de distribuição, observou-se que o genótipo A e D estão presentes nas regiões Centro e Oeste de Portugal, contudo no Alentejo só se verificou a presença do genótipo A. Aquando análise conjunta de resultados de estudos prévios com os que foram obtidos no presente trabalho, verificou-se que o genótipo A é o genótipo mais distribuído na Europa, possivelmente devido à introdução mais precoce do mesmo no continente Europeu. As técnicas de fingerprinting genómico utilizadas, nomeadamente rep- e MSP-PCR, permitiram a obtenção de perfis genómicos complexos, mas muito homogéneos entre si. Desta forma, os mesmos mostraram não ter poder discriminatório suficiente para a diferenciação dos isolados de E. amylovora a nível infraespecífico. Estes resultados são o reflexo de uma variedade genómica limitada característica desta espécie bacteriana, que culmina numa elevada homogeneidade entre os isolados. Os ensaios em peras imaturas revelaram que, à excepção dos isolados CPBF 142 e CPBF 544, 43 isolados são patogénicos, uma vez que possuíram a capacidade de induzir o aparecimento de pelo menos um dos sintomas típicos do fogo bacteriano no hospedeiro. Todos os isolados patogénicos provocaram lesão necrótica de cor castanha a preta, contudo apenas 28 isolados produziram exsudado bacteriano. A análise destes sintomas permitiu observar variabilidade na virulência, de tal modo que os isolados foram distribuídos em três categorias de virulência, nomeadamente baixa, média e alta. Estas categorias tiveram em consideração o tamanho da lesão necrótica provocada no hospedeiro. Adicionalmente, observou-se a existência de correlação entre a categoria de virulência baixa e o genótipo D. Os dados de CRISPR-PCR, patogenicidade, virulência e presença/ausência de exsudado bacteriano foram agrupados para uma análise polifásica de modo a adquirir mais informação acerca dos isolados. Os dados obtidos por fingerprinting genómico não foram considerados, uma vez que não tiveram o poder discriminatório adequado. A utilização desta abordagem permitiu separar os isolados em 11 grupos de estirpes distintos. Contudo, não foi possível fazer uma associação entre estes grupos e o hospedeiro original a partir do qual cada isolado foi obtido. Relativamente à citometria de fluxo, a utilização dos fluoróforos permitiu a distinção de diferentes populações relativamente à integridade celular e potencial membranar presentes na cultura pura de E. amylovora. De facto, esta técnica apresentou uma boa correlação entre a viabilidade esperada e a observada, o que significa que os três fluoróforos se mostraram apropriados para serem utilizados em estudos de viabilidade celular nesta bactéria. O estudo de viabilidade celular de E. amylovora em cultura mista ficou comprometido devido ao comportamento semelhante que as bactérias podem ter face aos fluoróforos. O protocolo de imuno-citometria de fluxo permitiu detetar E. amylovora em cultura pura a partir da observação de uma elevada intensidade de fluorescência verde. Em cultura mista, para além da intensidade de fluorescência verde característica de E. amylovora, verificou-se também a ocorrência de uma emissão de fluorescência verde menor que se supõe que resulte de ligações inespecíficas entre o anticorpo secundário e outros alvos que não o anticorpo monoclonal Ea7A IVIA. Contudo, uma vez que a intensidade da emissão de fluorescência verde adicional foi reduzida, a mesma não impossibilitou a detecção de E. amylovora. O presente estudo reforçou a paradoxalidade existente entre a homogeneidade genómica e a heterogeneidade de virulência em Erwinia amylovora. Adicionalmente, o mesmo mostrou que o desenvolvimento de um potencial método alternativo para a detecção desta bactéria, nomeadamente com recurso a citometria de fluxo e técnicas afins, pode significar uma melhoria considerável no processo de diagnóstico, bem como abrir portas para descobertas futuras relacionadas com o ciclo de vida de E. amylovora

The Impact of Collateral Evolution on Optimal Dosing Strategies and Evolution on Paired Fitness Landscapes

Drug resistance is an ever-growing threat to successful treatment of bacterial, cancer and viral infections. As pathogens and cancers continue to find evolutionary solutions to the drugs we treat them with, scientists have begun to focus on more evolutionary-based therapies such as drug cycling. These therapies aim to constrain or control evolution in a particular way such that intractable resistance never evolves. In the same vein, recent work has revealed collateral sensitivity as a promising avenue to guide evolution away from untreatable resistance states. Collateral evolution occurs when a population evolves resistance to the selecting drug and this mechanism of resistance confers "collateral" effects to different drugs it is not exposed to. In this work we show how collateral profiles might be used to slow the acquisition to resistance in a simplified laboratory-based evolution experiment. We demonstrate that intuitive cycling protocols often fail over long time periods, whereas mathematically optimized protocols maintain long-term sensitivity at the cost of transient periods of high resistance. We then extend this work to include nonantibiotic stressors such as pH, salt and food preservatives. This extension highlights that more work is necessary to understand the role these common environments have on the development of multidrug resistance. Finally, using the well-known fitness landscape paradigm, we explore how collateral effects influence the evolutionary dynamics of a pair of landscapes with tunable correlations. We show that alternating evolution in highly correlated environments can lead to higher mean fitness than evolution in either landscape alone, while alternating between two anti-correlated landscapes results in a lower mean fitness. We demonstrate this is due to the location and number of shared maxima between the two correlated landscapes, which change as a function of ruggedness (epistasis) and paired landscape correlation. Taken together, these results begin to answer many of the important questions required to translate collateral sensitivity into clinical treatments.PHDBiophysicsUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/163088/1/jamaltas_1.pd

Gut microbes in the prevention and onset of metabolic disorders

ABSTRACT: Gut microbes are suggested to play an essential role in maintaining humanhealth, and unfavorable alterations in gut microbiota have been associated with several human diseases and disorders. Despite the significant advancements in analysis tools, several sources of uncertainty still exist in the gut microbiota composition analyses, limiting the reproducibility and comparability of the results from distinct gut microbiota studies. The first main aim of this study was to set up a functional high-throughput pipeline for 16S rRNA gene sequencing -based gut microbiota composition analysis. This was achieved by evaluating the effects of fecal sample processing, i.e. two sample storage conditions and five DNA extraction kits, along with two different 16S rRNA gene sequencing protocols. The second main aim of this study was to explore the role of gut microbiota in the onset of obesity-related metabolic disorders such as non-alcoholic fatty liver disease. This was done by studying the effects of intragastric administration of two distinct gut microbes, Faecalibacterium prausnitzii and Enterobacter cloacae, on host health and metabolism of high fat diet -fed C57BL/6N mice. The results of this study showed that the 16S rRNA gene sequencing protocol had a significant effect on the analysis results. Meanwhile, the effect of sample pre-processing was more modest, yet still potentially important. These results indicate that careful design and adequate method optimization are required in order to produce reliable 16S rRNA gene sequencing results. Moreover, the results of this study revealed that the intragastric F. prausnitzii administration appeared to protect the C57BL/6N mice from liver steatosis, whereas the intragastric administration of E. cloacae seemed to induce liver damage. Further studies are needed in order to clarify the underlying mechanisms and to fully elucidate the possible therapeutic potential of F. prausnitzii.TIIVISTELMÄ: Suolistobakteereilla uskotaan olevan merkittävä vaikutus ihmisen terveydelle, ja suolistomikrobiston epätasapaino on yhdistetty moniin eri sairauksiin ja oireyhtymiin. Mikrobistotutkimuksessa käytettävät analyysimenetelmät ovat viimeisen vuosikymmenen aikana kehittyneet merkittävästi, mutta analyysitulokset ovat yhä vahvasti menetelmäriippuvaisia, mikä heikentää tutkimusten toistettavuutta sekä tulosten luotettavuutta ja vertailukelpoisuutta. Tämän tutkimuksen ensimmäisenä päätavoitteena oli pystyttää toimivat menetelmät 16S rRNA -geenisekvensointiin perustuvalle mikrobistoanalytiikalle. Tutkimuksessa vertailtiin viittä eri DNA-eristysmenetelmää sekä kahta eri sekvensointiprotokollaa, ja selvitettiin ulostenäytteiden pakastuksen vaikutusta mikrobistoanalyysin tuloksiin. Tutkimuksen toisena päätavoitteena oli kartoittaa hiirimallin avulla suolistomikrobiston roolia ei-alkoholiperäisen rasvamaksan synnyssä. Tutkimuksessa selvitettiin, miten suun kautta tapahtuva Faecalibacterium prausnitzii- tai Enterobacter cloacae -annostelu vaikutti korkearasvaista ravintoa syövien C57BL/6N-hiirten terveyteen jaaineenvaihduntaan. Tutkimuksen tulokset osoittivat, että 16S rRNA –geenisekvensointimenetelmällä oli merkittävä vaikutus mikrobistoanalyysista saataviin tuloksiin. Näytteiden esikäsittelyn vaikutus oli huomattavasti vähäisempi, mutta kuitenkin tulosten vertailukelpoisuuden kannalta mahdollisesti merkityksellinen. Tulokset osoittavat, että huolellinen suunnittelu ja asianmukainen menetelmäoptimointi ovat välttämättömiä luotettavien 16S rRNA –geenisekvensointituloksen tuottamiseksi. Lisäksi tämän tutkimuksen tulokset osoittivat, että F. prausnitzii -annostelu vaikutti positiivisesti hiirten aineenvaihduntaan ja vähensi hiirten maksan rasvoittumista, kun taas E. cloacae -annostelu aiheutti maksavaurioita. F prausnitzii -suolistobakteerin mahdollista terveyttä edistävää potentiaalia ja taustalla olevia tekijöitä tulisi selvittää tarkemmissa jatkotutkimuksissa

2011 progress towards implementation of a public health action plan to combat antimicrobial resistance

In 2001, the Task Force developed an initial Action Plan, outlining specific issues, goals, and actions important for addressing the problem of AR. This document, entitled A Public Health Action Plan to Combat Antimicrobial Resistance, Part I: Domestic Issues, reflected a broad- based consensus of participating federal agencies, which was reached with individual input from state and local health agencies, universities, professional societies, pharmaceutical companies, healthcare delivery organizations, agricultural producers, consumer groups, and other members of the public. Continued collaboration with these partners has been vital to achieving successful implementation of the Action Plan. The 2011 revision of the Action Plan was based in part on individual input obtained at a consultants' meeting held in Atlanta, Georgia, in December 2007. Present at the public meeting were consultants with wide-ranging expertise in areas such as human and veterinary medicine, pharmaceutical and diagnostics manufacturing, animal husbandry, clinical microbiology, epidemiology, infectious diseases and infection control, and state and local public health officials. The Action Plan includes action items organized into four focus areas: Surveillance, Prevention and Control, Research, and Product Development.Executive summary -- Introduction and overview -- The Focus Areas -- Focus Area I: Surveillance -- Focus Area II: Prevention and Control -- Focus Area III: Research -- Focus Area IV: Product Development -- Acronyms and abbreviationsInteragency Task Force on Antimicrobial Resistance ; co-chairs: Centers for Disease Control and Prevention, Food and Drug Administration.Title from caption (viewed on October 1, 2012).The Interagency Task Force on Antimicrobial Resistance (hereafter referred to as the Task Force) was created in 1999 to coordinate the activities of federal agencies in addressing antimicrobiala resistance (AR) in recognition of the increasing importance of AR as a public health threat. The Task Force is co-chaired by the Centers for Disease Control and Prevention (CDC), the Food and Drug Administration (FDA), and the National Institutes of Health (NIH) and also includes the Agency for Healthcare Research and Quality (AHRQ), the Centers for Medicare and Medicaid Services (CMS), the Department of Agriculture (USDA), the Department of Defense (DoD), the Department of Veterans Affairs (VA), the Environmental Protection Agency (EPA), the Health Resources and Services Administration (HRSA), the Department of Health and Human Services Office of the Assistant Secretary for Preparedness and Response (HHS/ASPR), and the Department of Health and Human Services Office of the Assistant Secretary for Health (HHS/OASH).Mode of access: Internet; PDF reader (Acrobat .pdf file: 459 KB, 69 p.).Text (electronic publication

Planetary Biology and Microbial Ecology: Molecular Ecology and the Global Nitrogen cycle

This report summarizes the results of the Planetary Biology and Molecular Ecology's summer 1991 program, which was held at the Marine Biological Laboratory in Woods Hole, Massachusetts. The purpose of the interdisciplinary PBME program is to integrate, via lectures and laboratory work, the contributions of university and NASA scientists and student interns. The goals of the 1991 program were to examine several aspects of the biogeochemistry of the nitrogen cycle and to teach the application of modern methods of molecular genetics to field studies of organisms. Descriptions of the laboratory projects and protocols and abstracts and references of the lectures are presented

Many-Objective Genetic Programming for Job-Shop Scheduling

The Job Shop Scheduling (JSS) problem is considered to be a challenging one due to practical requirements such as multiple objectives and the complexity of production flows. JSS has received great attention because of its broad applicability in real-world situations. One of the prominent solutions approaches to handling JSS problems is to design effective dispatching rules. Dispatching rules are investigated broadly in both academic and industrial environments because they are easy to implement (by computers and shop floor operators) with a low computational cost. However, the manual development of dispatching rules is time-consuming and requires expert knowledge of the scheduling environment. The hyper-heuristic approach that uses genetic programming (GP) to solve JSS problems is known as GP-based hyper-heuristic (GP-HH). GP-HH is a very useful approach for discovering dispatching rules automatically. Although it is technically simple to consider only a single objective optimization for JSS, it is now widely evidenced in the literature that JSS by nature presents several potentially conflicting objectives, including the maximal flowtime, mean flowtime, and mean tardiness. A few studies in the literature attempt to solve many-objective JSS with more than three objectives, but existing studies have some major limitations. First, many-objective JSS problems have been solved by multi-objective evolutionary algorithms (MOEAs). However, recent studies have suggested that the performance of conventional MOEAs is prone to the scalability challenge and degrades dramatically with many-objective optimization problems (MaOPs). Many-objective JSS using MOEAs inherit the same challenge as MaOPs. Thus, using MOEAs for many-objective JSS problems often fails to select quality dispatching rules. Second, although the reference points method is one of the most prominent and efficient methods for diversity maintenance in many-objective problems, it uses a uniform distribution of reference points which is only appropriate for a regular Pareto-front. However, JSS problems often have irregular Pareto-front and uniformly distributed reference points do not match well with the irregular Pareto-front. It results in many useless points during evolution. These useless points can significantly affect the performance of the reference points-based algorithms. They cannot help to enhance the solution diversity of evolved Pareto-front in many-objective JSS problems. Third, Pareto Local Search (PLS) is a prominent and effective local search method for handling multi-objective JSS optimization problems but the literature does not discover any existing studies which use PLS in GP-HH. To address these limitations, this thesis's overall goal is to develop GP-HH approaches to evolving effective rules to handle many conflicting objectives simultaneously in JSS problems. To achieve the first goal, this thesis proposes the first many-objective GP-HH method for JSS problems to find the Pareto-fronts of nondominated dispatching rules. Decision-makers can utilize this GP-HH method for selecting appropriate rules based on their preference over multiple conflicting objectives. This study combines GP with the fitness evaluation scheme of a many-objective reference points-based approach. The experimental results show that the proposed algorithm significantly outperforms MOEAs such as NSGA-II and SPEA2. To achieve the second goal, this thesis proposes two adaptive reference point approaches (model-free and model-driven). In both approaches, the reference points are generated according to the distribution of the evolved dispatching rules. The model-free reference point adaptation approach is inspired by Particle Swarm Optimization (PSO). The model-driven approach constructs the density model and estimates the density of solutions from each defined sub-location in a whole objective space. Furthermore, the model-driven approach provides smoothness to the model by applying a Gaussian Process model and calculating the area under the mean function. The mean function area helps to find the required number of the reference points in each mean function. The experimental results demonstrate that both adaptive approaches are significantly better than several state-of-the-art MOEAs. To achieve the third goal, the thesis proposes the first algorithm that combines GP as a global search with PLS as a local search in many-objective JSS. The proposed algorithm introduces an effective fitness-based selection strategy for selecting initial individuals for neighborhood exploration. It defines the GP's proper neighborhood structure and a new selection mechanism for selecting the effective dispatching rules during the local search. The experimental results on the JSS benchmark problem show that the newly proposed algorithm can significantly outperform its baseline algorithm (GP-NSGA-III)