ANDREA NOVELLI

I MODIFICATORI DELLA REATTIVITA’ BIOLOGICA

Andrea Novelli

Dipartimento di Scienze della Salute

Sezione di Farmacologia Clinica & Oncologia

I modificatori della reattività biologica

• Angelini• Menarini Group• MERCK

• Named• Valeas• Zambon Group

Transparency Declaration

Honoraria or grant support received from:

Some biological response modifiers

NATURAL SUBSTANCES SYNTHETIC REAGENTS

Physiological Exogenous Pro-host drugs

• Interferons• Thymic factors• Tuftsin • Lactoferrin• Probiotics

• BCG• B. clausii spores• RU41740 (Biostim)• MPL• Muramil-dipeptide

(MDP)• Bacterial lysates• OK-432• Bestatin• β-glucan

• Methisoprinol (Isoprinosine)

• Lentinan (β glucans)• RC-529• MDP derivatives• Liposomes• Imiquimod• Resiquimod• Pidotimod

Human lactoferrin and lactoferricin

lactoferricin within the protein = yellow; ferric ions = red

Farnaud S & Evans RW, Mol Immunol, 2003

• An 80-kDa iron-binding glycoprotein of the transferrin family

• In body fluids is found in the iron-free form, the monoferric form and in the diferric form.

• Plasma lactoferrin is predominantly neutrophil derived but indications are that it may also be produced by other cells.

• Is a component of exocrine secretions such as milk and saliva, and is present in neutrophil granules

Antimicrobial peptides (AMP)

Modes of action

Vincent JL et al., Front Immunol, 2015

Bovine lactoferrinMechanism of action underlying the host-protective effects

Tomita M et al., Biochimie, 2009

Recurrent respiratory infections possible alterations in immune system function

↓ IgA, IgM and IgG↓ CD4+, CD8+, CD19+ and NK-cells

↓ Il-4, IL-10

↓ IFN-g, IL-12 and IL-2

↑ T-reg/Th ratio

↓ Th1/Th2 ratioDefective phagocytosis and chemotaxis of neutrophils

↓ TLR function and ciliary function

Dendritic cell immaturity

Mahashur A et al., Lung India, 2019

Composition of typical bacterial lysate preparations used for treatment of respiratory infections

• Haemophilus influenzae• Neisseria catarrhalis• Klebsiella pneumoniae• Klebsiella ozaenae• Streptococcus pneumoniae• Staphylococcus aureus• Streptococcus pyogenes• Streptococcus viridans

Bacterial lysate cocktail

Masihi KN, Int J Antimicrob Agents, 2000

Lisati batterici liofilizzati

• OM-85 Lisato batterico chimico liofilizzato (PCBL) di: • Haemophilus influenzae, Diplococcus pneumoniae, Klebsiella pneumoniae e ozaenae,

Staphylococcus aureus, Streptococcus pyogenes e viridans, Neisseria catarrhalis:

– Broncho Munal®, Broncho Vaxom®, Ommunal®

• D53 Lisato di frazioni ribosomiali di: • Klebsiella pneumoniae, Streptococcus pneumoniae, Streptococcus pyogenes gruppo A,

Haemophilus influenzae. Frazioni di membrana di: Klebsiella pneumoniae:

– Biomunil®, Immucytal®, Biostim®

• Lisato batterico meccanico liofilizzato (PMBL) di: • Staphylococcus aureus, Streptococcus pyogenes, Streptococcus viridans, Klebsiella

pneumoniae, Klebsiella ozaenae, Haemophilus influenzae sierotipo B, Neisseria catarrhalis, Diplococcus pneumoniae

– Immubron®, Ismigen®, Imudon®, Paspat®

Bacterial lysates

Effect on dendritic cells

Spisek R et al., Vaccine, 2004

OM-85

BIOSTIMBIOSTIM

OM-85PMBL PMBL

PMBL protects mice against lethal pneumococcal challenge

Rial A et al., Microbes and Infection, 2016

Sub-lingual administration of a polyvalent mechanical bacterial lysate (PMBL) in patients with moderate, severe, or very severe chronic abstructive pulmonary disease (COPD) according to the GOLD spirometric classification: A multicentre, double-blind, randomised, controlled, phase IV study (AIACE study: Advanced Immunological Approach in COPD Exacerbation)

Braido F, Melioli G, Cazzola M, Fabbri L, Blasi F, Moretta L, Canonica GW, for the AIACE Study Group

• PMBL did not achieve the primary outcome of reducing the number of exacerbation by 25%

• However the number of days with fever (21 vs 40.15), the days of hospitalization (65 vs 162), the interval between the first and second exacerbation (123.9 vs 70.4) and the number of days of poor health (109 vs 171) were significantly better then in the placebo group

Pulmonary Pharmacology & Therapeutics 33: 75-80; 2015

OM-85 p.oProtection of mice against influenza virus and secondary bacterial infection

Pasquali C et al., Frontiers in Medicine, 2014

Influenza virus

OM-85 p.oProtection of mice against influenza virus and secondary bacterial infection

Pasquali C et al., Frontiers in Medicine, 2014

Influenza virus Bacterial infection

A randomized, placebo controlled, double blinded, single centre, ‑ ‑ ‑phase IV trial to assess the efficacy and safety of OM 85 in children ‑suffering from recurrent respiratory tract infections

Esposito S , Bianchini S, Bosis S, Tagliabue C, Coro I, Argentiero A and Principi N

• Methods: This study was a randomized (3:3:1), placebo-controlled, double-blind, single-center, phase IV trial carried out 288 children aged 1 to 6 years with a history of recurrent RTIs

• Results: The number of RTIs and of children who experienced at least one RTI were significantly lower among patients receiving OM-85 for 3 months than among those given placebo (33% vs 65.1%, p < 0.0001)

• No difference was seen between children who received OM-85 for 3 and those who received OM-85 for 6 month

• Benefit was maximally evident among children with a history of frequent recurrences

J Transl Med (2019) 17:284

Controindicazioni - Ipersensibilità al principio attivo o ad uno qualsiasi degli eccipienti.Generalmente controindicati o da evitare in gravidanza e allattamento. Malattie autoimmuni. Infezioni intestinali acute.

Avvertenze speciali e precauzioni d’impiego - Il trattamento deve essere sospeso in caso di febbre elevata in particolare all’inizio del trattamento.Il paziente deve essere informato della possibilità di questo evento indesiderato raro ed il tipo di febbre deve essere differenziato dalla febbre che insorge a seguito della patologia originaria.

Deve essere evitata l’assunzione concomitante di un altro immunostimolante.In alcuni casi è stata osservata l’insorgenza di attacchi d’asma in pazienti predisposti dopo l’assunzione di farmaci contenenti estratti batterici.

Interazioni con altri medicinali e altre forme di interazione - Si raccomanda un intervallo di 4 settimane tra la fine del trattamento e l’inizio della somministrazione di un vaccino.

LISATI BATTERICI

• L’ EMA ha avviato una revisione dei medicinali a base di lisati batterici per le patologie respiratorie

• Recenti studi hanno sollevato dubbi sull’efficacia dei lisati batterici negli adulti e nei bambini che soffrono di infezioni respiratorie ripetute. Inoltre, in casi molto rari, è noto che questi medicinali causano gravi effetti avversi legati al sistema immunitario

• Questa revisione è stata richiesta dall’Agenzia Italiana del Farmaco (AIFA). L’ EMA revisionerà ora tutte le informazioni disponibili e raccomanderà se mantenere, variare o sospendere l’autorizzazione al commercio di questi medicinali in tutta l’UE

EMA/351982/2018

29 Giugno 2018

medicinali a base di lisati batterici

• Sebbene i dati siano limitati, dalla loro rivalutazione sono emerse alcune evidenze di efficacia di tali prodotti nella prevenzione delle infezioni ricorrenti delle vie respiratorie ed il profilo di sicurezza è in linea con l’atteso per questa tipologia di prodotti

• L’Agenzia Europa per i Medicinali (EMA) raccomanda che i medicinali a base di lisati batterici autorizzati per le malattie respiratorie siano utilizzati soltanto per la prevenzione delle infezioni respiratorie ricorrenti, con l’esclusione della polmonite

01 luglio 2019

Structure formula: C14H16N4

Molecular weight: 276.8

Imiquimod (R-837)

Resiquimod (R-848)

Structure formula: C17H22N4O2

Molecular weight: 314.4

Pidotimod

Molecular formula: C9H12N2O4SRelative molecular mass: 244.26

Small Synthetic compounds

Pidotimod

Synthesis of compounds with a peptide-like structure tested as immunostimulant agents

Magni A et al., Arzneimittelforschung, 1994

Pidotimod

Plasma levels in 12 healthy volunteers

200mg IV200mg OS400mg OS800mg OS

Mailland F et al., Arzneimittelforschung, 1994

Pidotimod 800mg OS single dose

Effect of food in 12 healthy male volunteers

D’Angelo L et al., Arzneimittelforschung, 1994

Sachet after fastingSachet after standard mealVial after fastingVial after standard meal

*

FED

FASTING

PidotimodMechanism of action

Ferrario BE et al., Clinical and Molecular Allergy, 2015

Pidotimod (800mg BID) in 16 pts with CAP with concomitant antimicrobial therapy*

* Results after 3 days of therapy

Trabattoni D et al., Pulm Pharmacol Ther, 2017

Pidotimod

Increase in IFN-g production and OVA-specific IgG production

OVA50μg/ dose

Pidotimod (100μg/ dose)

PBS

Days 0, 14 and 21

Giagulli C. International Immunopharmacology 2009

Splenocytes

IFN-g

IFN-g

IgG

Bone marrow cellsIntranasal administration

p= 0.050

p= 0.015p= 0.016

Pidotimod up-regulates TLR expression on monocytes

In children suffering from CAP

Esposito S et al. J Transl Med, 2015, 13:288

p= 0.041

p= 0.035

p= 0.044

p= 0.050

p= 0.049

p< 0.001p= 0.007

p= 0.007

Pidotimod stimulates IL-12 and TNF-a in monocytes In children suffering from CAP

Esposito S et al. J Transl Med, 2015, 13:288

IL-12 TNF-a

Immunomodulatory activity of pidotimod administered with standard antibiotic therapy in children hospitalized for community acquired pneumonia‑

Esposito S, Garziano M, Rainone V, Trabattoni D, Biasin M, Senatore L, Marchisio P, Rossi M, Principi N and Clerici M

PDT administration might significantly increase the activity of the immune system for a long period of time, thus reducing the risk of early recurrences during CAP in children

J Transl Med, 2015, 13:288

The immunomodulatory molecule pidotimod induces the expression of the NOD-like receptor NLRP12 and attenuates TLR-induced inflammation

Fogli M, Caccuri F, Iaria ML, Giagulli C, Corbellini S, Campilongo F, Caruso A, Fiorentini S

• We decided to investigate whether PDT could modify the immune response triggered by TLR ligands

• Monocytic cells were exposed to PDT then stimulated with a panel of TLR agonists• We have observed that PDT specifically up-regulates the expression of the NOD-like

receptor NLRP12 mRNA in the absence of any further co-stimulation• Furthermore, in myeloid/monocytic cells we demonstrated that PDT treatment counteracts

the NLRP12 reduction induced by TLR agonists• Finally, the results obtained using NLRP12 silenced cells showed that down-regulation of

the pro-inflammatory function occurring in PDT-treated cells upon interaction with TLRs is associated with the increased levels of NLRP12 induced by PDT

• To our knowledge this is the first evidence of an immunomodulatory peptide that upregulates NLRP12 and, through this molecule, antagonizes the TLR-induced inflammatory response

J Biol Regul Homeost Agents; 28(4):753-66, 2014

To assess the effectiveness and safety of pidotimod (PDT) in children aged <14 yrs with RRTIs

29 RCTs (4344 pediatric pts) were included in meta-analysis. 10 RCTs were from Italy, Russia or Greece, and 19 RCTs were from Chinese groups

Patients treated with PDT had significant lower RTIs, decreased duration of cough and fever, than those with conventional treatment

Antibiotic use was also remarkably decreased in the PDT treatment group

PDT improved the levels of serum Ig and T-lymphocyte subtypes without increasing the risk of adverse events of any cause

PDT showed a good efficacy and safety in treatment of pediatric RRTIs. Further high-quality and large-scale RCTs are still required to provide confirmatory evidence

Pidotimod

Forest plot showing RR with 95% CI of the number of patients who were using antibiotics

Niu H et al., International Immunopharmacology, 2019

32 studies included24 in children (1 – 14 yrs)

8 in adults

17 studies in children with RRI (w/ or w/o asthma)

11 RCT studies in children without asthma worldwide from 1994 to 2015

2180 pts evaluated (1126 treated with pidotimod)

Pidotimod: in-depth review of current evidenceMahashur A, Thomas PK, Mehta P, Nivangune K, Muchhala S, Jain R

Lung India, 36(5), 2019

Conclusion

• Immune dysregulation involving innate and adaptive immune responses has been identified in a variety of diseases in children and adults. Being able to positively affect these immune responses, pidotimod proved to be effective in improving the outcomes. Current evidence convincingly established its efficacy in children with RRIs with or without asthma and in adults experiencing acute exacerbations of CB

Metabolomic profile of children with recurrent respiratory infectionsBozzetto S, Pirillo P, Carraro S, Berardia M, Cesca L, Stocchero M, Giordano G, Zanconato S, Baraldi E

• 13 children (age range 3–6 yeas) with RRI and 15 matched per age healthy peers were enrolled.

• Metabolomic urine samples were compared before and after treatment with pidotimod for 3 months.

• Out of 1502 time per mass variables, 138 were different between the two groups.• 35/138 variables (metabolites of bile and hippuric acid) persisted after

pidotimod treatment.

• After pidotimod treatment, the metabolic profile of the children with RRI was no longer very different from that of the healthy controls, except for the persistence of some microbiome-related variables.

Pharmacological Research, 2017

Effects of pidotimod and bifidobacteria mixture on clinical symptoms and urinary metabolomic profile of children with recurrent respiratory infections: a randomized placebo-controlled trial

Santamaria F, Montella S, Stocchero M, Pirillo P, Bozzetto S, Giordano G, Poeta M, Baraldi E

• Objective: To evaluate whether pidotimod and/or bifidobacteria can reduce RRI morbidity and influence the urine metabolic profile in 55 preschool children

• Materials and methods: Children aged 3–6 years with RRI were enrolled in a 4-arm trial and were randomly assigned to receive pidotimod +/- bifidobacteria, bifidobacteria + placebo or double placebo for the first 10 days of each month over 4 consecutive months

Pulmonary Pharmacology & Therapeutics, 2019

Effects of pidotimod and bifidobacteria mixture on clinical symptoms and urinary metabolomic profile of children with recurrent respiratory infections: a randomized placebo-controlled trial

Santamaria F, Montella S, Stocchero M, Pirillo P, Bozzetto S, Giordano G, Poeta M, Baraldi E

• Results: Compared to placebo, children receiving pidotimod, alone or with bifidobacteria, had more symptom-free days (69 versus 44, p=0.003; and 65 versus 44, p=0.02, respectively) and a lower percentage of days with common cold (17% versus 37%, p=0.005; and 15% versus 37%, p=0.004, respectively)

• Children treated with pidotimod present a biochemical profile with compounds related to the pathway of steroids hormones, hippuric acid and tryptophan

• No significant difference in the metabolic profile was found children receiving bifidobacteria

• Conclusions: Preschool children with RRI treated with pidotimod have better clinical outcomes and a different urine metabolomic profile than subjects receiving placebo. Further investigations are needed to clarify the connection between pidotimod and gut microbiome

Pulmonary Pharmacology & Therapeutics, 2019

Updated instructions for pidotimod following online debate

• Pidotimod is often prescribed by pediatricians, ENT doctors, and dermatologists in China to treat various infections, and even colds or diarrhea, in young children.

• The new directions must clarify that the drug can only be prescribed as supplementary medication to adults and children over 3 years old, and for no more than 60 days.

• The drug is also no longer endorsed for treatment of acute infections, ENT infections such as rhinitis, or gynecological infections.

China FDA March 13, 2018

Conclusion (I)

• The rationale for the use of biological response modifiers (mainly in children or elderly people suffering from RRTI) is related to the purpose of increasing the immune response or enhancing the innate defence mechanisms.

• The idea of improving and speeding infectious episode resolution using bacterial lysates raised almost one century ago.

• Bacterial lysates might be an interesting therapeutic option for adults and children with recurrent respiratory infections, but their clinical effect, despite being visible, is moderate.

• Moreover The long-term safety profile of these preparations raises concerns regarding their potential to induce or aggravate autoimmune phenomena.

Conclusion (II)

• Among synthetic BRMs pidotimod, a dipeptide molecule synthesized in Italy in 1990, has biological and immunological activity on both the adaptive and the innate immune responses.

• This molecule seems particularly indicated in reducing the rate of recurrent infections both in children and disease-prone patients.

• Pidotimod is also able to increase the concentration of salivary IgA directed against bacteria; furthermore, it can modulate airway epithelial cells functions up-regulating the expression of toll-like receptors and acting on adhesion molecules.

• Finally it seems that the compound could affect T-lymphocytes balance with a possible additional anti-allergic activity.