Neuron point-of-care stem cell therapy (N-POCST): Transplantation of autologous bone marrow-derived stem cells for

traumatic central nervous system disorders, the future of a new

method.

Situation

TRAUMATIC BRAIN INJURY

Each year 235 000 persons in US are hospitalized for nonfatal TBI and 50 000 die

(Corrigan et al. 2010)

The prevalence of US civilian residents living with disability following hospitalization

with TBI is 3.2 million (Corrigan et al. 2010).

Despite claims to the contrary, no clear decrease in TBI-related mortality or

improvement of overall outcome has been observed over the past two decades.

(Roozenbeek et al. 2013)

Corrigan, J.D., Selassie, A.W., Orman, J.A.L., 2010. The epidemiology of traumatic brain injury. J. Head Trauma Rehabil. 25, 72–80. doi:10.1097/HTR.0b013e3181ccc8b4.

Roozenbeek, B., Maas, A.I.R., Menon, D.K., 2013. Changing patterns in the epidemiology of traumatic brain injury. Nat. Rev. Neurol. 9, 231–236.

doi:10.1038/nrneurol.2013.22

Situation

SPINAL CORD INJURY

Every year, between 250 000 and 500 000 people become spinal cord injured. (WHO

2013)

“pi al ord i jur is a medically complex

and life-disrupting condition, notes Dr.

Etienne Krug, Director of the Department

of Violence and Injury Prevention and

Disability, WHO. However, spinal cord

injury is preventable, survivable, and need

not preclude good health and social

inclusion.

http://www.who.int/mediacentre/news/releases/2013/spinal-cord-injury-20131202/en/

Situation

STROKE

Is the leading cause of serious long-term disability. (Mozaffarian et al. 2015)

While I speak 5 persons in US will die consequence of stroke. (Mozaffarian et al.

2015)

Mozaffarian D, Benjamin EJ, Go AS, et al. Heart disease and stroke statistics—2015 update: a report from the American Heart Association.

Circulation. 2015 ;e29-322.

STROKE

SCI

TBI

http://data3.whicdn.com/images/11707674/original.jpg

Current treatment approach

Preventable diseases

Shin, S.S., Dixon, C.E., Okonkwo, D.O., Richardson, R.M., 2014. Neurostimulation for traumatic brain injury. J. Neurosurg. 121, 1219–1231.

doi:10.3171/2014.7.JNS131826

Fehlings, M.G., Vawda, R., 2011. Cellular Treatments for Spinal Cord Injury: The Time is Right for Clinical Trials. Neurotherapeutics 8, 704–720. doi:10.1007/s13311-011-0076-7

Burns, T.C., Verfaillie, C.M., Low, W.C., 2009. Stem Cells for Ischemic Brain Injury: A Critical Review. J. Comp. Neurol. 515, 125–144.

doi:10.1002/cne.22038

TBI: Durable treatments for neurological function deficits following TBI have been

elusive, as there are currently no FDA-approved therapeutic modalities for mitigating

the consequences of TBI. (Shin et al. 2014)

SCI: Despite advances in pre-hospital care, medical and surgical management, and

rehabilitation approaches, many SCI sufferers still experience substantial neurological

disability. (Fehlings et al. 2011)

STROKE: Unfortunately, although a plethora of neuroprotective compounds have

shown promise in animal models, no other treatment to date has shown efficacy in

clinical trials (Burns et al. 2009)

Rationale of aBM-SC

A scanning electron microscope picture of a nerve

ending. It has been broken open to reveal vesicles

(orange and blue) containing chemicals used to pass

messages in the nervous system. TINA CARVALHO

Secrete all kind of factors, which

influence the direct environment of

injured cells:

(Gnecchi et al. 2008, Caplan et al.

2006).

ASC in BM (HSCs and MSCs)

http://publications.nigms.nih.gov/insidethecell/chapter3.html

Induce NEUROPROTECTION,

inflammatory suppression and neural

repair, allowing

reconstruction of totally damaged tissues

or preventing partially damaged cells

from evolving to cell demise.

(Erceg and Stojkovic, 2009)

Gnecchi, M., Zhang, Z., Ni, A., Dzau, V.J., 2008. Paracrine mechanisms in adult stem cell signaling and therapy. Circ. Res. 103, 1204–1219.

doi:10.1161/CIRCRESAHA.108.176826

Caplan, A.I., Dennis, J.E., 2006. Mesenchymal stem cells as trophic mediators. J. Cell. Biochem. 98, 1076–1084. doi:10.1002/jcb.20886

Erceg, S.M.R. and M. Stojkovic, 2009. Human embryonic stem cell differentiation toward regional specific neural precursors. Stem Cells, 1: 78-87. DOI:

10.1634/stemcells.2008-0543

Rationale of aBM-SC

Zhang H, Zhang N, Li M, Feng H, Jin W, Zhao H, Chen X, Tian L (2008) Therapeutic angiogenesis of bone marrow mononuclear cells (MNCs) and peripheral blood MNCs:

transplantation for ischemic hindlimb. Ann Vasc Surg 22:238–247

Zeng, R., Wang, L.-W., Hu, Z.-B., Guo, W.-T., Wei, J.-S., Lin, H., Sun, X., Chen, L.-X., Yang, L.-J., 2011. Differentiation of human bone marrow mesenchymal stem cells into

neuron-like cells in vitro. Spine 36, 997–1005. doi:10.1097/BRS.0b013e3181eab76

Bareyre, F.M., 2008. Neuronal repair and replacement in spinal cord injury. J. Neurol. Sci. 265, 63–72. doi:10.1016/j.jns.2007.05.004

Kucia, M., Reca, R., Jala, V.R., Dawn, B., Ratajczak, J., Ratajczak, M.Z., 2005. Bone marrow as a home of heterogenous populations of nonhematopoietic stem cells. Leukemia

19, 1118–1127. doi:10.1038/sj.leu.2403796

aBM-SC are easy to harvest from the iliac crest and a gradient centrifugation can

separate the buffy coat layer, which contains the highest concentration

of HSCs and MSCs.

ASC in BM (HSCs and MSCs)

• Stimulate neovascularization and increase oxygenation (Zhang et al. 2008).

• Transdifferentiate into specific neuronal cells (Zeng et al. 2011).

• Promote synaptic connections (Bareyre et al. 2008).

• Promote neuroplasticity (Kucia et al. 2005).

NO Ethic or immune rejection concrens

Brain Injury

Cox CS, Baumgartner JE Jr, Harting MT et al (2011) Autologous bone marrow mononuclear cell therapy for severe traumatic brain injury in children. Neurosurgery 68:588–600

Tian, C., Wang, X., Wang, X., Wang, L., Wang, X., Wu, S., Wan, Z., 2013. Autologous bone marrow mesenchymal stem cell therapy in the subacute stage of traumatic brain

injury by lumbar puncture. Exp. Clin. Transplant. Off. J. Middle East Soc. Organ Transplant. 11, 176–181. doi:10.6002/ect.2012.0053

(Cox et al. 2005)

BMN MSC- Open label study in adults with sub acute Subacute sever TBI (N=97) 38 of 97 patients (39.2%)

HSC autolog (1 month) TBI; follow up 15 days. 1 × 106 cells/mL (5 ml.) with TBI improved.

Eleven of 24 patients

(45.8%) with persistent

vegetative state showed

post therapeutic

improvements in

consciousness (P = .024)

(Tian et al. 2013)

Spinal Cord Injury

Park DH, Eve DJ, Chung YG, Sanberg PR (2010) Regenerative medicine for neurological disorders. ScientificWorldJ 10:470–489

Sykova E, Homola A, Mazanec R et al (2006) Autologous bone marrow transplantation in patients with subacute and chronic spinal cord injury. Cell Transplant 15:675–687

(Park et al. 2005)

(Sykova et al. 2006)

Spinal Cord Injury

Yoon SH, Shim YS, Park YH et al (2007) Complete spinal cord injury treatment using autologous bone marrow cell transplantation and bone marrow stimulation with

granulocyte macrophage-colony stimulating factor: phase I/II clinical trial. Stem Cells 25:2066–2073.

Geffner LF, Santacruz P, Izurieta M et al (2008) Administration of autologous bone marrow stem cells into spinal cord injury patients via multiple routes is safe and improves

their quality of life: comprehensive case studies. Cell Transplant 17:1277–1293

(Yoon et al. 2007)

(Geffner et al. 2008)

ACUTE VS. CHRONIC ADMINISTRATION

Stroke

Denes, A., Coutts, G., Lénárt, N., Cruickshank, S.M., Pelegrin, P., Skinner, J., Rothwell, N., Allan, S.M., Brough, D., 2015. AIM2 and NLRC4 inflammasomes contribute with ASC

to acute brain injury independently of NLRP3. Proc. Natl. Acad. Sci. 201419090. doi:10.1073/pnas.1419090112.

Friedrich MA, Martins MP, Araujo MD et al (2012) Intra-arterial infusion of autologous bone-marrow mononuclear cells in patients with moderate to severe middle-

cerebral-artery acute ischemic stroke. Cell Transplant 21(Suppl 1):13–21

Savitz SI, Misra V, Kasam M et al (2011) Intravenous autologous bone marrow mononuclear cells for ischemic stroke. Ann Neurol 70:59–69

These discoveries identify the NLRC4 and AIM2 inflammasomes as potential therapeutic targets for stroke

and provide new insights into how the inflammatory response is regulated after an acute injury to the brain. (Denes et al. 2015)

(Friedrich et al. 2008)

(Savitz et al. 2008)

ACUTE TREATMENT PROTOCOL

http://www.tamug.edu/emergency/Emergency%20Procedures/Medical.html

The earl the etter ut if dela ed ot o trai di ated.

POint-of-Care Stem cell Therapy

(POCST)

200 cc of BM

Aktas, M., Radke, T.F., Strauer, B.E., Wernet, P., Kogler, G., 2008. Separation of adult bone marrow mononuclear cells using the automated closed separation

system Sepax. Cytotherapy 10, 203–211. doi:10.1080/14653240701851324.

Cox CS, Baumgartner JE Jr, Harting MT et al (2011) Autologous bone marrow mononuclear cell therapy for severe traumatic brain injury in children.

Neurosurgery 68:588–600

Autologous bone marrow-derived stem cells transplant with on-site cell concentration

for non-hematopoietic diseases.

The o e arrow olle tio i the a ute phase of the brain injury did not cause hemodynamic disturbances and

was well tolerated Co et al. TBI i hildre i the first 8 hours)

Minimal cell manipulation and effective separation

(Aktas et. al. 2008)

After 25 minutes

• IT: 8.5 million CD34+ cells in 12 millilitres.

• IV: 5.6 million CD34+ cells in 8 millilitres.

Matzner, Shaw C., Morrice J., Ruiz-Navarro F., Kobinia G., 2014. CD34+ cell counts and their relationship to clinical factors in 281 patients undergoing

Autologous Stem Cell Transplant, in: World Stem Cell Regenerative Medicine Congress.

Farivar, S., Mohamadzade, Z., Shiari, R., Fahimzad, A., 2015. Neural differentiation of human umbilical cord mesenchymal stem cells by cerebrospinal fluid.

Iran. J. Child Neurol. 9, 87–93.

CSF induces

differentiation of SC

into neurons

(Matzner et al. 2014)

(Farivar et al. 2015)

CD34+ according to the ISHAGE guidelines.

Viability was assessed with 7-Aminoactinomycin D (7-AAD) dye.

Lumbar Puncture?

Bakshi, A., Hunter, C., Swanger, S., Lepore, A., Fischer, I., 2004. Minimally invasive delivery of stem cells for spinal cord injury: advantages of the lumbar puncture technique. J. Neurosurg. Spine

1, 330–337. doi:10.3171/spi.2004.1.3.0330

Bakshi, A., Barshinger, A.L., Swanger, S.A., Madhavani, V., Shumsky, J.S., Neuhuber, B., Fischer, I., 2006. Lumbar puncture delivery of bone marrow stromal cells in spinal cord contusion: a novel

method for minimally invasive cell transplantation. J. Neurotrauma 23, 55–65. doi:10.1089/neu.2006.23.55

The use of LP and interventricular

routes allows more efficient

delivery of cells to the injured cord

compared with the intravenous

route. (Bakshi et al. 2004)

We found that MSC delivered by LP

reached the contused spinal cord tissues

and exerted a significant beneficial

effect by reducing cyst and injury size.

(Bakshi et al. 2006)

Our results in chronic patients

61 patients SCI

follow-up after 6 months

Mean time since diagnosis was 72 months

NO PATIENT PRESENT MAYOR SIDE EFFECTS

62% improved the muscular strength

50% improved the spasticity

37% improved the bladder control

93% of them referred a better general health

status

50% had an improved sensibility

Conclusions

Clinical safety

Reproducibility

Effectiveness

Future

• Imaging Stem cell trackers in humans.

• Protocol of treatment in the first 5 days after presentation.

READY FOR A Randomized

controlled Trial in acute

patients

www.regmedaustria.org

research@regmedaustria.org

Dr. med. Francisco Ruiz-Navarro

PI: Pof. Dr. Med. GEORG KOBINIA