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Host Defence Peptides
Technology
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Technology
Bacterial resistance to antibiotics and the unmet need for an efficacious first-line treatment option against respiratory viral pandemics, are both major and fast-growing health concerns
- Pharma Holdings is addressing the problem with its drugs derived from host defence peptides through Structure-activity-relationship studies on Lactoferricin.
- Pharma Holdings’ infection drug candidate degrades the membranes of microorganisms
- Pharma Holdings’ current focus is on topical treatment of both bacterial and viral infections
- Pharma Holdings’ infection drug candidate has proven efficacy against a broad range of resistant organisms
LTX-109 highlights:
- A synthetic protein fragment; a peptidomimetic
- High stability against degradation
- Produced in large scale
- Low propensity for resistance development
- Rapid mode of action
- Broad spectrum antimicrobial activity
LTX-109 is an investigational antimicrobial drug with a novel membrane-lysing mechanism of action, based on the biological principle of innate immune effectors, lytic peptides. The drug has a rapid bactericidal lytic activity. The drug has been tested in vitro and in vivo models and has undergone a comprehensive nonclinical safety and toxicology program and been studied in man in a Phase I study, three Phase I/IIa studies and one Phase II study.
LTX-109 structure
Mechanism of Action
The antimicrobial mode of action of LTX-109 represents a highly beneficial strategy for killing bacteria and other microorganisms. The diagram below demonstrates the mechanism of action for LTX-109 lysis of bacterial membranes.
The Shai-Matsuzaki-Huang model of the
mechanism of action of an antimicrobial peptide
Transient pores form at this stage. d, Transport of lipids and peptides into the inner leaflet. e, Diffusion of peptides onto intracellular targets (in some cases). f, Collapse of the membrane into fragments and physical disruption of the target cell’s membrane. Lipids with yellow headgroups are acidic, or negatively charged. Lipids with black headgroups have no net charge.
From the following article:
Antimicrobial peptides of multicellular organisms
Michael Zasloff
Nature 415, 389-395(24 January 2002)
doi:10.1038/415389a
Shai-Matsuzaki-Huang model
Zasloff M., Nature, 415,389-395
Pre-clinical highlights
Promising in Vitro results against respiratory viruses
Testing the virucidal activity of a 3% LTX-109 formulation against
respiratory viruses after 1 hour contact time
- The aim of these studies were to determine the antiviral activity of a 3% formulation of LTX-109 against Influenza A virus (IAV), SARS-CoV2 and
Respiratory Syncytial Virus (RSV) after 60 seconds contact time. - The viruses were incubated with 3% LTX-109 for 60 seconds, and the virucidal activity was determined by quantifying the number of infectious virus units
recovered. - Upon treatment with 3% LTX-109, over 2 log reduction in virus infectivity was detected compared to a negative control, after 60 seconds contact time. Cytotoxicity was observed under some conditions but did not affect the validity
of the assays. - Under the conditions tested, 3% LTX-109 displays a virucidal activity of over 2 log, corresponding to over 99% reduction of RSV infectivity and SARS-CoV2 infectivity, and a virucidal activity of over 1 log, corresponding to over 90% reduction of IAV infectivity.
Results of in vitro testing of the virucidal activity of a 1% LTX-109 formulation against respiratory viruses after 1 hour contact time and 3% LTX-109 for 60 sec.
Results of in vitro testing of the virucidal activity of a 1% LTX-109 formulation against respiratory viruses after 1 hour contact time and 3% LTX-109 for 60 sec.
Figure 1: The image shows Lenti-virus-like particles with intact envelope (control) and dissolved envelope (1% LTX-109). Top pictures before treatment and bottom pictures after treatment.
Figure 2: The loss of morphological integrity of the virus post-treatment with LTX-109, note the capsid of the virion without envelope. Top pictures before treatment and bottom pictures after treatment.
Lack of resistance development
In general, compounds acting on the cell membrane have low propensity for developing resistance. The occurrence of spontaneous resistance towards LTX-109 was studied by plating a heterogeneous mixture of bacteria. Neither in wild type, MRSA nor glycol peptide resistant Staphylococcus aureus was any spontaneous resistance observed. No signs of resistance development have been observed in 5 different Staphylococcus aureus strains after 14 passages
MIC Value
The minimum inhibitory concentration (MIC) is the lowest concentration of a chemical, usually a drug, which prevents visible growth of a bacterium or bacteria. MIC depends on the microorganism, the affected human being (in vivo only), and the antibiotic or antimicrobial peptide itself.
LTX-109 kills MRSA better than ‘Gold Standard’ drugs
- Murine skin infection model (tape-stripping, ATCC 33591)
- Read-out: bact growth +9 hours after teatment
- Retapamulin is a topical antibiotic developed by GlaxoSmithKline. It is the first drug in the new class of pleuromutilin antibiotics to be approved for human use. It is marketed as an ointment under the brand names Altabax and Altargo.
- Retapamulin was approved by the United States Food and Drug Administration in April 2007 for the treatment of bacterial skin infections such as impetigo. In May 2007, retapamulin received approval in the EU from the European Medicines Agency for the same indication.
- Fusidic acid is an antibiotic, derived from the fungus Fusidium coccineum and was developed by Leo Pharma in Ballerup, Denmark and released for clinical use in the 1960s.
Outstanding efficiency in animal models
In vivo efficacy has been investigated at Statens Serum Institut (SSI) in Copenhagen, Denmark, and NAEJA, Canada, using a well-documented skin infection model.
A skin lesion is infected on day one, and the infection allowed to develop for 24 hours. The drug and comparators are applied 3 times in a single day. LTX-109 is strongly bactericidal against all strains of Staphylococci tested, including both hospital-acquired and community-acquired MRSA (Strain USA 300).
A biofilm can be described as a microbial colony encased in a polysaccharide matrix which can become attached to a wound surface. This can affect the healing potential of chronic wounds due to the production of destructive enzymes and toxins which can promote a chronic inflammatory state within the wound. Biofilms can be polymicrobial and can result in delayed wound healing and chronic wound infection resistant to antibiotics, leading to prolonged hospitalisation for some patients. There appears to be a correlation between biofilms and non-healing in chronic wounds. It is suggested that biofilms are a major player in the chronicity of wounds
Green indicate living cells and red indicate dead cells
LTX-109 demonstrates a significant effect on MRSA wound biofilm
Based on the results of the first in vivo study, a second study was performed during which treatments were applied either once or twice daily. A lower concentration (1%) LTX-109 gel was also included. Dose-dependent changes in the appearance of wounds treated with LTX-109 formulations were observed. Two samples were taken from each group for histolopathology.
Microbiological analysis of the treatment groups revealed substantial biofilm efficacy for LTX-109 even with only once daily treatment. Relative to control, median log reductions in bioburden of > 4 log cfu/g were achieved with a single daily application at 1- 2% concentrations, and log reductions of ~6 for treatment daily with 5% LTX-109 or twice daily with 2% LTX-109. Mupirocin applied twice daily resulted in a median log reduction of 1.9. Placebo had a limited effect of 0.5. All LTX-109 gels were statistically significant in comparison to placebo. Due to the scatter and the removal of some samples for histology, only 2% LTX-109 applied twice daily achieved statistical significance compared to mupirocin, but the median value for mupirocin was within the 1-2 log reduction range typically observed.
LTX-109 reduces CA-MRSA (USA300) significantly faster than Mupirocin
Infection was established after tape-stripping and scalpel blade-cut injury of skin followed by inoculation with suspensions of community acquired MRSA (CA-MRSA) (USA 300). Treatment was performed TID (three times daily) at 3-hour intervals with 2 % LTX-109, 2 % mupirocin or placebo for either 1, 3 or 5 days. Bacterial tissue load was determined by colony forming units (CFU) from skin biopsy samples. The effect of LTX-109 on reducing bacterial tissue load was significantly better after 1 and 3 days treatment compared to mupirocin, indicating a faster resolution of the infection with LTX-109. The present model demonstrates the potential utility of LTX-109 as fast acting bactericidal drug in treatment of skin infections, even those caused by MRSA.
- A study investigated the fungicidal activity of LTX-109 by determining minimum fungicidal concentration (MFC) and by Time-Kill Kinetics.
- Amphotericin B is an antifungal medication used for serious fungal infections (which due to its extensive side effects, it is often reserved for severe infections
in critically ill patients) - Results shows Time-Kill Curves, measures as percentages survival of 3 different fungus (log10 % survival after 24 hours)
- The study concludes that LTX-109 is as fungicidal as Amphotericin B in Time-Kill experiments, an indicate that LTX-109 has a potential as treatment for fungicidal infections
LTX-109 is rapidly bactericidal
The onset of LTX-109 activity is rapid and dependent on concentration. Increased concentrations of up to 8x MIC (minimum inhibitory concentration) for LTX-109, compared to Vancomycin and Dicloxacillin against Staphylococcus Aureus and compared to Penicillin and Erythromycin against Streptococcus Pyrogenes, showed that the bactericidal effect of LTX-109 is dependent on concentration with a faster effect at higher concentrations, compared to Vancomycin, Dicloxacillin, Penicillin or Erythromycin.
LTX-109 acting on Staphylococcus aureus bacteria; Live (green) and dead (red). Time lapsing inminutes