Microbes in Cancer treatment

1. MEDICAL & PHARMACEUTICAL MICROBIOLOGY– MSDB6002
Course:
M.Sc. Microbiology
Course Instructor:
Dr. Mythily S.

2. Presented By:
SHIVAJI MISRA

4. INTRODUCTION
• Cancer is a group of diseases involving
abnormal cell growth with the potential to
invade or spread to other parts of the body.
• The cancerous cells repeatedly divide,
displacing normal tissue.
• The cancer or neoplasm may be either
benign or malignant: a benign cancer stays
confined to the tissue of origin, while
malignant cancer can spread to other
organs.
Cells showing metastasis

5. HISTORY
• The oldest descriptions of cancer date
back to Egypt, at about 1600 BC.
• Edwin Smith Papyrus describes eight
cases of what appears to be breast
cancer.
• The tumors of the breast were treated
by cauterization, with a tool called
"the fire drill."
Edwin Smith Papyrus

6. Cauterization is done with a tool called the fire drill.
In Left : Modern Tool
In Right: Traditional Tool

7. • The first occupational association with cancer was
noted in 1700 with the observation that nuns had
an elevated incidence of breast cancer.
• In 1775, the English physician and surgeon
Percivall Pott made the observation that exposure
to soot might explain the high incidence of
scrotum cancer in chimney sweeps.
• This was the first indication that exposure to
chemicals, in this case a complex mixture, could
cause cancer.
Percivall Pott

8. YEAR CANCER TYPE CAUSE
1775 Scrotal Cancer Soot
1822 Skin Cancer Arsenic
1879 Lung Cancer Uranium Mining
1895 Bladder Cancer Aniline Dyes
1902 Skin Cancer X-RAY
1908 Leukemia Filterable agents
1914 Experimental induction of skin cancers Coal Tar
1928 Experimental induction of skin cancers UV Light

10. INTRODUCTION:
According to the National Cancer institute, the most common types of
cancer are:
• Bladder, lung, melanoma, leukemia, kidney cancer, lymphoma, prostate,
colon & rectal, thyroid, pancreatic, breast and endometrial.
• Among these, lung, breast, prostate and colorectal were the most
common types of cancers in adult human beings.

11. GENERAL TYPES OF TUMORS:
Benign tumor:
• Benign tumors are not cancer.
• Grow only in one place.
• Do not spread.
Malignant tumor:
• Cancerous tumor.
• Abnormal cell division without control.
• Invades nearby tissues.

12. LUNG CANCER:
Causes:
• Smoking and secondhand smoke.
• Exposure to toxins.
• Family history.
Symptoms:
• Cough(often with blood).
• Chest pain and wheezing.
• Weight loss.

13. BREAST CANCER:
Causes (or rather risk factors):
• Family history.
• Being female(although rare cases of breast cancer
can also be diagnosed in men as well).
• Increasing age.
Symptoms:
• Lump in breast.
• Bloody dischrage from the nipple.
• Change in shape or texture of the breast or
nipple.

14. PROSTATE CANCER:
Causes of prostate cancer are unclear. It
occurs in the prostate gland.
Symptoms:
• Difficulty with urination.
• Sudden erectile dysfuntion.
• Blood in urine.

15. COLON AND RECTAL(COLORECTAL):
Causes:
• Family history.
• Older age.
• A sendetary lifestyle.
Symptoms:
• Rectal bleeding.
• Change in bowel habits.
• Abdominal discomfort.

16. CLASSIFICATIONS:
1. Carcinoma.
2. Sarcoma.
3. Melanoma.
4. Lymphoma.
5. Myeloma.
6. Central and nervous system cancers.

18. INTRODUCTION
• Cancer is a source of significant and growing mortality worldwide, with
an increase to 19.3 million new cancer cases per year projected for 2025.
• More than half of cancer cases and mortality occur in low- and middle-
income countries, and these proportions are expected to increase by
2025 .
• Current treatments for cancer include surgery, radiotherapy and systemic
treatments comprising cytotoxic chemotherapy, hormonal therapy,
immunotherapy, and targeted therapies.

19. THERAPUTIC APPROACH TOWARDS CANCER

20. SURGERY
• Surgery is used to treat many types of
cancer. It works best for solid tumors that
are contained in one area.
• Surgery, when used to treat cancer, is a
procedure in which a surgeon removes
cancer from your body.
• Surgeons are medical doctors with special
training in surgery.

21. HOW SURGERY WORKS AGAINST CANCER
• Remove the entire tumor
Surgery removes cancer that is contained in one area.
• Debulk a tumor
Surgery removes some, but not all, of a cancer tumor. Debulking is used when removing an entire
tumor might damage an organ or the body. Removing part of a tumor can help other treatments
work better.
• Ease cancer symptoms
Surgery is used to remove tumors that are causing pain or pressure.

22. RADIATION THERAPY
• Radiation therapy (also called radiotherapy)
is a cancer treatment that uses
high doses of radiation to kill
cancer cells and shrink tumors.
• At low doses, radiation is used in x-rays to
see inside the body, as with x-rays of patient
teeth or broken bones.
• At high doses, radiation therapy kills cancer
cells or slows their growth by damaging
their DNA.
• Cancer cells whose DNA is damaged
beyond repair stop dividing or die. When
the damaged cells die, they are broken
down and removed by the body.

23. • Radiation therapy does not kill cancer cells right away.
• It takes days or weeks of treatment before DNA is damaged
enough for cancer cells to die.
• Then, cancer cells keep dying for weeks or months after
radiation therapy ends.

24. • The type of radiation therapy that may have depends on many factors,
including:
1. The type of cancer
2. The size of the tumor
3. The tumor’s location in the body
4. How close the tumor is to normal tissues that are sensitive to radiation.
5. General health and medical history of patient
6. Whether patient has other types of cancer treatment.
7. Other factors, such as age and other medical conditions.

25. SIDE EFFECTS OF RADIOTHERAPY
• Radiation not only kills or slows the growth of cancer cells, it can also
affect nearby healthy cells.
• Damage to healthy cells can cause side effects.

26. CHEMOTHERAPY
• Chemotherapy (also called chemo) is a type
of cancer treatment that uses drugs to kill
cancer cells.
• Chemotherapy works by stopping or
slowing the growth of cancer cells, which
grow and divide quickly.
• Chemotherapy is used to:
 Treat cancer
Chemotherapy can be used to cure cancer,
lessen the chance it will return, or stop or
slow its growth.
 Ease cancer symptoms
Chemotherapy can be used to shrink
tumors that are causing pain and other
problems.

27. HOW CHEMOTHERAPY IS USED WITH OTHER CANCER TREATMENTS
When used with other treatments, chemotherapy can:
• Make a tumor smaller before surgery or radiation therapy. This is called adjuvant
chemotherapy.
• Help other treatments work better.
• Kill cancer cells that have returned or spread to other parts of your body.

29. SIDE EFFECT OF CHEMOTHERAPY
• Chemotherapy not only kills fast-growing cancer cells, but also kills or slows the growth
of healthy cells that grow and divide quickly.
• Examples are cells that line in the mouth and intestines and those that cause hair to
grow.
• Damage to healthy cells may cause side effects, such as mouth sores, nausea, and hair
loss.

30. IMMUNOTHERAPY
• Immunotherapy is a type of cancer treatment that helps
your immune system fight cancer.
• The immune system helps the body fight infections and other
diseases. It is made up of white blood cells
and organs and tissues of the lymph system.
• Immunotherapy is a type of biological therapy.
• Biological therapy is a type of treatment that uses substances
made from living organisms to treat cancer.

31. HOW DOES IMMUNOTHERAPY WORK AGAINST CANCER?
• As part of its normal function, the immune system detects and
destroys abnormal cells and most likely prevents or curbs the
growth of many cancers.
• For instance, immune cells are sometimes found in and around
tumors. These cells, called tumor-infiltrating lymphocytes or TILs,
are a sign that the immune system is responding to the tumor.
• People whose tumors contain TILs often do better than people
whose tumors don’t contain them.

33. • Intrusion of microorganisms into the body leads to the activation of immune mechanisms.
• It is assumed that the “mobilized” immune system, by intentionally introducing microorganisms
into the oncological patient, is able to at least limit the development of cancer.
• This is a method in which microbes indirectly lead to cancer regression—especially in those in
whom other commonly used treatments have failed.
• The safety of the used microorganisms is extremely important, because the aim of the therapy is
to fight cancer, not to harm the patient’s organism by infecting it with a pathogen.
INTRODUCTION

34. Bacteria can be applied in various forms for therapeutic purposes.
• Apart from whole, living attenuated cells, we can use genetically engineered bacteria expressing
particularly desirable factors.
• Microorganisms are also applied as vectors, which are carriers of specific antineoplastic agents (e.g.,
chemotherapeutics) or enzymes useful in cancer cell destruction.
• The use of bacteria as a vector to transfer a chemotherapeutic agent directly into the tumor allows a
significant reduction of the side effects of treatment that usually accompany traditional
chemotherapy.
• In addition, there is a therapeutic potential in using bacterial secretion products, for example, toxins.
Their presence in the tumor environment could have destructive effect on cancer cells.
• The use of sporangial bacteria, which can survive under unfavorable environmental conditions,
represents another approach, which has been applied with Clostridium novyi.
APPLICATION FORMS

35. • The beginnings of the use of microbes in cancer therapy date back to
the nineteenth century.
• Dr. William Coley (1862–1936) developed a mixture of bacterial
microbes and, for the first time in modern medicine, he successfully
treated certain types of cancer, thus becoming the father of
immunotherapy.
• He was employed at the New York Cancer Hospital and then at the
Hospital for Special Surgery in New York, as a surgeon specializing in
sarcoma, especially bone cancer.
• He was deeply shocked when one of his first oncological patients died,
and that was a reason he began seeking more effective forms of cancer
treatment.
• Coley studied in-depth the case report forms of his contemporary and
much earlier oncological patients. He came across information on
spontaneous regression of sarcoma in patients with severe bacterial
infection.
HISTORY
Dr. William Coley

36. • He worked on new treatment method for the next forty years,
preparing other variants of microbial mixture.
• This preparation could be called a vaccine because it
stimulated/activated the immune system by the introduction of
antigens (bacterial components).
• Twenty different versions of the vaccine (called the Coley’s toxin)
were devised at that time, and each of them had a different
effectiveness.
• It is also very important that the microbes were administered in
different ways: intramuscularly, intravenously, or directly into the
tumor.
• Coley’s toxin was given to hundreds of patients, and more than a
quarter of them were cured.
• After Coley’s death, due to the lack of systematic and precise
documentation on the research methodology and preparation of
the vaccines, such spectacular results were not replicated.

37. • The antitumor efficacy of microorganisms is extremely diverse.
• Results of clinical trials allow determining whether a particular product can be
intended for general use.
• Currently used anticancer bacterial microbial preparations have the status of a
therapy complementary to standard treatment, increasing the patient’s chances of
complete recovery.
BACTERIA USED AS ANTICANCER AGENTS

38. • Bacillus Calmette-Guerin (BCG) is a strain of Mycobacterium bovis developed by Albert
Calmette and Camille Guerin as a tuberculosis vaccine and has been used since 1921.
• In many countries, this vaccine has been induced in the mandatory vaccination schedule
and is administered to children within 24 hours after birth, in a single dose, intradermally.
• Mycobacterium bovis is an etiological agent of bovine tuberculosis. However, in certain
circumstances (e.g., after ingestion of untreated milk from an infected animal), it can
cause tuberculosis symptoms in humans as well.
• Calmette and Guerin have sub-cultured M. bovis (231 sub-culture in total) for 13 years on
a medium consisting mainly of cooked potato slices soaked in ox bile and glycerin. Only
then did it become safe for human use, as an avirulent but immunogenic strain.
MYCOBACTERIUM BOVIS BCG

40. • At the beginning of the twentieth century there were some links between the occurrence of
tuberculosis and cancer regression.
• However, only after Morales and his colleagues demonstrated in 1976 that the use of BCG
was accompanied with the cancer regression, the vaccine was approved as the
complementary treatment of bladder cancer.
• Treatment of this type of cancer with the M. bovis BCG strain requires the intravesical
infusion of the microbial suspension using urethral catheters.
• This therapy is most often used after resection to eliminate accurately the cancer cells and to
prevent recurrence.
• The dose and duration of treatment are strictly dependent on the stage of cancer.
• Clinical observations show that recurrence is much less likely to occur after tumor resection
or resection and chemotherapy when BCG is administered intravesically .

42. • BCG’s mechanism of action is based on stimulating the patient’s immune system.
• It appears that IFN-γ and effector cells, that is, CD4+ and CD8+ lymphocytes, play an
extremely important role in the recognition of tumor antigens.
• In addition, the pool of proinflammatory cytokines is increasing, which enhances the
immune response of the body by activating the phagocytosis of cancer cells.
• Providing the selected vitamins during therapy may increase the survival of M.
bovis BCG cells, which improves the quality of therapy.
MECHANISM OF ACTION

43. • Streptococcus pyogenes was originally used in the treatment of bone sarcoma by Dr. William Coley.
• Presently, the S. pyogenes OK-432 strain has been used in that way in many countries around the world.
• Lymphangiomas are tumors formed by excessive division of lymphatic vessels’ endothelial cells.
• They are most often found in the head and neck area of children under the age of two.
• The pathological development of lymphatic vessels is primarily associated with impaired lymph flow, which in
turn manifests itself in the formation of cysts.
STREPTOCOCCUS PYOGENES OK-432

44. Streptococcus pyogenes OK432
Lymphangioma
Changes in children resemble goiter, which is
associated with an enlarged thyroid gland.

45. • Treatment primarily involves surgical removal of the cyst, but this is not an easy task, and is often
burdened with numerous adverse effects, including death.
• An alternative and safer method of treatment is sclerotherapy.
• Streptococcus pyogenes OK-432 is injected into pathologically changed lymphatic vessels.
• In Japan, this microorganism has been successfully used in the treatment of lymphangiomas in
children since 1987.
• Studies show that the strain is safe and results in at least 50% reduction of cyst volume.

46. Sclerotherapy of Lymphangioma.
Intralesional injections of sclerosing agents such as 25%dextrose, hypertonic saline,
bleomycin, aethoxysklerol, or OK432 are recommended for the treatment of
Lymphangioma.

47. • The mechanism of action of the microorganism is also based on the sensitization of the
immune system.
• Activated cells destroy the neoplasm, further growth is inhibited, and the lymphangioma is
reduced.
• Studies using flow cytometry have shown that the first day after suspension administration,
the numbers of neutrophils and macrophages, as well as lymphocytes, rapidly increase. NK
CD56+ cells, TNFα, IL-6, IL-8, IFNγ, and VEGF (vascular endothelial growth factor) levels also
increase.
• Due to the appearance of inflammation immediately after the procedure, the lesion may be
swollen, but therapeutic effects are noticeable after a few months.
• Moreover, studies conducted in the years 2005–2015 showed the great effectiveness of this
strain also in the treatment of intraoral ranula. Complete regression occurred in 78.2% of
patients
MECHANISM OF ACTION

49. • Obligate anaerobes and facultative anaerobes have potential to
be used in anticancer therapies because they grow best under
conditions hypoxia.
• Oxygen is delivered to the cells through blood vessels which
penetrate mainly the tumor surface area.
• That results in impaired diffusion of oxygen into the tumor and
hypoxia.
• The anaerobic environment creates favorable conditions for the
development of anaerobic bacteria.
• The greatest advantage of using this microorganisms is that they
locate directly inside the tumor.
CLOSTRIDIUM NOVYI
CLOSTRIDIUM NOVYI

50. • Bacteria develop in the tumor’s necrotic areas and can directly damage tumor cells.
• The history of the use of Clostridium in the fight against cancer dates back to 1935, when Connell
published an article describing the regression of advanced cancer under the influence of enzymes
produced by Clostridium histolyticum.
• The attenuated strain of Clostridium novyi-NT has positively undergone phase I and phase II
clinical trials, giving extremely promising results for the treatment of leiomyoma.
• The mechanism of the anticancer activity of Clostridium spp. is unknown yet, but it is common
knowledge that bacterium is capable of producing specific enzymes and toxins that destroy cancer
cells.
• It also produces specific proteins that can be conjugated to specific chemotherapeutics which
allows the drug to enter the tumor.
• In traditional chemotherapy, drugs are not able to penetrate into the tumor precisely due to its
external vascularization and internal hypoxia.

51. A leiomyoma, also known as
fibroids, is a benign smooth
muscle tumor that very rarely
becomes cancer (0.1%). They
can occur in any organ, but the
most common forms occur in the
uterus, small bowel, and the
esophagus.

52. • Salmonella enterica serovar Typhimurium, an
etiological agent of typhoid fever, shows similar
features as Clostridium.
• It is a relatively anaerobic rod that can also be located
in the necrotic tumor regions.
• In the treatment of cancer, the attenuated
strain Salmonella typhimurium VNP20009 is used for
safety reasons.
• Clinical trials on the use of this microorganism for
melanoma treatment began in 2002.
• In addition, the VXM01 antitumor vaccine, which is
based on the attenuated strain of Salmonella typhi,
has successfully passed phase I clinical trials.
• This bacterium has a plasmid-encoding expression of
VEGFR2 (vascular endothelial growth factor receptor-
2).
• The vaccine blocks the angiogenesis process. The
formulation was tested in individuals with pancreatic
cancer
SALMONELLA ENTERICA SEROVAR TYPHIMURIUM
SALMONELLA ENTERICA SEROVAR TYPHIMURIUM
PHOTO: MICROBEWIKI.KENYON.EDU

53. • The most recent anticancer strategies use the achievements of various scientific disciplines,
like, nanobiotechnology.
• Nanoparticles (nanocapsules), lipid vesicles with a chemotherapeutic drug inside, are the
object of growing interest.
• Nanoliposomes are able to deliver the drug inside the tumor.
• However, they are not a perfect solution because many of the particles do not reach the target.
• The tumor is only vascularized from the outside, which makes it impossible for
chemotherapeutics to reach the inside of the lesion.
• Hence, the idea of delivering drugs directly to the tumor with vectors/carriers would allow for
more precise targeting of the cancer site.
• Limiting the spread of the drug only to the tumor area would significantly reduce the adverse
effects of chemotherapy.
RECENT UPDATE

54. • Magnetococcus marinus MC1 is a Gram-negative coccus, which is found in the Atlantic Ocean near
Rhode Island, USA.
• This microorganism has cilia, arranged in two bundles located at one pole, which enable the bacteria
to move.
• The unique feature of this bacterium structure is the presence of magnetosomes—special elements
which are magnetite particles (Fe3O4) surrounded by membranes, forming chains in the cytosol.
• The presence of magnetite orients the bacteria with the Earth’s magnetic field.
• In addition, this microorganism shows negative aerotaxis capacity, that is, prefers an environment
that is poor in oxygen.
• These properties make the Magnetococcus marinus a useful tool to destroy cancer cells.
• Using a powerful magnetic field, the same as in the MRI technique (magnetic resonance imaging), it
would be possible to direct bacteria containing magnetosomes to the site of the tumor.
• The bacteria will be located precisely in the areas of hypoxia, in that case inside the tumor, where
they would deliver a chemotherapeutic encapsulated in nanoliposomes attached to the bacteria
surface.
• Animal studies have shown that approximately 55% of nanoliposomal transmission cells reach tumors
MAGNETOCOCCUS MARINUS

55. Magnetococcus Marinus
Photo: microbewiki.kenyon.edu

56. • Anticancer therapy with the use of microorganisms is often marginalized and neglected.
• There is primarily a risk of developing infection and related consequences, including death.
• In experimental studies, laboratory animals have been used to show that the most effective strain actually
destroyed cancer, but animals died because of infection by pathogens.
• It is very important to ensure the safety of the patients, especially by using only adequately attenuated
microorganisms.
• A perfect balance between the attenuation of a microorganism and its immune stimulatory ability can
guarantee the proper effect.
• Costs associated with clinical trials and the introduction of a new product to the market are extremely
high.
• Legal regulations are also very complicated, due to the not fully known impact of microbes on cancer.
CHALLANGES

58. INTRODUCTION
• Cancer fungotherapy is a promising scientific field, which deals with antitumor substances
derived from mushrooms. It has been an integral part of the world traditional medicine since the
antiquity
• The concept of fungal treatment officially appeared in Traditional Chinese Medicine and can be
dated back to several thousand years ago.
• The ancient Chinese pharmacopoeia included hundreds of herbal and fungal species - the latter
were considered to be the most effective natural remedies for various types of tumors.
• In other countries of East and Southeast Asia, mushrooms were also highly valued and rated as
“beneficial to health” for centuries. Plant and fungal products were also widespread in Russia,
representing the main medicinal resources until the 18th century.

59. WHAT ARE MEDICINAL MUSHROOMS?
• Mushrooms are part of the fungus family and there are hundreds
of different species. They have been a part of traditional chinese medicine
for centuries and are used to treat illness. They are also known as medicinal
mushrooms.
• Some species of mushroom are not edible.
• Research has looked at different types of mushroom and mushroom extracts
or compounds to see if they can prevent or stop the growth of cancer cells.

60. WHY PEOPLE WITH CANCER USE IT?
• Mushrooms are used in Japan and China to treat lung diseases and are
sometimes given alongside cancer treatment. Research is looking at whether
mushrooms can help the immune system.
• It is thought that some of the chemical compounds in mushrooms might
strengthen the immune system. If they do, researchers wonder if this could
help fight cancer cells.
• Button mushrooms and flat mushrooms are commonly eaten in the UK. They
contain all the essential amino acids and are a good source of vitamins. So
they can be used as part of a healthy diet. But there is no evidence that they
can treat cancer.
• In the UK, powdered shiitake, maitaki and reishi (also called ganoderma)
mushrooms are available. You can also sometimes get preparations of their
juices. These are sold in health food shops.
• As far as we know, there is nothing in the mushrooms or compounds that
would be harmful but we don't currently know how helpful they are in cancer
care.

61. FUNGUS USED FOR CANCER THERAPHY
• The fields of cancer fungotherapy and of search for novel antitumor agents are by far
not limited to these species; however, these four can serve as typical representatives of
widespread medicinal mushrooms used both in traditional medicine and in modern
biomedical research.
• They belong to three different orders, and are a rich source of bioactive compounds
such as polyphenols, polysaccharides, glucans, terpenoids, steroids, cerebrosides and
proteins, which can be used for treatment of various cancers.
• These Basiodiomycota mushrooms are:
• Fomitopsis pinicola
• Hericium erinaceus
• Trametes versicolor
• Inonotus obliquus

62. FOMITOPSIS PINICOLA
• Fomitopsis pinicola, class Agaricomycetes, order Polyporales, family Fomitopsidaceae (common
name Red Belted Conk) is a brown rot fungus, a member of Basidiomycota. It is saprotrophic on
the dead wood of coniferous and broad-leaved trees, which are common throughout the
temperate Northern Hemisphere.
• F. pinicola fruiting bodies, which are considered to be nontoxic mushrooms in Europe, have been
used in Korean folk medicine as hemostatic and anti-inflammation agents.
• F. pinicola is known to contain a variety of primary metabolites (such as polysaccharides) and
secondary metabolites (such as triterpenes, esters, lactones and steroids).
• Recently, more studies have been performed on F. pinicola extracts in a search for more specific
anticancer activity.
• Chloroform extracts of the mushroom have demonstrated cytotoxicity, which was almost twice
more specific to colorectal cancer cells (SW-480) than to control HEK293 cells. The cytotoxic effect
took place through the ROS-mediated apoptotic mechanism.

63. • Moreover, the extracts were able to inhibit migration of the SW-480 cells in scratch wound and
transwell assays by means of downregulation of matrix metalloproteinases.
• Another study has revealed the potential of F. pinicola ethanol extracts not only to induce apoptosis
in various human and murine cancer cell lines, but also to significantly inhibit xenograft sarcoma-
derived tumor growth in mice, along with prolongation of their survival time and absence of severe
side effects, when given as a food supplement.
• Interestingly, combined treatment of mice with the extract and a common chemotherapeutic agent
cisplatin gave a synergistic effect on slowing down the tumor growth. Taken together, these findings
provide a stronger evidence that apart from the unspecific cytotoxic compounds, F. pinicola contains
substances possessing specific anti-oncogenic potential, probably acting through induction of
apoptosis.
• Regarding the fact that F. pinicola is known as an anticancer agent in the Chinese folk medicine, we
can conclude that this mushroom is of a certain interest for modern drug discovery as a potential
source of novel anticancer compounds, which are yet to be characterized.

64. THE ANTICANCER PROPERTIES
OF FOMITOPSIS PINICOLA
EFFECTS OF DIFFERENT
MUSHROOM DERIVATIVES
AND THEIR MECHANISMS OF
ACTIONS ON VARIOUS
MODELS ARE DEPICTED.
MOUSE AND CELL ICONS
INDICATE RESULTS OBTAINED
ON ANIMAL AND CELL
MODELS, RESPECTIVELY. ROS –
REACTIVE OXYGEN SPECIES,
MMP↓ – DOWNREGULATION
OF MATRIX
METALLOPROTEINASES.

65. HERICIUM ERINACEUS
• Hericium erinaceus, class Agaricomycetes, order Russulales, family Hericiaceae, is an edible medicinal
mushroom.
• It is also known under the name “Lion's mane” in English, “Yamabushitake” in Japan or “ Houtougu ” in
China.
• The mushroom is considered a saprotroph or a weak parasite. It is found on oak (Quercus) and beech
(Fagus) in Europe, North America, Japan, Russia, and China.
• H. erinaceus has attracted special scientific attention in recent four years, being intensively studied in
terms of its primary and secondary metabolites and their possible medicinal use.
• Therapeutic and anti-carcinogenic application of H. erinaceus is its salutary influence on the digestive
organs, including stomach, liver, intestine and colon. Water and ethanol extracts of the mushroom have
demonstrated growth inhibitory effects on gastric (NCI-87), liver (HepG2 and Huh-7), and colon (HT-29)
cancer cell lines in the MTT proliferation assay, with the highest efficacy against Huh-7 cells (IC50 of 0.8
mg/ml for the dried extract).

66. • Although not comparing these results with non-cancer cell lines of the respective tissues, the same study
describes efficient application of the extracts against xenograft tumors formed by aforementioned cancer cell
lines in SCID mice. The extracts, given orally, have demonstrated a tumor suppressing activity similar to that
of 5-fluoruracil, a most widely used drug clinically applied for the treatment of gastrointestinal cancers, but
demonstrated a much lower general toxicity than 5-fluoruracil.
• Another study shows that water extracts of H. erinaceus, given as a food supplement, possess an anti-
metastatic activity, strongly inhibiting the migration of CT-26 murine colon carcinoma cells to lungs after
intravenous injection into BALB/c mice, reducing the formation of tumor nodules in the lung by about 50%,
and preventing metastasis-caused increase in the lung weight. The mechanism of action involves suppression
of matrix melalloproteinases 2 and 9, as well as suppression of ERK and JNK kinase activation, also decreasing
the general tumor cell viability.
• Further research on tumor suppressing activity of the H. erinaceus extracts allowed to reveal the possible
spectrum of their action modes. Studies on CT-26 derived human colon cancer xenograft tumors in mice have
shown a significant reduction in tumor growth after treatment by H. erinaceus water extracts. It has been
demonstrated that the extracts stimulated the activities of natural killer cells and macrophages on one hand
and blocked angiogenesis on the other.

67. • Efforts to study anticancer effects of individual compounds isolated from H. erinaceus have produced
impressive results. Cyanthine diterpenoid Erinacine A, a mycelial derivative of H. erinaceus, has
demonstrated growth-inhibitory activities against different cancer cell lines and tumors related to the
digestive tract.
• H. erinaceus is not merely a source of low molecular weight biologically active compounds, but also of
some proteins that possess potential tumor-suppressive activities. Indeed, a Hericium-derived protein
HEP3, which demonstrated a complex immunomodulatory impact in mice, has also been able to strongly
reduce growth of CC531 cell xenograft tumors after intraperitoneal injection. The immunomodulatory
effect was induced through stimulation of the gut microbiota with the protein and involved activation of
the proliferation and differentiation of T-cells and stimulation of the intestinal antigen-presenting cells.
• Another example of bioactive protein from the same mushroom is a glycoprotein HEG-5 that was able to
induce apoptosis in a gastric cancer cell line SGC-7901, stimulating the expression of pro-apoptotic
factors such as p53, Bax, Caspase 8 and Caspase 3.

68. THE ANTICANCER
PROPERTIES OF HERICIUM
ERINACEUS
EFFECTS OF DIFFERENT
MUSHROOM DERIVATIVES AND
THEIR MECHANISMS OF
ACTIONS IN VARIOUS MODELS
ARE DEPICTED. MOUSE AND
CELL ICONS INDICATE
RESULTS OBTAINED ON
ANIMAL AND CELL MODELS,
RESPECTIVELY. ARROWS UP
AND DOWN REFLECT UP- OR
DOWN-REGULATION OF
RESPECTIVE PROTEINS OR
PATHWAYS. ROS – REACTIVE
OXYGEN SPECIES, MMP -
MATRIX
METALLOPROTEINASES, DC –
DENDRITIC CELLS, MP –
MACROPHAGES, TC – T-CELLS,
NK – NATURAL KILLERS.
OTHER PROTEINS/PATHWAYS
ARE MENTIONED UNDER
THEIR STANDARD NAMES.

69. CONCLUSIONS
• The complex anticancer potential of medicinal mushrooms may be embodied not only through
inhibition of certain cancer-specific processes or targeted activation of tumor-specific apoptosis,
but also through indirect actions such as immunomodulation.
• The polysaccharide-mediated antitumor immunomodulatory action seems to be rather common
for many medicinal mushrooms and gives a major input into the therapeutic potential of at least
three out of the four reviewed species, which is probably determined by similar carbohydrate
composition and thus similar effects on the immune system of different mushrooms.
• Fomitopsis pinicola, is closer to the initial stages of involvement into modern cancer treatment: it
is known to possess certain anticancer activities, and a set of compounds were isolated, but
experiments on animal models and clinical trials are lacking, as well as precise studies on the
molecular targets and signaling pathways affected by the fungus.

70. • Inonotus obliquus is a better-studied mushroom: here we have more data on mouse
xenograft experiments and more molecular targets, including the Wnt/β-catenin
pathway, a promising target for anticancer drugs of the future, but the medical
relevance is still to be improved by clinical trials.
• Hericium erinaceus and especially Trametes versicolor are much more advanced in
terms of medical applications due to their uncovered strong and complex
immunomodulatory potential provided by rich polysaccharide and proteoglycan
diversity.
• There are numerous clinical trials confirming applicability of these mushrooms and
their extracts as components of modern anticancer chemotherapy.

72. • Oncolytic virus therapy has recently been recognized as a promising new therapeutic approach for
cancer treatment. An oncolytic virus is defined as a genetically engineered or naturally occurring virus
that can selectively replicate in and kill cancer cells without harming the normal tissues.
• In contrast to gene therapy where a virus is used as a mere carrier for transgene delivery, oncolytic
virus therapy uses the virus itself as an active drug reagent.
• The concept of oncolytic virus therapy has existed for some time. Tumor regression has often been
observed during or after a naturally acquired, systemic viral infection.
• In 1949, 22 patients with Hodgkin’s disease were treated with sera or tissue extracts containing
hepatitis virus. Between 1950 and 1980, many clinical trials were performed in attempts to treat cancer
with wild type or naturally attenuated viruses, including hepatitis.
• West Nile fever, yellow fever, dengue fever and adenoviruses. However, these viruses were not deemed
useful as therapeutics reagents because, in those days, there was no known method to control the
virulence and yet retain viral replication in cancer cells.
INTRODUCTION

73. • It is now recognized, because protection mechanisms against viral infection (e.g. interferon-beta
signal pathway) are impaired in the majority of cancer cells, that most viruses can replicate to a
much greater extent in cancer cells than in normal cells.
• Attempts to achieve cancer cell-specific replication have been undertaken either by selecting a
virus that is non-virulent in humans or by engineering the virus genome.
• The latter strategy is, however, better suited to achieving strict control of viral replication. In
1991, Martuza et al.demonstrated that a genetically engineered herpes simplex virus type I (HSV-
1) with a mutation in the thymidine kinase (TK) gene replicated selectively in cancer cells and was
useful for treating experimental brain tumors.

74. • During the past two decades of thriving development, probably the most important
finding regarding oncolytic virus therapy was that a systemic tumor-specific immunity is
efficiently induced in the course of oncolytic activities.
• This phenomenon is now widely recognized as the common feature for all oncolytic virus
therapy that is expected to play a major role in prolonging the survival of cancer patients.
• To date, two genetically engineered oncolytic viruses have been approved for marketing
as drugs. One is Oncorine , an E1B-deleted adenovirus, which was approved in China for
head and neck cancer and esophagus cancer in 2005. The use and clinical data of
Oncorine is so far limited to China.
• The other is T-Vec, which was approved for melanoma by the FDA in the USA in October
2015 and was subsequently approved in Europe in January 2016 and in Australia in May
2016.

76. GENETICALLY ENGINEERED ONCOLYTIC VIRUSES
T-Vec.
• T-Vec is a double-mutated HSV-1 with deletions in the c34.5 and a47 genes, and the human
granulocyte-macrophage colony-stimulating factor (GM-CSF) gene inserted into the deleted
c34.5 loci.
• Because the c34.5 gene functions to negate the host cell’s shut-off of protein synthesis upon
viral infection, inactivation of c34.5 renders the virus unable to replicate in normal cells.
However, because cancer cells are in defect of the shut-off response, c34.5-deficient HSV-1
can still replicate in cancer cells.
• The a47 gene functions to antagonize the host cell’s transporter associated with antigen
presentation; therefore, the deletion of the gene precludes the downregulation of MHC class
I expression, which should enhance the antitumor immune responses.
• The deletion in the a47 gene also results in immediate early expression of the neighbor US11
gene, which results in enhanced viral replication in cancer cells.

77. G47Δ.
• G47Δ is a triple-mutated third-generation oncolytic HSV-1 that was developed by Todo et
al. by adding another deletion mutation to the genome of G207, a second generation HSV-
1.
• Two of the mutations of G47D are created in the c34.5 and a47 genes, the same genes that
T-Vec utilizes.
• G47Δ further has an insertion of the Escherichia coli LacZ gene inactivating the ICP6 gene.
• Because the immediate-early expression of US11 caused by the deletion within the a47
gene prevents the premature termination of protein synthesis that slows the growth of
c34.5-deficient HSV-1 strains such as G207, G47Δ shows augmented replication capability in
cancer cells, resulting in having a wider therapeutic window than any other oncolytic HSV-1.

78. JX-594.
• JX-594 (pexastimogene devacirepvec, Pexa-Vec) is a genertically engineered vaccinia virus that
has a mutation in the TK gene, conferring cancer cell-selective replication, and an insertion of
the human GM-CSF gene, augmenting the antitumor immune response.
• JX-594 also has a LacZ gene insertion as a marker.
• In a phase I study, intralesional injection of primary or metastatic liver tumors with JX-594 was
generally well tolerated in the context of JX-594 replication, GM-CSF expression and systemic
dissemination.
• Direct hyperbilirubinemia was the dose limiting toxicity. High dose JX-594 was used for a dose
escalation phase I trial to test the feasibility of intravenous delivery.

79. RECENT ADVANCES IN GENETIC MODIFICATION OF ADENOVIRUS VECTORS FOR CANCER TREATMENT
• Adenoviruses are widely used to deliver genes to a variety of cell types and have been used
in a number of clinical trials for gene therapy and oncolytic virotherapy.
• However, several concerns must be addressed for the clinical use of adenovirus vectors.
Selective delivery of a therapeutic gene by adenovirus vectors to target cancer is precluded
by the widespread distribution of the primary cellular receptors.
• Adenoviruses induce strong innate and acquired immunity in vivo.
• Furthermore, several modifications to these vectors are necessary to enhance their
oncolytic activity and ensure patient safety. As such, the adenovirus genome has been
engineered to overcome these problems.

80. • The adenovirus family consists of non-enveloped DNA viruses with a linear genome of
30–38 kb.
• There are 57 adenovirus serotypes, which are classified into categories A–G, based on
viral properties of agglutination.
• Adenovirus vectors based on human serotype 5 of species C are beneficial as gene
delivery vehicles that enable high-titer production and highly efficient gene transfer into
a wide spectrum of dividing and nondividing cells both in vitro and in vivo.
• These vectors have been used in a number of clinical trials utilizing gene therapy and
oncolytic virotherapy. However, several concerns need to be addressed before
widespread clinical use of adenovirus vectors is possible.

81. CANCER-TARGETING ADENOVIRUS VECTORS
The entry of serotype 5 adenovirus into cells requires two distinct and sequential steps.
• To date, several promising approaches have been used for target-specific gene delivery. For
example, CAR expression can decrease with tumor progression.
• Replacing part of the serotype 5 adenovirus capsid with non-serotype 5 adenovirus capsid
(fiber switching) can be used to circumvent CAR deficiency.
• The pseudo-typing of serotype 5 adenovirus with serotype 3 adenovirus fibers was shown
to result in CAR-independent infectivity and enhanced gene transfer to a broad range of
cancer cell types.
• Chimeric serotype 5 vectors with serotype 35 fibers exploit the CD46 receptor for cell
infection, as many cancers express high levels of CD46. As such, a chimeric serotype
3/serotype 11p virus was found have increased selectivity for colon cancer cells.

82. www.wileyonlinelibrary.com/journal/cas

83. www.wileyonlinelibrary.com/journal/cas

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